Volume 69 Number 1 Fishery Bulletin U.S. DEPARTMENT OF COMMERCE 69 '^'^ National Oceanic and Atmospheric AdministJ^gpj^g gj^i^gj^g, ^^^^^^^^^^ NatJona! Marine Fisheries Service LIBRA- MAY 171971 WOODS HOLE, .vJASS. Vol. 69, No. 1 January 1971 ROEDEL, PHILIP M. In Memoriam — Wilbert McLeod Chapman and Milner Baily Schaefer . 1 AHLSTROM, ELBERT H. Kinds and abundance of fish larvae in the eastern tropical Pacific, based on collections made on EASTROPAC I 3 SMILES, MICHAEL C, JR., and WILLIAM G. PEARCY. Size structure and growth rate of Euphausia pacifica off the Oregon coast '^9 THOMAS, WILLIAM H., and ROBERT W. OWEN, JR. Estimating phytoplankton production from ammonium and chlorophyll concentrations in nutrient-poor water of the eastern tropical Pacific Ocean 87 CLUTTER, ROBERT I., and GAIL H. THEILACKER. Ecological efficiency of a pelagic mysid shrimp; estimates from growth, energy budget, and mortality studies 93 ROTHSCHILD, BRIAN J., and JAMES W. BALSIGER. A linear-programming solu- tion to salmon management ^^'^ DUBROW, DAVID L., and BRUCE R. STILLINGS. Chemical and nutritional char- acteristics of fish protein concentrate processed from heated whole red hake, Urophycis chitss '■^'■ DUBROW, DAVID L., NORMAN L. BROWN, E. R. PARISER, HARRY MILLER, JR., V. D. SIDWELL, and MARY E. AMBROSE. Eflfect of ice storage on the chemical and nutritive properties of solvent-extracted whole fish — red hake, Urophycis cJuiss 145 CREAR, DAVID, and IRWIN HAYDOCK. Laboratory rearing of the desert pupfish, Cyprinodon inacularius ^^^ HAYDOCK, IRWIN. Gonad maturation and hormone-induced spawning of the gulf croaker, Bairdiella icistia l" ' SECKEL, GUNTER R., and MARION Y. Y. YONG. Harmonic functions for sea-surface temperatures and salinities, Koko Head, Oahu, 1956-69, and sea-surface temperatures, Christmas Island, 1954-69 181 JELLINEK, GISELA, and MAURICE E. STANSBY. Masking undesirable flavors in fish oils 215 COOK, HARRY L., and M. ALICE MURPHY. Early developmental stages of the brown shrimp, Penaeus aztecxts Ives, reared in the laboratory 223 KOURY, BARBARA, JOHN SPINELLI, and DAVE WIEG. Protein autolysis rates at various pH's and temperatures in hake, Merluccius productus, and Pacific herring, Clupea harengus pallasi, and their effect on yield in the preparation of fish protein concentrate 241 Notes EMILIANI, DENNIS A. Equipment for holding and releasing penaeid shrimp during marking experiments 247 TOPP, ROBERT W., and FRANK H. HOFF. An adult bluefin tuna, Thimnus thynnua, from a Florida west coast urban waterway 251 U.S. DEPARTMENT OF COMMERCE Maurice H. Stans, Secretary NATIONAL OCEANIC AND ATMOSPHERIC ADMINISTRATION NATIONAL MARINE FISHERIES SERVICE Philip M. Roedel, Director FISHERY BULLETIN The Fishery Bulletin carries technical reports on investigations in fishery science. The Bulletin of the United States Fish Commission was begun in 1881; it became the Bulletin of the Bureau of Fisheries in 1904 and the Fishery Bulletin of the Fish and Wildlife Service in 1941. Separates were issued as documents through volume 46; the last document was No. 1103. Beginning with volume 47 in 1931 and continuing through volume 62 in 1963, each separate appeared as a numbered bulletin. A new system began in 1963 with volume 63 in which papers are bound together in a single issue of the bulletin instead of being issued individually. Bulletins are distributed free to libraries, research institutions, State agencies, and scientists. Some Bulle- tins are for sale by the Superintendent of Documents, U.S. Government Printing Office, Washington, D.C. 20402. EDITOR Dr. Reuben Lasker Scientific Editor, Fishery Bulletin National Marine Fisheries Service Fishery-Oceanography Center La Jolla, California 92037 Editorial Committee Dr. Elbert H. Ahlstrom National Marine Fisheries Service Dr. William H. Bayliff Inter-American Tropical Tuna Commission Dr. Daniel M. Cohen National Marine Fisheries Service Dr. Howard M. P'eder University of Alaska Mr. .John E. Fitch California Department of Fish and Game Dr. Marvin D. Grosslein National Marine Fisheries Service Dr. J. Frank Hebard National Marine Fisheries Service Dr. John R. Hunter National Marine Fisheries Service Mr. John C. Marr Food and Agriculture Organization of the United Nations Dr. Arthur S. Merrill National Marine Fisheries Service Dr. Virgil J. Norton University of Rhode Island Mr. Alonzo T. Pruter National Marine Fisheries Service Dr. Theodore R. Rice National Marine Fisheries Service Dr. Brian J. Rothschild University of Washington Dr. Oscar E. Sette National Marine Fisheries Service Mr. Maurice E. Stansby National Marine Fisheries Service Dr. Majmard A. Steinberg National Marine Fisheries Service Dr. Roland L. Wigley National Marine Fisheries Service rfi"^ CONTENTS Vol, 69, No. 1 January 1971 ROEDEL, PHILIP M. In Memoriam — Wilbert McLeod Chapman and Milner Baily Schaefer 1 AHLSTROM, ELBERT H. Kinds and abundance of fish larvae in the eastern tropical Pacific, based on collections made on EASTROPAC I ^ SMILES, MICHAEL C, JR., and WILLIAM G. PEARCY. Size structure and growth rate of Euphausia pacifica off the Oregon coast ^^ •THOMAS, WILLIAM H., and ROBERT W. OWEN, JR. Estimating phytoplankton production from ammonium and chlorophyll concentrations in nutrient-poor water of the eastern tropical Pacific Ocean 87 CLUTTER, ROBERT I., and GAIL H. THEILACKER. Ecological eiBciency of a pelagic mysid shrimp; esti- mates from growth, energy budget, and mortality studies 93 ROTHSCHILD, BRIAN J., and JAMES W. BALSIGER. A linear-programming solution to salmon management . 117 DUBROW, DAVID L., and BRUCE R. STILLINGS. Chemical and nutritional characteristics offish protein concentrate processed from heated whole red hake, Urophycis chuss 141 DUBROW, DAVID L., NORMAN L. BROWN, E. R. PARISER, HARRY MILLER, JR., V. D. SIDWELL, and MARY E. AMBROSE. Effect of ice storage on the chemical and nutritive properties of solvent-extracted whole fish — red hake, Urophycis chuss 145 CREAR, DAVID, and IRWIN HAYDOCK. Laboratory rearing of the desert pupfish, Cyprinodon macularius . . 151 HAYDOCK, IRWIN. Gonad maturation and hormone-induced spawning of the gulf croaker, Bairdiella icistia 157 SECKEL, GUNTER R., and MARION Y. Y. YONG. Harmonic functions for sea-surface temperatures and sa- linities, Koko Head, Oahu, 1956-69, and sea-surface temperatures, Christmas Island, 1954-69 181 JELLINEK, GISELA, and MAURICE E. STANSBY. Masking undesirable flavors in fish oils 215 COOK, HARRY L., and M. ALICE MURPHY. Early developmental stages of the brown shrimp, Penaeus aztecus Ives, reared in the laboratory 223 KOURY, BARBARA, JOHN SPINELLI, and DAVE WIEG. Protein autolysis rates at various pH's and temper- atures in hake, Merluccius producUis, and Pacific herring, Clupea harengus pallasi, and their effect on yield in the preparation of fish protein concentrate 241 Notes EMILIANI, DENNIS A. Equipment for holding and releasing penaeid shrimp during marking experiments .... 247 TOPP, ROBERT W., and FRANK H. HOFF. An adult bluefin tuna, Thunnus thynnus, from a Florida west coast urban waterway "^l Wilbert McLeod Chapman Milner Baily Schaefer IN MEMORIAM Wilbert McLeod Chapman and Milner Baily Schaefer '0^: lea / - < Ilu I LIBRARY \':^\ «^. A ^ Fisheries science in particular and society in general suffered two tragic losses within the period of only a month with the deaths of W. M. Chapman on June 25, 1970, and M. B. Schaefer on July 26, 1970. It is indeed a strange and sad commentary that these two men whose careers were intimately entwined since college days should pass within such a short time of each other. Death is never easy to accept; it is particu- larly hard to do so when it occurs at an untimely age. Both of these brilliant men, we would have hoped, would have been with us for years to come. Both were unique, each in his own way, and while the world adjusts to such events, each is in a very real sense irreplaceable. I had the opportunity to work rather closely with both of them in California over most of the past two decades. Wib Chapman was a member of the California Marine Research Com- mittee for many years, during most of which I served as that body's secretary. It was a chal- lenge to try to capture the essence of his re- marks. The breadth of his knowledge and the incisiveness of his thinking stimulated all of us to higher goals, and we who were close to him are better today for the good fortune of his friendship. "Benny" Schaefer was equally brilliant. His expositions on the scientific method and pop- ulation dynamics before the Inter-American Tropical Tuna Commission were models of trans- lation into lay terms of highly complex mathe- matical theories applied to living resources. Again a personal note. A few years ago Benny was a consultant to the California Department of Fish and Game during the formation of that body's Fish and Wildlife Plan and we worked closely in developing the philosophy behind the sections concerned with living marine resources. His imprint is deeply ingrained in that docu- ment and in subsequent legislation, as well as in my thinking. And, as Wib Chapman had a large input in that task, so did Benny into the deliberations of the Marine Research Committee. Meantime both worked diligently as members of the Cal- ifornia Marine Advisory Committee on Marine and Coastal Affairs. While these men will right- fully be remembered for their major contribu- tions to national and international affairs, their energy and interests were such that they en- compassed an amazingly broad spectrum. Each of their contributions to the State of California is more than most men could accomplish in a lifetime devoted to that pursuit alone. One hears parallel stories throughout the scientific and fisheries communities. Wilbert McLeod Chapman was born in Ka- lama, Washington, on March 31, 1910. He died in San Diego, California, on June 25, 1970, and is survived by his wife of 35 years, Mary Eliza- beth, and five of their six children. He did both his undergraduate and graduate work at the University of Washington, obtaining his Ph.D. (fisheries) in 1937. His publications, ranging from morphology and systematic ich- thyology through fisheries economics and inter- national law, number some 250. One of these. Fishing in Troubled Waters, is a book recounting his experiences as a fisheries development oflicer in the South Pacific during World War II. It is fascinating reading and makes one regret all the more that the other books he had in mind will never be forthcoming. He was particularly proud of his papers on systematics and morphol- ogy and always spoke fondly of that part of his career. His honors were many: among them he was a Fellow of the Guggenheim Foundation and of the California Academy of Sciences, .Man of the Year of the National Fisheries Institute in 1966, and the recipient of the First Sea Grant College Award in 1968. He began his professional career in 1933 with the International Fisheries (now Halibut) Com- mission. He was later employed by the Wash- ington State Department of Fisheries, the U.S. Fish and Wildlife Service, and, in 1943, by the California Academy of Sciences where he was Curator of Fishes until 1947. It was during this period that he served in a civilian capacity in the South Pacific, his job being to develop sub- sistence fisheries at advanced island bases. In 1947, Dr. Chapman became director of the School of Fisheries at the University of Wash- ington. He left there in 1948 to become the first Special Assistant to the Under Secretary of State for Fish and Wildlife. In 1951 he be- came Director of Research for the American Tunaboat Association; a decade later he joined the Van Camp Sea Food Company as Director of the Division of Resources. When Van Camp was acquired by the Ralston Purina Company in 1968, he became Director, Marine Resources, of that firm, a position he held until his death. Milner Baily Schaefer was born in Cheyenne, Wyoming, on December 14, 1912. He died in San Diego, California, on July 26, 1970. He is survived by his wife, Isabella, and three children. Dr. Schaefer obtained his B.S. degree cum laude from the University of Washington in 1936 and his doctorate from the same institution in 1950. He worked for the Washington State Department of Fisheries from 1935 to 1938 and for the International Pacific Salmon Fisheries Commission from 1938 until 1942. Following wartime duty with the Navy, he joined the U.S. Fish and Wildlife Service in 1946, serving first as a fishery research biologist in the South Pacific Fisheries Investigations at Stanford, and from 1948 to 1950 as Chief, Research & Development, Pacific Oceanic Fish- ery Investigations in Honolulu. He became Director of Investigations of the Inter-American Tropical Tuna Commission in 1951, holding that post until he became Director of the Institute of Marine Resources and Pro- fessor of Oceanography, Scripps Institution of Oceanography, University of California, in 1962. He remained there until his death save for an 18-month period in 1967-69 during which he was Science Adviser to Secretary of the Interior Stewart Udall. Among other honors, he was a fellow of the California Academy of Sciences and a member of the National Academy of Sciences. He wrote more than 100 scientific papers, particularly in the area of population djTiamics and fisheries development and utilization. He served on a multitude of panels at the international, national and state levels. Despite his huge workload, he always found time to discuss individual problems with people both large and small, and to ad- minister and develop first the Inter-American Tropical Tuna Commission and later the Insti- tute of Marine Resources in an exemplarj' man- ner, setting standards for each that others will be hard-pressed to equal. This recitation cannot give a measure of these men: their unflagging energy, their knowledge in fields far apart from fisheries, their ability as raconteurs, their good fellowship. Nor does it give a measure of their contributions to the nation and to the world, contributions that will help make it a better place in which to live for a long time to come. Philip M. Roedel KINDS AND ABUNDANCE OF FISH LARVAE IN THE EASTERN TROPICAL PACIFIC, BASED ON COLLECTIONS MADE ON EASTROPAC I Elbert H. Ahlstrom' ABSTRACT This paper deals with kinds and counts of fish larvae obtained in 482 oblique plankton hauls taken over an extensive area of the eastern tropical Pacific on EASTROPAC I, a four-vessel cooperative survey made during February-March 1967. On the basis of abundance of larvae, the dominant fish group in oceanic waters are the myctophid lanternfishes (47 %), gonostomatid lightfishes (23 %), hatchetfishes, Stemoptychidae (6 %), bathylagid smelts (5 %). Scombrid larvae ranked fifth, and ex- ceeded 2 % of the count. Two kinds of larvae were outstandingly abundant : larvae of the lantemfish Diogenichthys latematus made up over 25 % of the total, while larvae of the gonostomatid genus Vinciguerria made up almost 20 %. More fish larvae were obtained per haul, on the average, in the eastern tropical Pacific than were obtained per haul in the intensively surveyed waters of the California Current region off Cal- ifornia and Baja California. EASTROPAC I was the first and most wide- ranging of a series of cooperative cruises made in tlie eastern tropical Pacific between February 1967 and April 1968. A vast expanse of the eastern tropical Pacific was surveyed on EAS- TROPAC I, extending from lat 20° N to 20° S, and from the American coasts ofi'shore to long 126° W (Fig. 1). Four research vessels par- ticipated in EASTROPAC I: Alaminos oper- ated by Texas A & M, occupied the inner pat- tern, while Rockaway operated by the U.S. Coast Guard, David Star?- Jordan operated by the Bureau of Commercial Fisheries (now the National Marine Fisheries Service), and Argo operated by the Scripps Institution of Ocean- ography, occupied patterns successively seaward. The oceanographic, biological, and meteorolog- ical data collected on EASTROPAC cruises will be graphically presented in a series of EAS- TROPAC atlases, including generalized charts dealing with fish eggs and larvae. The present paper is the result of a chain of events that began 2 decades ago, at the initiation of CalCOFI (California Cooperative Oceanic Fisheries Investigations) in which a large-scale sea program was set up to investigate the distri- ' National Marine Fisheries Service Fishery-Ocean- ography Center, La JoUa, Calif. 92037. bution and abundance of sardine spawning, and the factors underlying fluctuations in survival of the early life-history stages of sardines. The plankton collections not only contained eggs and larvae of sardine but those of most other pelagic fishes in the California Current region. A de- cision was made to attempt to identify and enu- merate all fish larvae in the collections in order to obtain more precise information about the eco- logical associates of the sardine. At that time few fish larvae, other than those of the sardine and anchovy, could be identified. Within a few years most kinds of fish larvae were identified to genus or species. Once the larvae were identified and enumerated, it be- came obvious that this was an exceptionally use- ful tool for evaluating fish resources. Most oceanic fishes have pelagic eggs and/or larvae that are distributed in or just below the photic zone, i.e. within the upper 150 to 200 m of depth. At no other time in their life histories are so many kinds of fishes associated together — deep- sea fishes (mesopelagic and bathypelagic) as well as epipelagic species — where they can be collected quantitatively with a single type of gear, a plankton net. Once the larvae of the pelagic fish fauna of a region, such as those in the California Cur- rent region, are known, there is a large trans- Manuscript received September 1970. FISHERY BULLETIN: VOL. 69, NO. I, 1971. FISHERY BULLETIN: VOL. 69. NO. I 90° 80° Figure 1. — Location of plankton stations occupied by four research vessels participating in EASTROPAC I. Symbols for vessels indicated in legend above. Samples collected from Argo are numbered as 11.000 series (as 11.022, 11.173), samples from David Starr Jordan as 12.000 series, Rockaway samples as 13.000 series and Alaminos samples as 14.000 series. f erence of the accumulated knowledge and skills for work in other areas, such as, in this in- stance, the eastern tropical Pacific. My study was undertaken to demonstrate the value of identifying all elements of the fish fauna of tropical regions, rather than restricting interest to scombrid larvae. Much information can be gained for little extra expense (a few percent of the cost of collecting the material at sea) . Of equal consequence, identification of all kinds of fish larvae can be made more critically in- cluding scombrid larvae. METHODS OF MAKING ZOOPLANKTON COLLECTIONS Three nets, differing in size and in coarseness of mesh, were employed to collect zooplankton and micronekton on EASTROPAC cruises. In this paper I am concerned primarily with oblique hauls made with the net of intermediate size and mesh — a net, 1-m mouth diameter, con- structed of 505 /J. nylon (Nitex) cloth, with ap- proximately a 5 to 1 ratio of effective straining surface (pore area) to mouth area. This net was paired in an assembly frame with a finer- 4 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC meshed net when hauled obliquely, but was used alone for taking surface hauls. The finer- meshed net was 0.5 m in diameter at the mouth, constructed of 333 /^ Nitex cloth, with approx- imately an 8 to 1 ratio of effective straining surface to mouth area. The third net, used for collecting micronekton, had a 5-ft square mouth opening and was constructed of mesh measuring approximately 5.5 X 2.5 mm; this net could not be operated from the research vessel Rock- away on EASTROPAC I but was employed from the other three vessels. Usually four zooplankton collections were made at each "biological" station: an oblique collection and a surface collection with the 1-m net, an oblique collection with the 0.5-m net, and an oblique collection with the micronekton net. In taking oblique plankton hauls, the 1-m net was paired in an assembly frame with the 0.5 m net. The assembly of nets was fastened to the towing cable by a bridle about 5 m above a 100-lb weight. The assembly was lowered to depth by paying out 300 m of towing cable at the controlled rate of 50 m of wire per minute. The assembly remained at depth for 0.5 min and then was retrieved at a uniform rate of 20 m per min. Total towing time was about 21.5 min. Towing speed was ca. 2 knots. The depth reached by the net was estimated from the angle of stray (departure from the vertical) of the towing cable. We sought to maintain an angle of stray of 45°, which lowered the assembly to a depth of approximately 210 m. Our con- cern was to sample the upper 200-m stratum. The average depths of hauls taken by the four research vessels are summarized in Table 1. Over 80 % of the hauls made on EASTROPAC I were lowered to depths of 200 m or more, and nearly 95 ''r reached depths of 180 m or greater. However, two hauls were exceptionally shallow (71-90 m) , and nine additional hauls were taken to depths of less than 150 m. Usually four paired net-assembly hauls were taken per day, spaced at about 6-hr intervals. Although the four hauls were planned to be taken at about midnight, dawn, noon, and sunset, the timing of hauls was not coordinated between research vessels. The middle-of-the-night hauls Table 1. — Depth of paired oblique plankton hauls taken by the four research vessels on EASTROPAC I. (Net lowered by paying out 300 m of towing cable) Number of houls token to eoch depth interval from Average depth of houl Argo D avid St Jordan arr Rockaway Alaminot All vessels M 70.1. 80.0 __ 1 80.1- 90.0 _, _. _. 1 90.1-100.0 __ __ .. __ __ 100.1-110.0 _^ 1 110.1-120.0 .. __ __ _. ._ 120.1-130.0 2 __ 3 130.1-140.0 1 __ __ 1 140.1-150.0 1 __ _. 150.1-160.0 __ __ 1 2 3 160.1-170.0 2 __ 2 2 6 170.1-130.0 2 2 2 1 7 180.1-190.0 15 5 4 5 29 190.1-200.0 21 10 11 10 52 200.1-210.0 41 59 58 30 188 210.1-220.0 26 44 57 41 168 220.1-230.0 9 _. 3 5 17 230.1-240.0 -' — 1 — 1 Toral 119 121 139 103 482 were all taken before midnight (2201-2400) on Rockaway, for example, while on Argo most hauls, were made after midnight (be- tween 0001 and 0400 hr). The time of day of occupancy of stations (based on the midtime of each haul) is summarized by hourly intervals in Table 2. At least some hauls were taken during every hour of the day, although fewer than 10 (2-8) were obtained during six of the hourly intervals. Fewest hauls were obtained between 0901 and 1000 hr (2 hauls) and be- tween 2101 and 2200 hr (4 hauls), whereas the largest number of hauls were taken between 2201 and 2300 hr (59 hauls) and between 1001 and 1100 hr (53 hauls). Hauls were made with equal frequency during the four periods of the day on Argo, Jordan, and Rockaway; most plankton hauls were taken near midnight or noon from Alaminos. The numbering system for observations em- ployed on EASTROPAC cruises made use of five digits divided into two groups, as 11.022, 12.002, etc. The outer digit preceding the period is the cruise number common to all vessels participat- ing in a given EASTROPAC cruise; for EAS- TROPAC I, this number is 1. The other digit preceding the period is the identifying number given to each research vessel, with the lowest FISHERY BULLETIN: VOL. 69. NO. I Table 2. — Hour of day that paired oblique plankton hauls were taken from the four research vessels par- ticipating in EASTROPAC I. (Midtime of haul used.) Hours of day Number of hauls token during each hour of the day frorr Argo David Starr Jordan Rockaway Alaminos All vessels 0001-0100 7 10 3 20 01 01 -0200 8 7 2 17 0201-0300 5 2 7 0301-0400 9 7 16 0401-0500 1 1 17 1 20 0501-0600 2 9 10 3 24 0601-0700 7 10 1 1 19 0701-0800 13 10 23 0801-0900 7 7 0901-1000 2 2 1001-1100 1 26 26 53 1101-1200 1 5 5 10 21 1201-1300 7 22 3 1 33 1301-1400 12 3 1 4 20 1401-1500 8 a 1501-1600 1 1 12 1 15 1601-1700 10 3 13 1701-1800 8 6 12 6 32 1801-1900 7 19 1 27 1901-2000 10 1 11 2001-2100 3 3 6 2101-2200 1 3 4 2201-2300 2 2 23 32 59 2301-2400 9 11 5 25 Total 119 121 139 103 482 number given to the offshore vessel. The three digits following the period are numbers given to observations made from each vessel during a cruise, numbered sequentially. Not all "stations" included obliqne plankton hauls; hence there are gaps in numbers applied to plankton collections. The locations of plankton stations occupied by the four research vessels participating in EASTROPAC I are showTi in Figure 1. Sam- ples collected from the Argo are designated as the 11.000 series, samples from the David Stan- Jordan as 12.000 series, Rockaway samples as 13.000 series and Alaminos samples as 14.000 series. In tables to follow, the series of samples taken by each vessel is designated by the above identifying series numbers. The aggregate of stations occupied by each vessel is referred to in text discussions as its pattern. PROCESSING SAMPLES ASHORE As noted above, only samples from 1-m oblique net hauls were sorted routinely for fish eggs and larvae. As a rule the entire sample was sorted; in fact only six collections out of 482 were aliquoted — four collections were split into 50 ^r aliquots, two collections into 2.5 '^r aliquots. The author made all identifications and counts of lan'ae from EASTROPAC I collections. Ac- tual counts of larvae rather than standardized values (see below) are used in tabulation throughout this paper, except one (Table 7). There are several reasons why I chose to do this. As indicated previously, all hauls were made in a roughly comparable fashion. In many studies the investigator is interested in the presence or absence of the larvae of a given species or as- semblage of species as such relate to water masses, community composition, time of day, etc. Such information is most readily obtained from records of actual counts. Some statistical tests require the use of original counts rather than standardized data. For persons interested in deriving standardized counts comparable with those employed for CalCOFI data (Ahlstrom, 1953), standard haul factors for the 482 oblique hauls taken with the 1-m net on EASTROPAC I are given in Appendix Table 7. Two major considerations in the quantitative sampling of fish larvae for resources evaluation are (1) how well has their depth range been covered and (2) how effectively have the larvae been sampled within this layer? We do not have direct answers to either of these questions from EASTROPAC cruises. No studies were made on depth distributions of fish eggs and larvae in the EASTROPAC area. As will appear, fewer fish larvae wei'e obtained during daylight hours than in night hauls; how- ever, we lack information on how completely larvae were sampled in night hauls. DEPTH DISTRIBUTION OF FISH LARVAE Although collecting methods used on EAS- TROPAC did not permit a study of depth distri- bution of fish larvae, such information for the California Current region off California and Baja California and in a less detailed way for the NORPAC Expedition of 1955 are available (Ahlstrom, 1959). In the California Current region, most fish eggs and larvae were distributed within the up- AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC per mixed layer or in the upper portion of the thermocline, between the surface and approxi- mately 125 m. Of the 15 most common kinds of fish larvae taken in vertical distribution ser- ies, 12 were so distributed (ibid., p. 134). Two of the kinds that occurred most commonly below the thermocline were bathylagid smelts, closely related to the two common bathylagid smelts taken on EASTROPAC I. On the NORPAC Expedition of August 1955, two depth strata were sampled at most stations ; a closing net, fastened to the towing cable 200 m below a standard open plankton net, sampled the level between 262 and 131 m on the average, while the upper net sampled from the surface to approximately 131 m deep. Only about one- ninth as many larvae were taken in the closing net hauls as in the upper net hauls; fully half of these were larvae of hatchetfish, family Ster- noptychidae, largely absent from upper net hauls. The two most abundant kinds of fish larvae taken on EASTROPAC I were those of the myctophid lanternfish, Diogenichthys kitem- atics, and of the gonostomatid lightfish, Vinci- guerria spp. In NORPAC collections, only 3 % of the larvae of D. laternatus were taken in the closing net hauls and only 2 % of the Vinciguer- ria larvae. Among the kinds of larvae common to both the NORPAC and EASTROPAC sur- veys that occurred in significant numbers in the deeper NORPAC collections were those of Chaul- iodus (72 % taken in closing net hauls), Proto- myctophum (48 %) and I diacanthus (32 %). Inasmuch as the vertical distribution studies in the California Current region had pointed up the importance of the thermocline in the depth distri- bution of larvae, the pattern of thermocline depth was analyzed for EASTROPAC I (Table 3). Thermocline depth was invariably shallow in the inner pattern occupied by Alaminos (data not included in Table 3) ; the greatest depth recorded was only 40 m, and the majority of observations were at depths shallower than 20 m. Along the six station lines covered in Table 3, thermocline depths were shallowest near the equator, and usually were deepest at the north- ern (20-15° N) and southern (15-20° S) ends of the lines. The thermocline also deepened off- shore; approximately three-fourths of the rec- ords of thermocline depths of 50 m or greater were from the tw^o outer lines, occupied by Argo. Most oblique plankton hauls taken on EAS- TROPAC I sampled to depths of 200 m or more (Table 2), hence sampled considerably deeper than the thermocline in all parts of the EAS- TROPAC area. EFFECTIVENESS OF SAMPLING FISH LARVAE IN DAYLIGHT HAULS AS COMPARED WITH NIGHT HAULS Fewer fish larvae were obtained in hauls made during daylight hours than at night (Table 4). Original (unstandardized) counts of larvae av- eraged 2.76 times as many in night hauls as in day hauls, 285 larvae per occupancy as compared with 103 larvae. Hauls made within 1 hr of sunrise or sunset contained intermediate num- bers of larvae, averaging 217 larvae per oc- cupancy. Table 3. — Summary of records of thermocline depths along six station lines occupied by the research vessels Rockaway, David Starr Jordan, and Argo on EASTROPAC I. Station line along longitude Range in depth of thermocline (m) at latitudes 15-10° N 5° N-0° 0-5" S 5-10° S 10-15° S 15-20° S All latitudes 92° W 0-1 S 7-14 5-29 0-16 15-40 24-45 30-54 0-54 98° 16-30 13-68 23-t4 5-13 2-27 13-32 20-48 40-60 2-68 105* „ 37-50 27-14 0-20 0-28 23-45 24-55 54-66 0^56 112° 8-42 41-79 32-58 0-37 2-22 33-52 .. 0-79 119° 3«hS7 44-90 42-55 0-85 0-65 34-76 50-73 30-71 0-90 126° 52-116 45-79 35-49 0^2 0-60 40-71 43-71 43-70 0-116 % obs. with T. D. shallower than 10. 1 m 17 % 8 % 7 % 46 % 43 % 20 % % obs. w T. D. deep thon 49.9 th m 56 % 46 % 9 % 11 % 9 % 25 % 35 % 63 % 26 % FISHERY BULLETIN: VOL. 69, NO. I 3 O 2 Average number per occupancy IP C O 1-0 Average number per occupancy i-_a III 3 o o> z IZ Average number per occupancy (N) II |a° "5 -C D a Average Total number per larvae occupancy (D) 111 3 o^ Z a i > ) n ■* »o rx ■* (N r*v ro 'O *o r^ ^ ul — Ul 03 D CO 1^ ■^ ro w — lO o ^ CM ^ "O r-^ - (M O^ CO <3 *o o. c^ _ _ -^j- O -O CNI — ^ O C^ — C3 r*. lo -v CM *o *o <> lo -d i< CN — d "S o> r^ rv -o wi — rv. (N CM CM O' 'O CM CO — rs. CO -o rCT^'O'O'O'OiM o o -c -K - "> > >- O « n I" 2 = CM to ; f DO p CO o^ "* n '^ CM -^ O* CO <— CO ^ CM CN -O — CM O^ hv CO ~ o- r-* ^ o CM i^ r^ "^ — CO ^ ■O — -. (O -^ IS. O; CM 00 «0 ■^ ^ CS ^ c^ CO — CM CO -fl- ^N CO O CO -^ CM — <0 CN rs 00 "* ■O O- r*^ CM — CM ■V — <>t in 0 •O lO UO O "^ CO -o CM ■v -(J- — rs ■v-^cjcno — o*-* CO CO CO CM CM CM CO ■O UO to ^s CO ^ CO O o _ lO — lO o CM CM -O rv CM O -O CM — — C3 CO -O CO «0 Co' cm" — CO • — CO "^ o- o- o- o — CO CO 00 lO CO . o «* 4) O ■a o o , S.S s = tt) i to * hv CO O* — CM lO c ^O O CM — CO CO c ^O "O CO ^ — ■^ ' lO *0 "O -e CO O; UO CO CO C> O >0 r^ rs CM rsi c*i CM wi XT) — — — ■— CO CO lO O — CN 'O CO o rs. ro CO o IN. — rs ■>}■ — ^ o lOiOCMt^COCMOCS 3 tT (x Tj- o, CM >0 5 CO CO Ov' " K "i CN CO " — --^ hs! ^ CO ** CO CO CO In. 'O 2 *0 CN O^ CM O^ ^O O O UO O* -^ ■«r "1 -O O O ' ■O ^O lO 00 — "■ >0 "O CM CO -^ CO — d N." 'O d — — 00 CO ■^ CM lo (^ lo -o CM rs — — -o "O o lO CO CO o v CM IN. CO ^ 00 p rs CO io — d —■ ■v d CO MD K O In O* rs — ^ 00 00 "O — o 0> O >0 CO CO &• CO CMCMO^|N.-^COIOCO corstocoooco-o ■^^^COCOCM — -^ lO CM CO CO — — CN "tr "O lo o« o* CO ^ CM >0 *0 -O — "O CO CO "* I o*' rv CM lo o* fN o CO ■v o — o- — o o«. o- -^ 0« "O '^ o- o. _ CM "sr CO p p ■^" CO CM CM — CO CS MO CO CO "O *o o 2 -o' o^ (d -^ ^* d O O 0> — fN N. CO rs. o CO o* o o« "O >0 — CO ^ CO CM 00 00 CO — ~ CO -,7? "O o o ! qSOwcoSwO'" to o - <; O o •; • -&S 1 X S o . U) HI S w) < AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Larvae of some families of fishes were sampled almost as well in day hauls as in night hauls — including Sternoptychidae, Bathylagidae, and Melamphaidae. In contrast, less than one- fourth as many gonostomatid larvae and one- third as many myctophid larvae were taken in day hauls, on the average, as in night hauls. Catches of scombrid larvae were more variable with regard to time of sampling — the night-day ratio in the outer half of the EASTROPAC area was only about 1.5 to 1, whereas the ratio jumped to about 7.5 to 1 in the inner pattern occupied by Alaminos. Larvae collected about in equal amounts in day and night hauls were those known to occur principally below the thermocline. Despite the lower abundance of larvae in day hauls as compared with night hauls, the per- centage of hauls containing larvae of most fam- ilies was only slightly lower (Table 5). The most marked day/night difference in frequency of occurrence was for scombrid larvae, these Table 5. — Percentage of hauls containing larvae of the more abundant fish families on EASTROPAC I, grouped by day, night and dawn or sunset. Family Day hauls Night hauls Dawn or sunset hauls (± 1 hr) All hauls % % % % Myctophidae 97.4 97.8 99.0 97.9 Gonostomctidae 92.7 97.3 95.2 95 Sternoptychidae .70.5 76.1 67.6 69.9 Bathylagidae 61.1 65.2 61.9 62.9 Melamphaidae 60.6 65.2 58-1 61.8 Scombridae 31.1 45.1 40.0 38.4 All others 94.8 99.5 97.1 97.1 Total 97.9 100.0 100.0 99.2 were taken in 45 % of night hauls, but in only 31 '/'c of day hauls. In the discussions that fol- low I make use of all collection data, irrespective of time of collection. NUMBERS OF FISH LARVAE OBTAINED ON EASTROPAC I Fish larvae were obtained in 478 of 482 oblique plankton tows made with the 1-m plank- ton net on EASTROPAC I. The number of larvae per collection ranged from to 2,197, averaging 197 larvae (actual counts). Differences in abundance of larvae with lat- itude are summarized for the four series in Table 6. Fish larvae were obtained in largest num- bers, on the average, in an equatorial band ex- tending from about lat 10° N to 5° S. The least productive waters for fish larvae were in the central water mass of the South Pacific, espe- cially between lat 15° and 20° S. Abundance of fish larvae also decreased off- shore,' averaging only 130 larvae per haul in the outer pattern, occupied by Argo, as com- pared with 246 larvae per haul in the inner pattern, occupied by Alaminos. Tropical waters and oceanic waters are usu- ally considered to be relatively unproductive, compared with temperate coastal regions such as the California Current region (Ryther, 1969). Hence, it is surprising to find that the average number of fish larvae obtained per haul on EASTROPAC I was larger than either on the CalCOFI cruises from the California Current region (Ahlstrom, 1969) or on NORPAC (un- Table 6. — Total catches of fish larvae (actual counts) taken by the four research vessels on EASTROPAC I, summarized by latitude. Argo n.OOO Series David S(flrr Jordan 12.000 Series Ro(kaway 13.000 Series Alaminos 14.000 Series Total EASTROPAC 1 Latitude No. hauls No. larvae No. hauls No. larvae No. hauls No. larvae No. No. hauls larvae No. houls No. larvae Average no. larvae per haul 20° N-15° N 16 1,070 20 4,128 5 462 __ __ 41 5,660 138.0 15° N-10° N 14 1,372 23 3,130 26 5,508 -- -- 63 10,010 158.0 10° N- 5° N 14 2,516 14 3,344 29 10,104 15 5,167 72 21,131 293.5 5° N- 0° 14 4,797 15 4,403 14 4,331 27 11,329 70 24,860 355.1 0° 5° S 14 2,089 18 5,454 14 4,350 17 5,042 63 16,935 268.8 5° S-10° S 13 1,370 15 1,051 14 2,360 16 2,113 58 6,894 118.9 10° 3-15° S 14 1,512 8 863 15 2,337 28 1,673 65 6,385 98.2 15° 3-20° S 20 793 8 513 22 1,928 — - 50 3,234 64.7 Total 119 15,519 121 22,886 139 31,380 103 25,324 482 95,109 197,3 FISHERY BULLETIN: VOL. 69, NO. 1 published data) . Standard haul totals of larvae are used in this comparison (Table 7) not ori- ginal counts. CalCOFI cruises repeatedly sur- veyed a coastal area extending 200 to 300 miles offshore between San Francisco, California, and Magdalena Bay, Baja California. NORPAC was the first comprehensive survey of the North Pacific, made in August-September 1955; the area surveyed by four CalCOFI vessels partici- pating in NORPAC was between lat 20° and 45° N and offshore to long 150° W. Table 7. — Comparison of the average number of fish larvae obtained per haul (standard haul values) EAS- TROPAC I, NORPAC, and CalCOFI cruises. Number hauls Averoge Total depth number of of hauls fish larvae' Average number larvae/haul EASTROPAC 1 1967 482 CO. 200 m 274,131 569 NORPAC 1955 196 CO. 260 m 27,000 "138 CalCOFI cruises 1956 1,407 CO. 140 m 408,140 290 1957 1,493 CO. 140 m 493,550 331 1958 1,852 ca. 140 m 456,020 246 1959 2,182 CO. 140 m 470,450 216 1960 1,826 CO. 140 m 504,980 277 ^ Standard houl totals. 2 Data from two net hauls combined: on overage of 124 larvae per haul were token in upper net hauls (0 to 130 m) and an average of 14 larvae per haul in closing net hauls,, sampling between co. 260 and 130 m. EASTROPAC hauls sampled a somewhat deeper stratum than hauls made on CalCOFI cruises, ca. 200 m as compared to ca. 140 m. As indicated previously, information is available for the majority of NORPAC stations on the rel- ative abundance of fish larvae in the level be- tween ca. 130 and 260 m (closing net hauls) as compai'ed with the level above, to 130 m. Only about one-ninth as many larvae were taken in the deeper hauls. The difference between catches of larvae on EASTROPAC I and NORPAC are particularly marked — four times as many larvae were taken per haul, on the average, on EASTROPAC I as on NORPAC (both nets combined). For com- parison with shallower CalCOFI hauls, I am as- suming that 10 % of the EASTROPAC larvae were obtained in the level between ca. 140 and 200 m. The adjusted value for EASTROPAC larvae, 512 larvae per haul, on the average, is 1.55 times as large as the highest CalCOFI val- ue listed (331 larvae per haul in 1957) and 2.35 times as large as the lowest value (216 larvae per haul in 1959) . The majority of EASTROPAC larvae were those of fishes which never attain a large size as adults — myctophids, gonostomatids, sternopty- chids, etc. — hence numbers of larvae, per se, cannot be considered reliable indices of biomass. The familial composition of larvae was not dis- similar on NORPAC and EASTROPAC, how- ever; hence this comparison of relative abun- dance of larvae is more relevant, as regards biomass, than the comparison with CalCOFI fauna. KINDS OF FISH LARVAE OBTAINED ON EASTROPAC I The kinds of larvae obtained on EASTRO- PAC I are summarized by family and vessel pattern in Table 8, the principal summary table in this paper. Larvae of more than 50 families are listed, but larvae of 10 families contributed 90 9f of the total. The myctophids were the dominant group with 47.2 % of the larvae oc- curring in nearly 98 % of the collections. Gono- stomatid lai-vae were about half as numerous, contributing 23.2 % of the larvae while oc- curring in 95 % of the collections. Hatchetfish larvae (Sternoptychidae) ranked third in abundance with 6 % of the larvae taken in 70 % of the hauls. Bathylagid larvae also exceeded 5 % of the total and occurred in 63 % of the collections. Scombrid larvae ranked fifth and exceeded 2 % of the count, followed by Breg- macerotidae, 1.9 %, Paralepididae, 1.7 %, Idia- canthidae, 1.0 Yc, Nomeidae, 1.0 %, and Mel- amphaidae, 0.9 %. About one-third of the re- maining larvae were too poorly preserved (dis- integrated) to identity. On the basis of larval abundance, the domi- nant orders of fishes in oceanic waters are the Myctophiformes and Salmoniformes, making up between 85 and 88 'li ; the latter value assumes a proportionate representation of larvae of these groups in the "disintegrated" category, i.e., larvae too damaged or disintegrated to identify with certainty. Despite the dominance of fishes of the above two orders, a number of other groups of fishes are represented in the oceanic pelagic fish fauna. The berycoid fishes are rep- 10 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC i < OJ M ^ 1 ri U e > 01 1 ' •a = « ■SSs ^ 1 3 M C 41 CO *" ' z i 3 3 5) J) 5 5 >. E D u COOOOCO^OO — "COO"^ — ^cow — — CN eqo-o — O^'O O^-O CO — CO CO — CIO CN CMcocTO — ooooo* — O — dC9CNeoN.cn — o-"^rvO« — 1 — eo ■V — to-^ -ow — — sS*o c*i Ovooco"^ •— — — Mi O. — C4 — COCN^O — ^ — iifOlOr) — OO'OO— NO'CM-* — coo O N"*OCO -O >0 CO ■OOO'OCN rv VCN fcO'O'ffrO-* lOCOCN*OC*)- - coV CN — lOOOUlO — O — OCN"0 CO — N OJCN — CN COCM CO f^ O CO— CNVl — CO -^ jiOO'^CO- ■^O'V- -COCNCtTj-O-OCNO-OO -M'^OCMCN'^'O'OkO- O^cO'O'^COCNOCN- O'OOO'O^COCNOO^OOOCNOOCO'OIV^O-O — O-OCOiOOOOO^^-iOrON. loo-co- NO'to rvio o . cm -o — cn'^ .© — — cnco— ■* — — — iocnco ■^fs^rv CO— cMuo — — CO n co co tta3"*00'000>ONOO — coKcoOcN — o— '^rooOOOco'O^'Orv OJ uo CO^OO oocM'^rcoco-orNNrsOOcocNO'^c — CO-OCN CO"*!-^ O'^CNO CN-OO ~ OCN -O-OCOOOOOCO — oo -00100000CON>0 -vn— CNiO -oo— oo^ "S" "^■^ — — - CNCMCMeNeNCNCNCSCNO4C0C0C0nC0C0COC0C0CT^"0>OioiOiO«0 11 FISHERY BULLETIN: VOL. 69, NO. I resented by Melamphaidae, a family of fishes that is almost as ubiquitous as the myctophids or gonostomatids. Fishes of the gadoid family, Bregmacerotidae, also are widely distributed in the warmer waters of all oceans. Among the ubiquitous epipelagics are the flyingfishes, Ex- ocoetidae. Only a moderate number of perciform fishes are widely distributed in offshore, oceanic wa- ters. Among the more important are fishes of the families Scombridae, Gempylidae, Trichiur- idae, Istiophoridae, Coryphaenidae, Bramidae, Nomeidae, Apogonidae, Chiasmodontidae, and Tetragonuridae. Larvae of some demersal fishes have a much wider offshore distribution than one would asso- ciate with the known distribution of adults. In- cluded in this group are larvae of bothid and cynoglossid flatfishes, and larvae of Scorpaeni- dae, Gobiidae, and Labridae. Another widely distributed gi-oup in oceanic waters are the bizarre ceratioid fishes. The rotund larvae of these fishes were taken in about 30 % of the EASTROPAC collections, always in small numbers. The basic data on the kinds and numbers of fish larvae obtained in the 482 EASTROPAC I collections are contained in six appendix tables, whose contents are summarized below, and keyed to Table 8 and to other tables in this report. Appendix Table 1. — Counts of fish larvae, tabulated by family, for all stations occupied on EASTROPAC I. This table contains 22 categories, mostly families, but for complete- ness, a category is included for "other identi- fied larvae," one for "unidentified larvae" and one for "disintegrated larvae" (i.e., larvae too damaged or disintegrated to identify with any certainty) . Appendix Table 2. — Myctophid larvae, tab- ulated by genus or species, for all stations oc- cupied on EASTROPAC I. Myctophid larvae are tabulated by species for 12 kinds, and by genus for 8 kinds. Also included are cate- gories for unidentified myctophids, and total myctophids. A summary of this appendix table is contained in Table 15. Appendix Table 3. — Counts of selected ca- tegories of fish larvae by station. Table con- tains 23 categories including 10 species, 10 genera, 2 families, and 1 suborder; 9 of these were included in the category "other identi- fied larvae" in Appendix Table 1. Appendix Table 4. — Summary of occur- rences and numbers of larvae of eight families limited in distribution to a broad coastal band or around offshore islands. Only positive stations are included. These eight families also were included in the category " other identified larvae" in Appendix Table 1. Appendix Table 5. — Numbers and kinds of larvae of Gempylidae-Trichiuridae obtained in EASTROPAC I collections. Only positive stations are included. A summary of this ap- pendix table is given in Table 19. Appendix Table 6. — Numbers and kinds of flatfish (Pleuronectiformes) larvae obtained in EASTROPAC I collections. Only positive hauls are included. A summary of this ap- pendix table is given in Table 22. Appendix Table 7.— Standardized haul factors for the 482 oblique 1-m net hauls taken on EASTROPAC I. These factors adjust ori- ginal counts of larvae to the comparable stan- dard of numbers of larvae in 10 m3 of water strained per meter of depth fished. I will not attempt to comment on all 58 cate- gories (family or larger grouping) summarized in Table 8, but will limit my discussion to 31 of these. In order to tie the text discussion closely to this table, I i-etain the numbers for categories as given in Table 8; those discussed in the text ai-e preceded by an asterisk in this table. COMMENTS ON LARVAE OF THE MAJOR FISH FAMILIES COLLECTED ON EASTROPAC I 1. CLUPEIDAE ( 10 occurrences, 81 larvae) Three species of clupeid larvae were taken in EASTROPAC I collections — Opisthonema sp. 12 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC (5 occurrences, 12 larvae), Etrumeus acumina- tus Gilbert (2 occurrences, 6 larvae), and Sar- dinops sagax (Jenyns) (3 occurrences, 63 larvae). The latter two species were collected in the vicinity of the Galapagos Islands. 2. ENGRAULIDAE (10 occurrences, 205 larvae) The majority of the engraulids (5 occurrences, 174 specimens) were those of the Peruvian an- chovy, Engraulis ringens Jenyns, collected at coastal stations between lat 6° and 13.5° S. Al- though larvae from only a few surface hauls have been sorted as yet, one haul was outstand- ing: the surface tow taken at station 14.069 contained 10,466 larvae and transforming speci- mens of Peruvian anchovy, E. ringens. Speci- mens ranged in size from 3.5 to 37.5 mm ; most were between 4.0 and 7.5 mm in length but even transforming specimens, 20.0 to 37.5 mm long, were rather common (83 individuals). In the oblique 1-m haul at this station, 97 anchovy larvae were obtained. 3. ARGENTINIDAE (43 occurrences, 87 larvae) Three kinds of argentinid larvae were ob- tained: Argentina sp. (1 specimen), Nansenia sp. A (84 lai-vae), and Nansetiia sp. B. (2 larvae) . The specific identities of the two kinds of Nansenia larvae are still uncertain. On EASTROPAC I, Nansenia sp. A was taken most commonly in an equatorial band between lat 5° N and 5° S (Fig. 2). Larvae of Nansenia sp. A also occur in the southern portion of the area surveyed on cruises of CalCOFI, particularly to the south of Point San Eugenio, Baja California. A Nansenia larva with markedly different pig- mentation pattern was obtained at station 11.154 in the central water mass of the South Pacific. A similarly pigmented Nanseyiia larva was ob- tained on NORPAC from the central water mass of the North Pacific. 4. BATHYLAGIDAE ( 304 occurrences, 4,880 larvae) Although two kinds of Bathylagus larvae were obtained, one species was taken in only two con- tiguous southern stations, 12.142 and 12.144. The eyes of the latter were carried on short stalks. The distribution of larvae of the com- monly occurring species, B. nigrigenys Parr (296 occurrences, 2,987 larvae), was almost identical with that of the myctophid, Diogenich- thys laternatus (Garman) (Fig. 3). The larvae of neither species occurred in the central South Pacific water mass; on the four outer lines, sur- veyed by Argo and Jordan, the occurrences of B. nigrigenys larvae ended at about lat 5° S. In the portion of the EASTROPAC area in which larvae of this species were distributed, they occurred in three-fourths of the stations occupied. In the innermost pattern occupied by Alami- nos, larvae of Leuroglossus stilbius urotranus (Bussing, 1965) were common (37 occurrences, 1,890 larvae). All but four specimens were obtained between lat 10° N and 10° S, and most within 300 miles of the coast (Fig. 2). 5. GONOSTOMATIDAE (459 occurrences, 22,046 larvae) Areal occurrence and relative abundance of gonostomatid larvae on EASTROPAC I are summarized in Table 9. They were obtained in 95 % of the hauls and made up approximately 23.2 % of the larvae. As noted earlier, gonostomatid larvae were markedly more abundant in night hauls than in day hauls: 4.35 times as many, on the aver- age. In contrast, larvae of the closely related hatchetfishes, Sternoptychidae, were taken in only slightly larger numbers at night (1.24 times as many as in day hauls). In the section dealing with depth distribution of fish larvae it was pointed out that the gonostomatid, Vinci- guerria spp. occurred no deeper than ca. 130 m in NORPAC collections, whereas sternoptychid larvae were inhabitants of the aphotic zone be- low 130 m. An interesting exception should be noted: gonostomatid larvae of the subfamily Maurolicinae had depth distributions similar to sternoptychid larvae on NORPAC. Larvae of two Maurolicinae, Mauroliciis and Araiophos, genera were taken on EASTROPAC. Although the depth distribution of these genera has not 13 FISHERY BULLETIN: VOL. 69, NO. I 90° 80" Figure 2. — Distribution of larvae of the argentinid, Nansenia spp., and of the bathylagid, Letiroglossiis stilbius urotranus (Bussing) on KASTROPAC I. Records of occurrence of A'awscnto larvae are shown as open circles with dot in center, while those of Leuroglossus larvae are open squares with dot (1 to 100 larvae) or closed squares (101 to 490 larvae). Small solid circles represent other stations occupied on EASTROPAC I. Table 9. — Areal occurrence and relative abundance of lari'ae of Gonostomatidae on EASTROPAC I. Argo David Starr Jordan Rod away jilaminoj Total 11.000 series 12.000 series 13.00C series 14.000 series EASTROPAC 1 Lalilude No. No. No. No. No. No. No. No. No. No. Average no. pOSitiVQ positivo positive positive positive larvae per positive haul hauls larvae hauls larvae hauls larvae hauls larvae hauls larvae 20° H\S° N 14 418 20 1,534 5 115 .. 41 2,067 50.4 15° N-10° N 14 380 22 745 24 607 __ 60 1,732 28.9 10° N- 5° N 13 185 13 242 27 2.085 14 417 67 2,929 43.7 5° N- 0° 14 2,112 IS 637 14 1,825 27 1,882 70 6,456 92.2 0° - 5° S 14 409 18 912 14 1,577 16 1,036 62 3,934 635 5° S-I0° S 13 202 14 161 14 799 10 647 51 1,809 35.5 )0° S -IS- S 14 635 8 368 IS 524 21 490 58 2,017 34.8 IS" S-20° S 20 322 8 183 22 597 — — SO 1.102 22.0 Totol lis 4,663 118 4,782 135 8,129 88 4,472 459 22,046 48.0 14 AHLSTROM : FISH LARVAE IN EASTERN TROPICAL PACIFIC 130* 120° 110° 100° 90° 80° Figure 3. — Distribution of larvae of Bathylagus nigrigenys Parr on EASTROPAC I. Two orders of abundance are shown: open circles with dot in center represent counts of 1 to 25 larvae, large solid circles represent counts of 26 or more larvae. Small solid circles represent negative hauls. been determined, they were sampled more fully during daylight hours than other gonostomatids; the night/day ratio for Maurolmis and Arai- ophos larvae was ca. 1.6 and 2.0 respectively. Larvae belonging to six gonostomatid genera were common to abundant (Table 10) and larvae of several additional genera were taken occasionally. Larvae of two genera were of outstanding importance in the EASTROPAC area — Vinciguerria and Cyclothone. Vinciguer- ria occurred in 87.5 % of the collections, Cyclo- thone in 62.4 %. Charts showing the distribution and relative abundance of larvae of Gonostomatidae and Sternoptychidae (combined) on EASTROPAC I will be included in the EASTROPAC Atlas. Araiophos eastropas Ahlstrom and Moser ( 18 occurrences, 529 larvae) Larvae of A raiophos eastropas were obtained only on the outermost pattern to the south of lat 10° S (Fig. 4). Within this limited area it was the most common gonostomatid. The spe- cies taken on EASTROPAC represented an un- described species in a genus that previously 15 FISHERY BULLETIN: VOL. 69. NO. 1 Table 10. — Frequency of occurrence and relative abundance of the kinds of gonostomatid larvae on EASTROPAC I. Argo DavU St arr Jordan Rork away AlaminoJ Total 11.000 series 12.000 series 13.00C series I4.00C series EASTROPAC 1 Gonostomatid larvae No. No. No. No. No. No. No. No. No. No. positive positive positive positive positive hauls larvae hauls larvae houls larvae hauls larvae hauls larvae liraiaphos eastropaj 18 529 18 529 Cydothone spp. 94 697 71 582 89 735 47 167 301 2,181 Diplopkos taenia IS 51 40 107 14 24 1 I 73 183 Ichthyococcu! spp. 7 9 11 16 18 31 5 5 41 61 Maurolicui muelleri 11 43 19 143 13 78 43 264 VincigutTria spp. 96 3,339 109 4,011 131 7,179 86 4,211 422 18,740 Other gonostomotids 13 38 9 23 12 17 8 10 42 88 Total IIS 4,663 118 4,782 135 8,129 88 4,472 459 22,046 FiGUKE 4. — Distribution of larvae of three species of Gonostomatidae on EASTROPAC I. Records of occurrence of larvae of Araiophos eastropas Ahlstrom and Moser are shown as triangles, Diplophos taenia (Giinther) as large open circles, and Maurolicus muelleri (Gmelin) as squares. Solid triangles and squares are for counts of 26 or more larvae. Small solid circles represent negative hauls. 16 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC was known from a single collection made off Hawaii (Grey, 1961). Adults and larvae were described by Ahlstrom and Moser (1969). Cyclothone spp. (301 occurrences, 2,181 larvae) Larvae of Cyclothone spp. were taken least frequently in the northern quarter of the EAS- TROPAC pattern (betweeen lat 10° and 20° N, and in the inner pattern occupied by Alaminos (Table 11 and Fig. 5). In the former area, less than 20 Sf of the hauls (20 of 103) con- tained Cyclothone larvae; in the inshore pat- tern only about 45 % of the hauls (47 of 103) contained Cyclothone larvae. Over the remain- der of the EASTROPAC I pattern Cyclothone larvae occurred at most stations (234 of 276). The lowest number of larvae per positive haul, 2.15 larvae, was obtained in the northern sec- tion; the next lowest, 3.55 larvae per positive haul, in the Alaminos pattern. Over the re- mainder of the pattern, 8.42 larvae were ob- tained per positive haul. No attempt was made to identify the larvae of Cyclothone to species, and our hauls did not extend deep enough to collect adults. Diplophos taenia Giinther (73 occurrences, 183 larvae ) A study was made of larval and adult speci- mens of Diplophos in an attempt to determine whether the Pacific specimens should be as- signed to D. taenia or retained as a distinct species, D. pacificus Giinther. Grey (1960) had placed Pacific specimens in D. taenia but later she (Grey, 1964, p. 89) developed reservations because of the consistently lower photophore count of the ventral series in Pacific specimens. Without detailing my observations on Diplophos, which I plan to publish separately, I have con- cluded that our eastern Pacific Diplophos is not separable from the Atlantic D. taenia. Larvae of Diplophos were taken most com- monly to the north of lat 10° N — 36 occurrences, 105 larvae (Fig. 4). The remaining 37 occur- rences, 78 larvae were distributed throughout the EASTROPAC I pattern. Ichthyococcus spp. (41 occurrences, 61 larvae) Two kinds of Ichthyococcus larvae were taken on EASTROPAC L The specific identity of the more common form has been determined as /. irregularis Rechnitzer and Bohlke; the other form has yet to be identified to species. Maurolicus muelleri (Gmelin) (43 occurrences, 264 larvae ) Larvae of this species were taken only on an equatorial band between lat 5° N and 5° S and were not taken in the outer pattern occupied by Argo (Fig. 4). This distribution, without additional information, could be misleading. Maurolicus is known to have a wide latitudinal distribution in the South Pacific. For example, Maurolicus larvae were obtained at lat 33° S on MARCHILE VL the portion of EASTRO- PAC II occupied by the Chilean vessel Yelcho. We also have collections from south of New Zealand, obtained on an Eltanin cruise. The species may be carried northward oflF South America in the Humboldt Current and then off- shore in the equatorial current system. Table 11.— Area occurrence and relative abundance of larvae of Cyclothone spp. on EASTROPAC I. Argo 1 1 .000 series David Starr Jordan 12.000 series Rockaway 13.000 series Alaminos 14.000 series Total EASTROPAC 1 Latitude No. positive hauls No. larvae No. positive hauls No. larvae No. positive hauls No. larvae No. positive hauls No. larvae No. positive hauls No. larvae Average no. larvae per positive haul 20° N-10° N 12 31 4 8 4 4 20 43 2.2 10° N- 0° 24 136 25 137 33 235 23 69 105 577 5.5 0° -10° S 24 179 29 246 20 117 13 43 86 585 6.8 10° S-20° S 34 351 13 191 32 379 11 55 90 976 10.8 Total 94 697 71 582 89 735 47 167 301 2,181 7.2 17 130° T — \ — I — I — r -| — I — I — I — I — I — I — I — r 100" -I — I — \ — TTT — I — I — I — I — I — r— T — I— T — I — I — I — 1—1 — I — I — I — I — I — I I I — r FISHERY BULLETIN: VOL. 69, NO. 1 90° 80° 20' 10' 10" ® e @ © © VM4NZANILL0 20" s Q®®® ® 1^ A 3® © 0001 I L_ I I I J 1—1—1 I I I I I I 130° 120" no* 100* 90* 80" Figure 5. — Distribution of larvae of the gonostomatid Cyclothone spp. on EASTROPAC I. Collections of 1 to 25 larvae are shown as circles with dot in center, collections of 26 or more larvae as large solid circles; neg- ative hauls are shown as small solid circles. Vinciguerria spp. (422 occurrences, 18,740 larvae ) Larvae of Vinciguerria occurred in more hauls than those of any other genus and ranked sec- ond in abundance to the myctophid genus Dio- genichthys. The distribution of Vinciguerria larvae is shown in Figure 6. Although most of the material unquestionably is V. bicetia (Garman) , some of the collections from offshore and particularly from the central South Pacific water mass between lat 5° and 20° S represent V. nimbaria (Jordan and Williams) . The larvae of V. nimbaria are indistinguishable from those of V. lucetia (Ahlstrom and Counts, 1958), hence identification must be made on meta- morphosing specimens, juveniles, and adults. The two species are closely allied, but readily separable from V. poweriae (Cocco) and V. attenuata (Cocco), the other two species of Vinciguerria, at all stages of development. A trenchant difference between the two "pairs" of species is the development of a pair of sym- physeal photophores under the lower jaw in V. lucetia and V. nimbaria and the absence of this pair in V. poweriae and V. attenuata. The two characters most readily used for distinguishing 18 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC 130' 120° MO" Figure 6. — Distribution of larvae of the gonostomatid, Vindguerria spp. on EASTROPAC I. Collections of 1 to 100 larvae are shown as circles with dot in center, collections of 101 or more larvae as large solid circles; negative hauls are shown as small solid circles. between V. lucetia and V. nimbaria are (1) number of gill rakers and (2) number of IV (and OV) photophores. Material of V. nim- baria studied from the eastern North Pacific (ibid.) had 5 to 6 +15 gill rakers and 23 to 24 IV photophores (13 to 14 OV photophores) whereas V. lucetia had 8 to 10 + 18 to 23 gill rakers and 20 to 23 IV photophores (10 to 13 OV photophores) . In the EASTROPAC area, V. lucetia maintained the high gill raker counts, but usually had 21 IV (11 OV) photophores. The offshore form referred to V. nimbaria usu- ally had 22 IV (12 OV) photophores (1 less per group than in V. nimbaria from the temper- ate North Pacific) and 6 to 7 + 15 to 16 gill rakers (a slightly higher count). In most areas the adults of the two species of Vindguerria did not co-occur, hence the larvae can be assigned with some assurance to one or the other. For example, all collections made between lat 5° and 20° S from Argo and Jordan patterns were exclusively V. nimbaria. On these patterns the plankton hauls were sup- plemented by micronekton net hauls, and the latter contained material of Vindguerria ju- veniles and adults from most stations occupied 19 FISHERY BULLETIN: VOL. 69. NO. I at night. Unfortunately, the micronekton net was not used on Rockaway (12.000 series), and insufficient numbers of older stages (metamor- phosing specimens and juveniles) were taken in plankton hauls to permit a meaningful separa- tion of the two species in waters to the south of lat 5° S in this series. Vinciguerria poweriae (Cocco) co-occurred with V. nimbaria in the central water mass of the North Pacific (Ahlstrom and Counts, 1958), but no material of V. poweriae was obtained in EASTROPAC collections. However, material of V. attenuata (Cocco) was obtained from farther south in the eastern Pacific on the "Downwind Expedition" — hence all four spe- cies of Vinciguerria do occur in the eastern Pacific. Other gonostomatids (42 occurrences, 88 larvae) Included in this category are larvae of two identified genera, Gonostoma and Woodsia, and several kinds of larvae that are unmistakably gonostomatid, but not identified as to kind. 6. STERNOPTYCHIDAE (337 occurrences, 5,687 larvae) Hatchetfish larvae ranked third in abundance (5.98 /f of total), exceeded by larvae of Mycto- phidae and Gonostomatidae. The majority of hatchetfish larvae were those of Sternoptyx di- aphana Hermann, and most of the remainder of Argyropelecus lychmis Carman. Because larvae of Sternoptychidae are more fragile than most other kinds and are usually in poor condition, no attempt was made to identify them to genus or species. Areal occurrence and relative abun- dance of sternoptychid larvae on EASTROPAC I are summarized in Table 12. Larvae were not only taken in markedly more collections between lat 10° N and 10° S— 94 9^ of the collections were positive as compared with only 41 % in the remainder of the pattern — but more larvae were taken per positive haul — 21.1 larvae as compared with 5.2. 7. ASTRONESTHIDAE (12 occurrences, 13 larvae) Several kinds of astronesthid larvae were collected in the EASTROPAC area: only one kind had heavy pigmentation on the body; the others were lightly, but characteristically pig- mented. Larvae of Astronesthidae are similar in appearance to other stomiatoid larvae; they have a slender, elongated body, and a long in- testine that underlies the body for about Yiq or more of the standard length, and usually has a free terminal, trailing portion that can be quite long, often trailing beyond the caudal fin. As- tronesthid larvae can be distinguished readily from other stomiatoid larvae by the forward po- sition of the dorsal fin in relation to the anal fin. Developmental series of astronesthid larvae have not been described in literature. Eleven of the 12 occurrences of astronesthid larvae were taken within 10° ± of the equator. 8. CHAULIODONTIDAE (80 occurrences, 165 larvae j Larvae of Chaidiodus are readily identifiable to genus, but are difficult to separate at the spe- Table 12. — Areal occurrence and relative abundance of larvae of Sternoptychidae on EASTROPAC I. Areo 1 1 .000 series David St 12.000 arr Jordan series Ro." Syacium has a distinctive larva with heavy opercular spination, a sphenotic spine on either side of 41 FISHERY BULLETIN: VOL. 69, NO. I the head, and 5 to 8 elongated anterior dorsal rays. Larvae of the closely related genus, Cy- clopsetta, also develop opercular and head spina- tion. The opercular spination Is more pro- nounced in Syachim — particularly an antlerlike spine that develops on the posterior border of the preoperculum. The three anterior rays of the left pelvic fin become only moderately elon- gated in Syacium larvae; the rays are of about equal length, firmly joined together by a mem- brane, and pigmented distally. The full com- plement of dorsal and anal fin rays usually are laid down before the larvae attain a standard length of 10 mm; the largest specimens studied, ca. 20 mm long, were undergoing metamor- phosis. Citharichthys-Etropus (26 occurrences, 50 larvae) Before discussing problems in identification of Citharichthys-Etropus larvae from the EAS- TROPAC area, some background information will be given on Citharichthys larvae in the Cal- COFI region. Illustrations of larvae of three spe- cies of Citharichthys were given in Ahlstrom (1965). Two species, Citharichthys sordidus (Girard) and C. xanthostigma Gilbert, develop 2 elongated dorsal rays and also 2 elongated vent- ral rays on larvae larger than about 5 mm ; the other species never develops such rays. Another species that occurs off central and southern Baja California, C. fragilis Gilbert, also develops 2 elongated rays on the dorsal and ventral fins. Two species of Citharichthys, C. gilberti Jenkins and Evermann, and C. platophrys Gil- bert, and the widely distributed Etropus cros- sotus Jordan and Gilbert are known to occur in the EASTROPAC area. Three kinds of larvae were taken in EASTROPAC collections refer- able to Citharichthys or Etropus. The most common kind developed 3 elongated dorsal rays, a less common form developed 2 elongated dorsal rays, and some specimens lacked elongated rays. The form with 3 elongated dorsal rays is almost certainly referable to Citharichthys. Larvae of a common Atlantic species, C. arctifrons Goode, develop 3 elongated dorsal rays, confirming the presence of this combination in Citharichthys larvae. A cleared and stained specimen from station 13.040 with 3 elongated dorsal rays pos- sessed 10 + 25 vertebrae, 78 dorsal rays, and 59 anal rays. The meristics of the dorsal and anal fins could fit either C. platophrys or C. gil- berti. Yet so little is known of C. platophrys that I would hesitate to refer the common Citharichthys larvae in EASTROPAC material to this species. A similar problem attends larvae of the form that lacks elongated dorsal rays. Two specimens, 11.5 and 12.0 mm, from station 14.014 each had 88 dorsal and 67 anal rays; vertebrae counts were 10 + 23 and 10 + 24. These counts best fit E. crossotus, except that the vertebral counts are low. No material of the form with 2 dorsal rays (undoubtedly a Citharichthys) has been cleared and stained for precise meristics. A definite identification has yet to be made on all three kinds of larvae. 55. CYNOGLOSSIDAE (63 occurrences, 304 larvae) Only one cynoglossid genus, Symphurus, oc- curs in the eastern Pacific. Five or more kinds of Symphurus larvae were obtained in EAS- TROPAC collections; these were obtained in more collections than larvae of bothid flatfishes (63 as compared with 56) , and made up a larger percentage of the total flatfish larvae (ca. 60 '^,'r ) . A moderate number of recently transformed specimens of Symphurus were obtained in EASTROPAC collections; in contrast, all spe- cimens of bothid flatfish were pretransformation larvae. The distribution of Symphurus larvae in EASTROPAC I is shown in Figure 13. ACKNOWLEDGMENTS I am indebted to a number of persons for as- sistance during the preparation of this manu- script. Kenneth Raymond prepared the distri- bution charts. Amelia Gomes helped in many facets of the work including the preparation of cleared and stained specimens of flatfishes and other groups. H. Geoff"rey Moser has worked closely in studies of larvae of Myctophidae and Gonostomatidae. W. L. Klawe has been help- 42 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC ful in many ways; he graciously has permitted me to include station information on occurrence and numbers of larvae of Auxis sp. and skip- jack tuna. I wish particularly to thank David Kramer and H. Geoffrey Moser for reviewing the manuscript. LITERATURE CITED AHLSTROM, Elbert H. 1953. Pilchard eggs and larvae and other fish larvae, Pacific Coast - 1951. U.S. Fish Wildl. Serv., Spec. Sci. Rep. Fish. 102. 55 p. 1959. Vertical distribution of pelagic fish eggs and larvae off California and Baja California. U.S. Fish Wildl. Serv., Fish. Bull. 60: 107-146. 1965. Kinds and abundance of fishes in the Cal- ifornia Current region based on egg and larval surveys. Calif. Coop. Oceanic Fish. Invest. Rep. 10: 31-52. 1969. Mesopelagic and bathypelagic fishes in the California Current region. Calif. Coop. Oceanic Fish. Invest. Rep. 13: 39^4. AHLSTROM, Elbert H., and Robert C. Counts. 1958. Development and distribution of Vinciguerria lucetia and related species in the eastern Pacific. U.S. Fish Wildl. Serv., Fish. Bull. 58: 363-416. AHLSTROM, Elbert H., and H. Geoffrey Moser. 1969. A new gonostomatid fish from the tropical eastern Pacific. Copeia 1969(3): 493-500. Alverson, Franklin G. 1961. Daylight surface occurrence of myctophid fishes off the coast of Central America. Pac. Sci. 15(3): 483. Beebe, William, and Mary Vander Pyl. 1944. Eastern Pacific expeditions of the New York Zoological Society. XXXIII. Pacific Myctophi- dae. (Fishes.) "Zoologica (New York) 29(2): 59-95. Berry, Frederick H., and Herbert C. Perkins. 1966. Survey of pelagic fishes of the California Current area. U.S. Fish Wildl. Serv., Fish. Bull. 65(3) : 625-682. Bruun, Anton Fr. 1937a. Monolene danae, a new flatfish from Pan- ama, caught bathypelagically. Ann. Mag. Natur. Hist, 10th Ser. 19(110): 311-312. 1937b. Chascanopsetta in the Atlantic; a bathy- pelagic occurrence of a flatfish, with remarks on distribution and development of certain other forms. Vidensk. Medd. Dansk Naturhist. Foren. 101: 125-136. Bussing, William A. 1965. Studies of the midwater fishes of the Peru- Chile Trench. In George A. Llano (editor). Bi- ology of the Antarctica Seas II, p. 185-227. Ant- arctic Res. Ser. 6. Nat. Acad. Sci. Nat. Res. Counc. Publ. 1297. d'Ancona, Umberto, and Geminiano Cavinato 1965. The fishes of the family Bregmacerotidae. Dana Rep. Carlsberg Found. 64, 92 p. Ebeling, Alfred W. 1962. Melamphaidae I. Systematics and zoogeogra- phy of the species in the bathypelagic fish genus Melamphaes Glinther. Dana Rep. Carlsberg Found. 58, 164 p. Ebeling, Alfred W., and Walter H. Weed III. 1963. Melamphaidae III. Systematics and distri- bution of the species in the bathypelagic fish genus Scopelogadus Vaillant. Dana Rep. Carls- berg Found. 60, 58 p. Ege, Vilh. 1953. Paralepididae I {Paralepis and Lestidium) . Dana Rep. Carlsberg Found. 40, 184 p. Fraser-Brunner, a. 1949. A classification of the fishes of the family Myctophidae. Proc. Zool. Soc. London 118(4) : 1019-1106. Garman, S. 1899. Reports on an exploration off the west coasts of Mexico, Central and South America, and off the Galapagos Islands, in charge of Alexander Agassiz, by the U.S. Fish Commission steamer "Albatross," during 1891, Lieut. Commander Z. L. Tanner, U. S. N., commanding. XXVI. The fishes. Mem. Mus. Comp. Zool Harvard Coll. 24, 431 p. Gibbs, Robert H., Jr. 1964. Family Idiacanthidae. In Fishes of the western North Atlantic, p. 512-522. Mem. Sears Found. Mar. Res. 1, Part 4. 1969. Taxonomy, sexual dimorphism, vertical dis- tribution, and evolutionary zoogeography of the bathypelagic fish genus Stomias (Stomiatidae). Smithsonian Contrib. Zool. 31, 25 p. Grey, Marion. 1955. The fishes of the genus Tetragonurus Risso. Dana Rep. Carlsberg Found. 41, 75 p. 1960. A preliminary review of the family Gonos- tomatidae, with a key to the genera and the de- scription of a new species from the tropical Pa- cific. Bull. Mus. Comp. Zool. Harvard Coll. 122(2) : 55-125. 1961. Fishes killed by the 1950 eruption of Mauna Loa, Part V, Gonostomatidae. Pac. Sci. 15 (3) : 462-476. 1964. Family Gonostomatidae. In Fishes of the western North Atlantic, p. 78-240. Mem. Sears Found. Mar. Res. 1, Part 4. KuME, SusuMU, and Milner B. Schaefer. 1966. Studies on the Japanese long-line fishery for tuna and marlin in the eastern tropical Pa- cific Ocean during 1963. Inter-Amer. Trop. Tuna Comm. Bull. 11(3): 101-170. 43 FISHERY BULLETIN: VOL. 69. NO. 1 Kyle, H. M. 1913. Flat-fishes (Heterosomata). Rep. Dan. Oceanogr. Exped. 1908-10 Mediter. Adjacent Seas 2(A.l), 150 p. MosEH, H. Geoffrey, and Elbert H. Ahlstrom. 1970. Development of lanternfishes (family Myc- tophidae) in the California Current. Part I. Spe- cies with narrow-eyed larvae. Bull. Los Angeles County Mus. Natur. Hist, Sci. 7, 145 p. Nafpaktitis, Basil G., and Mary Nafpaktitis. 1969. Lanternfishes (family Myctophidae) col- lected during cruises 3 and 6 of the R/V Anton Bruun in the Indian Ocean. Bull. Los Angeles County Mus. Natur. Hist., Sci. 5, 79 p. Norman, J. R. 1934. A systematic monograph of the flatfishes (Heterosomata). Vol. 1, Psettodidae, Bothidae, Pleuronectidae. British Museum (Natural His- tory), London, viii + 459 p. Perkins, Herbert C. 1963. Redescription and second known record of the bothid fish, Monolene asaedai Clark. Copeia 1963(2) : 292-295. Pertseva-Ostroumova, T. A. 1964. Come morphological characteristics of mycto- phid larvae (Myctophidae, Pisces). [In Russian]. (Transl., 1966, In T. S. Rass (editor). Fishes of the Pacific and Indian Oceans, biology and distri- bution, p. 79-97. (Available Clearinghouse for Federal Scientific and Technical Information, Springfield, Va., as 65-50120.) Rofen, Robert R. 1963. Diagnoses of new genera and species of alepisauroid fishes of the family Paralepididae. Aquatica 2, 7 p. Ryther, John H. 1969. Photosynthesis and fish production in the sea. Science 166(3901): 72-76. Strasburg, Donald W. 1964. Postlarval scombroid fishes of the genera Acanthocybium, Nealotus, and Diplospinus from the central Pacific Ocean. Pac. Sci. 18(2) : 174- 185. Taning, a. Vedel. 1918. Mediterranean Scopelidae (Saurus, Aulopus, Chlorophthalmus and Myctophiim) . Rep. Dan. Oceanogr. Exped. 1908-10. Mediter. Adjacent Seas 2(A.7), 154 p. Voss, Nancy A. 1954. The postlarval development of the fishes of the family Gempylidae from the Florida Current. I. Nesiarchiis Johnson and Gempylus Cuv. and Val. Bull. Mar. Sci. Gulf Carib. 4(2) : 120-159. 44 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 1. — Counts of fish larvae, tabulated by family, for all stations occupied on EASTROPAC I. s § V3 % E o CD s o o u 1 s CO i n a> c o u < H c o 1 1 o s c o 2 'o i 3 w u O a; '2 % o s 0} ■5 a 2 u > a E ca u 1 s E ■a .060 5 26 59 4 1 18 2 3 2 120 .062 7 8 1 44 5 1 1 8 5 3 83 .064 15 71 6 2 274 21 1 2 1 1 5 7 406 .065 2 72 6 4 31 6 1 1 4 1 128 .067 7 54 3 I 34 6 1 1 107 .069 37 60 33 1 6 99 7 7 1 3 3 257 .071 37 572 8 1 8 6 318 9 8 1 14 6 3 1 13 7 3 1015 .073 42 167 53 2 27 1 172 27 11 3 7 10 1 1 16 7 8 555 .075 8 21 3 1 39 25 6 1 2 1 1 1 4 113 .077 59 38 2 89 14 2 1 5 3 3 1 2 2 36 257 .079 135 43 69 3 3 2 1 5 6 I 268 .081 2 164 13 1 16 2 1 8 4 1 3 215 .083 5 43 4 17 1 6 1 1 78 .085 2 1 17 20 .087 66 6 1 37 1 1 3 115 .089 26 29 105 2 1 2 3 15 183 .091 11 2 1 1 49 6 3 2 2 1 1 3 2 17 101 .093 3 8 2 13 .095 6 146 4 1 29 3 195 1 2 4 7 10 7 6 12 2 4 439 .097 3 103 11 7 2 205 2 2 6 4 1 5 1 4 9 2 1 368 .099 16 7 1 1 48 1 1 6 1 1 83 13.101 3 11 1 45 2 3 1 7 7 4 1 85 .103 1 162 6 1 3 4 255 5 5 3 7 3 14 3 7 479 .105 50 4 1 1 166 2 3 1 2 1 4 5 2 242 .107 1 13 1 1 16 .109 12 27 2 41 .111 18 1 49 1 1 1 71 .113 30 2 1 72 2 2 1 2 112 .115 8 25 1 1 1 1 37 .117 9 4 4 52 1 2 1 3 1 1 1 2 81 .119 36 4 86 4 4 134 .121 17 3 22 1 2 45 .123 3 2 3 1 1 2 2 14 .125 1 1 2 1 5 .127 20 2 3 39 1 1 1 1 1 69 .129 11 1 2 14 28 .131 6 7 3 16 .133 5 4 9 1 2 21 .135 56 I 1 78 2 1 1 140 .137 40 1 50 1 2 2 96 .139 12 1 8 1 1 1 24 .141 4 8 12 .143 76 2 86 1 1 I 6 173 .145 20 2 2 44 2 2 72 .147 17 33 3 4 2 3 3 65 .149 14 6 29 2 1 3 59 .151 22 3 1 72 1 2 1 1 103 .153 103 1 1 3 1 394 2 1 1 5 3 1 2 17 1 3 539 .155 8 4 16 2 2 2 4 38 .157 8 8 1 45 1 1 1 2 67 .159 12 5 8 53 1 1 1 4 2 2 2 1 92 .161 24 6 65 3 2 2 102 .163 5 2 14 1 22 .165 4 17 24 1 2 48 50 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 1. — Counts of fish larvae, tabulated by family, for all stations occupied on EASTROPAC I.- Continued. •c 3 n s z z o p < S5 8 g 3 § o O s s i i u to < c o 1 1 0} 1 5 'a 1 a u to 3 E CO SI (0 o H 13.334 37 21 17 1 116 2 3 1 1 5 3 1 208 .338 9 49 26 295 1 2 5 8 24 3 422 .340 4 11 23 2 47 4 4 1 4 100 .342 9 24 21 2 76 4 1 3 4 1 2 147 14.001 39 97 1 1 1 6 867 4 5 2 194 9 1 12 261 15 195 1710 .006 32 19 38 1 66 2 3 2 10 15 25 8 226 .008 34 4 32 2 1 86 1 2 1 1 2 25 4 2 197 .010 14 19 40 4 2 198 1 2 3 1 12 2 3 301 .012 6 1 7 1 1 57 1 3 1 2 2 4 4 90 .014 42 4 9 1 67 1 9 2 30 5 28 198 .016 19 1 20 8 2 4 2 44 16 5 121 .017 17 2 16 1 61 1 2 4 1 105 .018 41 48 64 2 2 424 4 5 24 1 19 634 .020 6 10 12 1 229 1 2 10 54 325 .022 7 22 14 3 80 5 1 1 32 4 169 .024 6 47 1 1 29 5 22 111 .027 23 31 42 3 387 2 7 9 6 87 34 19 650 .029 24 42 25 5 382 1 1 2 6 1 3 1 119 47 26 685 .031 30 43 46 9 2 594 15 2 6 1 1 3 75 5 43 875 .033 21 5 2 26 1 1 9 3 3 71 .040 48 2 2 36 8 3 4 6 1 21 3 21 155 .043 65 17 2 1 159 8 8 1 22 1 3 15 7 4 313 .047 111 3 4 4 22 3 2 3 9 6 7 44 218 .051 225 27 1 1 5 78 3 1 1 46 11 1 I 3 25 429 .055 154 2 2 40 4 8 210 .060 139 18 1 2 54 2 15 231 .066 13 1 II 1 26 .069 20 2 97 119 .076 3 1 11 15 .073 .081 2 16 I 18 37 .084 2 3 4 1 I 2 13 .086 2 1 7 3 13 .088 3 3 .091 2 40 1 43 3 2 91 .095 2 3 1 50 1 1 I 59 .099 2 3 1 15 21 14.103 .106 3 11 14 .110 8 27 1 1 37 .112 1 2 1 1 5 .114 2 2 .115 2 6 1 5 1 2 17 .117 2 I 1 1 5 .118 .120 1 6 1 15 1 1 25 .122 2 11 1 1 19 I 1 36 .123 7 23 7 51 1 1 1 2 9 102 .124 7 76 6 152 4 2 3 2 12 4 268 .126 3 20 1 2 6 53 1 3 2 3 4 1 15 114 .127 5 5 1 22 2 3 3 3 44 .128 5 60 3 9 145 5 6 1 ] 13 248 .130 3 44 1 1 7 45 2 2 4 3 2 1 5 3 4 15 142 52 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 1. — Counts of fish larvae, tabulated by family, for all stations occupied on EASTROPAC I.- Continued. a s t % « S Z § < -a 1 o g a o 1 IS :o § (0 < c o 1 1 o % c n 3 2 "3 o 1 O u O o s 2 rt -5 S. V 0. o u d •B o c o C in o c c g to 3 ■a u "9 c a, in s rt §■ 3 & rt o c rt 2 c 3 a a to s 3 JZ a o 2 > > c a « U & (0 d E J 8 o > H o £ > w s o 8 1 Q. & g 3 o JZ t >> E •a a> c o o £ U bo Q^ c CO •a 2 a o O E ■< c 2 s a g u c s o be S e a 3 s o "o o p £ a ■a lo ^ S o a u s o O S ^ rt ►J 3 2 g o Z O Z 0. ^ h c 3 I 11.022 1 1 c 1 1 4 .025 1 1 1 4 4 11 .027 1 1 4 4 C 1 1 14 .030 c c 1 .032 6 13 C c 19 .034 2 10 16 C ' 35 .036 2 1 c 3 .038 2 10 4 C 1 2 2 21 .040 5 4 39 C 1 4 2 55 .044 1 4 C c 5 .046 4 41 ) C 4 1 50 .048 1 6 c 13 20 .050 33 c 1 1 I 36 .052 56 c 2 58 .054 3 147 c c 159 .056 56 c 8 3 67 .058 27 c c 1 28 .060 4 53 1 3 c 1 1 72 .062 16 1 2 1 I 21 .064 2 43 ) c 2 3 1 51 .066 4 14 ) 2 9 4 20 3 4 I 2 63 .068 67 33 14 2 4 7 4 21 32 1 21 8 14 229 .070 1 15 8 e C 1 48 6 2 4 5 96 .072 28 6 22 1 12 7 44 21 2 24 7 4 178 .076 23 ) c c 2' 20 5 1 1 13 90 .080 1 18 2 1 c e 3 2 1 1 1 36 .084 63 0' i: 1 3 2 42 131 .088 2 10 £ c 6' 1 7 2 2 3 104 .094 5 8 4 c 32 9 1 1 4 2 66 .098 3 107 2 50 lOS 1 404 12 1 180 8 10 9 1 1 9 907 11.102 21 3 26 12 26 1 10 99 .106 6 c C 7 4 1 22 .110 41 8 c 7 I 1 1 1 57 .114 182 c 31 1 11 1 1 2 2 243 .118 70 c ; 3 84 .120 8 c 1 9 .124 37 c r 1 3 I 66 .128 1 31 c 2£ 2 6 16 1 8 98 .130 1 11 c 4 1 8 1 29 .132 4 3 C c 1 4 16 .134 26 39 3C 2 6 6 109 .136 30 39 4 8 2 60 15 168 .138 10 • 2 1 1 2 1 2 21 .140 2 c ( 3 1 12 .142 4 6 le 31 2 9 69 .146 10 3 c 1 2 1 17 .148 5 ) 1 c 4 3 13 .150 4 3 15 13 38 .152 10 13 i IS 1 2C 1 3 43 2 115 .154 1 e 5 3 15 54 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 2. — Myctophid larvae, tabulated by genus or species, for all stations occupied on EASTROPAC I. — Continued. t K 11.156 .158 .159 .161 .163 .167 .169 .171 .173 .175 .177 .179 .181 .183 .185 .187 .189 .191 .195 .197 .199 11.201 .203 .205 .207 .209 .211 .213 .215 .217 .219 .221 .223 .226 .228 .234 .238 .242 .246 .250 .254 .258 .262 .266 .270 .278 .282 .285 .287 .289 1 i % a ^ % 9 (0 a CB s B CO O o & o 4 8 19 3 2 1 1 1 1 3 2 3 4 3 14 1 5 7 3 17 4 1 3 4 2 9 1 1 (D 3 1 "3 ED >i > 5 fl •a U S « Q a 14 14 5 1 1 1 1 5 13 4 9 1 16 2 26 2 1 1 1 2 4 3 9 17 33 I I i g t s *• eo o Use I i 5 I g 8 111 U X s a -2 s s Z bo 14 3 58 62 17 18 32 57 19 8 51 14 7 2 17 1 2 1 1 1 2 2 1 4 « 2 1 2 1 5 2 2 9 23 16 2 1 3 6 2 13 6 13 5 11 3 4 7 1 9 14 15 30 1 2 3 7 6 14 19 1 I 8 9 7 17 2 6 1 1 2 1 7 6 1 11 6 2 7 3 1 4 3 1 5 5 5 a 2 2 4 2 1 2 2 13 98 8 4 20 5 1 20 19 6 4 1 3 4 2 2 2 1 1 1 4 1 1 2 2 1 2 1 3 1 3 4 5 3 2 1 1 2 1 1 2 16 11 85 4 3 7 26 6 16 1 2 8 2 6 4 3 1 2 3 3 13 3 1 18 12 1 4 2 5 27 25 4 1 1 1 6 2 1 2 8 3 4 2 17 7 5 3 19 6 2 3 1 2 1 4 36 1 1 8 2 5 14 1 1 2 5 1 4 1 1 11 2 2 2 1 29 19 103 22 117 10 2 20 3 1 5 4 3 13 13 4 24 19 2 4 60 9 8 17 40 21 12 28 71 44 16 10 74 20 7 16 46 10 95 6 198 57 149 108 85 38 17 1 116 82 8 30 3 87 12 131 55 FISHERY BULLETIN; VOL. 69, NO. I Appendix Table 2. — Myctophid larvae, tabulated by genus or species, for all stations occupied on EASTROPAC I. — Continued. Q •B s c c a (0 o E E a o o C bi bi o > >, O X X en CO H O c en n a H £ ffl rt J hJ - 2 11.291 .293 .295 .297 .299 11.301 .303 .306 .308 .310 .312 .314 .316 .318 .320 .322 .324 .326 .328 13 36 94 263 23 5 42 37 6 13 16 15 2 26 1 21 2 2 3 6 2 4 3 2 2 55 1 3 16 1 2 18 3 2 1 2 2 1 1 5 1 12 4 17 6 7 I 1 20 1 1 1 2 4 1 6 14 2 39 50 130 297 29 8 44 40 13 15 26 27 8 34 11 115 13 31 55 12.002 .004 .006 .008 .010 .012 .014 .016 .018 .020 .022 .024 .026 .028 .030 .032 .033 .035 .037 .039 .041 .043 .045 .047 .049 .051 .053 .055 .057 63 7 13 6 24 18 8 7 20 60 8 1 24 29 6 10 21 14 22 17 107 82 33 48 53 35 1 2 12 69 22 45 73 12 13 38 65 60 13 72 80 1 22 6 1 6 13 1 2 9 7 13 6 6 32 1 10 22 11 2 4 6 3 1 1 9 37 85 33 88 121 31 23 69 137 74 16 97 121 13 24 20 87 23 36 17 108 94 33 61 61 51 6 7 37 56 AHLSTROMt FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 2. — Myctophid larvae, tabulated by genus or species, for all stations occupied on EASTROPAC I. — Continued. as u s e o c o i 1 Si m O , >i c c bj (fl Q P Q CD 3 E 3 B 3 3 C i u 1 a a DQ > fi e JZ c u o > O a: X f^ & 3 U a> > Sa n a fc; t: en J vJ s- rt E c o 5 3 o > s s ■2 S 12.059 .061 .063 .065 .067 .069 .071 .075 .077 .079 .081 .084 .087 .090 .092 .094 .097 12.100 .103 .106 .109 .112 .115 .118 .120 .122 .124 .126 .128 .130 .132 .134 .136 .138 .140 .142 .144 .146 .148 .150 .152 .154 .156 .158 .160 .162 .164 .184 .186 .188 28. 2 5 1 10 6 19 4 3 1 7 3 11 1 26 32 104 555 4 337 195 2 25 16 204 6 65 18 80 7 103 27 127 38 5 12 62 36 33 67 10 277 4 41 1 12 5 14 5 19 28 2 2 3 1 6 11 21 2 5 47 7 2 e 1 7 15 2 3 25 48 7 1 1 1 1 7 14 4 1 4 1 1 5 1 1 3 13 1 2 1 1» 1* 1 1 2 2 25 3 2 4 1 2 2 1 1 1 1 1 2 1 2 2 12 7 9 2 12 42 3 2 1 1 5 10 1 7 5 3 1 56 5 25 17 14 15 11 1 5 37 30 12 7 20 140 27 5 35 25 8 1 17 54 7 6 14 6 1 1 2 3 2 3 4 13 1 1 4 5 2 6 10 1 1 1 1 48 3 2 11 22 2 6 3 2 6 10 4 12 11 1 2 6 34 9 3 3 3 2 2 1 3 1 1 1 2 3 3 2 1 1 4 2 1 2 1 7 2 3 17 4 6 11 11 10 10 16 13 2 4 35 6 1 3 1 3 5 1 1 1 4 3 3 6 2 13 1 2 4 2 1 3 20 2 6 3 39 8 2 3 1 1 3 7 3 1 2 2 14 2 1 3 3 1 11 1 5 4 7 5 1 3 3 18 6 3 193 1 1 1 7 20 2 3 2 3 5 3 1 1 1 6 4 2 5 2 2 4 2 1 99 41 109 614 366 227 71 294 110 129 389 207 64 18 71 377 101 56 124 319 65 32 72 107 15 30 29 108 49 12 8 20 19 10 69 84 72 11 11 86 37 12 14 41 19 5 36 45 5 57 FISHERY BULLETIN: VOL. 69. NO. I Appendix Table 2. — Myctophid larvae, tabulated by genus or species, for all stations occupied on EASTROPAC I. — Continued. c c t % n o V £ s 0} 09 O o s; <0 n u =1 H g i 3 E 1 CO a > h h 5 J D a c bl O tq E 5) m B U 0) > rt l3 a c f-i H ^ ^ a g- e to rt 2 £ rt rt c i (fl u > (D 0) 3 > o O o o 2; 2: s t 12.190 .192 .194 .196 .198 12.200 .203 .212 .215 .218 .221 .224 .227 .230 .233 .235 .238 .240 .242 .244 .24fi .248 .250 .252 .254 .256 .258 .260 .262 .264 .265 .268 .270 .272 .274 .276 .278 .280 .282 .284 13 1 2 1 1 1 1 1 80 10 28 92 92 22 102 315 22 126 194 2 145 10 7 6 7 9 5 127 3 17 57 12 1 17 7 6 2 23 14 17 205 43 38 13 22 11 38 26 24 15 31 44 35 61 25 8 19 5 10 11 2 5 1 1 1 1 1 1 1 1 1 1 1 4 3 63 5 1 1 1 1 1 1 1 54 11 16 40 3 13 2 2 3 1 2 1 2 13 39 4 1 5 3 17 5 8 9 2 13 10 21 35 176 98 61 47 471 30 16 107 61 42 5 1 10 16 5 6 12 16 2 1 40 2 3 1 1 7 I 2 1 1 1 1 6 18 105 10 2 2 3 16 8 1 7 2 8 1 5 5 2 6 6 3 4 17 11 2 2 15 2 2 2 6 1 2 6 12 5 30 2 1 4 3 2 2 9 1 3 2 12 1 5 6 1 1 1 12 2 5 35 3 1 7 2 1 1 1 3 2 6 2 3 11 38 4 1 6 3 6 1 1 2 4 1 1 1 6 45 16 22 12 36 91 36 70 300 202 127 209 1089 162 49 250 280 225 54 25 105 248 51 54 44 84 23 43 74 161 26 54 156 60 81 85 20 45 5 8 16 13.001 .003 .005 .007 .009 .011 31 315 1020 115 470 372 6 34 21 6 21 2 12 24 7 1 10 1 41 385 1075 133 494 374 58 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 2. — Myctophid larvae, tabulated by genus or species, for all stations occupied on EASTROPAC I. — Contimied. < El o s CQ e 1 01 CO o o CO Q) PQ u QQ r) 3 E 1 rt rt tn 00 > > .c ^ ^ F F 4= x: a >i o £1 £^ « CO e 0) rt > CO en 3 3 , (0 o o ^ 2 S bo 13.013 .015 .017 .019 .021 .022 .028 .030 .032 .034 .036 .038 .040 .042 .044 .046 .048 .050 .052 .054 .056 .058 .060 .062 .064 .065 .067 .069 .071 .073 .075 .077 .079 .081 .083 .085 .087 .089 .091 .093 .095 .097 .099 13.101 .103 .105 .107 .109 .111 .113 432 407 9 25 13 45 20 15 6 29 30 2 1 6 17 1 1 3 2 4 5 1 2 2 1 12 I 11 21 4 10 46 19 1 15 17 26 186 477 550 715 2 24 44 142 11 122 408 271 79 44 160 71 42 36 33 50 54 22 191 12 24 73 284 122 28 60 50 11 13 9 1 9 2* 18 2 I 12* 6 4 1 2 3 1 4 2 1 3 1 15 2 2 2 6 3 2 1 1 1 1 3 2 1 3 2 10 1 45 33 5 6 42 22 2 2 5 1 20 2 5 9 16 2 18 47 25 14 28 76 25 14 20 43 16 1 7 18 8 2 10 20 40 5 23 8 3 2 3 7 3 1 19 5 1 1 3 5 2 2 5 2 1 5 1 6 1 1 2 15 18 3 11 5 2 7 1 1 1 3 7 8 19 11 5 12 8 1 1 1 4 7 1 21 37 7 4 16 7 1 2 1 1 1 1 1 21 1 3 5 4 7 13 1 1 1 1 7 9 4 4 2 4 2 2 1 1 7 1 3 6 18 34 8 2 30 24 5 10 26 33 1 1 2 1 10 4 4 12 5 3 10 1 2 5 5 3 2 1 3 6 6 5 6 24 1 4 2 2 1 4 2 1 5 3 21 5 2 3 1 I 32 3 5 1 10 4 2 1 2 207 479 559 13 1219 409 2 24 44 158 13 142 469 307 109 105 300 133 79 77 144 83 59 44 274 31 34 99 318 172 39 89 69 16 17 17 37 105 49 8 195 205 48 45 255 166 13 27 49 72 59 FISHERY BULLETIN: VOL. 69. NO. I Appendix Table 2. — Myctophid larvae, tabulated by genus or species, for all stations occupied on EASTROPAC I. — Continued. a H < ■3 $ Si s a u CO a a F F 3 3 X o O X en « « R c O ■s !5 a. O £: «! -1 _; 5 o £ 2 S M =■ C OJ C 13.115 .117 .119 .121 .123 .125 .127 .129 .131 .133 .135 .137 .139 .141 .143 .145 .147 .149 .151 .153 .155 .157 .159 .161 .163 .165 .167 .169 .171 .173 .175 .179 .183 .187 .191 .195 .199 13.203 .207 .211 .215 .219 .223 .227 .231 .235 .237 .239 .241 .243 2 4 4 2 1» 2 3 1 4 2 25 2 20 18 7 1 1 3 52 7 56 2 3 6 3 1 4 4 86 6 1 1 1* 4 I 2 1 2 2 1 22 2 1 3 1 1 2 7 3 3 2* 4 4 3 5 1 1 4 2 39 7 4 1 2 14 2 1 2 2 7 2 1 2 1 I 2 9 12 35 1 1* 1 11 7 1 5 2 2 78 5 13 3 2 2 6 2 6 9 2 50 3 2 1 2 8 1 1 1 3 1 1 8 15 21 6 2 4 5 7 2 5 I 1 13 2 2 86 16 6 4 2 4 2 2 8 44 2 5 2 2 2 3 2 3 1 10 1 33 2 2 2 1 1 2 3 5 5 3 1 2 29 43 6 2 4 3 2 1 1 3 1 I 72 83 172 7 30 23 9 7 26 1 n 8 3 14 394 4 2 2 4 4 16 11 19 1 1 4 2 1 2 4 45 12 16 3 1 7 7 2 5 53 27 11 1 4 1 4 4 10 1 2 65 1 2 3 1 6 1 14 3 6 1 2 3 8 I 24 16 2 2 3 1 8 1 33 115 1 8 2 20 1 1 17 4 169 5 13 54 1 7 2 8 3 11 104 2 98 13 1 2 1 4 121 1 168 1 52 1 2 18 2 245 122 1 7 2 1 133 64 3 7 1 2 2 1 80 49 2 11 1 3 I 67 78 2 43 3 126 4 34 125 2 1 16 182 I 31 4 2 12 3 2 55 7 1 3 3 4 1 1 20 1 72 12 8 1 24 1 2 1 4 3 129 3 40 5 1 9 3 1 2 4 68 2 1 « 2 2 7 6 1 3 1 11 18 I 9 1 2 1 32 2 90 1 3 15 2 1 1 6 121 29 2 39 4 3 1 2 2 82 1 64 1 3 14 1 5 7 3 1 1 5 106 133 4 34 8 1 1 2 1 5 189 131 1 29 7 4 2 5 179 52 1 1 1 1 1 2 59 67 1 21 1 3 93 60 AHLSTROM: FISH LARVAE IN ELASTERN TROPICAL PACIFIC Appendix Table 2. — Myctophid larvae, tabulated by genus or species, for all stations occupied on EASTROPAC I. — Continued. % s o s o cd E DQ 1 1 % c (0 m 1 E © m o i E I E 3 £ X d d ID z % 09 u > 0. CL > OQ % c i d m E 1 K d oa ■0 i y E -0 4) c -0 c 3 BQ •a §■ CO 1 CJ >^ E H 1 < QQ 3 § CJ m o ed Q> O a a (0 CO p 1 5 (D > c bd o 5 CO > o c g ■q & n g w c (h £ § bx >, X £ a E I 1 & CO c •0 E n ►J z D s C4 ►J ■D ! & .2 c « t U .5 Q. M E D f 8 > > i .c "o Z CD .2 & u w z 1 £ 5 % 1 E CO CO s B t u ■a a 'rt u tc CJ ;s CO 5 13.245 37 20 57 .247 17 9 26 .249 43 1 2 46 .251 15 10 25 .253 5 1 6 .255 12 11 4 2 29 .257 4 5 1 10 .259 16 2 1 4 23 .261 8 4 1 13 .263 33 5 8 46 .265 11 2 1 14 .266 14 14 .268 12 19 2 33 .270 5 3 1 9 .272 11 2 13 .274 26 33 4 63 .276 38 48 D 1 2 89 .278 2 4 1 7 .280 23 6 2 31 .282 80 7 1 1 89 .284 13 7 13 1 34 13.318 1 2 3 .320 3 11 4 18 .322 ,0 C .324 c 25 1 26 .326 5 39 44 .328 28 7 35 .330 24 24 .332 62 1 63 .334 111 1 4 116 .338 274 15 1 1 2 1 1 295 .340 33 5 3 3 1 1 1 47 .342 62 6 2 3 1 1 1 76 14.001 4( 1 725 1 49 18 5 17 6 867 .006 9 43 3 1 2 8 66 .008 78 3 5 86 .010 6 179 2 4 6 1 198 .012 47 2 7 1 57 .014 6£ 1 67 .016 8 8 .017 54 1 1 4 1 61 .018 246 I 148 23 4 1 1 424 .020 22E 2 1 1 229 .022 7C 3 2 1 80 .024 34 13 47 .027 372 11 4 387 .029 ) 371 6 382 .031 ) 54C 1 1 19 13 6 2 1 7 594 61 FISHERY BULLETIN: VOL. 69. NO. I Appendix Table 2. — Myctophid larvae, tabulated by genus or species, for all stations occupied on EASTROPAC I. — Continued. 1 1 c o c a a a o 1 s c 1 tn 3 s o o o a a m m z n g > o c OQ 3 ■i in o d rt § to o 3 c o Hi > S n 3 g 2 o E 5 a. s o to V) 3 o > c to 3 "o .Q g > a tn § s d £ g a ^> > to > •a c o i s (0 & d to E s § 1 cS > w E O .C s i (A 2 1 to f o >^ E -a c to ■o f V e to •a i H X J= (U O w g s Q Q ^ rt (A o ^ JZ 0) < c a D ti tl o c o O E B D '£ ^ y "o O O £ Q •D « rt H u o o > > M rt flji o > o O L. ^ 'u C o m n u b Q b U o K X J ,J J ►J '^ s 2 2 0. (- 3 Q H 14.033 21 c C 2 G G 1 2 26 .040 1< i 4 5 G 9 G G 1 36 .043 2 ; s 40 G 41 25 G 2 3 1 9 159 .047 ) c 4 8 G 1 G 22 .051 ) 3 c G 27 12 G 1 1 6 78 .055 ) 3 1 1 G G 2 G G G G 40 .060 ) 1 1 c G G G G G G 1 G 18 .066 c G G G G G G G 1 .069 ) c G G G G G G G G .076 c G G G 1 .078 ) c G G G G G .081 ; c 8 G G 3 1 1 18 .084 1 c G G 4 .086 c c ( G G G 1 7 .088 ) c c G G G G 3 .091 ) c G 40 G 1 1 43 .095 ) 1 1 c G C 1 G 1 G 19 12 50 .099 7 c 7 G 1 15 14. 103 ) c c G .106 ) c G G .110 5 c 14 G G 1 6 27 .112 ) c c 2 G G 2 .114 c 1 G G 2 .115 ) c 4 G G 1 5 .117 ) c c G 1 1 .118 ) c ( G G .120 • G G 4 2 15 .122 1 G G I G 9 1 1 19 .123 ] 6 4 1 G 1 16 1 13 51 .124 1 1 40 18 1 2 1 2 51 1 10 152 .126 5 1 5 3 21 1 13 S3 .127 I 2 1 G 1 5 9 1 22 .128 I 1 33 c 2 G G 4 3 72 9 2 145 .130 3 G G 5 2 G 21 5 45 .131 GOO G G G 2 2 .132 GOO 2 G 3 1 I 8 .134 C G G 1 2 11 .136 ) 4 G G 43 .138 ) 1 ] G 11 G G G 2 29 .142 ) 10 G G G G 1 1 110 .146 ) 15 5 13 G 6 G 176 .150 ) 3 6 G G G G G 2 5 52 .154 4S G G G 16 8 2 3 92 .158 G G G 2 G G G 4 30 .164 3C G G 5 1 2 G 2G 135 .172 2 6 G G G 1 1 G G 26 .174 ] 4 G G 2 3 1 G 26 .177 C G G G G G G G G G .183 1 G G G 1 G G I 4 .188 3 18 3 4S G G 6 2 9 5 6 261 62 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 2. — Myctophid larvae, tabulated by genus or species, for all stations occupied on EASTROPAC I. — Continued. a n s 2 Z o c V c a c E o tn o 5 c o (0 c CO § V ta O 3 en £ > o c Vi 3 O i rt rt to 1 X u c & c en > E s a u rt E 3 E £ D 1 a a « rt c a -a « B > c rt 1 s s D o rt u el u rt c a. & 6 3 c rt > "rt > s o (0 c •g c i £ to o u g- E 8 1 c c rt > » e o & a a w « g 3 O £ O §• u £ 2 rt u to CO T3 1 O O >> E H ^ J= 0) CJ u c o" a 1^ ^ 5 3 w o ^ J= o c < c 2 i g § o C s bf £ £ '5 3 s o o o p £ Q ■o rt H a o o > > rt rt o o 2 > o o ^ U c o u n U S s s u o E .J ►J J J s Z z s. H 3 s H 14.194 ) 155 I C 7 C 158 2 12 4 8 5 5 1 1753 .195 24. ! 2 c c 3 4 252 .199 1 ) c 2 1 2 16 14.203 9( ) C 1 6£ 3 1 3 4 4 1 7 179 .209 2. ) c c 2 25 .213 18] c D C 2E 206 .218 16f C 8 1 177 .220 ) 2( ) ( c 2 22 .220 4 ; 1 c 2 49 .224 9 ) G c 21 2 2 115 .228 2f c 1 1 30 .230 ) 3 ) c c c 1 1 41 .232 2 c c c 1 31 .234 9« c c c 81 1 48 228 .236 3( ) c c 4 34 .240 7( ) c 4 c 12 1 20 2 2 1 1 3 116 .243 2 i c 2 C 14 2 2 1 44 .247 ) 4 c c c 2' 5 2 1 6 1 86 .251 ) 1 ) c 1 c c :; 1 1 25 .255 21 3 C 1 c c 2 3 3 1 227 .259 ) 4 ; c c e 2 1 3 1 53 .263 9- 1 c c c IC 1 105 .267 1 c c c : 1 8 .276 ) c c 1 6 2 1 13 .280 ) 5 C 2 C c 2 26 3 5 44 .283 ) 1 ! C c c c 8 1 1 22 .287 ) C c c 4 1 2 4 1 15 .291 ) ) C c c c .295 ) 4 ) C c c 15 1 65 .300 ) 1 i C I c c c 1 18 .303 ) 18 i C c c 5 3 3 200 .306 ) 2 J c c 1 4 32 .310 6 c 2 C c 8 1 5 4 14 96 .314 ) 3i i c c c I'S 5 2 59 .318 ) 49 c 7 C c 38 23 5 2 566 .323 14 ) c c c 2 3 3 165 .326 i 57 ) c c c 7: 1 4 5 659 .330 ) ( ) 2 ! C c c 1 c 23 63 FISHERY BULLETIN: VOL. 69, NO. 1 Appendix Table 3. — Counts of selected categories of fish larvae, tabulated by station, EASTROPAC I. n 10 u tt E n 2 3 E g 1 u z o > u u u 'c m *> .s & 1 1' 1 C 2 _3 § 1 c B a 1 I s o u E i 1 i w 3 D E « t o "3 'u 1 a u Q. o. tA 1 < CD 1 "o o CO 1 i 1 1 c 1 o o 3 s 2 CO si E E CTi Q a "Z X o. CO ca E i ■3 2 11.022 ) ( 10 .025 ) f 46 1 .027 ) J 34 1 I .030 ) ( 1 .032 1 ) ( 15 D .034 1 ) ( 88 .036 ) ( 26 .038 4 ) ( 22 e .040 ) 1 1 18 c 1 1 .044 2 ) ( 9 .046 3 ) 1 57 c 1 .048 12 ) 41 c .050 24 ) 1 22 c .052 15 ) 2 c .054 U ) ^ .056 10 ) 8 .058 10 ) 2 1 .060 13 ) 1 ' 1 3 1 .062 ) C .064 ) 1 C 2 .066 ) 2 c 2 2 1 1 11 1 .068 5 ) 6 1 7C 1 3 3 4 .070 1 ) 6 1 1" 3 1 1 8 1 .072 2 ) 2 6 2 6£ • 1 9 1 .076 6 ) 5 2 682 1 4 .080 7 ) 2 1 139 1 .084 11 ) 3 358 1 3 2 .088 1 ) 9 31E 1 1 2 .094 ) 9 41 1 1 1 2 .098 2 ) 2 ) 8- I 1 11.102 6 ) 3 8 4 2 .106 1 1 K 4 2 .110 8 ) c c .114 1 J 1 2 c 44 3 1 .118 4 ) 2 e 4 c 2 .120 1 3 2 1 D 1 1 .124 5 7 1 c r 1 .128 3 6 c 95 2 1 .130 3 6 c 1 .132 1 5 1 c 2 1 .134 3 3 24 c 1 1 166 2 .136 3 2 I 2 c 20 2 .138 3 1 3 c 2 1 .140 3 4 c ] .142 3 9 c 4 1 .146 3 20 2 c 3 .148 3 61 1 5 c 1 .150 a 70 1 c 1 4 .152 3 89 3 i 1 c 9 1 1 1 2 1 .154 » 2 5 3 c 1 .156 B 83 J c .158 3 20 1 3 f 1 .159 70 2 I c 6 3 .161 3 1 3 D c 2 1 64 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 3. — Counts of selected categories of fish larvae, tabulated by station, EASTROPAC I. — Continued. 1 m u E n 3 0> 1 § CO > c .§ t. tl c CO ">, 3 u i o g 3 5 c c nl z « o % < o o >, o & (0 o I a S 3 o o o 3 E M 3 "o u 3 3 "a c c £ .2 s oil 2 1 s o u « to 1 i m E 3 « o 1 E 1 ■e o % 1; u Q. a 1 < u E "Z to 1 i 1 n i c 1 o o u o o -a z J2 E Q m % 1 & E 3 c o u u s 2 u o 11. 163 8 a 1 .167 25 9 4 1 .169 3 1 1 r .171 11 .173 9 1 .175 1 .177 5 1 .179 4 1 1 .181 13 1 .183 3 2 D .185 4 .187 19 3 c 1 .189 20 1 c .191 1 c .195 4 10 c .197 8 32 5 2 1 c c 1 1 c 3 .199 5 1 2 c c 11.201 2 1 c c .203 8 12 3 2 I c c .205 22 20 2 9 3 c c .207 10 5 c c 1 .209 3 3 1 1 .211 35 4 1 c 1 .213 13 10 1 12 1 3 1 .215 3 2 D 1 3 1 .217 3 5 1 1 .219 1 .221 18 2 36 1 1 .223 8 5 .226 2 3 .228 1 2 c 1 .234 2 7 2 8 1 2 1 1 1 .238 3 5 1 D .242 4 13 1 36 c 4 .246 2 15 36 } 1 I 1 .250 1 1 12 8 3 1 1 c 1 .254 5 15 1 c 2 .258 3 1 8 46 3 c 1 .262 2 10 58 D c c .266 3 3 30 3 c .270 4 1 17 D 7 : c .278 13 1 13 142 ) 1 c c .282 1 6 1 20 1 ] ] c .285 9 5 5 2 1 1 1 c 1 .287 1 4 14 1 2 3 2 C 2 .289 7 4 5 3 1 IS c 6 .291 1 1 I 3 4 C .293 7 1 1 2 3 6 14 1 .295 10 5 10 3 3 1 1 3 2 .297 5 3 9 D 3 2 1 1 4 .299 27 2 3 5 3 1 1 11.301 1 13 3 1 3 3 .303 12 2 45 1 3 1 .306 4 64 1 3 1 65 FISHERY BULLETIN: VOL. 69, NO. I Appendix Table 3. — Counts of selected categories of fish larvae, tabulated by station, EASTROPAC I. — Continued. o s E E o g § p 1 fa (0 9 10 3 m O t 3 rt c & m O JZ g s > CO 1 1 in 3 g g 3 E [0 1 s 3 i 'C ,1 u c u _3 1 1 C & E u n a 1 o u 1 d. & E 3 to o 1 1 V s 3 ■§. u o V JZ (0 1 1 re o CO 1 1 d & c JZ g o o o 3 ■a (0 B rt o s E E ri a s §■ E 3 o s o m (0 ^J Z < o ia s > B s m M !- < X o z B f~ u 11.308 1 J 4 2 1 .310 4 J ( 6 1 .312 ) 1 59 1 1 1 .314 5 1 32 1 1 .316 ) 3 1 1 .318 ) 2 .320 1 ) 10 1 .322 ) 33 .324 ) 19 1 .326 ) 36 .328 ) 31 12.002 3 ) 11 1 .004 3 ) n 3 .006 2 ) 3 12 5 2 .008 3 ) 6 59 2 1 .010 8 ) 97 .012 4 ) 8 16 1 1 .014 4 ) 5 33 2 .016 8 ) 4 178 4 1 2 .018 ) 199 7 1 .020 3 ) 3 20 4 1 .022 ) 4 11 1 1 .024 3 ) 8 234 32 I .026 4 ) 2 99 3 .028 ) 31 2 3 1 .030 6 ) 12 1 3 .032 ) 13 1 7 3 .033 3 ) 36 3 6 1 .035 1 ) ] 72 4 5 2 2 .037 9 1 10 1 2 .039 4 3 3 .041 3 2 20 3 3 1 .043 28 2 3 1 .045 7 3 c .047 1 2 2 3 1 1 .049 2 2 52 1 2 .091 68 1 .053 1 18 2 1 .055 c 7 2 .057 2 c 13 .059 21 12 66 1 .061 8 6 .063 8 .065 11 44 .067 2 A 31 2 19 7 3 1 .069 5 6 c 1 11 1 2 1 7 .071 9 2 16 I 1 .075 19 6 1 119 1 2 1 2 1 1 2 .077 13 ; c 23 2 1 1 4 .079 17 24 c 2 21 2 42 3 2 .081 29 15 c 59 2 30 2 1 .084 14 5 6 8 .087 1 3 ( ) 6 10 1 66 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 3. — Counts of selected categories of fish larvae, tabulated by station, EASTROPAC I. — Continued. ED tn 10 E £ E 2 -> 3 e o a .1 u 1 3 (0 3 m 1 3 1 n c c « 2 1 'rt < i o 1 a 5 ! 3 a o o o > s 3 E 3 s i « i- u .5 > O n c B s E en 1 n s 1 1 C/3 a. a. E « o I i u 3 o 1 s a a a m 1 < CO E re Ij a i 1 1 o o o u 3 •o QJ rt u i 2 rt 4) E a ri & E 3 c o t 1 "3 ■3 rt o u 12.090 4 1 7 e c .092 4 41 c 1 .094 3 3 t 1 124 c .097 3 8 46 c D 1 1 12.100 1 2 ( 1 10 c 1 1 1 1 .103 ( 1 33 c .106 2 B ( ) f 35 1 1 3 .109 16 .112 ) 1 4 .115 1 J ( 12 36 3 1 .118 ) 18 141 1 1 11 2 3 1 .120 4 3 .122 ) 1 4 .124 5 D 1 .126 ) 4 18 1 D .128 ) C 8 .130 ) C •) .132 ) 1 6 .134 ) IC 18 .136 13 .138 ) c 12 .140 18 2 2 .142 ) 22 86 1 1 .144 ) IE 147 1 1 .146 ) C 12 1 .148 ) C 3 .150 ) 28 6 1 16 1 .152 ) E 5 2 .154 ) 8 3 D .156 ) 7 1 3 .158 ) 42 8 2 1 1 .160 ) 19 .162 ) 1 1 .164 24 15 1 .184 ) 31 17 1 .186 ) 1 1 1 .188 2 1 .190 ) 10 2 .192 14 1 5 .194 4 1 .196 1 .198 2 12.200 3 3 1 1 13 D .203 1 3 1 .206 ) D 15 1 1 .209 3 2 1 2 .212 1 17 17 5 1 .215 16 2 27 1 2 12 1 I .218 9 2 11 6 4 .221 IS 4 9 16 1 .224 ) C 48 2 201 2 2 .227 C 8 5 81 1 1 1 1 1 .230 i C 1 4 1 .233 4 i C ( ; 1 2 46 67 FISHKRY BULLETIN: VOL. 69, NO. I Appendix Table 3. — Counts of selected categories of fish larvae, tabulated by station, EASTROPAC I. — Continued. at c aj u M E £ e o n 3 E 1 a u > c c *£ EC 3 n m O 1 a a. 8- C i z Q. « o 1 < to Vi o § Q Q I o 1 3 £ w 3 O £ S i .5 'C u u p 1 n c E u HI & £ 1 u a o £ s o u m n 2 3 CO s, D o & o E 1 •£ o •a u 1: i i < 1 a c o ffl i c s o o u o o -o w s o « z to rt "a! E E rt D 1 d m e 3 c 1 o 12.235 25 ) 1 9 c 49 11 1 .238 17 ) 7 31 1 1 17 .240 11 ) 1 3 2 14 1 23 1 2 1 .242 ) 2 ' 2 12 1 3 .244 ) f 2 4 2 1 9 2 22 5 .246 11 ) 1 I 8 1 7 2 .248 ) 6 f 1 : 2 1 .250 ) 6 C .252 ) 2 c .254 ) 2 c .256 3 ) 1 2 1 .258 ) C .260 2 ) 1 15 C 2 2 .262 2 ) 5 97 1 .264 2 ) 6 14 1 1 .265 3 ) 7 11 1 .268 ) 2 103 1 2 2 .270 1 ) 29 C .272 4 ) 5 38 c 2 .274 ) 4 36 I .276 ) 138 c .278 1 ) 1 164 c .280 ) 2 20 c .282 1 ) 2 21 c .284 ) 1 118 c 13. 001 8 ) 3 90 c 1 .003 14 ) 3 J 1130 1 1 2 1 .005 54 ) 6 1 300 c 3 1 .007 9 ) 2 11 c 7 I .009 8 ) 9 c 7 .011 (i 2 1 .013 13 ) 8 c 1 1 .015 4 ) c 4 .017 16 ) 61 1 I 8 1 .019 9 ) 82 c 9 33 1 1 .021 4 ) 2 c 20 .022 1 ) c 1 .028 4 ) 6 c 1 .030 7 ) c 7 2 .032 ) 24 c 52 8 .034 23 ) 60 c 4 41 1 3 .036 8 ) c .038 28 ) c 2 .040 20 ) 5 10 1 20 1 .042 11 ) 5 6 c 1 4 .044 10 1 3 11 c I 1 .046 10 ) 5 73 4 1 1 .048 29 ) 1 ) 291 2 1 1 1 4 .050 10 ) 5 36 1 .052 i:i ) 1 J 1 181 c 4 1 5 .054 14 ) I 9 1 5 76 4 3 11 1 .056 33 ) I 2 ) 2 27 454 16 1 1 6 .058 6 ) J 3 8 I 10 319 1 7 68 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 3. — Counts of selected categories of fish larvae, tabulated by station, EASTROPAC I. — Continued. c IS s g s m :3 s 1 u 2 O 1 1 c e 1 3 3 ta 3 o "5 g 1 ffl c o c d 2 d n o 1 < i 1 o "o >, o & o g a Q d a m 3 i o f — 3 E 3 £ d a ft u 1 > u w 3 m 1 1 o ffl B 1 a o i m E a o w B IS 3 •g. t o u ! 1 < 1 1 o » c s 3 d D. c s 1 o o u o 3 •a o u rt u u 3 §. IS u B i o rt % 1 d at E 3 o o 13.060 5 2 2 14 8 17 a .062 7 c 3 1 .064 15 5 18 43 1 .065 2 4 1 11 56 1 .067 7 2 c C 52 .069 37 1 2 c 13 45 1 1 .071 37 17 c C 553 6 1 14 4 1 .073 42 13 c C 153 1 3 4 5 .075 8 3 c 18 1 e 1 .077 c 59 2 5 2 .079 1 c 134 5 1 .081 2 c 164 1 .083 5 c 43 .085 c 2 .087 2 c 64 2 .089 9 c r 2 .091 11 2 I 1 .093 : .095 6 25 1 120 2 1 10 1 .097 3 11 1 87 1 1 1 c .099 4 1 11 13.101 3 1 7 1 .103 1 36 125 1 2 2 6 .105 20 30 1 2 .107 1 .109 1 11 1 .111 7 1 .113 12 18 1 1 1 1 .115 4 4 1 .117 4 1 4 1 1 1 .119 26 3 6 .121 10 6 1 .123 3 .125 1 c .127 13 7 1 c .129 9 2 c .131 4 c .133 4 c .135 46 10 1 c .137 12 28 c .139 3 9 c .141 2 2 c .143 21 55 1 5 .145 12 8 .147 1 16 3 .149 6 8 1 1 .151 7 15 .153 44 59 3 .155 8 2 .157 3 3 1 .159 8 1 3 1 4 .161 11 13 .163 ) 5 c .165 3 4 c 69 FISHERY BLLLETIN: VOL. 69, NO. I Appendix Table 3. — Counts of selected categories of fish larvae, tabulated by station, EASTROPAC I. — Continued. 1 CD h B E e g o u g p < 00 >, c bi ca CO 1 a 3 ca _3 ■£ 3 CO o t 1 i a c s 5 z 09 1 o CO u < i g 6 d CQ g 1 a 5 a. & to a i — 3 s CA 3 y cfl s C8 r at g i > u t£ en 3 5 CQ a "3 c c E e m Q O 2 1 .2 6. & E to § CO 1 u E (0 3 o f 1; CO a. Q. 1 < 03 E a c « i c o x; o o Li o 3 •a a 3 CO 2 m rt E a — 1 X E 3 c o a 1 2 13.167 5 14 .169 7 c 76 .171 2 c 16 13 .173 7 ( c e 4 1 6 .175 22 6 c 169 1 1 45 1 1 .179 15 6 c 105 1 7 1 .183 50 3 c c 36 c 6 .187 43 2 c 26 c 1 .191 10 6 2 lis c 1 7 1 .195 10 f c 194 c .199 5 2 2E c 1 13.203 3 1 2 9 5 c .207 11 1 ' 14: c .211 5 6 1 27 c 1 .215 6 1 c .219 9 8 c .223 2 3 £ 15 c 1 .227 9 1' I 4 20 3 1 .231 9 1 5 C 29 c .235 9 1 C 31 c 1 1 .237 6 1 c 3: 1 7 1 1 .239 3 7 c 34 c .241 2 c 4 c 1 .243 11 c 13 c 1 .245 7 c 13 c 1 .247 9 c 11 c 1 2 1 .249 1 c c .251 6 c 10 2 c 3 .253 20 c 2S 4 c .255 7 c 29 c 1 15 1 3 .257 8 c 3 2 c .259 17 c le c 5 .261 8 c 36 c .263 19 c 54 c 1 11 .265 6 1 c c 12 8 c 1 1 .266 c 2 c 2 .268 3 26 c 1 .270 4 2 18 3 c 12 1 1 .272 2 c 3 2 c .274 1 3 49 6 3 1 1 .276 5 2 89 3 c 3 19 .278 1 1 20 2 c .280 8 3 5 c .282 3 33 6 c I I .284 18 60 2 1 13.318 13 1 c 1 1 .320 8 16 c 13 .322 C 5 c 1 9 1 .324 10 4 c 2 4 .326 13 9 c 18 1 .328 2 9 c 7 .330 4 12 c .332 1 1 ) 0-2 c 2 1 70 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 3. — Counts of selected categories of fish larvae, tabulated by station, EASTROPAC I. — Continued. — QJ •c a V u 6. & 3 a £■ h cd ^ 05 ? 8 O > 3 JZ S o o o p <0 6 o "H ^ 3 (8 ^ £ < U u t; > « CO rt d o ft a ta u h o o a i5 1 o "E £ 3 "o u i a C u s > Li m 1 o H n 1 o 3 E CO CO 13 1 ri D 3 3 1 3 *S u: U Z EC H 14. 131 14 .132 8 17 1 1 2 .134 1 3 1 .136 1 18 .138 9 2 90 9 .142 1 53 4 I 1 .146 16 30 111 1 8 .150 33 1 2 1 45 4 1 5 .154 28 6 9 1 372 1 7 3 .158 4 1 219 1 1 .164 8 4 1 48 3 6 1 .172 1 2 5 90 12 .174 3 1 3 33 .177 3 .183 1 1 .188 34 4 9 155 1 2 36 1 1 .194 161 7 22 98 1 22 1 4 .195 54 16 2 .199 15 5 196 1 20 3 14. 203 3 2 1 1 .209 5 2 10 .213 34 10 86 11 1 .218 27 1 38 2 1 4 2 .220 2 3 13 .222 2 4 2 I .224 9 3 6 31 1 7 1 .228 2 5 2 .230 1 5 9 .232 9 49 12 .234 9 2 867 1 161 1 .236 3 16 1 10 .240 21 1 4 31 85 1 .243 24 1 7 19 5 .247 4 13 39 1 .251 8 3 1 1 23 1 6 .255 1 209 10 76 59 1 6 .259 16 40 1 5 .263 2 2 231 2 .267 3 33 1 2 3 .276 9 6 52 2 1 .280 7 3 59 2 2 6 .283 3 13 1 1 .287 5 3 59 9 .291 .295 4 1 63 2 5 .300 1 286 1 .303 3 490 2 8 1 1 1 .306 12 22 2 .310 70 21 9 12 1 .314 24 1 24 .318 32 2 2 66 1 1 .323 n 24 1 .326 15 12 1 65 8 .330 1 72 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 4. — Summary of occurrences and numbers of larvae of eight families, limited in distribution to a broad coastal band or around offshore islands. « S z o H < CO 0) eg •o a u "a 1 c o c >v CO -a '5) u o 4) ■V 1 ►J "B 1 o a; a "B "c & o o CO a m z z c < CO '3 c o o "a be c W n c o 1 CO 8 CO 1 u u eg V rt 'u Si rt .J B o O "5 c & o eg 11.076 1 14.001 5 1 1 84 55 11 11.246 1 .006 1 5 1 .008 2 5 12. 020 1 .010 3 2 .024 1 4 .012 1 1 1 .026 10 .014 6 4 11 1 .028 3 3 .016 34 .030 1 1 .017 1 .031 7 .018 3 10 4 .033 1 2 1 2 1 2 1 1 .020 n 1 1 1 .035 6 5 3 1 .022 14 1) 2 1 .041 1 1 .024 4 6 5 6 .059 1 .027 n 3 5 1 4 36 9 12.221 CI 2 .029 7 2 12 9 37 .256 1 .031 3 3 1 18 4 .262 1 .033 1 1 1 2 .264 1 .040 6 5 1 .268 1 e .043 1 2 .047 1 1 3 13.003 1 1 6 .051 1 .005 1 1 .055 1 .007 1 .060 52 2 .011 9 1 .066 11 .019 13 1 70 49 2 47 12 .069 97 n .021 2 7 11 3 3 1 .076 11 .030 1 2 14.106 3 .032 e 12 8 .110 1 .034 9 23 3 .154 5 .040 1 .158 5 1 8 3 .042 1 .164 9 a .054 1 .172 2 11 22 3 n .056 1 .174 1 9 4 2 1 .062 « 11 2 .177 60 a 2 6 1 1 13.235 1 .194 1 1 1 9 .237 1 n .195 2 .239 2 .199 9 2 1 .245 13 14.209 1 a .247 5 .213 3 .249 1 .220 I 1 .253 72 .222 1 .255 2 1 2 2 3 .224 3 1 .257 1 .228 1 4 .261 41 .230 2 2 .263 3 2 .232 2 .265 1 .234 9 3 8 3 .266 1 .236 1 .268 a 1 .240 1 1 .274 3 .243 1 .276 2 .247 2 13. 320 1 1 7 3 14.303 1 1 .328 2 1 .314 9 1 .330 4 .318 10 4 2 43 5 .334 1 .323 1 3 1 .338 17 .326 13 2 23 73 FISHERY BULLETIN: VOL. 69, NO. I Appendix Table 5.— Numbers and kinds of larvae of Gempylidae-Trichiuridae obtained in EASTROPAC I collections. 03 3 « IS i K "E m s § .1 u m B o c & 03 3 a "a E 01 3 C S 03 u a m s 1 u 2 o D3 ta 3 1 03 3 £ 03 a o ta u < 13 £ O 5 < Z 1 a S 3 i 11.056 1 13.107 1 .064 1 .119 1 3 .072 1 .137 2 11.114 1 .139 1 .138 1 .147 1 e .140 1 .153 1 .146 1 .159 2 8 .158 1 .167 1 fl .159 1 .171 1 ( 11.213 1 .173 1 .219 1 .175 I .228 1 .179 3 .234 1 .187 2 .295 2 .191 1 .297 2 13.235 1 11.318 5 .245 1 .320 1 .280 1 .324 1 .326 2 14.001 1 .010 1 12.004 1 .012 1 .014 1 .029 1 1 1 .020 1 .031 1 .047 2 .095 .081 1 14.122 12.115 1 .123 .118 6 1 .124 1 1 .120 1 .126 1 .144 1 .127 3 .150 3 .128 6 .152 1 .130 3 .158 1 .131 1 .188 1 .134 1 1 12.246 2 .138 3 .260 1 .142 2 .262 1 .146 2 .272 1 .150 1 .276 1 .164 1 .188 1 13. 048 1 2 .194 2 .054 17 .195 1 .056 2 14.222 1 .071 6 e .224 1 .073 7 .234 8 1 .075 2 .240 1 .077 3 .259 3 .081 8 .280 1 4 1 .083 1 .283 1 .095 7 .287 1 2 .097 1 4 .295 1 .101 1 6 14.318 1 2 .103 7 .326 1 .105 2 .330 1 74 AHLSTROM; FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 6.— Numbers and kinds of flatfish (Pleuronectiformes) larvae obtained in EASTROPAC I collections. 5 t- w 1 t3 w a, en 1 C3 a £ U O U W Cfi s 12.028 1 14.001 1 5 1 35 .030 1 .006 1 2 .031 2 e .008 1 3 .033 4 1 6 6 .010 1 1 1 .035 3 .014 2 4 .045 1 .016 5 1 .017 2 .013 1 1 13.007 1 .020 5 .009 1 .022 1 3 1 .011 1 .024 1 9 .013 5 .027 1 6 1 3 9 .015 1 1 .029 1 5 2 24 .019 6 1 1 25 31 1 .031 1 30 .021 2 2 13 8 .033 1 2 .030 4 .040 1 2 4 .032 8 .047 1 .034 2 4 9 .055 1 2 .036 1 14.164 1 1 .040 1 2 1 3 .174 1 .042 1 1 6 1 .183 1 .054 1 .194 1 13.245 1 .195 1 .251 1 .199 1 .253 4 4 14.209 1 .255 3 1 9 .213 1 .257 1 .220 3 .259 1 1 .228 1 3 .261 1 1 1 .230 1 .263 1 3 8 .232 1 .265 2 1 .234 2 2 3 1 13.318 2 1 1 .236 1 2 .320 2 2 .240 1 .322 5 .259 2 .324 1 .295 1 .326 1 2 14.300 1 .328 1 .303 1 1 1 .334 1 .306 1 .314 1 .318 1 2 1 19 .323 1 3 2 2 .326 1 4 3 .330 1 75 FISHERY BULLETIN: VOL. 69, NO. 1 Appendix Table 7. — Standardized haul factors (SHF) : These factors are used to adjust original counts of larvae to the comparable standard of numbers of larvae in 10 m" of water strained per meter of depth fished. Station SHF Station SHF Station SHF Station SHF Station SHF 11.022 3.06 LI. 156 2.74 11.291 3.46 12.061 3.33 12.192 3.27 11.025 2.87 11.158 3.12 11.293 2.93 12.063 3.27 12.194 3.45 11. 027 2.38 11.159 2.64 11.295 3.16 12.065 3.23 12.196 3.32 11.030 2.47 11.161 3.35 11.297 2.86 12.067 3.36 12.198 3.40 11.032 3.01 11.163 2.64 11.299 3.57 12.068 3.39 12.200 3.18 11.034 3.64 11.167 2.97 11.301 3.31 12.071 3.34 12.203 3.29 11.036 3.04 ] 11.169 3.27 11.303 3.19 12.075 3.33 12.206 3.53 11.038 2.80 11.171 2.92 11.306 3.22 12.077 3.42 12.209 3.51 11.040 3.32 ] 11.173 2.94 11.308 3.15 12.079 3.56 12.212 3.32 11.044 2.81 ] 11.175 3.47 11.310 3.19 12.091 3.53 12.215 3.27 11. 046 3.24 ] 11.177 1.36 11.312 3.42 12.084 3.73 12.218 3.02 11.048 3.08 ] 1.179 3.37 11.314 3.18 12.087 3.86 12.221 3.07 11.050 2.36 ] LI. 181 2.74 11.316 2.84 12.090 3.10 12.224 2.58 11.052 2.86 ] 1.183 2.92 11.318 3.27 12.092 2.55 12.227 2.96 11.054 2.54 ] 1.185 3.19 11.320 3.34 12.094 2.29 12.230 3.72 11.056 2.90 ] 1.187 2.75 11.322 3.01 12.097 3.01 12.233 2.66 11.058 3.28 : 1.189 3.00 11.324 3.02 12.100 2.48 12.235 3.56 11.060 3.15 ] 1.191 3.79 11.326 2.84 12.103 3.28 12.238 3.21 11.062 3.72 ] 1.195 3.11 11.328 2.62 12.106 3.55 12.240 3.22 11.064 3.01 ] 1.197 3.14 12.109 3.39 12.242 3.41 11.066 2.12 ] 1.199 2.46 12.002 3.12 12.112 3.43 12.244 3.36 11.068 2.62 ] 1.201 3.27 12.004 3.02 12.115 3.48 12. 246 3.14 11.070 2.25 ] 1.203 3.09 12.006 3.31 12.118 2.45 12. 248 3.07 11.072 3.43 ] 1.205 3.20 12.008 3.08 12.120 3.46 12.250 2.49 11.076 2.92 ] 1.207 3.65 12.010 3.13 12.122 3.43 12.252 2.33 11.080 2.45 ] 1.209 3.06 12.012 3.17 12.124 3.17 12. 254 3.30 11.084 2.70 ] 1.211 3.39 12.014 3.28 12.126 3.47 12.256 3.26 11.088 3.19 ] 1.213 2.87 12.016 3.17 12.128 3.30 12.258 3.26 11. 094 3.61 ] 1.215 3.13 12.018 3.13 12.130 3.35 12.260 3.51 11.098 1.78 ] 1.217 2.90 12.020 3.12 12.132 3.38 12.262 2.98 11.102 2.72 ] 1.219 3.36 12. 022 3.43 12.134 3.29 12.264 3.38 11.106 1.36 ] 1.221 2.92 12.024 3.11 12.136 3.22 12.265 3.27 11.110 2.95 ] 1.223 3.71 12.026 3.30 12.138 3.38 12.268 3.35 11.114 3.35 ] 1.226 3.05 12.028 3.44 12.140 3.00 12.270 3.36 11.118 4.65 ] 1.228 3.29 12.030 3.44 12.142 3.42 12.272 3.12 11.120 3.68 ] 1.234 3.65 12.032 3.32 12.144 3.20 12.274 3.28 11.124 3.67 ] 1.238 3.41 12.033 3.21 12.146 4.36 12.276 3.34 11.128 2.85 ] 1.242 3.77 12.035 3.35 12.148 3.21 12.278 3.00 11.130 3.80 ] 1.246 3.01 12.037 3.20 12.150 3.14 12.280 3.39 11.132 3.37 ] 1.250 2.77 12.039 3.47 12.152 3.17 12.282 3.58 11.134 3.22 ] 1.254 2.51 12.041 3.42 12.154 3.27 12.284 3.41 11.136 3.24 ] 1.258 2.86 12.043 3.33 12.156 3.28 11.138 3.38 ] 1.262 3.23 12.045 3.35 12.158 3.22 13.001 2.26 11.140 2.77 ] 1.266 2.91 12.047 3.42 12.160 3.49 13.003 2.45 11.142 3.35 ] 1.270 3.69 12.049 3.39 12.162 3.21 13.005 1.42 11.146 3.25 1 1.278 3.09 12.051 3.31 12.164 2.98 13.007 2.42 11.148 2.54 ] 1.282 3.99 12.053 3.27 12.184 3.22 13.009 2.56 11.150 3.45 ] 1.285 3.20 12.055 2.84 12.186 3.22 13.011 3.68 11.152 2.59 ] 1.287 3.45 12.057 3.22 12.188 3.35 13.013 2.29 11.154 3.40 ] 1.289 3.12 12.059 3.41 12.190 3.39 13.015 2.76 76 AHLSTROM: FISH LARVAE IN EASTERN TROPICAL PACIFIC Appendix Table 7. — Standardized haul factors (SHF) : These factors are used to adjust original counts of larvae to the comparable standard of numbers of larve in 10 m' of water strained per meter of depth fished. — Continued. Station SHF Station SHF Station SHF Station SHF Station SHF 13.017 2.16 13.119 2.67 13.249 3.46 14.047 4.10 14. 203 3.15 13.019 1.88 13.121 3.14 13.251 3.46 14. 051 2.93 14. 209 3.23 13.021 2. 12 13.123 3.06 13.253 3.13 14. 055 3.77 14.213 3.26 13.022 2.72 13.125 3.50 13.255 3.58 14. 060 3.58 14. 218 2.87 13.028 1.53 13.127 3.30 13.257 3.68 14. 066 3.81 14.220 3.42 13.030 2.50 13.129 4.01 13.259 3.42 14. 069 3.65 14. 222 3.64 13.032 3.05 13.131 3.64 13.261 1.85 14.076 3.61 14. 224 3.77 13.034 3.21 13.133 3.84 13.263 3.49 14. 078 3.64 14. 228 3.87 13.036 2.34 13.135 2.51 13.265 3.29 14. 081 3.39 14. 230 2.96 13.038 2.25 13.137 2.58 13.266 3.31 14.084 3.86 14. 232 2.70 13.040 2.85 13.139 3.57 13.268 3.47 14.086 3.95 14.234 0.72 13. 042 2.74 13.141 3.36 13.270 3.30 14. 088 3.54 14. 236 2.96 13. 044 2.58 13.143 3.23 13.272 3.06 14. 091 3.08 14. 240 3.43 13. 046 3.08 13.145 3.49 13.274 3.73 14.095 3.87 14. 243 3.55 13. 048 2.71 13.147 3.58 13.276 3.54 14.099 3.70 14. 247 3.52 13.050 3.02 13.149 3.56 13.278 3.16 14.103 3.57 14.251 3.49 13.052 2.91 13.151 3.11 13.280 3.48 14.106 3.68 14.255 3.64 13.054 3.07 13.153 3.25 13.282 3.37 14.110 3.55 14.259 3.54 13.056 2.87 13.155 3.34 13.284 3.36 14.112 3.66 14.263 3.68 13.058 2.75 13.157 3.40 13.318 3.17 14.114 4.84 14.267 3.04 13.060 3.62 13.159 3.00 13.320 2.93 14.115 3.24 14.276 3.47 13. 062 3.15 13.161 3.30 13.322 3.22 14.117 4.29 14. 280 3.56 13.064 2.76 13.163 2.70 13.324 3.12 14.118 4.03 14. 283 3.60 13.065 2.81 13.165 3.22 13.326 3.05 14.120 3.76 14.287 3.53 13.067 2.67 13.167 3.64 13.328 3.15 14.122 3.78 14. 291 3.11 13.069 2.12 13.169 3.25 13.330 3.03 14.123 3.51 14.295 2.28 13. 071 2.61 13.171 3.12 13.332 3.13 14.124 3.38 14.300 3.58 13.073 3.11 13.173 2.80 13.334 2.85 14.126 3.69 14.303 3.48 13.075 3.42 13.175 2.71 13.338 3.02 14.127 3.89 14.306 3.29 13.077 2.72 13.179 2.46 13.340 3.00 14.128 3.66 14.310 2.85 13.079 2.53 13.183 3.39 13.342 3.03 14.130 3.62 14.314 3.60 13. 081 2.75 13.187 3.31 14.131 3.56 14.318 3.51 13.083 3.06 13.191 3.53 14.001 0.99 14.132 3.56 14.323 3.15 13.085 4.11 13.195 3.02 14. 006 2.94 14.134 3.67 14.326 1.51 13.087 2.87 13.199 2.77 14. 008 3.56 14.136 3.47 14.330 3.49 13. 089 2.65 13.203 2.60 14.010 5.83 14.138 3.83 13.091 2.97 13.207 3.31 14.012 3.50 14.142 3.69 13.093 2.87 13.211 3.01 14.014 3.51 14.146 3.75 13.095 2.81 13.215 2.97 14.016 3.28 14.150 3.60 13.097 3.02 13.219 2.44 14.017 4.19 14.154 4.24 13.099 2.64 13.223 3.01 14.018 3.13 14.158 2.45 13.101 2.75 13.227 3.32 14.020 2.89 14.164 1.01 13.103 2.77 13.231 2.42 14.022 3.45 14.172 3.55 13.105 2.77 13.235 3.05 14.024 3.55 14.174 3.57 13.107 2.76 13.237 3.56 14.027 3.55 14.177 3.88 13.109 2.90 13.239 3.51 14.029 2.63 14.183 3.94 13.111 2.88 13.241 3.55 14. 031 2.03 14.188 2.15 13.113 2.85 13.243 3.42 14.033 5.05 14.194 1.57 13.115 3.46 13.245 2.98 14. 040 3.65 14.195 1.39 13.117 2.99 13.247 3.27 14.043 3.53 14.199 1.54 77 SIZE STRUCTURE AND GROWTH RATE OF Euphausia pacifica OFF THE OREGON COAST' Michael C. Smiles, Jr.^ and William G. Pearcy' ABSTRACT Euphaiisia pacifica (Hansen) oflf Oregon has a maximum life expectancy of about 1 year. During this time it grows rapidly to a length of 22-24 mm. Furcilia larvae were found throughout the year but were most abundant during the autumn months. The population density and the proportion of juve- niles was higher within 25 miles of the coast than in offshore oceanic waters. Growth rates off Oregon are about twice those previously reported for this species from other re- gions. Spawning also appears to be later in the year. All these features may be explained by the high primary production which is extended throughout the summer by coastal upwelling and by the lack of wide seasonal fluctuations of water temperatures along the Oregon coast. Euphausia pacifica is one of the most abundant euphausiids in the North Pacific Ocean. Dense populations are found in Subarctic and Transi- tional waters (Brinton, 1962a; Ponomareva, 1963) and off the Oregon coast (Hebard, 1966; Osterberg, Pearcy, and Kujala, 1964 ; Pearcy and Osterberg, 1967). Euphausiids are important food for many marine carnivores (see Mauchline and Fisher, 1969, and Ponomareva, 1963, for reviews) , and Euphausia pacifica is no exception. It is preyed upon by salmon (Ito, 1964), baleen whales (Ne- moto, 1957, 1959; Osterberg et al, 1964), her- ring (Ponomareva, 1963), sardine and mack- erel (Nakai et al, 1957, as cited by Ponomareva, 1963; Komaki,1967),rockfish ( Pereyra, Pearcy, and Carvey, 1969), pasiphaeid and sergestid shrimp (Renfro and Pearcy, 1966), pandalid shrimp (Pearcy, 1970), and myctophid fishes (Tyler, 1970). Studies on the growth of several species of euphausiids are reviewed in the monograph by Mauchline and Fisher (1969). Data on the ' This research was supported by the National Science Foundation (GB-5494) and the Atomic Energy Com- mission (AT (45-1) -1750; RLO 1750-50). ° Formerly, Department of Oceanography, Oregon State University; present address: Biology Depart- ment, State University of New York, Farmingdale, N.Y. 11735. ' Department of Oceanography, Oregon State Uni- versity, Corvallis, Oreg. 97331. growth and life history of E. pacifica are lim- ited. Nemoto (1957) presented some growth data for E. pacifica from the Japanese-Aleutian area. Ponomareva (1963), in her study on the distribution and ecology of euphausiids of the North Pacific, estimated the growth of E. po/- cifica from plankton samples collected during the winter and spring. Lasker (1966) deter- mined the growth of E. pacifica reared in the laboratory. Preliminary growth rates of E. pa- cifica based on some of our data were also pre- sented by Small (1967). Because growth rates are needed to under- stand the ecology and energetics of a species, we undertook this study on the abundance, size structure, and growth rate of E. pacifica off Oregon. COLLECTION METHODS We made a total of 174 collections using 1-m mouth diameter plankton nets between June 1963 and July 1967 at stations located 15, 25, 45, and 65 miles off Newport, Oreg. In addition, 25 collections were obtained from stations 85-285 miles off Newport. These provided samples of E. pacifica for all seasons of the year over a 4-year period. Nets had 0.57 1-mm mesh open- ings and were used with a flowmeter placed in Manuscript received September 1970. FISHERY BULLETIN: VOL. 69, NO. I, 1971. 79 FISHERY BULLETIN: VOL. 69. NO. 1 the mouth to measure the amount of water filt- ered. The first 20 samples were from oblique tows, and the other 154 were from vertical tows. This change to vertical tows was made to ensure equal sampling at all depths throughout a tow. Com- parison of the catches of several oblique and vertical tows taken during the same night indi- cated little difference in the number and size of E. pacifica per unit volume filtei'ed. Because euphausiids may avoid nets in the daytime, all tows were taken during nighttime when visual avoidance would be minimal (Brint- on, 1967) . This is also a period when E. pacifica presumably has migrated into the upper 100 m of the water column. E. pacifica captured in several 6-ft Isaacs-Kidd midwater trawls were measured to see if large eui:)hausiids that were possibly avoiding the small vertical meter net could be captured. There was no indication that the maximum size of trawl-caught was larger than meter net-caught euphausiids. The maximum depth of our tows was usually 200 m. Because Ponomareva (1963) suggested that E. pacifica adults inhabit the 200-500-m layer in their second winter and no longer mi- grate daily to the surface, tows were taken to 1000 m with both the midwater trawls and vertical meter nets. These deeper tows, how- ever, did not contain any larger animals. Twelve vertical meter net samples from de])ths of 200 m or 1000 m to the surface did not show appre- ciable differences in size structure. Therefore, we assumed that a representative sample of the E. pacifica population was caught in the upiier 200 m at night. The entire plankton sample was preserved at sea in neutralized 10 'r Formalin. In the lab- oratory ashore, all euphausiids were removed from each sample unless the number of euphau- siids was large (more than 200 individuals). In such cases the samj^le was usually divided in half with a Folsom plankton splitter (Mc- Ewen, Johnson, and Folsom, 1954), and euphau- siids were sorted from only one-half the sample. Males and females were not differentiated. The length of each individual E. pacifica was measured to the nearest 0.1 mm from behind the eye to the posterior margin of the carapace, and each animal was then assigned to a 0.3-mm size-group. Total lengths (from the posterior of the eye to the tip of the telson) were also measured from randomly selected individuals of various lengths to enable comparisons of our data with those of others. A least squares fit of 146 comparisons gave the equation: Y = 2.54 X + 0.66 where Y = total length and X = carapace length. The variance was 248.19. Our measure- ments are all given as total lengths. RESULTS RECRUITMENT AND ABUNDANCE Although larval E. pacifica occurred during almost all months of the year, definite trends in abundance were evident over the 4-year per- iod ( Fig. 1 ) . Larvae were usually most abun- dant between October and December. During some years recruitment began as early as June and was also prominent in the summer months. No major concentrations of larvae were found during winter or spring. These larval forms of E. pacifica were f urcilia of about 7 mm or less, agreeing with Boden's (1950) size measurements and description of E. pacifica furcilia. Furcilia are found 16-18 days after spawning, usually within the upper 100 m of the water column (Ponomareva, 1963; Brint- on, 1967). Catch curves (Fig. 2) show the average num- ber of different size-groups of E. pacifica col- lected during the entire study. All sizes of E. imcifica were much more abundant i)er m^ in- shore over the continental shelf than in oceanic offshore waters. Individuals larger than 15 mm were rare at station 65 miles or farther offshore. Our finding that larvae were less abundant at offshore than inshore stations agrees with Brinton (1962b), who also noted that E. pa- cifica was more abundant inshore than oflFshore of California. Thus, despite the wide oceanic distribution of E. pacifica, the density of near- 80 SMILES and PEARCV : GROWTH RATE OF Eufhauna farifia 4000 3000 2000 1000 O 3000- 5 2000- (t 1000 g 3000 2000 1000 1000 I I I I I I I I I I I I I I I I I NH-15 I I I I I I I I I I I I I I I I I I I I I I I H I I I I I I I 10 o o oi— I oHo OOP OO P SlSL Us) 13.900 NH-25 o|— 10 0_0 NH-45 X\ r— lO I — I Q I — IQ Q □ Q .J3. TO Q^ 27000 _Q ^-~0 L. n Ql IQ 0. Or- lO O- n^ 1963 1964 oelUl |io| |i2| loil |04| ioeTloal fio] |i2 |o2| M W H MoMi^ lo^lW W M lio| jiF |oi| H M 1965 pa] [iol [IF |o2| |o4| 1966 1967 Figure 1. — Number of furcilia of E. pacifica collected at four stations off Newport, Oregon (NH-15, 25, 45, 65) during 1963-67. "0" indicates no sample taken for that month. • INSHORE {NH-15, NH-25) ALL STATIONS ▲ OFFSHORE (NH-65. NH-> 65) TOTAL LENGTH (mm) Figure 2. — Catch curves : the logarithm of the average number of various sizes of E. pacifica caught per lO'^ m'^ for all samples during the study. shore populations may be considerably higher than offshore populations in the same region. Although inshore tows were generally made only to 50 m and 130 m at the 15- and 25-mile stations respectively because of depth of water, euphausiid abundance at these stations was ap- proximately 10 times greater than at offshore stations. This difference is too great to be ex- plained by the differences in sampling depths even assuming that all euphausiids were con- centrated in the upper 50 m at night. GROWTH RATE The extended spawning season and variability of catches of E. pacifica made interpretation of growth difficult. Three related methods, all based on progressions of size-frequency histo- grams, generally gave similar growth rates (Table 1) and led to the same conclusion: E. pacifica lives for a period of about 1 year and attains a maximum size of about 22-24 mm total length. We tenuously assumed for all these analyses that we sampled the same population, or populations with similar age structures and growth rates. Two illustrations of growth based on monthly 81 FISHERY BULLETIN; VOL. 69. NO. 1 Table 1. — Summary of average growth rate estimated from the progression of modes or means (see Figs. 3 and 4). Year class Recruitmenf month Number monHis followed Growth rotes Modes (Fig- 3 for 1965 and 1966 year classes) Modes (Fig. 4) Mm/month 1963 09 10 1.6 1.9 1.6 1964 10 9 2.0 2.0 1.9 1965 10 8 2.1 2.2 2.0 1966 11 5 2.9 2.S 2.4 1967 03 3 2.6 2.5 2.5 size-fre(iuency histograms of all stations com- bined (Fig. 3) illustrate the increasing modal lengths between December and June for the 1965 and 1966 year classes. Recruitment of small E. pacifica is also obvious during the spring of 1966 and 1967 and also shows a shift in modes with time. The 1963 and 1964 year classes (not shown here) showed similar trends. A modified histogram plot (Fig. 4) was used to show the data for all 4 years and all 4 stations together. The advantage of this method is that one can follow the main modes of different sizes throughout the 4-year period. A disadvantage is that these plots are distorted by the arbitrary constraints that (1) at least 50 individuals per 103 m3 of water within one size-group had to be present for plotting and (2) concentrations above 5000/103 m3 were plotted only as 5000/ 103 m3. All of the years represented in Figure 4 show some similarity. The main recruitment pulses are in the fall and summer, and the max- imum size attained is approximately 22-24 mm length. After about 1 year, late in the second summer or fall, these large individuals dis- appeared from our collections. Interestingly, many of the modes that were composed of small euphausiids during the spring and early summer disappeared or were undiscernible by the fall. Either these individuals were subjected to high- er mortality than the fall recruits or were trans- ported out of the area. Apparently they made no major contribution to the local adult popu- lation. Average lengths of size modes were also calculated for each collection using the com- puter techniques described by Hasselblad (1966). The means were generally close to the values for the modal lengths of various col- lections shown in Figures 3 and 4 and, therefore, are not illustrated here but are given in Table 1. Our estimates of the growth of E. pacifica by all these methods are summarized in Table 1. As expected, estimates are similar for the same year classes. Growth varied from 1.6 to 2.9 mm per month among year classes, averaging about 2.0 mm per month. Growth rates were fastest for young stages. Year-classes 1963 and 1964 had slower average rates (1.6 and 2.0 mm/ month) and were calculated over a longer period. Year-classes 1966 and 1967, on the other hand, were represented for the shortest periods of time and had the fastest average rates (2.9 and 2.6 mm month) . This deceleration of growth at the larger sizes is also apparent in Figure 3 where the growth rate from January to March 1966 was about 3.2 mm/month, while from March to June it was about 2.0 mm/month. Our estimates are biased in several ways. They favored the recruitment pulses of the fall because the smaller modes of young that ap- peared earlier (June through September) did not comprise a good series of modal sequences. Moreover, the modes and means of the smaller sizes of E. pacifica are probably slightly over- estimated since catch curves (Fig. 2) indicate escapement from our nets of individuals below 6 mm. This may cause an underestimation of growth rates. DISCUSSION Generalized growth curves of E. pacifica for three regions of the North Pacific are con- trasted in Figure 5. On the basis of bimodal size-frequency distributions of winter and spring samples, Ponomai-eva (1963) concluded that E. pacifica lives for a period of 2 years. She found predominantly 8 and 14-15 mm indi- viduals in the winter and 12-13 mm (her 1- yearolds) and 19 mm (2-year olds) in the spring. Off Oregon not only were 12-13 mm individuals rare or absent in sirring samples, but also 13-14 mm individuals, the size that Ponomareva would expect to find in the summer and fall, were ab- sent. Moreover, our data, unlike Ponomareva's, show no large seasonal fluctuations of growth with retarded growth of the 13-14 mm sizes 82 SMILES and PEARCY: GROWTH RATE OF Euphamia padfica 3S00- 20CX)- 1000- 400- 100- 10- 0-^ I I I I I I I I I I I I I I I I I OCTOBER 1965 _ \m\^ 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 NOVEMBER 1966 mn 3500- NOVEMBER 1965 n rfT}n->-. r^ rn 1^ - J~ '-. DECEMBER 1966 _ 3500- Id DECEMBER 1965 _ _ \im. 3500- I § I JANUARY 1966 _ _ rniiiirrrh E im^ FEBRUARY 1967 _ ~h-h-r-i 3500- 0- 3500- FEBRUARY 1966 rfTTrmrrm MARCH 1967 "hrhm rrTil il>n 3500- MARCH 1966 APRIL 1967 TffHi m I ITTti n F 0- 3500- APRIL 1966 On MAY 1967 THth F =HTlTTTT^>rTTTnn JUNE 1966 rm ii nmTn — — _ — — Cg(\J(sj I I I I I i I I I T IT tf>O)Or>Jrnio^<0ooO0)OO (\J (NJ OJ TOTAL LENGTH (mm) Figure 3. — Size frequency distributions of E. pacifica from all stations for the 1965 year class (left) and the 1966 year class (right). 83 FISHERY BULLETIN: VOL. 69, NO. 1 361 |0e| 1 10 1 |I2| b2| M |06| |08| 1 10 1 ||2| |02| |04| |06| 1963 ' 1964 ' 1965 1966 Toil lo4l bel 1967 Figure 4. — Size frequency histograms for all stations, 1963-67. Dashed lines are an estimate of average growth of individual year classes. "0" indicates no samples for that month. 22 20 ^'^ § 12 -J 8 6 2 S OUR RESULTS OUR RESULTS >'l I I I I I.I I I I I I M I I I I I I I I I I 6 8 10 12 2 4 6 8 10 12 2 4 6 8 10 I MONTHS I Figure 5. — Comparison of generalized growth curves of E. pacifica. in the summer and fall. Nemoto (1957 and per- sonal communication) believes that E. pacifica grows rapidly, reaching a length of 17-18 mm after 1 year. Many individuals spawn after 1 year and then may continue to live for another year, reaching a maximum of 22 mm after 2 years. We find no convincing evidence, how- ever, for continuation of large adults through a second year. Large euphausiids disappeared from our samples by the winter (Fig. 4). Thus our results indicate a faster growth rate and shorter life cycle than those of Ponomareva and Nemoto for the northwestern Pacific but a similar maximum size. Our growth rates off Oregon averaged 0.065 mm/day for the entire life span, about twice those for the other field studies of E. pacifica. Maximum rates for rap- idly growing juveniles were 0.095 mm /day. These rates are higher than Lasker's (1966) maximum rates for juvenile E. pacifica reared in the laboratory, suggesting that growth in nature may exceed "optimal" conditions in the laboratory. Although our estimates of the growth of E. pacifica are higher than previousl.v reported, they approximate the estimates for .sevei'a! other species of euphausiids. A length of about 22 mm after 1 year was also found by Mauchline ( 1966) for Thysanoessa raschii; by Ruud (1936), Mauchline (1960), and Einarsson (1945) for Meganyctiphanes norvegica; by Einarsson (1945) for Thysanopoda aciitifrons; by Ruud 84 SMILES and PEARC\- : GROWTH RATE OF Eupham,a pa„fica (1932), Bargmann (1945), and Marr (1962) for Euphausia superba; and by Baker (1959) for Enphau^ia triacantha. Most of these species have a maximum life expectancy of 2 years, reproduce each year, and grow slowly during the winter. Other species are known to have a life expectancy of 1 year (Mauchline and Fisher, 1969). Development, growth, and sexual maturity of the same species of euphausiid are known to vary among geographic iropulations (Einarsson, 1945; Nemoto, 1957; Ponomareva, 1963; Mauchline and Fisher, 1969). Mauchline and Fisher (1969) stress that this variability is probably directly related to differences in food and temperature. Hence, the rapid growth of E. pacifica off Oregon may be related to the high productivity of the region and the lack of large seasonal temperature fluctuations in nearshore waters. Small, Curl, and Glooschenko' report high values for primary productivity in the coastal waters off Oregon. Curl and Small' found that standing stocks of chlorophyll-n averaged high- est inshore and steadily decreased offshore. High production and stocks persist through the summer, the upwelling season, in inshore waters, whereas offshore waters have a tyijical summer productivity minimum (Anderson, 1964). Note that those seasonal and inshore-offshore gradi- ents in phytoplankton are correlated in time and place with the spawning of E. pacifica off Ore- gon, mostly inshore and protracted over the summer and fall months. Ponomareva (196?.) believes that phytoplankton is not only im- portant as food for euphausiid larvae, but also may be necessary in the diet for development of reproductive products of E. pacifica. Water temperatures along the Oregon coast are fairly uniform throughout the year and lack the extremes found along the eastern coasts of continents at similar latitudes. Advection of cool water to the surface (upwelling) during the summer and warm water toward shore dur- * L. F. Small, H. Curl, Jr., and W. A. Glooschenko. Seasonal primary production in a region of upwelling. III. Effects of solar radiation and upwelling on daily production. Unpublished MS. ^ H. Curl, Jr., and L. F. Small. MS. ing the winter moderates the usual seasonal variations. Pattullo, Burt, and Kulm (1969) observed that the seasonal range of heat con- tent was twice as large offshore as inshore (with- in 65 miles) of the Oregon coast. The absence of severe winter temperatures may help to ex- plain the rapid growth of E. pacifica through- out the year off Oregon. Conversely the slow and seasonally variable growth of E. pacifica found by Ponomareva (1963) was in the Far Eastern Seas of Asia where temperatures are often lower and where thermal variations are greater. The fact that E. pacifica is the only widespread euphausiid that spawns in the sum- mer, when the phytoplankton bloom was almost over, indicates that this boreal species may be poorly adapted to the cold marginal Far Eastern Seas (Ponomareva, 1963). The main pulses of larvae, hence spawning, of E. pacifica were in the fall, and not in the spring and summer as found by Ponomareva (1963), Nemoto (1957) off Japan, and Barham (1957) in Monterey Bay, Calif. Brinton (per- sonal communication) notes larval recruitment throughout the year off Southern California. The later spawning off Oregon, like the rapid growth, may again be related to the prolonged production cycle caused by upwelling off Oregon and the moderate fall and winter water temper- atures. ACKNOWLEDGMENTS We are grateful to J. Mauchline for his sug- gestions and to T. Nemoto for providing his growth curve for E. pacifica. LITERATURE CITED Anderson, George C. 1964. The seasonal and geographic distribution of primary productivity off the Washington and Oregon coasts. Limnol. Oceanogr. 9(3) : 284-302. Baker, A. de C. 1959. The distribution and life history of Euphaus- ia triacantha Holt and Tatersall. Discovery Rep 29: 309-340. Bargmann, Helens E. 194.5. The development and life-history of ado- lescent and adult krill, Euphausia superba. Dis- covery Rep. 23: 10,3-176. 85 FISHERY BULLETIN: VOL. 69. NO. 1 Barham, Eric George. 1957. The ecology of sonic scattering layers in the Monterey Bay area, California. Ph.D. Thesis, Stanford Univ. 192 p. Univ. Microfilms, Ann Arbor, Mich. Publ. 21, 564. BoDEN, Brian P. 1950. The post-naupliar stages of the crustacean Euphausia pacifica. Trans. Amer. Microsc. Sec. 69(4) : 373-386. Brinton, Edward. 1962a. The distribution of Pacific euphausiids. Bull. Scripps Inst. Oceanogr. Univ. Calif. 8(12) : 51-270. 1962b. Variable factors affecting the apparent range and estimated concentrations of euphausiids in the North Pacific. Pac. Sci. 16(4) : 374-408. 1967. Vertical migration and avoidance capability of euphausiids in the California Current. Limnol. Oceanogr. 12(3): 451-483. EiNARSsoN, Hermann. 1945. Euphausiacea. 1. North Atlantic species. Dana Rep. Carlsberg Found. 27, 1-185. Hasselblad, Victor. 1966. Estimation of parameters for a mixture of normal distributions. Technometrics 8(8) : 431- 444. Hebard, J. F. 1966. Distribution of Euphausiacea and Copepoda off Oregon in relation to oceanic conditions. Ph.D. Thesis, Oregon State Univ., Corvallis. 85 p. Ito, Jun. 1964. Food and feeding habit of Pacific salmon (genus Oncorhynchits) in their oceanic life. Bull. Hokkaido Reg. Fish. Lab. 29: 85-97. Komaki, Yuzo. 1967. On the surface swarming of euphausiid crustaceans. Pac. Sci. 21(4): 433-448. Lasker, Reuben. 1966. Feeding, growth, respiration, and carbon utilization of a euphausiid crustacean. J. Fish. Res. Bd. Can. 23(9) : 1291-1317. Marr, James. 1962. The natural history and geography of the Antarctic krill {Euphausia superba Dana). Dis- covery Rep. 32: 33-464. Mauchline, J. 1960. The biology of the euphausiid crustacean, Meganyctiphaves norvegica (M. Sars). Proc. Roy. Soc. Edinburgh, Sect. B. (Biol.) 67: 141-179. 1966. The biology of Thysanoesaa raschii (M. Sars), with a comparison of its diet with that of Meganyctiphanes norvegica (M. Sars). 7n Harold Barnes (editor), Some contemporary studies in marine science, p. 493-510. George Allen and Unwin Ltd., London. Mauchline, John, and Leonard R. Fisher. 1969. The biology of euphausiids. In Frederick S. Russell and Maurice Yonge (editors), Advances in marine biology. Vol. 7, 454 p. McEwEN, G. F., M. W. Johnson, and Th. R. Folsom. 1954. A statistical analysis of performance of the Folsom plankton sample splitter, based on test observations. Arch. Meteorol. Geophys. Bioklima- tol., Ser. A. 7: 502-527. Nemoto, Takahisa. 1957. Foods of baleen whales in the northern Pa- cific. Sci. Rep. Whales Res. Inst. 12: 33-90. 1959. Food of baleen whales with reference to whale movements. Sci. Rep. Whales Res. Inst. 14: 149-290. Osterberg, Charles, William Pearcy, and Norman Kujala. 1964. Gamma emitters in a fin whale. Nature (London) 204(4962): 1006-1007. Pattullo, June G., Wayne V. Burt, and Sally A. KULM. 1969. Oceanic heat content off Oregon: Its vari- ations and their causes. Limnol. Oceanogr. 14 (2) : 279-287. Pearcy, William G. 1970. Vertical migration of the ocean shrimp, Pa7idalus jordani: A feeding and dispersal mech- anism. Calif. Fish Game 56(2): 125-129. Pearcy, William G., and Charles L. Osterberg. 1967. Depth, diel, seasonal, and geographic vari- ations in zinc-65 of niidwater animals of Oregon. Int. J. Oceanol. Limnol. 1(2): 103-116. Pereyra, Walter T., William G. Pearcy, and Forrest E. Carxtiy', Jr. 1969. Sebastodes flai'idus, a shelf rockfish feeding on mesopelagic fauna, with consideration of the ecological implications. J. Fish. Res. Bd. Can. 26(8) : 2211-2215. Ponomareva, Larisa Natal'evna. 1963. Euphausiids of the North Pacific, their dis- tribution and ecology. Akad. Nauk SSSR Inst. Okeanol. (Translated by Israel Program for Sci- entific Translations, Jerusalem 1966, IPST cat- alog no. 1368, 154 p.) Renfro, William C, and William G. Pearcy. 1966. Food and feeding apparatus of two pelagic shrimps. J. Fish. Res. Bd. Can. 23(12): 1971- 1975. Ruur, John T. 1932. On the biology of southern Euphausiidae. Hvalradets Skr. 2. 105 p. 1936. Euphausiacea. Rep. Dan. Oceanogr. Exped. 1908-1910 Mediter. Adjacent Seas 2D6(Biol.), 86 p. Small, Lawrence F. 1967. Energy flow in Euphausia pacifica. Nature (London) 215(5400): 515-516. Tyi-er, H. R., Jr. 1970. The feeding habits of three species of lant- emfishes (Myctophidae) off Central Oregon. Master's Thesis, Oregon State Univ., Corvallis. 64 p. 86 ESTIMATING PHYTOPLANKTON PRODUCTION FROM AMMONIUM AND CHLOROPHYLL CONCENTRATIONS IN NUTRIENT-POOR WATER OF THE EASTERN TROPICAL PACIFIC OCEAN"° William H. Thomas" and Robert W. Owen, Jr.* ABSTRACT Previous work has shown that nitrogen is the limiting nutrient in poor (nitrate-free) water in the eastern tropical Pacific Ocean and has suggested that ammonium is the principal nitrogen source for phytoplankton in this water. Enrichment and uptake experiments with various concentrations of ammonium have provided values for the half-saturation constant, Kg, and the maximum growth rate, /i^ax' which can be used to calculate actual growth rates with the hyperbolic model relating growth rate to limiting nutrient concentration. At two stations, growth rates calculated from ammonium con- centration agreed well with those calculated from chlorophyll and 14c production, and the hyperbolic equation could be combined with that using production and chlorophyll to calculate production alone. In this paper calculated production rates are compared with those observed from 14c uptake mea- surements for a number of EASTROPAC cruises. The regression between calculated production and observed production is highly significant and the slope is close to 1.0, indicating reasonable agreement, particularly when all of the errors in the calculation, especially in Ks, are considered. The results suggest rather close control of phytoplankton production by the limiting nutrient, ammonium, in these near-surface, nutrient-poor waters. This paper describes how concentrations of a limiting nutrient in sea water and some mea- sure of the standing crop of phytoplankton can be used to estimate phytoplankton production. Estimated production is compared with observed i'*C production, and the two sets of values are shown to agree reasonably well when all the errors in the estimation are considered. The EASTROPAC Expedition series has de- lineated particularly well areas that are rich in nutrients and that are nutrient-poor in the eastern tropical Pacific Ocean. Rich areas in- ' Contribution from the Scripps Institution of Ocean- ography. " This work was part of the research of the STOR (Scripps Tuna Oceanography Research) Program. It is also a result of the EASTROPAC Expedition, a co- operative study of the biological, chemical, and physical oceanography of the eastern tropical Pacific Ocean. The work was supported by National Science Foundation Grant No. GB-8618 to the senior author and by contracts #14-17-0007-963 and #14-17-0007-989 between the Bu- reau of Commercial Fisheries (now the National Marine Fisheries Service) and the Institute of Marine Resources. ' Institute of Marine Resources^ Scripps Institution of Oceanography, University of California, San Diego, La Jolla, Calif. 92037. * National Marine Fisheries Service Fishery-Ocean- ography Center, La Jolla, Calif. 92087. Manuscript received September 1970. FISHERY BULLETIN: VOL 69, NO. I, 1971. elude the Peru Current, the Costa Rica Dome, and an area of equatorial upwelling extending across the EASTROPAC area (from the Amer- ican coast to long 119° W). Poor areas lie to the north and south of the equatorial upwelling zone and to the west of the Peru Current and Costa Rica Dome. Rich and poor near-surface waters were mapped in previous papers (Thom- as, 1969, 1970b) and will be shown in detail in the EASTROPAC Atlas (Thomas, unpub- lished data) . Nutrient values for rich and poor water are also given in Table 1 of Thomas (1970a). Corresponding areal and seasonal changes in the phytoplankton production in this region have been observed and attributed in part to mecha- nisms of nutrient supply (Owen and Zeitzschel, 1970). No accounting has been possible, how- ever, for the variations observed within the nutrient-poor surface layer of the region. Near-surface water in poor areas is especially low in nitrate-nitrogen; this nutrient is gener- ally not detectable (<0.1 /ng-at./liter). Ammo- nium-N is present in concentrations ranging up to 1 /ng-at./liter and organic nitrogen can reach 87 FISHERY BULLETIN: VOL. 69, NO. 1 concentrations of 17 /.ig-at. /liter, but this latter nitrogen source is probably not utilized by phy- toplankton (Thomas, Renger, and Dodson, in press). Prior to EASTROPAC (pre-1967) low ni- trate/phosphate ratios in tropical Pacific poor water suggested that nitrogen was a limiting nutrient although ratios were increased when ammonium was included along with nitrate, and it was suggested that this latter nutrient alle- viated N deficiency (Thomas, 1966). Recent EASTROPAC enrichment experi- ments provided direct evidence for N limitation. Phytoplankton growth occurred in experiments where nutrients were added singly to sea water samples only with N addition, and if N was de- leted from an otherwise complete enrichment, little or no growth resulted (Thomas, 1969, 1970b). The fact that photosynthetic assimi- lation ratios were only slightly (but signifi- cantly) decreased in poor water as compared with rich water testified further to the allevi- ation and control of deficiency by ammonium (Thomas, 1970a). Having established which nutrient is com- monly limiting, one can use a quantitative nu- trient requirement in an appropriate math- ematical model to estimate growth rates (pro- duction) from concentration of the limiting nu- trient. Recent work (Caperon, 1967; Dugdale, 1967) indicates that the best model is hyperbolic: (1) K, + S where /j. is the phytoplankton specific growth rate, Mj is the maximum rate which is un- limited by low nutrient concentration, S is a measured limiting nutrient concentration in sea water, and Kg is the "half-saturation constant" — a nutrient concentration that supports a rate equal to /:tma.x/2. This equation is equivalent to the Michaelis-Menton formulation for enzyme kinetics and was first applied to bacterial growth rates by Monod (1942). Many biological pro- cesses follow the hyperbolic model and since growth is the result of a series of coupled en- zymatic reactions, the hyperbolic model is the model of choice. A previous paper (Thomas, 1970b) provides information on /umax and Kg (for ammonium) from which /i can be calculated. To obtain these values we enriched samples of nutrient-poor Pa- cific sea water from a depth of 10 m with a com- plete mixture of non-nitrogenous nutrients to which various concentrations of ammonium were added. The samples were then incubated in natural light approximating the intensity that would be found at 10 m depth. Growth was es- timated by successive daily measurements of in vivo chlorophyll (Lorenzen, 1966) in each treatment, and rates integrated over a daily peri- od were calculated from the maximum increases in chlorophyll. These rates were plotted against ammonium concentrations to fit a hyperbolic model and values of A',, and /xmax were obtained from the plot. These values and their 95% con- fidence limits are given in Table 1 for two such experiments. Kg values can also be determined from uptake experiments since recent work has shown that A'., values for growth and uptake are equivalent (Eppley and Thomas, 1969). Also included in Table 1 are uptake Kg values obtained by Maclsaac and Dugdale (1969) for nutrient-poor tropical Pacific water. Their val- ues for Vniax. the maximum uptake rate, are not equivalent to /imax ^'id thus are not included Table 1. — Rate parameters for growth and uptake on ammonium in nutrient-poor tropical Pacific sea water. Cruise Station ilM) 95 percent limits ''max 95 percent limits (Doublings/day) EASTROPAC 76 007 1.68 ± 3.28 1.12 ± 0.83 Thomos (1970b) EASTROPAC 76 173 1.47 i: 0.91 1.22 ± 0.27 Thomas (1970b) Thompson 26 IS 0.10 -- - - Maclsoac and Dugdale (1969) Thompson 26 3« 0.5S - ~ ~ Maclsoac and Dugdale (1969) Te Vega 13 651.a 0.62 ~ — ~ Maclsaac and Dugdola (1969) Meon volues 0.88 1.17 95 % limits of mean 1.33 O.U 88 THOMAS and OWEN; ESTIMATING PHVTOPLANKTON PRODUCTION in Table 1. It will be noted that confidence lim- its for Ks values in given experiments are large as is the confidence limit for the mean of all five values which is used in subsequent calculations (see Results and Discussion). This can be at- tributed to lack of precision in measuring either growth or uptake; even in controlled experi- ments with laboratory cultures, A'., values are imprecise (Eppley, Rogers, and McCarthy, 1969; Eppley and Thomas, 1969). The integrated daily growth rate, fi, can also be calculated from ^^C production estimates and chlorophyll concentrations using the following equation: 3'.32 [log,o(/? • chl + Prod ) - logio(/? • chl)] ^ = 1 day (2) as has been done for laboratory cultures by Thomas (1964) and McAllister, Shah, and Strickland (1964). In this equation R is the carbon/chlorophyll ratio; R chl thus is the standing stock of phytoplankton carbon. The constant 3.32 converts logarithms to the base 10 to logarithms to the base 2 and allows /x. to be expressed as doublings of phytoplankton carbon per day. In the previous paper (Thomas, 1970b), ^ calculated from ammonium (equation 1) was compared with /j, calculated from l^c production and chlorophyll (equation 2) for the two EAS- TROPAC stations where Ksand /^max were de- termined from enrichment experiments. At station 76.007, /x calculated from ammonium was 0.385 doublings/day while that calculated from •^■^C uptake and chlorophyll was 0.365 doublings/ day. At station 76.173 both values were iden- tical — 0.276 doublings/day. For the calcula- tion we used an R value of 98, that found by Eppley (1968) for nitrate-free water off La Jolla. This excellent agreement suggested that we could set equation (1) equal to equation (2) and solve for production as a function of ammonium and chlorophyll using A's and fimax ^s constants. The new equation thus derived is /q\ Prod = chl • R antilog [^max L \3.32 3.32 Ks+ S 1 This expression allows a direct comparison cal- culated and measured ^'*C production (see Re- sults and Discussion). METHODS Methods for determining A's and ^ma.x were given previously (Thomas, 1970b; Maclsaac and Dugdale, 1969) — see also the previous sec- tion. Chlorophyll and production samples were taken from the depth of the 50 % light level, which was always in the upper mixed layer and varied from 9 to 16 m. This depth was determined by multiplying the depth at which the Secchi disc disappeared by 0.38. This factor employs the assumption that the Secchi disc disappears at 16 % of surface light in- tensity (Strickland, 1958). Chlorophyll was determined in these samples by filtration on glass fiber filters, followed by 90 % acetone extraction of the filters, and mea- surement of fluorescence of the extract ( Yentsch and Menzel, 1963; Holm-Hansen, Lorenzen, Holmes, and Strickland, 1965) using equations developed by Lorenzen (1966). Simulated in s'itw production was measured by adding 20 ^c Na^^'^COs solution to the samples (Steemann Nielsen, 1952) and incu- bating them in a tubular shipboard incubator space in which natural light intensity was at- tenuated to 50 % of that incident. Incubation was started at noon and continued until sunset at sea surface temperature. Following incuba- tion the samples were filtered through HA Mil- lipore®' filters and their radioactivity assayed ashore by G-M counting of the filters. The l^c solution was standardized by liquid scintillation counting and the efficiency of the G-M counter for these filters was determined by combusting some of these and measuring the evolved ^^C02 with an ionization chamber. Daily uptake was determined by multiplying the activity by 2; we also corrected for the isotope effect by mul- tiplying by 1.05. Darkened samples were incu- bated with illuminated samples and dark uptake was subtracted from light uptake. No cor- ° The use of trade names is merely to facilitate de- scriptions: no endorsement is implied. 89 FISHERY BULLETIN: VOL. 69, NO. I rections for respiration by phytoplankton were made. Ammonium was measured ashore in frozen samples from a depth of 10 m by the method of Richards and Kletsch (1964). Some labile amino-N which is probably available to phyto- plankton is measured along with ammonium by this method. RESULTS AND DISCUSSION For the comparison of calculated and mea- sured ^''C production, we have used samples from 10 m incubated at light intensities approx- imating those at 10 m to determine Ks and fimax. and actual ^^C values from the 50% light level. We did this so that light intensities would not be a factor in the comparison — that is, light was presumed to be at saturating intensities but not inhibitory, which would be the case if surface samples had been incubated in the growth experiments and compared with surface production. Ammonium was not determined at all pro- duction stations, and we selected those pro- duction values where data were available for / / / / / / - Calcu loted Prod. = 1.057 (Observed Prod.)^ / / ^~7 / 10 / / / y / ''' / / / ^ y / / / m / / • X / / / " / y • /^ 5 — • • • • • / / • // • • . // • '/ • * / • //• • . • • • • /^ • • • • • • // • • • /y *• A - • • • • — 1 — i — 1 — 1 , , , , 1 . OBSERVED '*C PRODUCTION {Mg C/m'/Day) 10 Figure 1. — Phytoplankton production calculated from ammonia and chlorophyll con- centrations at 10 m compared with simulated in situ ^^C production at the 50 7o light level in northerly nutrient-poor water in the eastern tropical Pacific Ocean. The dashed line is the regression that would be expected if agreement between the two sets of pro- duction values were perfect. 90 THOMAS and OWEN : ESTIMATING PHYTOPLANKTON PRODUCTION ammonium and where nitrate was undetectable. One hundred and five such production stations were available from 10 EASTROPAC cruises in this nutrient-poor water. Production calculated from equation 3 is com- pared with measured ^*C production in Figure 1. There is a highly significant (P<.01) rela- tionship between the two sets of values. The slope of the regression line is 1.057, which is very near to the value 1.0 which would be ex- pected if agreement were perfect. Nevertheless, there is a large amount of scatter in the values of Figure 1; that is, the calculation overesti- mates in some cases and underestimates in others. Table 2. — Errors in the calculation of production. Parameter Standard errors Reference Chlorophyll ± 12% Holmes, Schaefer, and Shimada (1958) R ± 17% Eppley (1968) ± 6% Table 1 (this paper) S ± 5% Richards and Kletsch (1964) K, ± 76% Table 1 (this paper) Total ± 79% 95% confidence limits ± 152% Errors in the values used in the calculation are given in Table 2. To figure total error these have been converted to variances and summed. The 95% confidence limit shows that any cal- culated production value can vary by ± 1.5 fold. Thus, one would expect quite a large scatter in Figure 1. Most of the error is in Ks. When only the Ks values of Thomas (1970b) are used the cal- culation generally underestimates the observed •''*C production. Use of the mean of the Ks values of Maclsaac and Dugdale (1969) results in an overestimation. Since there is no reason to doubt either set of Kg values, we have used the overall mean Kg from Table 1. In applying this method to any other nutrient-limited waters, it would be well to obtain several values of Kg so that the error due to lack of precision in measuring Kg can be recognized. Part of the scatter in Figure 1 may also be due to the fact that the parameter Kg is species — and temperature — dependent (Eppley, Ro- gers, and McCarthy, 1969) and that variations in species composition of the crop or slight var- iations in temperature may have affected the calculation. The parameters ^max and R are also probably dependent upon the species com- position of the crop and on temperature. Be- cause of these factors, which are unknown, it is perhaps surprising that the relationship be- tween calculated and observed production is so good when constant values of Kg, ^ma.x.and R are used. This evidence supports the hypothesis that phytoplankton production in the upper mixed layer is controlled by the limiting nutrient, ammonium, and shows that the hyperbolic model describes this control very well. In this latter connection it should be noted that if a linear model having a term "S/Smax" in equation 3 (where Smax is that concentration supporting a maximum growth rate and which has a value near 10.0 ^M from the data of Thomas, 1970b) is used rather than the term "S/ (Kg + S)," the calculation very much underestimates the ^^C production. The linear model was used previously by Riley (1963) and Steele (1958) but should now be considered obsolete in view of more recent work using the hyperbolic model. ACKNOWLEDGMENTS We appreciate the assistance of many persons in gathering these data. Ammonium analyses were performed by Mr. Edward Renger, and Mrs. Anne Dodson aided in the determination of /-max and Kg. Sampling and incubation for production measurements and determination of chlorophyll concentrations were carried out by the following: Messrs. Tapuni Mulitauaopele, Michael Kruse, David Justice, James McCarthy, Lawrence Klapow, David Judkins, Gerald John- son, Eric Forsbergh, and Jack Metoyer. Dr. Bernt Zeitzschel and Mr. Michael Kruse helped to process and edit the ^'*C and chlorophyll data. Most of these data were collected aboard the NMFS vessel David Starr Jordan and we appre- ciate the assistance of Capt. C. W. Forster and his crew. 91 FISHERY BULLETIN: VOL. 69. NO. 1 LITERATURE CITED Capeeon. John. 1967. Population growth in micro-organisms lim- ited by food supply. Ecology 48(5) : 715-722. DUGDALE, R. C. 1967. Nutrient limitation in the sea: Dynamics, identification, and significance. Limnol. Oceanogr. 12(4): 685-695. Eppley, Richard W. 1968. An incubation method for estimating the carbon content of phytoplankton in natural sam- ples. Limnol. Oceanogr. 13(4) : 574-582. Eppley, Richard W., and William H. Thomas. 1969. Comparison of half-saturation constants for growth and nitrate uptake of marine phytoplank- ton. J. Phycol. 5(4): 375-379. Eppley, Richard W., Jane N. Rogers, and James J. McCarthy. 1969. Half-saturation constants for uptake of ni- trate and ammonium by marine phytoplankton. Limnol. Oceanogr. 14(6): 912-920. Holmes, R. W., M. B. Schaefer, and B. M. Shimada. 1958. SCOPE measurements of productivity, chloro- phyll "a", and zooplankton volumes. In Robert W. Holmes, and other members of the Scripps Cooperative Oceanic Productivity Expedition, Physical, chemical, and biological oceanographic observations obtained on Expedition SCOPE in the eastern tropical Pacific, November-December 1956, p. 59-68. U.S. Fish Wildl. Serv., Spec. Sci. Rep. Fish. 279. Holm-Hansen, Osmund, Carl J. Lorenzen, Robert W. Holmes, and John D. H. Strickland. 1965. Fluorometric determination of chlorophyll. J. Cons. Cons. Perma. Int. Explor. Mer 30(1): 3-15. Lorenzen, Carl J. 1966. A method for the continuous measurement of in vivo chlorophyll concentration. Deep-Sea Res. Oceanogr. Abstr. 13(2): 223-227. MACISAAC, J. J., AND R. C. DUGDALE. 1969. The kinetics of nitrate and ammonia uptake by natural populations of marine phytoplankton. Deep-Sea Res. Oceanogr. Abstr. 16(1) : 45-57. McAllister, C. D., N. Shah, and J. D. H. Strickland. 1964. Marine phytoplankton photosynthesis as a function of light intensity: A comparison of methods. J. Fish. Res. Bd. Can. 21(1) : 159-181. Monod, J. 1942. Recherches sur la croissance des cultures bacterienne. Hermann et Cie, Paris. Owen, R. W., and B. Zeitzschel. 1970. Phytoplankton production: seasonal change in the oceanic eastern tropical Pacific. Mar. Biol. 7(1) : 32-36. Richards, Francis A., and Richard A. Kletsch. 1964. The spectrophotometric determination of ammonia and labile amino compounds in fresh and sea water by oxidation to nitrite. In Yasuo Miyake and Tadashiro Koyama (editors). 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Bull. 63(2): 273-292. 1966. Surface nitrogenous nutrients and phyto- plankton in the northeastern tropical Pacific Ocean. Limnol. Oceanogr. 11(3): 393-400. 1969. Phytoplankton nutrient enrichment experi- ments off Baja California and in the eastern equatorial Pacific Ocean. J. Fish. Res. Bd. Can. 26(5): 1133-1145. 1970a. On nitrogen deficiency in tropical Pacific oceanic phytoplankton: Photosynthetic param- eters in poor and rich water. Limnol. Oceanogr. 15(3) : 380-385. 1970b. Effect of ammonium and nitrate concen- tration on chlorophyll increases in natural trop- ical Pacific phytoplankton populations. Limnol. Oceanogr. 15(3): 386-394. Thomas, W. H., E. H. Renger, and Anne N. Dodson. In press. Near-surface organic nitrogen in the eastern tropical Pacific Ocean. Deep-Sea Res. Oceanogr. Abstr. Yentsch, Charles S., and D. W. Menzel. 1963. A method for the determination of phyto- plankton chlorophyll and phaeophytin by fluor- escence. Deep-Sea Res. Oceanogr. Abstr. 10(3): 221-231. 92 ECOLOGICAL EFFICIENCY OF A PELAGIC MYSID SHRIMP; ESTIMATES FROM GROWTH, ENERGY BUDGET, AND MORTALITY STUDIES ' Robert I. Clutter^ and Gail H. Theilacker^ ABSTRACT The net ecological efficiency (yield/assimilated) of a population of Metamysidopsis elongata (Crus- tacea, Mysidacea) is estimated to be 32 %. The gross ecological efficiency (yield/ingested) is probably between 19 % and 29 %. Energy use by the field population was calculated from estimates of age specific natural mortality rates and data on growth, molting, reproduction, and respiration. Average growth and molting rates were determined by rearing the mysids in the laboratory. Size specific fecundity was determined from field and laboratory observations. The calorie contents of the mysids, their molts, eggs and larvae were estimated by bomb calorimetry and in part from biochemical composition. The energy used in metabolism was calculated from size specific respiration and data on body composition. Biological systems are organized by the flow of energ-y. Trophic structui-e, numbers of steps in food chains, and numbers of conjunctions in food webs depend on the amount of energy passed through populations to other populations. Energy units provide a means of expressing productivity in terms common to all organisms. The energy produced in the breakdown of biomass by organisms is stored as chemical en- ergy in the pyrophosphate bonds of adenosine triphosphate (Horowitz, 1968). The overall thermodynamic efficiency of this process is sim- ilar in all animals, about 60 to 70 '}t according to Krebs and Romberg (1957). It has been suggested (e.g. Slobodkin, 1961, 1962) that the efficiency of energy transfer between popula- tions of animals is also fairly constant. This efficiency is necessarily of lower order because, for example, there are losses involved in syn- thesizing macromolecules, in continually resyn- thesizing proteins that undergo thermal dena- turation, in transforming foodstuff energy into work energy (about 65 ^r efficiency), and in the degradation of energy during the perform- ' This research was supported in part by NSF Grant GB 7132. ' Formerly of National Marine Fisheries Service Fish- ery-Oceanography Center, La Jolla, Calif. 92037. ^ National Marine Fisheries Service Fishery-Oceanog- raphy Center, La Jolla, Calif. 92037. Manuscript received September 1970. FISHERY BULLETIN: VOL. 69. NO. 1, 1971. ance of work. All energy that passes through a population is either lost as heat or passes on to another trophic level. If one assumes that all mortality is caused by predation, the gross ecological efficiency (Phillipson, 1966) of energy transfer through that population is the ratio of the energy yield in mortality to the energy ingested. Through laboratory studies of growth, molt- ing, reproduction, respiration, body composition, and energy content, we have constructed an energy budget for the pelagic mysid shrimp Metamysidopsis elongata (Holmes). Various aspects of the distribution, behavior, and pop- ulation biology of this species have been de- scribed by Clutter (1967, 1969) and Fager and Clutter (1968). The energy budget data, to- gether with estimates of natural population mortality rates, are used to estimate net and gross ecological efficiencies for the field popu- lation. GROWTH AND DEVELOPMENT Metamysidopsis elongata is a member of the Mysidae, a family that is ubiquitous and often very abundant in most of the neritic zones of the world ocean. This species is free-swimming and occurs in shoals and swarms just above the sand bottom in areas where surf is common 93 FISHERY BULLETIN: VOL. 69, NO. 1 (Clutter 1967, 1969). As is characteristic of mysids, the eggs and larvae are held by the oostegites (brood pouch) of the adult females until they develop into juve- niles that are similar in form to the adults. The juveniles grow by shedding their exoskel- etons (ecdysis) at intervals that become pro- gressively longer until they reach maturity. Males and females develop distinguishable morphological features during the period of rapid growth prior to maturity. Growth be- comes progressively slower after maturity. Al- though there is no evidence that death occurs because of physiological aging, the maximum age observed was about 9 months. Most animals survive less than 3 months in the natural en- vironment . We assume that most of the na- tural mortality is caused by predation, especially by fishes. Some growth experiments have been reported for other species of Mysidae. Blegvad (1922) determined the growth rates of a few individu- als of My sis inernils from first stage juveniles through early maturity. Nouvel and Nouvel (1939) made disjunct determinations of time between molt stages for some size groups of Praunus flexuosis. Nair (1939) observed the time sequence in the egg and larva development of Mesopodopsis orientalis, determined the size and age at liberation, and noted the size at sex- ual maturity of males and females. In his review of growth in some marine Crustacea, Kurata (1960) presented the results of growth studies made by Ishikawa and Oshima on Neo- mysis japonica and by Matsudaira et al. on Gastrosaccus vulgaris. Mauchline (1967) main- tained adult Schistomysis spiritus in the labora- tory, estimated the time they take to attain sex- ual maturity, and estimated the minimum incu- bation time. Considering differences between species, sizes, and environmental temperatures, these reported patterns of development and size increase per molt are compatible with the results of our study. CULTURE METHODS Experimental animals were collected during the day from the middle of their habitat with nets (Clutter, 1965; Fager, Flechsig, Ford, Clutter, and Ghelardi, 1966). They were placed in large (20-50 liter), opaque plastic containei's with covers and transported to the laboratory within 1 to 2 hr after the time of capture. The culture methods were about the same as those described by Lasker and Theilacker (1965) for euphausid shrimps. Individual animals were placed in rectangular clear plastic containers in about 500 ml of sea water. The small con- tainers were partly immei-sed in trays of run- ning sea water. Since the running sea water was pumped continuously into the aquarium from midwater ofl'shore, within the Metamysi- dopsis habitation zone, the laboratory temper- atures ( 14°-20° C) were about the same as those that the animals would have experienced in their natural environment. Animals of both sexes and of several sizes were selected for the e.xperiments. Young ju- veniles were procured by j^lacing pregnant fe- males in containers and recovering the young on the day following their release from the brood pouch, which occurred at night. These young were then placed in separate containers. To determine the incubation time, i.e. the time from fertilization of the eggs to release from the brood pouch as juveniles, pregnant females with known times of fertilization were placed in in- dividual containers so that larval development could be observed. Mysids of all ages were fed freshly hatched nauplius larvae of brine shrimp, (Artemia sa- lina). Twice each week the mysids were re- moved while their containers were emptied of excess food and cleaned with hot fresh water followed by a sea water rinse. They were then provided with excess quantities of fresh nauplii in clean sea water. The containers were examined every day for the presence of molts or, occasionally, carcasses. The molts and carcasses were removed and placed individually in small vials of 5 % Forma- lin for subsequent microscopical examination and measurement. OOGENESIS AND INCUBATION Since Metamysidopsis has a transparent cara- 94 CLUTTER and THEILACKER: PELAGIC MYSID SHRIMP pace and body wall, it is possible to observe the late stages of oogenesis in live animals without dissecting them. The ovary (cf. Nair, 1939, for description) is situated in the interspace be- tween the alimentary canal and the pericardial floor. Its most obvious feature is the pair of larger tubes that lay side by side. It is in these tubes that the eggs to be extruded into the brood pouch are invested with yolk. The process of yolk formation takes about a week in Metamysi- dopsis and is completed just before the female molts and copulates. By observing the ova in these tubes it is possible to estimate the size or age at first reproduction in maturing females, and to count the number of eggs that will be spawned by reproducing females of all ages. Copulation occurs at night within 2 to 3 min after the mature female molts, during which time sperm are passed into the empty brood pouch by the attending adult male. The eggs are subsequently extruded into the brood pouch where they are fertilized. The eggs hatch from the vitelline membrane after 2 to 3 days. Ac- cording to Manton (1928) and Nair (1939) a larval ecdysis occurs in the brood pouch shortly before the larvae are liberated. These late stage larvae have movable appendages and pigmented eyes that show through the transparent ooste- gites of the brooding female. The small quantity of yolk that is present after the larval ecdysis is absorbed, or nearly so, prior to liberation from the brood pouch. After liberation the larvae tend to sink, then, according to Nair (1939), they undergo a sec- ond larval ecdysis after which the statocysts appear and they are capable of swimming. The mysids assume this highly mobile juvenile form within a few minutes after liberation. Although we did not attempt to distinguish sexes of larvae and juveniles, the observations of Nair (1939) indicate that dimorphism is exhibited by the ant- ennules and abdominal appendages even though neither the brood pouch nor the penis is de- veloped. Incubation time was determined in the lab- oratory. Adult females and adult males were observed in an aquarium during molting and copulation. Ten females were caught after be- ing observed in copulo and were placed in sep- arate containers of sea water at the temperature of their natural environment at that time (17°- 19° C). Five of them were removed, at var- ious times, to determine the stages of develop- ment of the young. The remaining five all re- leased their young as juveniles on the tenth day after fertilization. In addition, a large number of nonpregnant adult females were kept in separate containers for various periods up to 157 days. The range of intermolt periods in 218 observations was 5 to 13 days; the median and modal values were both 10 days. There was no obvious temper- ature effect. The adult females molt just before fertilization and just after liberation of the young; therefore, the average incubation time was taken to be 10 days. This is intermediate between incubation times given for Mysidae that live and reproduce at higher and lower temper- atures. Nair (1939) determined the incubation time of Mesopodopsis orientaUs to be 4 days at 25° to 29° C. Mauchline (1967) reports a min- imum incubation time of 3 weeks for Schisto- mysis spiritus at 12.5° C. MOLTING To avoid handling and possible injury of the experimental animals, the growth rates were determined by measuring molts. The molts suf- fered no appreciable decomposition because they were collected on the day following ecdysis. The morphological development of the animals was usually discernable from their molts. But the molts are fragile, split just back of the cara- pace where the animals emerge, and easily stretched out of shape. Therefore, to measure growth it was necessary to measure a part of the molt that always retained its form and bore a consistent relationship to the body length. Uropod-Body Length Relationship The exopod of the uropod (tail fan) was used to estimate the body length of each animal for its previous intermolt period. The uropods were measured from the base (end of last abdominal segment) to the tip, not including spines, which were sometimes broken, with an ocular micro- 95 FISHERY BULLETIN: VOL. 59, NO. I meter, at 27.5 x magnification. The relationship between uropod length and body length was established from a selected ser- ies of 94 animals that had been collected in the field and pi-eserved. The series included ani- mals that ranged in body length from 0.8 mm to 7.2 mm, and included late stage larvae, juveniles, immatures, and adults. Both sexes were in- cluded; there was no difi'erence between sexes in this relationship. The body length was measured from the end of the last abdominal segment (base of uropod) to the anterior edge of the carapace, behind the insertion of the eyestalk. Mysids tend to curl when preserved, and they can be distorted to appear longer if they are stretched when meas- ured. To avoid this we chose specimens that were at most only slightly curved, and measured the length of the arc through the midline of those that had significant curvature, rather than the straight line distance between head and tail. As shown in Figure 1, the relationship be- tween uropod length and body length is linear. The body length is 4.5 times the uropod length. Table 1. — Frequency of molting periods observed for Metamysidopsis in the laboratory. FiGUKB 1. — Relationship between uropod length and body length of Metamysidopsis. Molting Frequency Average intermolt periods were estimated from 414 observations, 146 on males and 268 on females. In many cases several observations were made on the same animal. The maximum period of laboratory survival for a single ani- mal was 157 days, and the maximum number of molts observed for a single animal (not the same Infermo t Period (dc ys) Sex and length 3 4 5 6 7 8 9 10 11 12 13 o c D 1 c o Females 2 3 4 4 4.0 1 3 6 2 4 4 4.3 jr 4 5 3 1 4 4-5 4.6 ai £ 5 3 7 7 1 5 2 5-6 6 6.2 i 6 10 7 9 13 13 28 17 5 8 10 9-10 9.2 " 7 11 8 6 9 16 12 28 9 9 11 10 9.4 Males E 2 11 3 3 3.0 ^ 3 2 II 1 4 4 3.9 JZ ? '^ 2 15 8 1 1 4 4 4.4 ~ 5 6 29 20 4 5 5-6 5.4 m 6 6 II 8 7 3 5 5-6 5.7 animal) was 21. The molting frequency data for animals reared in the laboratory are sum- marized in Table 1. The sex of the juveniles was established after they had grown large enough to develop obvious morphological dif- ferences. Supplementary data on molting frequency in the field population were obtained indirectly. Over a period of 3 days, 1,211 juveniles + im- matures and 2,979 adults were brought into the laboratory late in the day and placed in large aquaria. The following morning all the animals and their molts were collected and counted. Of the juveniles + immatures 218 or 18 % had molted, and of the adults 356 or 12 % had molted. The recii)rocal of the relative number molting is an estimate of molting period. The observed reciprocals were 5.6 for juveniles + immatures and 8.3 for adults. Since these values are mid- way in the ranges shown by laboratory animals (3-8 days for juveniles + immatures and 4-13 days for adults) we assume that the laboratory observations are valid estimates of molting fre- quency in the population as a whole. Although our observations were made from February to October, and the water tempera- tures in the rearing troughs varied from 14° to 20° C, we were unable to detect any obvious effects of temperature or time of year on molting 96 CLUTTER and THEILACKER: PELAGIC MYSID SHRIMP frequency or growth rates. Nouvel and Nouvel (1939) stated that the intermolt period for Praunus flexiiosis is least during the warmest months, and the incubatory period is 15 days in August and 3 to 4 weeks in September. Las- ker (1966) showed that Euphausia pacifica in- termolt ijeriods varied as the water temperature fluctuated, and that the intermolt period was shortened by an artificially produced warm per- iod, but that temperatures above 12° C did not accelerate molting further. Since we do not have evidence to the contrary, we must assume that our laboratory observa- tions on molting frequency provide adequate average values. From the median values given in Table 1 and estimated average growth rates (see below) we have estimated the molting schedules of females and males from juveniles to mature adults as follows: Females: first six molts — 4 days seventh molt — 5 days eighth molt — 6 days ninth molt — 8 days tenth molt and thereafter — 10 days IVIales: first four molts — 3 days fifth to eighth molts — 4 days ninth and tenth molts — 5 days eleventh molt and thereafter — 6 days GROWTH AND MATURATION Evidence of the temporal sequence of growth and maturation can be obtained from following peaks of abundance of size groups in natural populations. We sequentially sampled the my- sids in the field and observed some shifting peaks. But we consider that the results are not very reliable because of temporal changes in age- specific mortality rates (Fager and Clutter, 1968). Therefore, all the age-specific growth estimates presented here were obtained from laboratory studies. Observed Gro'wth A few mysids were reared in the laboratory from fertilized egg to adult. Several were reared from egg through the juvenile stage. In addi- tion, larger numbers of various sizes were col- lected in the field and kept in the laboratory for several molts. The growth data from these animals were combined as shown in Figure 2 (females) and Figure 3 (males). The sexes were separated because the growth and molting rates of males and females are different. As they are shown in Figures 2 and 3, these individual growth curves are simplified and slightly incorrect rep- resentations of true growth, for two reasons. First, the growth of the body integument is represented to be continuous, whereas it actually occurs in discrete increments. Second, the age Figure 2. — Observed growth in length (from molts) of Metamysidopsis females in the laboratory. Figure 3. — Observed growth in length (from molts) of Metamysidopsis males in the laboratory. 97 FISHERY BULLETIN: VOL. 69, NO. I shown is the age of the animal at the time it molted, rathei- than the age at the time that the molted integument was first formed. The pro- cedure for combining the various growth curves of individual animals was to first plot the growth of the animals of known age, and then plot the other gro\vth curves (actual ages unknown) in a manner that showed the least variation from the apparent trend. Some of the apparent variability in growth rates may be attributable to differences in the temperature at which the growth occurred, but we did not detect any obvious temperature efl^ect. Considerable individual variability occurred among animals of the same size or age that were reared simultaneously. Maturation Changes in morphology in relation to size, and known or estimated age, wei'e observed in the molts of animals reared in the laboratory. Ob- servations were made on live females collected from the field population to determine the size at which yolk invested ova first appear in the ovaries. Supplementary observations on the re- lationship between size and body form were made on preserved animals that had been col- lected in the field. There is some evidence from previous samples taken for other purposes (Clutter, 1967, 1969) that the relationship be- tween size and stage of development may vary seasonally. But during the period of observa- tions reported here, this did not appear to be significant. In particular, we wished to determine (1) the size (and subsequently the age) at which males and females were easily distinguishable by their secondary sexual characteristics, (2) the size at the onset of maturity, and (3) the size at which spawning and brooding of eggs and larvae occurs. The external characteristics that most obviously separate males from females of this species are the enlarged oostegites (brood pouches) of the females and the enlarged pleo- pods (abdominal legs) and antennae of the males. There is some variability in the size at which the stages of development occur. Therefore, our estimates are average values. The larvae are released and juvenile form is attained at age 10 days; at this time both sexes are about 1.2 mm long (body length; excluding antennae, eyes, and tail fan). Males exhibit sub-adult morphology when about 3.7 mm long, and be- come mature at 4.3 mm. Females exhibit sub- adult form at 4.0 mm, the ova become infused with yolk at 4.5 mm, and the eggs are extruded into the brood pouch, fertilized, and incubated at slightly less than 5.2 mm. Average Growth in Length Average continuous growth curves were fitted by eye to the combined growth data plotted in Figures 2 and 3. These curves are represented by the lower curves (fine, continuous unbroken lines) in Figure 4 (females) and Figure 5 (males) . These continuous curves represent the size of the molt at the time — days from fertili- zation — that the molt was shed. Actually the integument of the animal had attained that size by the beginning of the intermolt period in question. The true growth of the integument of the average animal is represented by the stair- step pattern, which is based on the molting fre- quency analysis. The broken curved line of con- tinuous growth (Fig. 4 and 5) represents the probable pattern of temporal change in average organic weight of the animal. This curve con- nects the points halfway betw^een the beginnings and endings of the intermolt periods. Since the average sizes at various stages of development were determined, it was possible to estimate the average time schedules of ma- turation and reproduction for females and males on the basis of the growth curves. The average female begins to develop a brood pouch at the seventh molt, 39 days after becoming a fertilized egg. Yolk invested ova begin to be formed at 45 days, during the ninth intermolt period; the ova are extruded into the developed brood pouch and fertilized at the beginning of the tenth in- termolt period, at 53 days; and reproduction can occur at 10 day intervals thereafter. Males and females gi-ow at rates that are in- distinguishable up to the age of about 30 days, even though the juvenile males molt more fre- quently than juvenile females. After that the 98 CLUTTER and THEILACKER: PELAGIC MYSID SHRIMP Figure 4. — Average growth in length of female Meta- mysidopsis in the laboratory. The lower curve (fine continuous line) was fitted to molt size data (Fig. 2). The steps represent changes in integument size. The upper curve (heavy broken line) represents the average size of the animals, assuming that the addition of body tissue is continuous. - alongalion of pleopodi ond or'enna* 1 3B dart I Figure 5. — Average gro%vth in length of male Meta- mysidopsis in the laboratory. The lower curve (fine continuous line) was fitted to molt size data (Fig. 3). The steps represent changes in integument size. The upper curve (heavy broken line) represents the average size of the animals, assuming that the addition of body tissue is continuous. males grow more slowly. The males develop easily recogriized secondary sexual character- istics at an average age of 38 days and become sexually mature after about 48 days. Average age at maturity was estimated from observa- tions of testes and copulatory behavior in the laboratory as well as from external morphology. Average Growth in Weight To estimate gro\vth in terms of energy it is necessary to translate growth in length into growth in dry weight. This growth in dry weight is then translated into growth in organic (ash-free) weight and thereafter into calories. The dry weights of Metamysidopsis of body lengths ranging from 1.9 mm to 6.5 mm were determined. The animals were captured alive, measured, washed very briefly with distilled water, and dried at 60° C in an oven for 24 hr. They were then weighed individually on a Cahn electrobalance immediately after they were re- moved from the oven. The observed relationship between body length and dry weight is shown in Figure 6. Figure 6. — Relationship between body length and dry weight of Metamysidopsis. The equation for the relationship was deter- mined empirically by fitting a straight line to the logarithms of body length and dry weight by the method of Bartlett (1949). The rela- tionship is: log,, (weight) = -5.436 + 2.77 log,, (length) or weight = 0.00436 (length)^ " where weight is expressed in mg and length in mm. It is common to assume that body weight and body volume have a linear relationship, and that body volume is proportional to the third power of length. Therefore dry weight is expected to 99 FISHERY BULLETIN: VOL. 69, NO. 1 be proportional to the third power of body length (Bertalanffy, 1951). The observed relationship does not quite conform to the expected. The re- lationship between body length and body di- ameter appears to be linear (Fig. 7); therefore the body volume must be proportional to the third power of the body length. The observed relationship between weight and length could be the result of orthogonal growth of the ap- pendages, which become progressively larger as the animals mature. From the average length-weight relationship and the average continuous growth in length curves (Fig. 4 and 5) we have calculated the average growth in weight curves shown in Fig- ure 8. The average continuous growth in length curves represented by the heavy broken lines in Figures 4 and 5 were used to calculate growth in weight, because we assume that growth in organic weight is continuous during intermolt periods even though growth of the integument occurs in discrete steps. The estimated growth in weight of males was extrapolated by eye from age 175 days to age 204 days. We do not have laboratory growth estimates for these larger males, but they occurred in the field population. The average dry weight per egg (140 eggs in sample) was 5.5 fig. Larvae weigh slightly less than this because they lose weight through metabolism while in the brood pouch, even though their ash content is slightly higher than that of the eggs. Figure 7. — Relationship between body length and body diameter of Metamysidopsis. Figure 8.— Average growth in dry weight of Meta- mydisopsis females and males in the laboratory. REPRODUCTION Data on reproduction and associated energy use are easier to obtain for Mysidae than for most pelagic invertebrates. The eggs and larvae are carried in the brood pouch of the female, and the incipient eggs can be counted prior to their full development and extrusion because the body walls of the mysids are transparent. In addition, copulation and fertilization can be ob- served in the laboratory, and frequency of preg- nancy among mature females can be observed in the natural population through sequential sampling because all stages live in the same area while gestating as they do when not reproducing. Nevertheless, average reproduction rate in these animals is not easy to assess with absolute certainty. FECUNDITY Minimum Estimate The most straightforward way to estimate fecundity is to collect animals in the field, pre- serve them, and count the number of eggs or larvae carried by females of diff"erent sizes. Figure 9 shows the relationship between body lengths and number of young for 310 females collected in the field at various times during the year. The data include 125 females bearing eggs and 185 bearing larvae; we excluded ani- mals that had obviously lost young during cap- ture and preservation. For both eggs and lar- 100 CLUTTER and THEILACKER: PELAGIC MYSID SHRIM' number roung : 4 9(body langth] - 14 5 numb*r vgo* : S.4[body Unglh) -16.0 . the time of extrusion of the eggs and the esti- mated average age at which they were counted (7 days) was estimated to be about 0.91. The number of brood pouch young per female was adjusted to the equivalent number of eggs ex- truded per female by multiplying the number of young by 1/0.91 = 1.10. The relationship (Fig. 9) then becomes: number of eggs = 5.4 (body length, mm) - 16.0, which is shown in Figure 9 as the upper, dashed line. We consider this to be a minimum estimate of fecundity, because some females that had lost eggs and larvae from the brood pouches during collection and preservation were probably in- cluded, despite our attempt to exclude them. Figure 9. — Relationship between body length and num- ber of brood pouch young (eggs and larvae) of preserved animals that were collected in the field. The lower line (continuous) was fitted to the points by the method of Bartlett (1949). The upper line (dashed) represents the equivalent relationship for newly laid eggs, assuming a brood pouch mortality of 0.013/day (see text). vae, the number of young per female is highly variable. The average relationship between the size of the female and the number of young, calculated by the method of Bartlett (1949), is represented by the straight line: number of young = 4.9 (body length, mm) - 14.5. This estimate of fecundity is not quite cor- rect because it was made from counts of eggs and larvae that were a few days old. Some eggs and larvae apparently are lost from the brood pouch during the incubation period. Therefore, we adjusted the relationship to account for the mortality which occurs during the incubation period. To estimate the mortality during incu- bation, counts were made of the maturing ova in the ovaries of 40 adult females and counts were made of late stage larvae in the brood pouches of 27 females of the same size, collected at the same time. The ratio of mean number of larvae/mean number of ova was 0.90. The larvae were estimated to be 8 days old, giving an instantaneous mortality rate of 0.013/day. The average age of the eggs and larvae from the 310 preserved females (Fig. 9) was esti- mated to be 7 days. Therefore, the relative sur- vival of the young in the brood pouch between Maximum Estimate We observed that the females that had re- leased young during the laboratory experiments had a higher apparent fecundity than those that were collected and preserved in the field. It is possible that there was some bias in selecting animals for the laboratory experiments, but we were not aware of any. The number of young released per female is plotted against the body length of the female for those 17 specimens in Figure 10. The average relationship between K) numbe. oggt -- 5 5 ( bod, lengihl-ll? , ^ "^ 2i \,^ o I 30 ^■^ ^^^-"''''^ £> E IS 3 '^ z -^^^ \ Int^mb*' lo'.a* '- d.Slbody l*nglh ) - 10.4 10 s 1 1 l^ Figure 10. — Relationship between body length and num- ber of young released by experimental animals in the laboratory. The lower line (continuous) was fitted to the points by the method of Bartlett (1949). The upper line (dashed) represents the equivalent relationship for newly laid eggs, assuming a brood pouch mortality of 0.013/day (see text). 101 FISHERY BULLETIN: VOL. 69, NO. 1 body length and number of young, calculated by the method of Bartlett ( 1949) , was: number of young = 4.8 (body length, mm) -10.4. This is represented by the lower, unbroken straight line in Figure 10. This relationship gives estimates of fecundity that are about 1.5 to 2 young per female higher than the relationship calculated from preserved animals. But this is not quite a maximum esti- mate of fecundity because it does not include the reduction from mortality that occurs during incubation. As already demonstrated, we can assume a brood pouch mortality rate of 0.013 per day. The relative survival of young in the brood pouch during the 10 days between the extrusion of eggs and the release of larvae was therefore estimated to be 0.87. The number of young released per female was adjusted to the equiv- alent number of eggs extruded per female by multiplying the number of young by 1/0.87 = 1.15. The relationship (Fig. 10) then becomes: number of eggs = 5.5 (body length, mm) -11.9, which is shown in Figure 10, as the upper, dashed line. This relationship gives estimates of fecundity that are about four eggs per female higher than the minimum estimates calculated from pre- served animals. We consider this to be the max- imum estimate of fecundity. It is the same as that used by Fager and Clutter (1968). COPULATION AND FERTILITY The fecundity estimates given above apply only to the females that engage in copulation and are fertilized. Mature females that are not fertilized apparently extrude some eggs, but only about one-half the usual number. Many observations of copulation were made in the laboratory (Clutter, 1969). It occurs in artificial light as well as in the dark, but only at night, between about 2000 and 2400 hr. It oc- cui-s within only 2 to 3 min after the mature females molt, and apparently only when the fe- male exudes a pheromone to attract adult males of the same species. Ten females were captured immediately after they were observed in copulo and kept in sep- arate chambers for 10 days. Impregnation had been successful and the usual number of eggs were extruded in every instance. Some adult females that molt do not stimulate males to at- tend them. Ten adult females were captured after they had been observed to be unattended by males during molting and recovery. They later extruded only about one-half of the normal number of eggs, which eventually disappeared from the brood pouch, presumably because they were infertile. Therefore, the unfertilized fe- males expended only about half the amount of energy in eggs that the fertilized females ex- pended. Since the mature females are subject to fertil- ization for only a few minutes following molting, and they apparently do not always attract males during the time, copulation does not always oc- cur. Therefore, not all produce young every 10 days. In a large number of field collections during all seasons, the observed fraction of ma- ture females carrying eggs or larvae in their brood pouches varied from 18 ^r to 78 Sr I the mean was 51 '}r . We are not certain of the source of this variability; there is some evi- dence that it could be related to population den- sity (Clutter, 1969) . We have assumed an aver- age value of 50 fr for the purpose of calculating the amount of energy used in reproduction. On the average, mature females extrude the usual number of eggs about one-half of the time, and they otherwise extrude only one-half of the usual number of eggs. Therefore, the effective average fecundity, in terms of energy used in reproduction (but not in terms of the number of viable young produced), is 0.5 + (0.5) (0.5) = 75 % of the fecundity estimated from counts of young produced/female. For the purpose of calculating the amount of energy used in repro- duction the fecundity equations are: minimum — number of eggs = 4.1 (body length, mm) - 12.0 maximum — number of eggs = 4.1 (body length, mm) -8.9 The second of these relationships is used in the ensuing energy budget calculations. 102 CLUTTER and THEILACKER; PELAGIC MYSID SHRIMP RESPIRATION A polarographic oxygen electrode (Kanwish- er, 1959) was used in a closed system to measure the respiration rates of Metamysidopsis. Both temperature and oxygen were recorded contin- uously on a strip chart. The experimental animals were taken from large constant-flow holding tanks (temperature 14°. 17° C) and acclimated overnight at the tem- perature used in the experiments (13.8°-18.1° C) , to avoid the overshoot in oxygen consumption described by Grainger (1956). They were then washed in millipore-filtered seawater, counted, and transferred to previously filtered seawater in the oxygen electrode system. In each experi- ment an attempt was made to use animals of a limited size range. During the run they were held within a 10- ml chamber, baffled at each end with silk screen cloth of 282 /i mesh aperture size. The water in the closed system circulated through this chamber and then past the electrode at a constant rate. The whole system was im- mersed in a temperature-controlled water bath. Oxygen use by bacteria was measured by mak- ing blank runs with the same water both befoi-e and after each test run. Bacterial use amounted to less than 2 ^c . Oxygen consumption by the mysids was corrected for bacterial uptake. The decrease in relative oxygen tension with time was nearly linear in both the blank runs and the test runs. The results of the respiration experiments are shown in Table 2. Observed weight-specific Table 2. — Summary of respiration experiments on Metamysidopsis. Specimens Number Mean dry weight Water temper- ature Weight-specific resp ration rote Uncorrected Corrected^ Juveniles 99 Ml. 003 ° C. 13.8 (ill Oi/mg 7.71 dry wt hr) 7.54 Juvenile ond immature moles and females 176 0,07 18.0 5-40 4.76 " 297 297 132 0-08 0.08 0.14 18.1 18.1 138 5.03 6.78 3.92 4.48 5.93 3.82 Immature females 85 0.28 15.2 1.95 2.46 Males Brooding females 51 27 0.31 0.47 138 13.8 3.60 3.22 3.53 3.16 27 0.66 13.3 2.65 2.59 lOO BO _-«■ ^ : ^ . ""i^j... „,. cadu'*) -' ^^ \ T B = 27wO" -X V E {Boxlan p'ocaduia 1 V. 6- NJ i 30 X S. \^ 10 Figure 11. — Relation between respiration rate of Met- amysidopsis and size at 16° C. The symbol if' rep- resents respiration rate per dry unit weight (R/W). The lines were fitted to the circle points by two sta- tistical pocedures. The x points are values calculated from published data on other species of Mysidae: 1- Neo7nysis americana (RajTnont and Conover 1961) ; 2-Neomysis integer (Raymont, Austin and Linford 1966); 3-Hemimysis labornae (Grainger 1956). respiration rates (/tl Oj/mg dry weight hr) were corrected for the initial percent oxygen satura- tion and for temperature. In correcting for temperature, a Q,o of 1.9 was used (Grainger, 1956). All values were corrected to 16° C, which is about the median of the year-round temperatures that occur in the natural environ- ment of the mysids. The corrected weight-specific respiration data are plotted in Figure 11 on log-log scales. The symbol R' (Conover, 1960) represents the res- piration rate per unit dry weight (R/W). The average relationship between mean dry weight and R' was estimated by two statistical pro- cedures. First, a straight line was fitted to the logarithmically transformed data by the median procedure (Tate and Clelland, 1957). This gave the relationship: or R' = 2.0 H'-o-3« R = 2.0 (^"-ss ^ Corrected for oxygen saturation level and corrected to temperature of 16.0° C by using Qio = 1.9 (Grainger, 1956). where R = respiration rate in fi\ Oj/hr and W = mean dry weight in mg. Second, a straight line was fitted to the logarith- mically transformed data by the method of 103 Bartlett (1949). This gave the relationship: R' = 2.2 W-"-'^- or R = 2.2 f^»-«» Theoretically, the respiration rate is expected to be proportional to the % power of weight. Since our estimates are slightly above (0.68) and slightly below (0.62) the expected value of 0.67, we consider that the % power relationship is the best estimate for Metanujsidopsis and that the best estimate of respiration rate (/nl Oj/hr) is given by the equation: /? = 2.1 1^"" Estimates of weight -specific respiration for three other, somewhat larger, species of Mysidae are compared with Metamysidopsis in Figure 11. The upper four points ("1" on Fig. 11) represents results for Neomysis americana from Ravmont and Conover (1961) that were ad- justed from 4° C or 10° C to 16° C by using a Q,o value of 1.6 that was estimated from their data. The intermediate point is an estimate of the median value oxygen consumption rate cal- culated from 12 determinations on Neomysis integer (Raymont, Austin, and Linford, 1966) that had been adjusted to 16° C by using a Q,o of 1.9 (Grainger, 1956). The lower point was estimated from the results of Grainger (1956) for Hemimysis lamomae. The ranges of values for these three larger species are about the same as the range (1-3 ^il/hr) calculated from the seasonal change data of Raymont et al. (1966) that had been adjusted to 16° C. The estimates for Metamysidopsis and the other three Mysidae FISHERY BULLETIN: VOL. 69. NO. I all lie well above the relationships calculated for marine planktonic Crustacea by Conover (1960) . BODY COMPOSITION AND ENERGY CONTENT To estimate the amounts of energy used in respiration, molting, and reproduction it was necessary to determine the body composition of the mysids, their molts, and their young. For these analyses the animals were captured alive and, within 2 hr, placed in a constant-flow hold- ing tank at 15° to 17° C where they were kept for a short time prior to analysis. BODY COMPOSITION The estimates of body composition of dried animals and molts are summarized in Table 3. The estimates for ash, protein, lipid, carbohy- drate, and chitin are not considered to be accu- rate past the first decimal point. The fractional percentage values are entered so that the sums will equal 100 ^/c. The methods by which these values were determined will be explained item by item. To determine dry weights, the animals were washed very briefly with distilled water while still alive, then were oven-dried to constant weight at 60° C. Materials that were available only in small quantities were weighed on a Cahn electrobalance. Ash Ash content was estimated by incinerating Table 3. — Average composition and energy content of dry Metamysi- dopsis bodies, molts, eggs, and larvae. Tabulated values for composition are %, and for energy content are cal/mg. The sums of % ash, "pro- tein", lipid, carbohydrate and chitin = 100 %. Nitrogen Carbon Ash "Protein"* Lipid Carbo- hydrate Chitin Energy % % % % % % % Cal/mg Body, whole II.5 36.8 12.5 69.0 10.0 1.5 7.0 4.60 Body, organic 13.2 42.0 79.0 11,4 1,6 8.0 5.24 Molt, v^hole 23.5 44 8 30.9 24.3 2.48 Molt, organic 42.S 56 44.0 4.49 Egg, whole 58.0 6.0 35.2 588 7.16 Egg, organic 61.8 37.5 62.5 7.62 Lorva, whole 45.7 66 60.8 28.9 3.7 5.78 Larva, organic — 48.8 65.0 31.0 4.0 6.20 * "protein" may include free omino acids. 104 CLUTTER and THEILACKER, PELAGIC MYSID SHRIMP whole animals or molts in a muffle furnace at 500° C and weighing the residue. Ash determi- nations were made on six samples composed of mixed animals, juveniles, immatures, adult males, and adult females. The samples con- tained from 2.7 to 7.3 mg of dried animals ; the mean ash content was 12.5 % of the dry weight, and the range was 9.4 to 13.3 % . There was no obvious difference between age groups or sexes. This ash content is within the range, but slightly higher than the mean, of values re- ported for other Mysidae: Mysis flexuosa — 16 ^f (Hensen, 1887) and 11.9 '^h (Delff, 1912, quoted by Vinogradov, 1953); Neomysis integer — 7.9 % (Raymont, Austin, and Linford. 1964) ; Siriella aequiremis — 10.2 ^r (Omori, 1969). Molts used for ash determinations were col- lected in the laboratory immediately after they were shed. Two samples, weighing 1.1 and 0.6 mg, composed of molts from a wide size range of mysids of both sexes had ash contents of 44.4 % and 45.7 Sf; the mean was 44.8 %. Lasker (1966) reported a similar value (46 %) for Euphausia pacifica. This high ash content in the molts suggests that a large fraction of the total body ash resides in the integuments of the whole animals. From 10 observations, we have found that the dry weight of the molt is on the average 13 "^r of the dry weight of the animal that sheds the molt. Assuming that the ash content of the molt is the same as the ash content of the integument of the whole animal, we estimate that 47 % of the body ash resides in the integument. Ash content of brood pouch young was esti- mated fi-om a large number of specimens taken from live females. A dry sample of 0.6 mg of newly hatched larvae had an ash content of 6.1 %. A sample of 1.2 mg of late stage larvae had an ash content of 6.6 ',? Ash content of eggs was not determined; we assume that the ash content is slightly less than that of the newly hatched larvae, and we have used a value of 6.0 %. Nitrogen and Carbon Nitrogen content was determined by the micro-Kjeldahl method from three samples of mixed juvenile-adult animals. The dry weights of the samples were 12, 24, and 63 mg, and contained 13.1 %, 11.7%, and 11.2 % nitrogen respectively; the mean was 11.5 % of total dry weight. From a large number of determina- tions, Raymont et al. (1964) found a value of 11.4 % for Neo?nysis integer. Omori (1969) reported 11.0 % for Siriella aequiremis, and Jawed (1969) found 11.9 % for Neomysis rayii. Carbon content was determined with an F and M carbon analyser model 180, described by Lasker (1966). We assume that all organic carbon, including that in chitin, is liberated by this method. Three samples of females, without young, that weighed 0.2 to 0.4 mg, had carbon fractions be- tween 35.6 9f and 38.1 % of dry weight; the mean was 36.8 %. This estimate is intermedi- ate among other values reported for mysids: Lophogaster sp. (family Lophogastridae) — 46.8 % (Curl, 1962a); Neomysis integer — 30.2 % and 29.5 % (Raymont et al., 1964, 1966) ; mixed mysids and euphausids — 40.7 % (Beers, 1966); Siriella aequiremis — 42.4 (Omori, 1969). From his analysis of several kinds of arthropods, Curl (1962a) found an average of about 38 ':? of the dry weight as carbon. He points out that this is about % of the commonly assumed value of 50 % (Krogh, 1934). In our carbon analysis of molts and young, we found that a 0.2-mg sample of fresh dried molts had 23.5 ^'r carbon, a 0.4-mg sample of eggs had 58.0 % carbon, a 0.4-gm sample of midstage larvae had 47.1 % carbon. The carbon contents of the ash-free organic fractions of the material were calculated from these values. Lasker (1966) found 17 % carbon in the molts of Euphausia pacifica and 50 % carbon in the eggs. Macromolecular Components We assume that the body nitrogen of our spe- cies, Metamysidopsis , is present as protein, free amino acids, and chitin (Raymont, Austin, and Linford, 1968). We made no evaluation of chitin content, but used the value of 7 % de- termined for Neomysis integer by Raymont et al. (1964). The percent "protein" (may include free amino acids) was estimated by the follow- ing relationship, given that 16 % of "protein" 105 FISHERY BLLLETIN: VOL, 69, NO- I is nitrogen, 6.5 "^r of ciiitin is nitrogen, and 7 '/c of the dry body is chitin: 0.16 ("protein") + (0.065) (0.07) = 0.115. From this relation- ship, the "protein" content of the whole dry body was estimated to be 69 %, which is sim- ilar to the value to 71 % protein estimated di- rectly by Raymont et al. (1964) for Neomysis integer. According to the estimates of Raymont et al. (1968) , the percent nitrogen in proteins of Mysidae may be lower than the value of 16 % commonly assumed for animal tissues. They found 13.3 9r N in the body protein of Neomysis integer, and estimated that about 17 ^/c of what we would have designated as "protein" nitrogen was actually free amino acid nitrogen. They suggest that the amino acids may function in osmoregulation for Neomysis integer, which is a euryhaline-brackish water species. We know nothing directly about this for Metamysidopsis. Our species lives in a constant oceanic salinity, and we estimated the ash content to be higher than that of N. integer. Therefore, a high con- centration of free amino acids may not be ne- cessary for osmoregulation in our species. What- ever the ratio of protein/free amino acids may be in Metamysidopsis , our energy calculations should not be affected materially. The lipid content of the mysid bodies was esti- mated by placing samples of dried, crushed bodies successively for 1 hr in each of two 10-ml portions of ethyl alcohol and two 10-ml washes of petroleum ether. The lipid content was esti- mated as the difference in dry weight before and after extraction. Two dry samples of mixed animals, weighing 62.9 mg and 13.4 mg, gave values of 9 % and 11 % lipid respectively. A third sample, containing 24.1 mg of brooding females that had full complements of young in their brood pouches, gave a value of 19 Cr lipid. Linford (1965) found that large females of Neomysis integer carrying young had higher lipid contents than males. From our knowledge of the number of young per female and the esti- mated percent lipid in the young, we calculate that i/i. to 1/2 of the 19 ^r lipid value could be contributed by the brood pouch young. There- fore, we have excluded the 19 ^r value from our estimate, and we have used 10 '/c as the estimate of average lipid content of the dry bodies. This is slightly less than the value of 13 7r estimated for Neomysis integer by Raymont et al. (1964), but within the range of means for three species estimated from a large number of determina- tions by Linford (1965): Mesopodopsis slavveri — 9.0 % ; Neomysis integer- — 10.1 S^ ; Praimus neglectus — • 9.3 %. The carbohydrate content of the mysids was estimated as the amount of macromolecular material remaining after the average estimates for ash, protein, chitin, and fat are subtracted from the dry weight. This remainder is 1.5 %. Apparently the carbohydrate fraction is low in all pelagic Crustacea. Raymont and Conover (1961) found that 1 '? of the dry weight of Neomysis aniericana was glucose; Raymont and Krishnaswamy (1960) found 1.3 ^r carbohy- drate in dry Neomysis integer; and Raymont et al. (1964) found 2.4 9( carbohydrate in dry Neomysis integer. We did no detailed analyses of the composition of molts, but we assume that the molt is com- posed of structural materials rather than energy storage materials. Since we consider that carbo- hydrates and lipids are virtually absent, we entered zero values for them in Table 3. The "protein"/chitin relationship was determined in- directly. First, we estimated the smiount of carbon in the average protein of the mysids from the relationship: (■■f C as protein) = C/c C in body) — (% C as chitin) — (% C as lipid) — (% C as carbo- hydrate) . The percent carbon in the organic fraction of the body is 42 ""f , the chitin fraction is taken as 8 '( , the chitin is assumed to be 50 % carbon (Curl, 1962a), the lipid content of the organic fraction is 11 ""r , the lipid is assumed to be 77 ^c carbon (Lasker and Theilacker, 1962), the car- bohydrate fraction is about 2 % . and the carbo- hydrate is assumed to be 40 % carbon (Curl, 1062a). Therefore, the percent carbon in the 106 CLUTTER and THEILACKER: PELAGIC MYSID SHRIMP mysid protein is calculated as: % C (0.08) (0.50) -(0.11) (0.77) L[0.42 -0.79 — (0.02) (0.40)] = 0.364 = 36.4 % This is considerably less than the average value of 52 '( carbon in protein given by Hawk, Oser, and Summerson (1954), but similar to an esti- mate of 37 % made from the data of Lasker (1966), and higher than an estimate of 23 Sr made from the data of RajTiiont et al. (1964). The second step in finding the relationship between chitin and protein in the molts was to estimate the chitin fraction from the following relationship: (chitin fraction) (9f C in chitin) + (protein fraction) (9f C in pi-otein) = (% C in molt) where chitin fraction + protein fraction 1.0. The chitin fraction calculated from this rela- tionship is 44 % for the organic molt. The protein fraction is therefore estimated to be 56 ^,x . This result suggests that a large fraction of the chitin may be reabsorbed by the animals before molting. This seems reasonable because in Crustacea the new endocuticle is formed dur- ing the intermolt period (between 2 % and 46 % of the time between molts, according to Passano, 1960). To estimate the protein content of eggs and larvae, we have made some arbitrary assump- tions that seem reasonable, and that do not measurably affect our energy calculations in any event. We have assumed that the eggs do not contain a measurable amount of carbohydrate, and that they contain little or no chitin because the integument is not yet formed. Therefore, we have assumed that the organic fraction of the eggs is either protein or lipid. For late stage larvae we have also assumed that carbo- hydrate is absent, but that some chitin is pre- sent because they form integument and molt once before they are released. We have as- sumed that the organic fraction of the larvae contains half the amount of chitin as the adults, or 4 %. The protein-lipid composition of the eggs was calculated from the carbon content of the ash- free fraction. We have estimated (above) that 36.4 % of the mysid protein is composed of carbon, that 77 % of the lipid is carbon, and that 61.8 '~r of the ash-free egg is carbon. By using these values we calculate that the organic fraction of the eggs is 62.5 % lipid and 37.5 % protein. The carbon content of intermediate age brood pouch young (about 5 days old) was less than that of eggs and more than that of late stage larvae. For these intermediate age young we calculate a lipid content of 43 %. ENERGY CONTENT Juveniles - Adults The ash-free calorie content of Metamysidop- sis was determined in a Parr non-adiabatic cal- orimeter. The data, converted to ash-free values, are given in Table 4. Three of the samples contained so little material that Nujol supplement had to be added to raise the heat of combustion to a measurable level. All three of these measurements fell outside the 95 % con- fidence limits of the six determinations made without the Nujol supplement. The variability among the three supplemented determinations can be attributed to the ± 2 % variation of the caloric content of the Nujol supplement (10,791 ± 200 cal/g) , because the weight of the supple- ment greatly exceeded the weight of the sample material in each case. Table 4. — Ash-free' caloric content of Metamysidopsis. Specimens Dry weight Calorie content Ms Cal/i Young juveniles 1.0S 23028.9 Juveniles 2.45 =6462.6 Young females 4.80 =4242.3 Advanced juveniles 12.55 5021.7 Immature males 1 7 JO 5049.0 Immature moles 17.30 5358.0 Mature males 15.75 51238 Mature females 12.40 5185.7 Mature females 17.25 5699.1 ^ Ash content 12.5 % used in all calculations. 2 Nujol supplement used in determinotions. 107 FISHERY BULLETIN: VOL. 69, NO. 1 The mean for the six nonsupplemented sam- ples is 5,240 cal/g (shown as 5.24 cal/mg in Table 3). No significant differences in energy content among developmental stages nor be- tween sexes were found. This mean calorie content estimate is some- what lower than those reported for other Crus- tacea. Slobodkin and Richman (1961) gave values of 5.4 to 5.6 cal/ash-free mg; Lasker (1965) reported a range of 4.9 to 5.4 cal/mg (in- cluding ash) for two species of copepods. Our mean value is also lower than the value that can be calculated from the information on body composition, together with reported average values of the calorie content of animal pi-otein, fat, and carbohydi'ate. Conversion factors given by Horowitz (1968) are: protein, 5.5 cal/mg; fat, 9.3 cal/mg; and carbohydrate, 4.1 cal/mg. Since chitin is glucosamine, we have assumed that it, like carbohydrate, has a calorie content of 4.1 cal/mg. From these conversion factors and the composition data given in Table 3, we calculated an expected value of about 5.77 cal/ash-free mg. We use the empirical value, 5.24 cal/ash-free mg, in our subsequent energy budget calcula- tions. We consider this to be a conservative estimate, because it assumes that the mysid pro- tein has an energy content of only 4.8 cal/mg. This lower than expected estimate may be re- lated to the empirical observation that the mysid protein contains only 36 9^ carbon, rather than about 50 % as is commonly assumed for animal protein. The juvenile and adult Metamysidopsis con- tained 12.5 % ash; therefore, the energy in the whole dry body of an adult or juvenile is esti- mated to be: (4.6 cal/mg) x (dry weight, mg). Molts We estimated the energy content of molts in- directly, because it was difficult to obtain enough material for calorie measurements. The ash- free fraction (55 '.i ) of the molts was estimated to be composed of 44 % chitin and 56 9ir protein. By assuming that chitin has an energy content of 4.1 cal/mg, and that the mysid protein has an energy content of 4.8 cal/mg, we calculate that the ash-free fraction of the molts has an energy content of 4.5 cal/mg. From a sample of 10 animals and their molts we found that the dry weight of molts is on the average 13 % (range 9-19 /r) of the dry weight of the animals that shed them. Lasker (1964, 1966) and Jerde and Lasker (1966) found that the dry molts of a euphausiid were about 10 % of the dry weight of the animals that produced them (range 4-14 %). The energy lost by molting Metamysidopsis is thei'efore proportional to the size of the animal: (0.13) (0.55) (4.5 cal/mg) X (dry weight of animal, mg) or (0.32 cal/mg) x (dry weight of animal, mg). Eggs and Larvae We estimated that eggs were 6 ^r ash, 35 % protein, and 59 % lipid. The energy content of an egg is estimated to be: (0.35) (4.8 cal/mg) + (0.59) (9.3 cal/mg) = 7.16 cal/mg. A sample of 140 eggs was dried and weighed; the mean dry weight per egg was 0.0055 mg. The energy content per egg is tlierefore 0.039 cal- orie. We estimated that, just before being released from the brood pouch, the larvae are about 6 % ash, 61 '"r protein, 29 Sr lipid, and 4 ''/c chitin. The energy content of a late stage larva is esti- mated to be: (0.61) (4.8 cal/mg) + (0.29) (9.3 cal/mg) + (0.04) (4.1 cal/mg) = 5.78 cal/mg. The mean dry weight per larva, esti- mated from 110 individuals, was 0.0051 mg. The energy content per larva is therefore 0.029 calorie. ENERGY BUDGET AND EFFICIENCY OF ENERGY TRANSFER From the data on average growth, age-spe- cific fecundity, respiration rate, and energy con- tent we have calculated cumulative curves of energy use by individual mysids in attaining various stages of development. Data on age- specific natural mortality rates (Fager and Clut- ter 1968) were used to estimate Ix (probability 108 CLUTTER and THEILACKER: PELAGIC MYSID SHRIMP of animal being alive at age x) schedules and average generation time of the field population. The field and laboratory data were combined in an analysis of the efliciency of energy transfer through the Metamysidopsis population to the organisms that feed on them. CUMULATIVE ENERGY CURVES At age zero the egg contains about 0.04 cal. Ten days later, at the time it is released from the brood pouch, the larva contains about 0.03 cal. Thereafter the average calorie content in- creases in proportion to the dry weight (4.6 cal/ mg). The average schedules of energy incor- 16 - U - 1 12 - / 10 - / 5 a / >. 1 c 1 respiration 6 - / / 4 2 " ,^ / y//^^.^^'^ growth 1 1 1 10 30 50 70 90 Aga Idon) Figure 12.— Cumulative energy used by individual Meta- mysidopsis females. The curves are additive, i.e. the space between the lower two curves represents the cumulative energy lost in molts, the next higher space represents energy used to produce eggs (both fertilized and unfertilized), etc. — so that the upper curve repre- sents cumulative energy used for all processes. poration differ between males and females after about 30 days; the rate of incorporation becomes lower and levels off sooner in males. The ac- cumulation of body energy is shown as the low- est curves in Figure 12 (females) and Figure 13 (males). The amount of energy lost in molts varies with age because the size of the molt increases and the molting frequency decreases. Females and males have different cumulative losses of energy from molting because their growth rates are different after age 30 days, and their molting frequencies are different (Table 1.) Although the actual loss of energy in molting occurs at discrete intervals, we have plotted the cumula- tive energy loss as smooth curves, because the accumulation of energy for integument forma- tion probably is continuous. Cumulative energy loss in molting is shown as the second curve in Figure 12 (females) and Figure 13 (males). The cumulative energy curves are additive, i.e. the area between the first curve (body energy) and second curve (molting energy) represents the cumulative energy loss in molts. Figure 13. — Cumulative energy used by individual Meta- mysidopsis males. The curves are additive (see Fig. 12) . 109 FISHERY BULLETIN: VOL. 69. NO. 1 Males use a small amount of energy in pro- ducing sperm, but we assume that this is negli- gible. In females, the ova begin to be infused with yolk about age 45 days. The actual dis- charge of eggs occurs at discrete intervals of about 10 days, beginning at age 53 days. We assume that the accumulation of energy for re- production is more continuous than this, there- fore we have shown reproductive energy use as a smooth curve. The reproduction energy curve shown in Figure 12 is based on the maximum fecundity estimate given previously [number of eggs = 4.1 (body length, mm) — 8.9]. A repro- duction energy curve based on our minimum es- timate of fecundity [number of eggs = 4.1 (body length, mm) — 12.0] would be 0.12 cal (3.1 eggs) lower per spawning. This would make the minimum estimate 72 Sr of the maximum estimate at the age of first spawning (53 days) and progressively higher in percentage there- after, e.g. 85 9f at the age of fifth spawning (93 days). All our reproduction energy cal- culations take into account the observation that, on the average, mature females extrude the usual number of eggs only one-half of the time and otherwise extrude only one-half the usual num- ber of eggs. The amount of energy used in respiration was calculated from the weight-specific respiration equation: R' — 2.1 (dry weight, mg)-"'^^ and from energy conversion factors based on our estimates of body composition. We do not know what substrate Metamysidop- sis catabolizes. The organic fraction of the body is largely protein; the storage ])roduct (carbo- hydrate and lipid) content is low. Raymont and Krishnaswamy (1960) observed that the carbohydrate content of Neomysis integer de- creased slightly, from about 1.30 % (of dry weight) to 1.06 ^(, when a marked reduction in feeding occurred. For the same species, Lin- ford (1965) found no significant change in lipid level whether the animals were starved, fed a lipid-free diet, or fed a high lipid diet. Raymont et al. (1968) asserted that N. integer uses pro- tein as an energy source. We agree with Linford (1965) that it seems likely that the mysids must live largely on their daily ingestion. We think that the food they ingest has composition similar to their bodies. Therefore, our energy calculations assume that they use catabolic substrates in proportion to their presence in the body. This is supported by the results of Jawed (1969). To convert the amount of oxygen used in respiration into the equivalent energy lost as heat we have used the following values for calories lost//il Oq con- sumed (Hawk et al., 1954; Prosser, 1950): protein, 4.5 X 10"^; lipid, 4.7 X lO"'; car- bohydrate, 5.0 X 10 ~^ Therefore, our esti- mate of the average amount of energy used in respiration is about 4.5 X 10""'cal /A O2. The cumulative energy used in respiration is shown as the uppermost curve in Figure 12 (females) and Figure 13 (males). The area between that curve and the next lower curve represents the catabolic heat loss. These res- piration data were calculated for a temperature of 16° C, which was the median temperature of the natural environment of Metamysidopsis. Our respiration measurements were made in flowing water during the daylight hours. There- fore, they represent basal metabolism + energy expended in active swimming. There is some evidence (Clutter, 1969) that the mysids may be less active at night, even though they con- tinue to swim at all times. For this reason we think that the field population may use some- what less than this amount of energy in respir- ation. Our estimated rate of energy loss in catabo- lism is higher than that estimated by Jawed (1969) in his study of nitrogen excretion in Neoviysis rayii. He suggested that protein is catabolized in relatively large quantities, there- fore nitrogenous excretion may provide a good estimate of catabolism. He found an average catabolism of about 2.5 % of body nitrogen per day in adult animals that were probably 8 to 10 mg dry weight, that were held at 10° C. The rate for adult Metamysidopsis of average size (0.6-0.8 mg) was 5 to 6 % of the body energy i)er day. This disparity in catabolism may result from diff'erences between the size and between the environmental temperatures of the two species. Jawed (1969) showed that about 15 % of the nitrogen was excreted as amino acids. We did 110 CLUTTER and THEILACKER: PELAGIC MYSID SHRIMP not investigate this in Metamysidopsis, there- fore, our estimate of total catabolism could be slightly low because it includes only losses of heat energy. NET ECOLOGICAL EFFICIENCY Mortality and Generation Time Estimates of natural mortality in the field population were made during the same period that the laboratory growth experiments were done (Fager and Clutter, 1968). Brood pouch mortality rate was estimated to be 0.013/ day (maximum of 0.017/day). Mor- tality rates for juveniles, immatures, and adults were estimated from consecutive series of field collections. The field mortality rates varied during the year. Survival curves (Ix = proba- bility of being alive at age x) for periods of at least mortality, median mortality, and great- est mortality are shown in Figure 14. The mor- tality rates that we used to calculate these Ix curves are shown in Table 5. The greatest mortality rate results in a declining population; at the median mortality rate the population size remains about constant; and at the least mor- tality rate the population increases. An average female first reproduces at about age 53 days. The generation length for the population is somewhat longer because the fe- males reproduce more than once. The gener- ation length for the field population varied be- tween 67 days and 71 days; the median was 68 days (Fager and Clutter, 1968). Figure 14. — Age specific survival (l^ = probability of being alive at age x) of Metamysidopsis calculated from estimates of greatest, median, and least mortality in the field population (Table 5). Table 5. — Mortality rates (per day) used to calculate Ir schedules for the Metamysidopsis field population. Least Median Greatest Specimens mortality mortality mortality Brood pouch young 0.013 0.013 0.017 Juveniles 0.02 0.06 0.15 Immatures 0.02 0.05 0.14 Adults 0.02 0.04 0.13 Relative Energy Use by Individuals We determined the calories of energy used by average individual female and male mysids, and the fractions used for growth, molting, re- production, and respiration from the estimates of cumulative energy use (shown in part in Figures 12 and 13). The amounts and the per- centage distributions required to reach selected stages of development are shown in Table 6. Table 6. — Energy used by individual Metamysidopsis to reach selected stages of development. Age Energy Relative use Respira- tion Repro- duction Molt, ing Growth Days Cat % % % % Females: Egg yolk production First reproduction Generation i^ = 0.0 1 45 53 68 1103 2.7 4.6 8.7 18.4 52 49 50 55 9 15 19 8 7 7 7 40 35 28 19 Males: Maturity /^ = 0.01 48 1103 3.0 12.8 54 67 10 12 36 21 * Approximate age at which l^ = 0.01 in a nearly stable population (r-».0). Ill FISHERY BULLETIN: VOL. 69. NO. 1 The indicated age at which the probability of being alive reaches 0.01 applies to the stable population (median death rates). The males require less energy to reach ma- turity than females, but relatively more of this energy goes into molting and respiration and less is incorporated. Two-thirds of the energy used in reproduction remains in the population; one-third is lost as unfertilized eggs. The estimates of relative use of assimilated food by Metamysidopsis females during a life span are compared with estimates for a copepod and a euphausid (Corner, Cowey, and Marshall, 1967) in Table 7. The mysids apparently use a fraction of assimiliated energy for growth that is intermediate between the other two species, a lower fraction for metabolism, and a higher fraction for producing eggs. Table 7. — Use of assimilated food by Metamysidopsis females (life span 103 days) compared with the copepod Calanus finmarchicus^ (life span 10 weeks) and the euphausid Euphausia pacifica' (life span 20 months). Assimiloted energy used by Metamysidopsis Assimilated N used by Calanui Assimiloted C used by Euphausia Growth % 19 % 25.3 % 10.1 Metabolism 55 61.4 72.3 Molts 7 0.9 \6.6 Eggs 19 12.4 1.0 * From Corner, Cowey, and Morshcll (1967). - From Losker (1966), revised in Corner et al. Relative Energy Use by the Population The values of relative energy use given in Tables 6 and 7 apply to individuals, or to pop- ulations wherein all members live a full life span. They do not apply to the natural popu- lation, because some die during all stages of growth. We have estimated the relative amounts of energy that would be lost by populations in res- piration, production of infertile eggs, molting, and mortality at the observed minimum, median and maximum mortality rates shown in Table 5. This was done by calculating the fraction of the population that died during each intermolt period (A/^), and multiplying this times: (1) the mean body energy content for the midpoint of that period, (2) the quantity of cumulative energj' lost in infertile eggs \x\) to the midpoint of that period, and (4) the quantity of cumula- tive energj' used in respiration up to the mid- point of that period. The product values for each of these loss categories (mortality, molting, etc.) were then summed over all ages (to Zx ^ 0.001). The relative energy use values were calculated as fractions of the overall sum for all categories combined. We excluded fertilized eggs because this reproduction energy is retained in the population. The age specific distribution of energy use (representing energy loss, because fertilized eggs are excluded) by a population (females and males) of Metamysidopsis at the median mortality rate is illustrated in Figure 15. All the curves are plotted with reference to the base line, zero. The rate of energy loss is low among eggs and larvae, and much higher among the juveniles that have just emerged from the brood pouch and begun to swim. In the larger animals, the respiration per unit weight is low- er, but the respiration per animal is higher, so that the respiration rate per day is highest among the animals that are about 25 days old. The loss of energy per day from all causes is highest among the animals that are about 30 days old. After this the curve declines because the effect of larger size becomes less than the effect of smaller numbers. The estimated relative amounts of energy lost by the population of females, males, and both sexes combined, for each loss category and Ao* Id Figure 15. — Age specific distribution of energy loss by a Metamysidopsis population at the median mortality rate. Production of fertilized eggs is excluded. 112 CLUTTER and THEILACKER: PELAGIC MYSID SHRIMP for each of three mortality rates, are shown in Table 8. The percentages for females and males combined are not quite the same as the means of the separate pei-centages for females and for males. At the minimum death rate 55 S^ of the energy loss would pass through the female half of the population (58 ^'r if fertile eggs are in- cluded). At the median death rate 52 Sr would pass through the females, and at the maximum death rate, 50 %. Table 8. — Relative amount (%) of energy lost by Metamysidopsis populations in respiration, production of infertile eggs, molting, and mortality; at minimum, median and maximum mortality rates. Sex Death rate Respira- tion Infertile eggs Molting Mortality % % % % Females minimum 63.7 6.7 8.6 20,9 median 55.6 3.7 7.7 33.0 maximum 45.4 0.1 6.1 48.4 Moles minimum 67.4 0.0 12.6 20,0 median 58.3 0.0 9.9 31.8 maximum 47.9 0.0 6.5 45,6 Females and minimum 64.5 3.7 10.4 20,5 Males median 56.9 1.9 8.8 32,4 maximum 46.7 0.1 6.3 47.0 If we assume that all the mortality is yield to predators (Odum and Smalley, 1959; Engel- mann, 1961), our mortality fractions are an estimate of net ecological efficiency (energy yield/energy assimilated). Apparently some Crustacea regularly die from natural causes other than mortality (e.g. Daphnia, Slobodkin, 1959). Many mysids of all ages died in our laboratory cultures, but we do not attribute this to senescence. In the field and in the laboratory we observed Metamysidopsis much older than the oldest animals that are involved significantly in our energy calculations. Our best estimate of the net ecological efficiency of the mysid pop- ulation, for transfer of energy to a higher troph- ic level, such as fishes, is about 32 'Jr. The net efficiency of transfer to all trophic levels is 1 — respiration fraction = 43 S^- ASSIMILATION AND GROSS ECOLOGICAL EFFICIENCY Assimilation Efficiency Gross ecological efficiency (energy yield/en- ergy ingested) is the product of net ecological efficiency (energy yield/energy assimilated) X assimilation efficiency (energy assimilated/en- ergy ingested). Therefore, an estimate of as- similation efficiency is required to estimate gross ecological efficiency for the mysid population. We attempted to estimate the assimilation ef- ficiency of Metamysidopsis directly by a carbon- 14 method described by Lasker (1960). This failed because .the mysids did not filter sufficient amounts of radioactive phytoplankton. An ex- periment with another member of the family Mysidae, taken from the same area, was suc- cessful. This gave an estimate of 90 Tr assim- ilation efficiency. Lasker (1966) obtained a similar high value (84 Cf ) for the morphologically similar Eiiphau- siapacifica; and Marshall and Orr (1955) found values greater than 90 % for the copepod Cal- ami^ finmarchicm. In his detailed reviews of assimilation in zooplankton, Conover (1964, 1966) suggests that these values probably are too high. The very large number of observa- tions, many of them his own, that are cited by Conover seem to be evidence that, although var- iable, the mean assimilation efficiency for crus- tacean zooplankton is at least 60 % and perhaps greater. Gross Ecological Efficiency From the information presently available we consider that the assimilation efficiency of the mysids is between 60 9f and 90 ^r. Our best estimate of net ecological efficiency (yield/as- similated) is 32 Sf. Therefore, the minimum estimate of gross ecological efficiency (yield/in- gested) is 19 9r and the maximum estimate is 29 ^c. These estimates are well within the broad range of available estimates of gross ecological efficiency (see reviews by Patten, 1959; Slobod- kin, 1961; Phillipson, 1966; Reeve, 1966), and within the range of 8 '^'r to 30 'c that Engel- mann (1961) considers to be acceptable. They are about 2 to 3 times as high as the median value of 10 % that is suggested by Slobodkin (1961, 1962) , but lower than the values of 30 % to 50 % suggested for marine zooplankton by 113 FISHERY BLXLETIN: VOL. 69, NO. I Ketchum (1962), Steemann Nielsen (1962), and Curl (1962b). LITERATURE CITED Bartlett, M. S. 1949. 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Rapp. Proces-Verbaux Reunions, Cons. Perma. Int. Explor. Mer 153: 178-182. Tate, Merle W., and Richard C. Clelland. 1957. Nonparametric and shortcut statistics. In- terstate Publishers and Printers, Inc., Danville, 111. ix 4- 171 p. Vinogradov, A. P. 1953. The elementary chemical composition of marine organisms. Mem. Sears Found. Mar. Res. 2, xiv -f 647 p. 115 A LINEAR-PROGRAMMING SOLUTION TO SALMON MANAGEMENT' Brian J. Rothschild^ and James W. Balsiger" ABSTRACT A linear-programming model was constructed to allocate the catch of salmon among the days of the salmon run. The objective of the model was to derive a management schedule for catching the salmon which would result in maximizing the value of the landings given certain constraints. These con- straints ensured that cannery capacity was not exceeded, and that escapement of both male and fe- male fish was "adequate." In addition to considering the allocation of the catch in the primal problem, the dual problem considered the shadow prices or marginal value of the various sizes of fish, eggs, and cannery capacity, thus enabling the manager to view his decisions in light of the marginal values of these entities. As an example, the model was applied to a run of sockeye salmon in the Bristol Bay system. In the particular example, which was chosen to replicate the 1960 run, the additional value of the catch owing to optimality amounted to an ex-vessel value of a few hundred thousand dollars. In addition it appeared that the required processing time could be reduced by several days. The op- timum allocation was obtained through conformance to the linear-programming model. The cost of this conformance was not, however, determined. The Pacific salmon fisheries have been cited as an example of irrational conservation (Crutch- field and Pontecorvo, 1969). Much of this ir- rationality is reflected in the dissipation of a sizable fraction of the available economic rent, a situation which results from the open- access nature of the fishery and legislated in- efficiency. The remedy for this situation is to alleviate the open-access and inefficiency problem. Such alleviation would require the dissolution of rather formidable institutional problems. In the present paper, we examine the salmon problem from a slightly different vantage point than Crutchfield and Pontecorvo. We examine the salmon problem under the status quo; we do not consider the optimal amount of gear or its efficiency (this should not, however, be construed as reflecting any diminution in the importance of these prob- lems); rather we consider, as an interim ap- proach, whether it is possible, under the strin- gent condition of knowing in advance the structure of the run, to increase the value of the fish on the dock by optimally allocating the ' Contribution No. 333, College of Fisheries, Uni- versity of Washington. " Center for Quantitative Science and Fisheries Re- search Institute, University of Washington, Seattle, Wash. 98105. ' Fisheries Research Institute, University of Wash- ington, Seattle, Wash. 98105. catch among the days of the run. The traditional approach to salmon manage- ment might be considered, at the risk of several simplifications, as consisting of (1) forecasting the magnitude of the run ; (2) setting an escape- ment goal and a catch implied by the forecast and the escapement; and (3) daily fishing closures and other devices which allocate the catch in varying quantities to the days of the run. The traditional approach, then, also in- volves an allocation of the catch to the days of the run. In the traditional approach, the allocations are usually based on the experience of management biologists. Although the ob- jectives of their allocations are not always clearly and explicitly stated, there is a tendency for the primary objective of management to be simply the attainment of the escapement goal. Our approach is to use the theory of linear pro- gramming to advise on a non-intuitive optimum allocation of the salmon catch among the days of the run where the objective of management does not explicitly involve escapement. Rather, we develop our allocation strategy to maximize the value of the catch on the dock given a va- riety of constraints which include the necessity for a given number of fish to escape the fishery. The objective of maximizing the value of the fish on the dock and the constraints explicitly define the objectives of the management scheme. Manuscript received October 1970. FISHERY BULLETIN: VOL. 69. NO. I, 1971. 117 FISHERY BULLETIN; VOL, 69. NO. I We consider these problems in three additional sections. In the first, we describe the linear- programming allocation model, which we be- lieve to be applicable, with simple modifications, to a variety of salmon management situations. In the second, we consider how the model might be applied to a run of salmon in the Naknek- Kvichak system of Bristol Bay, Alaska. As an example, we choose data from the 1960 run to that system and obtain an optimum allocation of large and small, male and female fish, on each day of the run to the daily catch. This optimum allocation served to maximize the value of the fish on the dock subject to constraints which ensured that the catch did not exceed the daily run, that the catch would be less than the can- nery capacity, and that an "adequate" escape- ment, both in terms of the number of eggs and sex ratio, passed the fishery. Thus, in addition to managing the run by a non-intuitive optimum allocation and satisfying an escapement goal, we also considered the quality of the run in terms of its sex and age composition. In order, how- ever, to achieve this optimum allocation we needed certain data on the structure of the run in advance and we also needed a mechanism by which we could select large and small male and female fish. It would most likely be im- practical to have either a precise prediction of the daily run or an ability to select, with high precision, large or small, male or female fish. We show that even if we had the necessary data, a technique for precise selection of the various entities of fish, and maintained the 1960 escape- ment and sex-ratio conditions, optimum alloca- tion would yield us a catch having a value of several hundred thousand dollars more than the actual catch. Thus given the cost of obtaining the necessary information to perform the op- timum allocation and the constraints extant in 1980, it is questionable whether biological man- agement could yield a better allocation than that which was obtained. This serves to re-empha- size the approach of Crutchfield and Pontecorvo, indicating that the system is most sensitive to variables which lie outside the objective and constraint equations specified in the present paper. On the other hand, our results show that it is possible, at least in terms of the model. to reduce the number of days during which the cannery operates and yet process the same num- ber of fish. Furthermore as previously indicated, we constrained our example to fit the statistics of the 1960 run and thus we had, in our ex- ample, a nearly 1:1 sex ratio; but as we indi- cate later, we could have caught a considerably larger number of male fish and still would have had sufficient male fish in the escapement to en- sure the efficient production of fertilized eggs. And finally the model was quite sensitive to de- creasing the escapement but unfortunately there is little guidance in the literature which would indicate the optimum escapement for the Nak- nek-Kvichak system and furthermore there ap- pears to be little hope of learning the magnitude, in the reasonably near future, of the optimum escapement for the Naknek-Kvichak system. Thus evaluation of the cannery processing time, catch problem, and relaxation of sex ratio and escapement constraints might result in an ad- ded value to the catch which would make some attempts at allocation practical. We also, in the second section, place some stress on in- terpretation of the shadow prices of the var- ious variables in the problem. This is of in- terest to operations researchers because it provides an example, in addition to those con- ventionally used, of an application of the inter- pretation of the linear-programming primal- dual relation. The shadow prices are of interest to the fishery manager because from them it is possible to impute values to the various resources under the manager's control, and, in making a decision, the manager can thus consider these values which, as we show, are not always intui- tively obvious. In the third and final section we conclude the paper with a general discussion of salmon management in a linear-programming setting. MODEL Most linear-programming models generally involve finding values A'j which maximize (or minimize) an objective function i;r,A',-, subject to a set of constraints each of which has the form :^PiXi « Lj, where the inequality can be in either direction or can, in fact, be an equality. 118 ROTHSCHILD and BALSIGER: LINEAR-PRCXSRAMMING SOLUTION The Pi's and the L/s are constants appropriate to a particular problem. The details of the LP (linear-programming) procedure can be found in the many treatises on the subject (e.g., Gass, 1964) or in most texts on operations research (e.g., Hillier and Lieberman, 1967). In our application of the LP model, we max- imize the following objective function A/ N Z = I Z CijXij, (1) where M refers to the total number of age-sex categories and N refers to the days of the run. The variable A',,- is the number of fish caught in the ith entity on the jth day of the run and cij corresponds to the value of the fish caught in the ith entity on the yth day (Table 1). The age-sex category classification results from the fact that salmon runs are comprised of a va- riety of age-groups. Because each age-group is usually of a different average size, the indi- Table 1. — Linear program model notation. Af — The total number of age-sex categories. N — The total number of days in the run. Xij — The number of fish in the t'th oge-sex category which are caught on day ; of the run. C-- — The value of a fish caught in the tth oge-sex category on day y of the run. R — The number of fish in the ith age-sex category which run past the fishery on day ; of the run. Kj — The copocity, in numbers of fish, of the canneries on day ;' of the run, K — The total seosonol copocity of the canneries in numbers of fish, J¥^j — The number of fish of the ith oge-sex category in the escape- ment on day ;' of the run. a — The overage number of eggs in each fish of the ith oge-sex category. T — The total number of eggs contained in the escopement and catch. £ — The minimum number of eggs required in the escapement, ^ — The totol number of moles in the escapement and catch. F — The average fecundity of the female oge-sex cotegories, ex- pressed in number of eggs, f{ — The sex ratio desired in the escapement, expressed as the number of females per mole. /.,- — The number of fish of the ith oge-sex category desired in the season's escapement. S — The number of fish in the totol season run of the ifh oge-sex category, P(j) — The proportion of the run thot arrives by doy ; of the run. P' (j) — The proportion of the run that orrives on day > of the run. viduals in each age-group also have a different average value which we denote by cij. It should be mentioned that size is not the only criterion which can be used for classification. For ex- ample, in the Naknek-Kvichak run of Bristol Bay, the sex of the fish can also be used be- cause within an age-group the male fish tend to be larger than the female fish and thus more valuable in terms of weight of fish-flesh; but, on the other hand, the eggs of the females are a valuable commodity and thus the per-pound value of females may be greater than the per- pound price of males. If the value of the fish were constant during the course of the run, we could replace the C;j with Cj and the allocation problem would become rather uninteresting. But the value, however, does tend to vary dur- ing the course of the run. One reason for this is a deterioration of the quality of fish, as in- dicated by declining oil content and reduction in color intensity with the progression of the run. Another way in which Cy could vary is that the average value of the fish on a par- ticular day would tend to vary during the course of the run because of a within-entity trend in the average size of the fish during the course of the run; this, however, is not considered in the present paper. It is obvious that, if we had sufficient information, we could establish a large number of different c,/s. As indicated previously, equation (1) is max- imized subject to a variety of constraints. For the salmon problem, the first set of constraints is rather obvious and constrains the catch, of any entity, on any day, to be less than, or equal to, the number of fish in that entity in the run. These constraints are of the form -namic programming or apply the outlined procedure in real time to handle the static nature of the programming problem, but unfortunately these approaches will present what can be quite complicated computational difficulties which may, in some instances, be in- surmountable. It is thus clear that we have made certain approximations, trading off real- ism for an easily computable solution which certainly provides management guidance. As we implied previously, we do not consider our departures from realism to seriously affect the utility of the model to provide guidance for decision making. Thus we believe that, for ex- ample, fixing the cannery capacity independent of the entities involved (or we could consider the cannery capacity to be fixed at a level which would accept a reasonable mixture of the en- tities) or using a simple average fecundity of the female entities to represent the average fecundity of the spawning females materially affects our conclusions. These, however, can be evaluated in direct applications by a sensi- tivity analysis. Having outlined some cautions with respect to assumptions, we can now examine some of the indications provided by the various trials of the procedure. These involve the value of the fish on the dock, a reduction in processing-season length, changing value of entities during the run, and finally future data needs. First with respect to the value of the total catch on the dock, we experimented with three value functions which set the daily value of each entity. Using the value functions to determine the value for each entity and day, and the actual distribution of the catch over the 1960 season, a total value of the catch was calculated which corresponds to the use of each of the three value functions. These values of the actual allocation of the catch were compared with the value of the optimal allocation as determined by the linear program as an indication of the value of op- timally allocating the catch over the season. The increased value of the optimally allocated catch ranged from approximately $350,000 to $420,000 dependent on which value-function curve was considered. Table 3 shows these results. In the table, a fourth value function is indicated, which is simply a straight-line function such that the value of each entity remained constant through the season. Each of the other value functions was determined such that the average value of each curve was equal to the constant value for that entity for the season. Table 3. — Comparison of the value of the optional al- location with the value of the actual allocation of the catch for the 1960 season. Value function P Value function 2^ Value function 3^ Value function 4* Optimal allocation Actual allocation $13,787,050 13,378,650 $13,927,860 13,506,250 $13,792,555 13,439,825 $13,517,870 13,517,890 Increased value $ 408,400 $ 421,610 $ 352,730 $ 'i-20 ^ After doy 6, the price dropped 3(f per pound. 2 The price was reduced by subtracting a logistic curve that reduced the price of eoch entity by 3tf per pound over the season. ^ The price was reduced by subtracting a quodratic curve that reduced the price of each entity by 3^ per pound over the season. * The price for each entity remained constant through the season (actual situation.) ^ Difference due to rounding in the linear programming algorithm. All three value functions had the effect of placing emphasis, in the optimal solution, on catching fish on the early days of the season. For tw'o of the functions the value for any entity of fish on a given day is less than the value for that entity on the previous day. This is not true in the step function and thus we do not have a unique allocation, but rather a set of alloca- tions under the high values and a set of allo- cations under the low values. But results are exactly the same; optimal allocations of fish are identical under the three value functions, al- though the total value of the catch changes some- what, according to the exact shape of the value- function curve. Again, we emphasize that these gains from allocation can only be obtained by 135 FISHERY BULLETIN: VOL. 69. NO. 1 knowing in advance of the run the information that we actually used in the allocation and having the ability to select the entities in the run as they are selected in the allocation. Next, an examination of the 1960 optimal allocation reflects that this optimal allocation not only increases the value of the fish on the dock, it also shortens the length of time which a cannery needs to operate. Thus, the same amount of fish could be processed in a shorter period of time, by the same labor force, etc. In the optimal allocation for the 1960 run, all of the fish could have been processed in the first 13 days of the season, 5 days less than the actual operation. Naturally, we need to assume that a policy of catching salmon only from the early part of the run would not aff"ect the genetic constituency of the stock. Furthermore, we must be careful here because, as we have em- phasized in several places, by our LP assump- tions, we cannot, a priori, let the cannery oper- ations on day ;/-l, for example, affect the can- nery operations on day / and we cannot at least in our formulation allow operating at peak ca- pacity to affect quality of the fish or overtime payments since the variables are external to our model. Another indication is that the values of fish change during the course of the season and that these values change in rather subtle ways de- pending upon the "rules" that we set forth (e.g., contrast Figures 8 and 9) and that in the fishery the marginal value of less valuable entities in Table 2 can be greater than the more valuable entities in Table 2. These changes in values need to be acknowledged in any management scheme. Thus, it appears that we have the opportunity to increase the economic efliciency of some salm- on runs. This is, of course, not a new concept, having been treated in some detail by, for ex- ample, Crutchfield and Pontecorvo (1969). Our approach is slightly different, however, in that we have concentrated on oiitimality only from the point of view of increasing the value, as we have defined it, of the fish on the dock. Any full treatment of the management problem must, of course, consider the distribution of fishing eff'ort and its ancillary fishing and economic implica- tions. Now if we accept the premise that conserva- tion is "optimum" allocation of resources in the times-space stream (c.f. Crutchfield and Ponte- corvo, 1969), and if we observe that mathema- tical programming provides guidance for optimal allocation, and note that LP is a special case of mathematical programming, and suggest that the kinds of information required to allocate salmon among the days of the run in an LP model are not going to be much different from the kinds of information required for more so- phisticated programming procedure, then we are led to the conclusion that perhaps we have not addressed ourselves to asking, in our research, the "right questions" concerning salmon man- agement. Following our argument, it would then be implicit that the right questions are con- tained in our formulation of the LP model. These answers must be feasible to obtain and they would contain either needed data or doc- umented policies which would be reflected in the right-hand side of the constraint ecjuations and, more importantly, provide an opportunity for enlightened dialogue. There is unfortu- nately a cost associated with asking right ques- tions. This cost involves the cost of doing new work, or that which inevitably results when ex- isting research activities are reallocated. Are these costs worth the expenditure? These, how- ever, are the kind of questions, the answers to which can be guided by the LP problem. For the salmon management model, we impute values to units of cannery capacity, etc., but, and per- haps of equivalent importance, we impute a val- ue, in meaningful terms, to information. Thus, for our salmon jn'oblem, we have cleverly avoid- ed indicating how we could catch Xij fish for some /,,/. But it is well known that catching can be approximated because it is i)ossible to catch salmon in traps (although this has never been done to any large extent in Bristol Bay) and, upon visual inspection, to distinguish between large and small, male and female fish, and doing this by virtue of ceteris paribus, the allocative process, we could add about 0.5 million dollars to the value of the salmon on the dock. This is, of course, not the full picture, because we would have to trade off the added value of salmon (it 136 ROTHSCHILD and BALSIGER: LINEAR-PROGRAMMING SOLUTION is a common opinion that salmon caught in traps are of better condition and higher vahie than the salmon which are taken by gill netting, for example), the reduction in cannery days used to process the fish, the cost of building traps, and the political problems which are described in some detail in Crutchfield and Pontecorvo (1969). It would not, however, be dithcult to determine the discounted present value of the various alternate ijrocedures and thus evaluate the wisdom of engaging in any. In this eval- uation, we need not be bound by what are per- haps extreme solutions such as traps, but we could examine the value of other selectivity pro- cedures such as modifying gill net selectivity, etc. In general, then, we can evaluate the value of information by approximating that informa- tion, employing it in the model, and contrasting the change in the objective function with the objective function when the information is not in the model. Additional information is needed on the pat- tern of the run. For the earlier years, this is available in Royce (1965), a publication which needs to be updated and implemented to obtain even rough estimates of the temporal movement of the fish of various entities through the fishery. This might be quite difficult to accomplish with present concepts, and the feasibility of a system which would acoustically monitor the passage of salmon through the entire river system and developing a central computer-oriented unit which would process the signals from all acoustic units and provide, in real time, through appro- priate algorithms, rules for catching fish and making observations on escapement is presently being explored. In our model, because of a lack of information, we used the total run and allocated this propor- tionately among the days of the fishery to de- termine the daily run. This emphasizes the need to have, for the purpose of management, a fairly accurate preseason guess of the total magnitude of the run and the Xij's. These guesses are already being made and the predictions need to be judged on the basis of whether the pre- dictions do better than simply averaging the run for cycle years and simply averaging the run for noncycle years and applying these aver- ages as predictions. The trick then may not be to estimate the average catch but rather to de- termine which years are cycle years. We have included cannery capacity in a rather simple way in our model and this is a subject that also needs additional data since the can- nery capacity constraint can be formulated in a variety of ways. It would be interesting to explore in a simulation setting the behavior of the slack variables in the cannery constraint. This is because it seems quite likely that there is a positive correlation between the cost of op- erating a cannery and the magnitude of the slack variable in the cannery constraint. If the run was constant from year to year, then it would be relatively easy to determine an optimal value for the magTiitude of the slack variable in the canneiy constraint. But the run varies con- siderably from year to year, and so in those years when the cannery constraint might be too low, we have an opportunity cost which appears as a slack variable in the dual formulation of the cannery constraint. It would seem then that the best value of the cannery constraint would be somewhere in between the capacity for a maximum run and a minimum run and that this might be investigated by employing the LP model in a simulation setting. We have also emjjloyed egg and sex ratio con- straints in our model. The egg constraints re- quire information on fecundity and escapement. There is not much information on fecundity but this should be either easily obtainable or easily approximated. Again, the static nature of the LP problem makes it difiicult to attribute a val- ue to an egg for years in the future. This is, of course, important, emphasizing the need of thinking not, as is conventionally done, in terms of the forthcoming year, but rather in terms of, for example, a series of years maximizing (c/. Riff'enburgh, 1969) economic benefits. In other words, the utilizers of resources may not be interested (even though they may think they are) in management on a year-to-year basis; rather, they are interested in some long-run sat- isfactory behavior of the time stream of economic benefits. Alternatively, though, we must be cautious of on-the-average management schemes which are typically presented in fishery appli- 137 FISHERY BULLETIN: VOL. 69, NO. 1 cations. This is because a particular manage- ment scheme might be on-the-average quite prof- itable in the long run but might frequently completely bankrupt the system for the first 20 years of operation. The problem of sex ratio is quite important because it appears that the objective function would be quite sensitive to selectively decreasing the number of males in the escapement and thus increasing the catch perhaps substantially. As indicated previously, Mathisen's study (1962) gives us some guidance on this subject and it would appear that, in some instances, the 3:1 ratio might be conservative. Furthermore, it should be mentioned that a year-to-year modi- fication of sex ratio might be a useful cushion for approaching stability for some economic as- pect of the fishery. Finally, the problem of escapement eludes us because in the wealth of literature on the subject there appears to be very little that is useful in setting the egg-min- imum constraint. It is generally agreed that the stock-recruitment relation for salmon is the fa- miliar Ricker-type curve. It is well known that the variability in these relations is quite large (in the case of the Naknek-Kvichak run, at- tempting to draw similarities between stock and recruitment places tremendous stresses on the imagination anyhow) and as a consequence, if the dome-shaped model holds, a minimum escape- ment set sufficiently, but not unreasonably high, could, on the average be reducing the return rather than increasing the return. It might be difficult even after several years of setting the minimum escapement value at too high a level, to detect, owing to the variability in the system, the effect of this policy. If this is true, then again we are asking the wrong questions by studying the stock and recruitment model per se. We are faced with a system that is so variable, either intrinsically or in terms of measurement techniques, or both, that a large number of data points is required before we can evaluate the relation between the empirical data and the theoiy and then use the theory to jjredict. There is but one point a year and so we are asking nature to "stand still" for a large number of years. Given these observations and our past experience, we wonder whether it might not be more appropriate for management purposes to avoid looking at stock and recruitment per se, to intensify study of the physiology and behavior of very young stages of fish, and thus examine fundamental problems of cause and eff'ect, vis- a-vis the variables that influence the magnitude of egg production and survival of these eggs and larvae or other young stages through the first several months of their life. And finally, in the meantime, would it be more appropriate to consider measuring stock and recruitment in terms of transition probabilities which might be estimated by computing the median stock and the median recruitment? Stock sizes which are below the median would be poor, those which are above, good, and similarly with recruitment. The empirical data could then be used to esti- mate probabilities of good-good, good-poor, poor- good, and pooi'-poor transitions. We need not in this procedure be restricted to medians, but could in fact use any fractile, and in fact we need not be restrained by fractiles because we might want to place the dividing line at some "optimal value" and explore the consequences. In conclusion, then, we have formulated a LP model for salmon runs and have shown how it might be related to the Naknek-Kvichak run. We see in this relationship that, given informa- tion on the structure of the run, we can both in- crease the value of the fish on the dock and at the same time reduce processing time. Whether it is worth obtaining the information in terms of the indicated data and the ability to select fish from the run to approach this allocation and whether decreased processing time is, in fact, a saving, are questions that must be an- swered by the processing industry in light of the increased value of salmon on the dock. If our estimate of increased value is approximately correct, we can see that allocation can add an interesting value to the catch, but far greater additions could come from reducing the escape- ment, if this is possible, and alleviating the open- access related problems. Perhaps the most in- teresting feature of the model is the richness of interpretations that LP aflfords in the salmon situation and the nature of questions and data needs raised by the model. Finally, we emi)ha- size that, as Hillier and Lieberman (1967) note. 138 ROTHSCHILD and BALSIGER: LINEAR-PROGRAMMING SOLUTION "A practical problem which completely satisfies all of the assumptions of LP is very rare indeed. However, the LP model is often the most accu- rate representation of the problem, which will yield a reasonable recommendation for action be- fore implementation is required." ACKNOWLEDGMENTS Much of the data used in this paper was un- available in the literature. We obtained un- published information on cannery operations from several members of the salmon industry. Bruce B. Bare was kind enough to advise us on several aspects of the linear-programming tech- nique. We also thank Donald E. Rogers for sup- plying us with unpublished biological data and considerable advice. We appreciate the critical reviews which were given by Robert L. Burgner, Gardner M. Brown, Douglas G. Chapman, and Allan C. Hartt, all of the University of Wash- ington, and we appreciate as well the various suggestions made by anonymous referees. LITERATURE CITED Burgner, Robert L., Charles J. DiCostanzo, Robert J. Ellis, George Y. Harry, Jr., Wilbur L. Hartman, Orra E. Kerns, Jr., Ole A. Mathisen, and William F. ROYCE. 1969. Biological studies and estimates of optimum escapements of sockeye salmon in the major river systems in southwestern Alaska. U.S. Fish WildL Serv. Bull. 67(2): 405-459. Crutchfield, James A., and Giulio Pontecorvo. 1969. Pacific sahnon fisheries: A study of irra- tional conservation. Johns Hopkins Press, Balt- imore, Md., 220 p. Gass, Saul I. 1964. Linear programming. McGraw Hill, Inc., New York, N.Y., 280 p. Hillier, F. S., and G. J. Lieberman. 1967. Introduction to operations research. Hold- en-Day, Inc., San Francisco, Calif., 639 p. Mathisen, Ole A. 1962. The effect of altered sex ratios on the spawn- ing of red salmon. In Ted Swei-yen Koo (ed.), Studies of Alaska red salmon, p. 137-245. Uni- versity of Washington Press, Seattle, Wash. 1969. Growth of sockeye salmon in relation to abundance in the Kvichak district, Bristol Bay, Alaska. Fiskeridir. Skr. Ser. Havunders. 15(3): 172-185. Mathews, Stephen Barstow. 1966. The economic consequence of forecasting sockeye salmon (Oncorhynchus nerka Walbaum) runs to Bristol Bay, Alaska: A computer simu- lation study of the potential benefits to a salmon canning industry from accurate forecasts of the runs. Ph.D. Thesis, University of Washington, Seattle, Wash., 238 p. Ripfenburgh, Robert H. 1969. Stochastic model of interpopulation dynam- ics in marine ecology. J. Fish. Res. Bd. Can. 26(11): 2843-2880. RoYCE, William 6. 1965. Almanac of Bristol Bay sockeye salmon. Univ. Wash., Fish. Res. Inst., Giro. 234, 48 p. 139 FISHERY BULLETIN: VOL. «9. NO- 1 Appendix Table 1. — Constants used in value-function equations for each entity and day of the 1960 run. Function P Function 2- Function 3^ Entity I 1.382 .153 1.365 .153 1.331 .000417 Entity 2 IP2 i2 1.998 .222 1.974 .222 1.925 .000583 Entity 3 1.470 .135 1.455 .135 1.425 .000472 Entity 4 1.964 .186 1.944 .186 1.903 .000694 For entity i on day ;, Value i,j = !Pi. for ; < 6. Value ,-,/ = !Pi - .03X (weight of entity i in pounds), for / > 6. For entity t on day /, Value a = IP: - • (4.5 + .5J) ^ For entity i on day ;, Value i^j = IPi - A,.V'2 Appendix Table 2. — Total run, total catch, and value functions for each entity and day of the 1960 season. Entity Day Total Totol Value Value Value Entity Day Total Total Value Value Value run catch function n function 2- function 3^ run catch function P function 2- function 3^ 1 I 280,351 119,149 1.382 1.362 1.331 3 1 211,080 53,192 1.470 1.453 1.425 2 368,062 156,426 1.382 1,361 1.329 2 277,120 69,834 1.470 1.451 1.423 3 475,118 201,925 1.382 1.358 1.327 3 357,724 90,146 1 470 1.449 1.421 4 600,110 255,046 1.382 1.353 1.323 4 451,832 113,861 1.470 1.445 1.418 5 737,489 313,432 1.382 1.347 1.319 5 555,267 139,927 1.470 1.439 1.415 6 876,389 372,465 1.382 1.337 1.314 6 659,846 166,281 1.470 1.430 1.410 7 1 ,000,833 425,354 1.229 1.324 1.308 7 753,542 189,892 1.335 1.419 1.405 8 1,092.325 464,238 1.229 1.307 1.301 3 822,427 207,252 1.335 1.404 1.398 9 1,134,780 482,281 1.229 1.238 1.293 9 854,393 215,307 1-335 1-387 1.391 10 1,120,032 476,014 1.229 1.270 1.284 10 843,289 212,488 1-335 1-371 1.383 11 1 ,050,967 446,661 1,229 1.253 1.274 11 791,289 199,404 1-335 1-356 1.375 12 940,395 399,668 1.229 1.240 1.264 12 708,038 178,426 1-335 1-345 1.365 13 806,344 342,696 1.229 1.230 1.251 13 607,110 152,992 1-335 1-336 1.355 14 666,514 283,268 1.229 1.224 1.238 14 501,829 126,460 1-335 1.330 1.343 15 534,438 227,136 1.229 1.219 1.225 15 402,387 101,401 1-335 1.326 1.331 16 418,185 177,729 1.229 1.216 1.210 16 314,858 79,344 1-335 1.324 1.318 17 321,003 136,426 1.229 1.215 1.195 17 241,688 60,905 1.335 1.322 1.304 18 242,797 103,188 1.229 1.214 1.178 18 182,805 46,066 1.335 1.321 1.290 2 1 9,590 5,427 1.998 1.970 1.924 4 1 24,985 17,289 1.964 1-941 1.902 2 12,590 7,126 1.998 1.967 1.922 2 32,803 22,700 1.964 1-939 1.901 3 16,252 9,199 1.998 1.963 1.919 3 42,344 29,302 1.964 1-935 1.898 4 20,528 11,618 1.998 1.957 1.914 4 53,484 37,010 1.964 1-930 1.894 5 25,227 14,278 1.998 1.948 1.908 5 65,727 45,483 1.964 1-922 1.888 6 29,978 16,967 1.998 1.934 1.900 6 78,106 54,050 1.964 1-910 1.882 7 34,235 19,377 1.776 1.914 1.891 7 89,197 61,724 1.778 1-894 1.874 8 37,365 21,149 1.776 1.890 1.881 8 97,351 67,367 1.778 1,874 1.866 9 38,817 21,970 1.776 1.863 1.669 9 101,135 69,985 1.778 1.851 1.856 10 38,313 21,968 1.776 1.836 1.856 10 99,820 69,075 1.778 1.828 1.845 11 35,950 20,347 1.776 1.812 1.841 11 93,665 64,816 1.778 1.808 1.832 12 32,168 18,207 1.776 1.792 1.825 12 83,811 57,997 1.778 1.792 1.819 13 27,582 14,479 1.776 1.778 1.808 13 71,864 49,730 1.778 1.780 1.804 14. 22,800 12,905 1.776 1.769 1.789 14 59,402 41,106 1.778 1.772 1-789 IS 18,281 10,347 1.776 1.763 1.769 15 47,631 32,960 1.778 1.767 1.772 14 14,305 8,096 1.776 1.759 1.747 16 37,270 25,790 1.778 1.763 1.754 17 10,980 6,215 1.776 1.756 1.724 17 28,609 19,797 1.778 1.761 1.735 18 8,305 4,700 1.776 1.754 1.700 18 21,639 14,974 1.778 1.760 1.714 ' After day 6, the price dropped 3tf per pound. 2 The price was reduced by subtracting a logistic curve that reduced the price of each entity by 3(f per pound over the season. ^ The price wos reduced by subtracting a quadratic curve that reduced the price of each entity by 3^ per pound over the season. 140 CHEMICAL AND NUTRITIONAL CHARACTERISTICS OF FISH PROTEIN CONCENTRATE PROCESSED FROM HEATED WHOLE RED HAKE, Urophycis chuss David L. Dubrow and Bruce R. Stillings' ABSTRACT This study was to determine whether cooking lean, whole fish before they are extracted by solvent af- fects the chemical and nutritional characteristics of the resulting fish protein concentrate. When red hake were heated at 100° and 109° C for as long as 80 min, the chemical and nutritional properties of the fish protein concentrate were not adversely affected significantly. The nutritional quality was slightly lower, however, in fish protein concentrate produced from red hake that were heated at 121° C for 10 to 80 rain. Fish protein concentrate (FPC) contains pro- tein tliat is high in quality. It tlierefore can be used to supplement diets that contain inadequate amounts of high-quality protein. Fish protein concentrate is prepared by re- moving most of the lipids and water from whole fish. Several methods for preparing FPC have been investigated. They can be classified as chemical, biological, and physical. Most inves- tigators have used chemical methods in which solvents extract the lipids and water from whole fish. In the United States, two processes for making FPC have been approved by the Food and Drug Administration. Both of these are chemical processes in which solvents are used. In the overall program of the National Marine Fisher- ies Service National Center for Fish Protein Con- centrate, various approaches to processing are being investigated. One such approach is cook- ing and pressing fish prior to solvent extraction. This procedure would tend to reduce the volume of solvent required for extraction, inasmuch as water and lipids would be expressed during the pressing stage. Raw fish are diflicult to press because of their physical consistency. The processor can over- come this problem by cooking the fish before ' National Marine Fisheries Service National Center for Fish Protein Concentrate, College Park, Md. 20740. pressing them. If he subjects the fish to a high temperature for a long time, however, undesir- able chemical reactions may occur that decrease the quality of the protein. The purpose of this study therefore was to find whether or not the chemical composition and nutritional quality of fish protein coilcen- trate are altered when the FPC is produced by solvent extraction of fish that have been cooked at different temperatures for varying periods of time. CHEMICAL COMPOSITION Reported here are both the proximate compo- sition and amino acid composition of the FPC produced from cooked fish. PROXIMATE COMPOSITION We used red hake, Urophycis chuss, which are lean fish. They were caught off the coast of New England in the area of Block Island, situ- ated south southwest of Point Judith, R.I. The hake were iced on board the vessel and were then frozen in 25-lb. wax laminated cartons at the dock. The hake were kept frozen while being shipped to the National Marine Fisheries Service National Center for Fish Protein Con- centrate in College Park, Md. The shipment contained about 96 cartons. From these 96 boxes, 15 cartons (375 lb.) were picked at random for the investigation and were stored Manuscript received August 1970 FISHERY BULLETIN: VOL. 69, NO. I, 1971. 141 FISHERY BULLETIN: VOL. 69, NO. 1 at — 20° C (The other cartons were used in another experiment.) The hake were used with- in 1 month after storage. About 17 to 18 hr before studying each pro- cessing variable, we placed one carton of fish in a refrigerated room at a temperature of 5° to 6° C. This treatment allowed the fish to thaw sufliciently so that they could be handled indi- vidually. The fish were ground through a Ho- bart meat grinder," which was equipped with an end plate containing holes that were one-quarter inch in diameter. After the hake were ground, they were thoroughly mixed, and a sample that weighed 20 lb was removed. The sample was divided into three equal portions, and each por- tion was placed in a 2-inch-deep tray lined with aluminum foil. (This procedure was used in order to permit existing equipment to be used.) The trays were placed in an autoclave and were heated at 100°, 109°, or 121° C for 10, 20, 40, or 80 min. Thermocouples were used to measure the temperature of the samples. After being heated, the trays were removed from the autoclave, were covered with aluminum foil, and were cooled in a refrigerated room at 5° to 6° C. A control sample was also prepared, which con- sisted of raw, unheated ground hake. The entire contents of the trays were mixed with solvent at a 2:1 (w/w) ratio of solvent to solid. The samples were extracted by the "cross-current" batch-extraction procedure de- scribed by Brown and Miller (1969). The solvent used for extraction was 91 % , by volume, isopropyl alcohol. The extracted and dried samples of FPC were ground in a Rietz Disintegrator. The samples were analyzed for crude protein, volatiles, and ash by the methods described by Horwitz (196.5). Lipids were determined by the method of Smith, Ambrose, and Knobl (1964). Table 1 shows the concentrations of crude protein, ash, and lipids found. The concentration of crude protein in the samples that were heated was slightly lower than in the sample that was not heated. The Table 1. — Proximate composition, expressed on a mois- ture-free basis, of samples of FPC prepared from hake that were heated for var>'ing times and temperatures before being extracted with solvent. Sompla Crudo protein Ash Lipid Nonheated control % 88.7 % 13.8 % 0.16 Heated sompies: 100° C for: 10 min 86.0 15.3 0.16 20 min 8S.3 15.4 0.16 40 min 063 15.0 0.20 80 min 85.9 15.5 0.15 Mean 85.9 15.3 0.17 109° C for, 10 min 87.5 14.4 0.12 20 min 85.6 15.8 0.12 40 min 86.8 15,7 21 80 min 87.4 13.8 0.15 Mean 86.8 14.9 0.15 121° C for: 10 min 87.3 13.6 0.12 20 min 85.9 15.2 0.14 40 min 86.5 13.7 0.16 80 min 86.8 14.3 0.16 Mean 86.6 14.2 0.14 ' The use of trade names is merely to facilitate de- scriptions; no endorsement is implied. concentration of crude protein, however, was not significantly affected by the temperature at which the samples were heated. Also, the time of heating did not significantly aflFect the con- centration of crude protein in the samples, except for the 20-min treatment. The samples that were heated for 20 min had a slightly low- er concentration of crude protein than did those that were heated for the other intervals of time. The concentration of ash was slightly higher in the samples that were heated than in the sample that was not heated. The concentration of ash in the heated samples was not afl!'ected by either the temperature of heating or the length of time of heating. The concentration of lipid in the samples was somewhat variable, but it was not significantly affected by the treatments. AMINO ACID COMPOSITION Es.sential amino acids, excejjt for tryptophan, were determined with an automatic amino acid analyzer by the method described by Moore, Spackman, and Stein (1958). Tryptophan was determined chemically by the method of Spies and Chambers ( 1949) . Cystine was determined 142 DUBROW and STILLINGS : FPC PROCESSED FROM HEATED RED HAKE microbiologically by the method of Henderson and Snell (1948). Available lysine was deter- mined by the method described by Carpenter (1960). We found only slight changes in the concen- trations of amino acids in the samples (table 2). The treatments that we used did not consistently affect the concentrations of amino acids, except for cystine and available lysine. The concen- tration of cystine was reduced in the sample heated at 121° C for 80 min. The concentrations of available lysine were slightly lower in the samples heated at 100° and 109° C for 20 min. The reason for these decreases is not apparent to us. Evans and McGinnis (1948) previously re- ported that cystine was reduced when soybeans were autoclaved at 130° C for 60 min. NUTRITIONAL QUALITY The nutritional quality of the samples was determined in a feeding study using rats. Diets were prepared that contained 10 ''r protein from the heated samples, the nonheated sample, or casein. The diet that contained the nonheated sample served as a control, and the one that contained casein served as a reference standard. The composition of the basal diet was described earlier by Stillings, Hammerle, and Snyder (1969). Male weanling rats of the Carworth Farms CFE strain were received when they were 22 days old. The rats were housed individually in cages with screen bottoms and were kept in an air-conditioned room maintained at about 23° C. During the first 2 days, the rats were fed a basal diet containing 15% casein. They were then allotted to groups on the basis of weight, and the groups were randomly assigned to different diets. Each group contained 10 rats, and the rats were offered feed and water ad libitum for 4 weeks. The amount of feed consumed was recorded three times each week, and the gains in weight were determined once each week. At the end of the experiment, the protein efficiency ratio was determined by dividing the gain in weight by the weight of protein consumed. The data were analyzed statistically. Dif- ferences between means were determined by Tukey's procedure as described by Steel and Torrie (1960: 109). Table 3 shows the data on the nutritive qual- ity of the FPC samples. Based on the gain in weight, intakes of feed, and protein efficiency ratios, the quality of the samples that were heated at 100° and 109° C was not significantly different from the quality of the control, which was not heated. Samples that were heated at 121° C, however, had a lower quality than the control sample. In general, the quality of the samples heated at 100° and 109° C was equal to that of casein or was slightly higher than Table 2. — Amino acid composition of FPC samples prepared from hake that were heated for varying times and temperatures before being extracted with solvent. acid Concentration of amino acid in: Unheated control sample Samples h eated at: Amino 100° C for: 109° : for: 121° : for: 10 20 40 80 10 20 40 80 10 20 40 80 mm. mm. mm. mm. mm. mm. mm. mm. mm. mm. mm. mm. — - Grams per 16 s rami 0/ n itrogfil - 6.2 6.7 6.4 6.8 6.4 5.7 7.0 6.1 6.4 6.9 6.1 6.6 6.3 Histidine 1.8 2.0 1.9 1.9 1.8 1.7 1.9 1.7 1.8 2.1 19 1.8 2.0 Isoleucine 4.5 4,3 4.4 4.5 4.3 4.3 4.6 4.5 4.5 4.9 4.6 4.4 4.8 Leucine 7.4 7.2 6.9 7.1 6.9 7.0 7.5 7.0 7.2 8.0 7.2 7.3 7.6 7.7 8-3 7.8 8.0 74 7.1 77 7.1 7.5 8.4 7.5 7.9 7.6 Methionine 3.2 3.1 3.1 3.1 3.0 3.0 3.0 3.1 3.0 36 3.1 3.1 3.4 Phenylaion ne 4.1 4.1 4,0 4.1 4.0 4.0 4.2 4.1 4.1 4.5 4.1 4.2 4.3 Threonine 4.3 4.2 4.1 4.2 4.0 4.0 4.4 4.2 4.0 4.6 4.1 4.2 4.4 Tryptophan 1.0 1.2 1.2 1.0 1.3 1.2 1.1 l.I 1.2 1.1 1.2 1.3 1.2 Valine 5.1 4.9 50 5.0 5.1 4.8 5,2 4.6 5,0 5.5 5.0 5.0 5.3 Cystine 09 1.2 1.2 1.2 0.9 1.1 1.1 1.1 1.1 1.1 1.1 — 0.7 Avoilable ysine 7.9 7.8 6.9 7.2 7.9 7.7 6.9 7.2 7.6 7.8 7.1 7.2 7.6 143 FISHERY BULLETIN: VOL. 69, NO, 1 Table 3. — Weight gain, feed intake, and protein effi- ciency ratio of groups of 10 rats fed diets of FPC samples prepared from red hake that were heated for varying times and temperatures before being extracted with solvent. Sample Average daily weight gain Averog© daily feed intake Protein efficiency ratio Grami Grams Nonheated control 4.85 13.8 3.37 Heated samples: 100° C for: 10 min 4.96 14.0 3.41 20 min 4.37 13,0 3.25 40 min 4.50 13.1 3.37 80 min 4.29 13.3 3.16 Mean 4.53 13.4 3.30 109° C lor; 10 min 4.37 12.7 3.35 20 min 4.31 13.1 3.11 40 min 4.45 13.4 3.20 80 min 4.20 12.6 3,28 Mean 4.34 13.0 3,24 121° C for: 10 min 3.21 10,6 3.01 20 min 4.01 12,2 3.13 40 min 3.52 11,1 3.10 80 min 3.20 10,3 3.02 Mean 3.49 11.0 3,07 Casein 3.85 12.0 3.18 Tukey's W (P<0.051 0.81 2.0 0.26 that of casein. When the temperature was in- creased to 121° C, the quality of the samples was slightly lower than that of casein but not sig- nificantly so. At each temperature, the temper- ature at which the samples were heated had a more significant effect on the quality of the samples than did the length of time of heating. SUMMARY AND CONCLUSIONS We conducted a study to determine the chem- ical composition and nutritional quality of FPC produced from fish that are heated before they are extracted with solvent. Red hake, which are lean fish, were heated at 100° C for 10, 20, 40, or 80 min. Other samples were heated for these same lengths of time at 109° or 121° C. The samples were then e.xtracted with isopropyl alcohol. The FPC produced from the samples of hake that were heated contained slightly less crude protein and more ash than did the FPC produced from the samples that were not heated. The amino acid composition of samples that had been heated did not differ markedly from the composition of those that were not heated. The nutritive quality of the samples that were heated at 100° and 109° C was not significantly affected. Samples heated at 121° C, however, were lower in quality than was the control sample. We conclude that red hake can be heated at temperatures of 100° and 109° C for as long as 80 min before being extracted by solvent with- out the quality of the protein being affected sig- nificantly. LITERATURE CITED Brown, Norman L., and Harry Miller, Jr. 1969. Experimental production of fish protein con- centrate (FPC) from Mediterranean sardines. Commer. Fish. Rev. 31(10): 30-33. Carpenter, K. J. 1960. The estimation of available lysine in animal- protein foods. Biochem. J. 77(3): 604-610. Evans, Robert John, and James McGinnis. 1948. Cystine and methionine metabolism by chicks receiving raw or autoclaved soybean oil meal. J. Nutr. 35(4) : 477-488. Henderson, L. M., and Esmond E. Snell. 1948. A uniform medium for determination of amino acids with various microorganisms. J. Biol. Chem. 172(1) : 15-29. HoRwiTZ, William (chairman and editor). 1965. Official methods of analysis of the Asso- ciation of Official Agricultural Chemists. 10th ed. Association of Official Agricultural Chemists, Washington, D.C., xx -f 957 pp. Sections 22.003, 22.010, and 22.011. Moore, Stanford, Darrel H. Spackman, and William H. Stein. 1958. Chromatography of amino acids on sulfo- nated polystyrene resins. Anal. Chem. 30(7): 1185-1190. Smith, Preston, Jr., Mary E. Ambrose, and George N. Knobl, Jr. 1964. Improved rapid method for determining total lipids in fish meal. Commer. Fish. Rev. 26(7) : 1-5. Spies, Joseph R., and Dorris C. Chambers. 1949. Chemical determination of tryptophan in proteins. Anal. Chem. 21(10): 1249-1266. Steel, Robert G. D., and James H. Torrie. 1960. Principles and procedures of statistics with special reference to the biological sciences. Mc- Graw-Hill Book Company, Inc., New York, xvi -|- 481 pp. Stillings, B. R., O. a. Hammerle, and D. G. Sntoer. 1969. Sequence of limiting amino acids in fish pro- tein concentrate produced by isopropyl alcohol extraction of red hake {Urophycis chtiss). J. Nutr. 97(1) : 70-78. 144 EFFECT OF ICE STORAGE ON THE CHEMICAL AND NUTRITIVE PROPERTIES OF SOLVENT-EXTRACTED WHOLE FISH-RED HAKE, Urophycis chuss David L. Dubrow, Norman L. Brown, E. R. Pariser, Harry Miller, Jr., V. D. Sidwell, and Mary E. Ambrose' ABSTRACT Because red hake that are to be used in the future production of fish protein concentrate will be caught in quantity, the preservation of the hake during periods of glut will present a problem that possibly can be solved by storage of the hake in ice. In our study of this problem, whole red hake were held in ice for 2, 6, 8, and 11 days. Organoleptic tests on the fresh fish showed that they were edible on the 8th day but were not edible on the 11th day. Samples of fish were removed during each period of storage and were processed ( 1 ) by f reeze-drying to produce a reference sample (2) by solvent extraction with isopropyl alcohol to produce a fish protein concentrate. Proximate composition, amino acid composition, and nutritive quality were determined comparatively on both of these two kinds of processed samples. From the data obtained, we concluded that red hake stored in ice for 8 days are suitable for use in the production of fish protein concentrate and that they would be suitable for this use up to the point of spoilage of the fish, which occurs sometime between 8 and 11 days. In the period between the capture and processing: of fish that are to be used in products for human consumption, they must be preserved in a man- ner that maintains their food-grade quality. This requirement applies to the production of fish protein concentrate (FPC) as well as to that of more common fish products. The preservation of fish is a problem not only aboard the harvesting vessel but at the shore processing plant as well. The problem ashore becomes especially important during periods of glut when the fresh fish must be held several days before being processed. In the manufacture of FPC by the method we use, oil and moisture are removed from the fish with isopropyl alcohol. We therefore in- vestigated the possibility of holding fish in this solvent (Dubrow and Hammerle, 1969). We found the method to be entirely suitable for pe- riods of holding up to 11 days. Although storage in isopropyl alcohol was satisfactory, more conventional means of holding ' National Marine Fisheries Service National Center for Fish Protein Concentrate, College Park, Md. 20740. the fish, such as storing them in ice, are likely to be used in commercial operations. During the time fish are held in ice, however, consider- able change may occur in the components of the fish tissue. Endogenous and bacterial en- zymes may break down protein into water-sol- uble and volatile components, causing off-flavors and odors in the fish. In addition, the highly unsaturated lipids of the fish may oxidize rapidly, causing the fish to become rancid. While these changes are taking place in iced fish, the water from the melting ice is leaching out some of the compounds that are forming. Furthermore, the subsequent extraction with alcohol during the production of FPC, if ade- quate, removes most of the undesirable com- Ijounds that were not leached out by the melt water. Just what effect the enzymatic and oxidative changes have on the various components of the tissues as well as on the nutritive quality of the protein in the finally processed FPC is not known. Accordingly, solubilization of the com- ponents of the fish tissues could alter the com- position of the finally processed FPC. We should Manuscript received August 1970. FISHERY BULLETIN; VOL. 69, NO. 1. 1971. 145 FISHERY BULLETIN: VOL. 69, NO. I know, of course, what occurs, because FPC is of value solely as a protein supplement of high quality. The aim of this study therefore was to de- termine the effect that storage of food-grade fish in ice has on the chemical composition of the components of the tissue and on the nutritive quality of the protein. We accomplished this aim by comparing FPC made from samples of the ice-stored fish with reference samples made by freeze-drying samples of the fish. We used freeze-drying because we believe that this meth- od of production results in minimum alteration in the samples during drying. CHEMICAL COMPOSITION Both the proximate composition and the amino acid composition of the samples were determined. PROXIMATE COMPOSITION As indicated earlier, we used standard refer- ence samples produced under ideal conditions, as a basis on which to evaluate our samples of FPC. Standard Reference Sample About 600 lb of red hake were caught on Jan- uary 6, 1965, in 25 to 26 fathoms of water oflf the coast of Rhode Island. The fish were di- vided randomly into lots of 100 lb each, were iced immediately, and then were taken to the Bureau of Commercial Fisheries (BCF) (now National Marine Fisheries Service) Technological Lab- oratoiy at Gloucester, Mass., where they were held in ice. During the next 11 days, each lot of fish was inspected periodically for freshness by exper- ienced BCF fish inspectors at Gloucester. The factors they considered were (1) damage to the fish, (2) conditions of the skin, eyes, and gills, and (3) texture, odor, and flavor of cooked samples. A numerical score ranging from one to four was used to rate fish of varying quality for each of the factors. Fish of perfect or nearly perfect quality were assigned a value of 1, whereas those at the limit of acceptability or beyond the limit were assigned a value of 4. Table 1 shows the data on the subjective eval- uation of the raw fish. The samples of fish tested after storage for 11 days in ice were judged to be at the limit of acceptability. The fish that had been stored in ice for 8 days were of acceptable quality and were considered to be of food grade. Table 1. — Freshness evaluations of raw red hake stored in ice for periods up to 11 days. [Each sample had 50 fish.] Storoge Averc ge sub ecfive evaluations of: Time Damage Sk.n Eyes Gills Texture Odor Flavor Day! 2 1.00 1.00 1.02 1.40 1.50 1-08 1.0 d 2.26 2.22 2.04 2.32 260 2.38 2.0 8 2.44 2.50 3.00 2.92 3.18 3.10 2,5 11 3.10 4.00 3.86 3.92 3.98 4.00 — Fish of perfect or nearly perfect qualify were assigned a value of I; ose of unacceptable quality were assigned a value of 4. those After the iced fish had been inspected for quality, they were shipped in ice to College Park, Md. Each box of fish, upon receipt at College Park, was divided into two gi-oups and were processed immediately— one into a standard reference sample and the other into FPC. One portion of 20 lb was selected at random from the group of fish to be used as a standard reference sample. The standard reference sample was prepared by freezing the fish in liquid nitrogen and grinding the whole fish through a Rietz Disintegrator' under a stream of liquid nitrogen, and then freeze-drying the liquid-nitrogen slurry of ground fish. The freeze-drying step was carried out under a pres- sure of 500;* of mercury and at a platen tem- perature of 40° C. The dried samples were then removed from the freeze dryer in an at- mosphere of nitrogen and were sealed in con- tainers. The containers were maintained at — 40° C until the samples were needed. The freeze-dried samples were analyzed for crude protein, ash, and volatiles in accordance with standard procedures (Horwitz, 1965). To- tal lipids were determined by the method of Smith, Ambrose, and Knobl (1964). Table 2 shows the ])roximate composition of ' The use of trade names is merely to facilitate de- scription; no endorsement of products is implied. 146 DUBROW ET AL, : EFFECT OF ICE STORAGE Table 2. — Proximate composition of freeze-dried, ground whole hake (standard reference samples) stored in ice for periods up to 11 days. Storage time Volatiles Lipids! Ash' Crude protein^ Days 2 6 8 11 3.80 2.49 2.46 4.70 »■(. % 15.30 14.06 14.34 15.07 in. % 13.44 12.84 12.43 12.49 (ff. % 74.47 77.34 77.38 77.01 ' The data on lipids, osh, and protein were based on the dry weight of sample. 2 Crude protein was calculated as N X 0.25. the various samples of freeze-dried whole fish. Data are presented on a dry-weight basis to re- veal possible losses during storage. The concentration of lipid varied between 14 andl5';r; that of ash, between 12 and 13^c. The data indicate that the nitrogen fraction did not change greatly. The crude protein remained rel- atively constant at about 77 Sr (on a dry- weight basis) except on the second day of sampling. This deviation on the second day was probably the result of a sampling error. Analyses for nonprotein nitrogen would have been helpful for interpretive purposes. Unfortunately, they were not made. Dassow" has reported that the nonprotein nitrogen fraction of whole Pacific hake stored in ice did not change significantly over a period of 11 days. Fish Protein Concentrate From the remaining portion of each lot of fish, 20 lb were selected at random and were ex- tracted with isopropyl alcohol according to established procedures (Brown and Miller, 1969). In brief, the fish were ground through a Hobart meat grinder, were slurried with 15 liters of 91% (v/v) isopropyl alcohol for 30 min, and were centrifuged. The centrifuged solids were then extracted continuously with hot isopropyl alcohol at 60° to 70° C and at a rate of flow of 0.2 gal per minute. After 2 hr the solids were removed by centrifugation and were desolventized under vacuum at 60° C. = Dassow, John A. 1966. Statement of project ac- complishment, Utilization of fishery resources program. In Quarterly progress report of the BCF Technological Laboratory, Seattle, Wash., July 1 - September 30, 1966. Unpublished report, 6 p. This method of processing was not intended to be representative of commercial methods. It was used in our laboratory at that time solely as an experimental technique to evaluate selected variables in the preparation of FPC by solvent extraction. It has since been replaced by a sev- eral-stage countercurrent extraction system, which is both much more economical in the vol- ume of solvent needed and is more representa- tive of commercial processing methods. A com- parison of FPC made by each system has shown no significant differences, however, either in chemical composition or in nutritive value. The proximate composition was determined by the same method used with the freeze-dried fish. Table 3 lists the proximate compositions of the FPC's prepared from the fish stored in ice for various periods. The concentrations of lipids Table 3. — Proximate composition of FPC prepared from raw fish stored in ice for periods up to 11 days. Storage time Volatiles Lipids! Ash' Crude protein^ Days Wl. % Ifl. % Ifl. % »■(. % 2 4.25 0.18 12.30 89.70 6 5.10 0.13 12,44 89.65 3 5.12 0.10 13.19 89.30 11 4.10 0.21 16 04 86.94 ! The dato on lipids, ash, and protein were based on the dry weight of sample. - Crude protein was calculated as N X 6.25. and volatiles remained essentially unafi'ected by storage. The concentration of ash increased, however, and that of protein (that is, of nitro- gen) decreased. The major change occurred after the 8th day of storage. Because the con- centration of protein in the standard reference samples did not drop in the same manner as the concentration of protein did in the FPC's, the loss of protein could not have occurred dur- ing storage but must have occurred during pro- cessing. This conclusion could be accounted for by the formation, during storage, of soluble ni- trogenous products resulting from enzymatic breakdown or bacterial breakdown, or from both, that were not leached out of the fish during storage but that were subsequently leached out during the extraction process used in making the FPC. This conclusion was further support- 147 FISHERY BULLETIN: VOL. 69. NO. 1 ed by the observed decrease in yield after pro- cessing — namely, 12.0 percent of 2-day-old fish to 10.0 percent of 8-day-old fish. Storage of whole red hake in ice up to 11 days did not influence the extractability of the lipids. A slight loss of nitrogen occurred, however, dur- ing the processing of whole fish stored for 11 days as compared with fish stored for shorter periods. AMINO ACID COMPOSITION Standard Reference Sample Amino acids were determined by the method of Spackman, Stein, and Moore (1958). Table 4 shows that the recovery of amino acids was relatively constant at about 92'^^'c of the protein. The essential amino acids for which analyses were made ranged between 45.5 and 46.3 "^r of the total. No major change in the pattern of any one particular amino acid resulted from storage. In general, this finding agrees with those by Cohen and Peters (1963) on whiting, Mer/«cc»« bilinearis, that were stored in ice. These auth- ors reported, however, that methionine de- creased after the 13th day with a subsequent Table 4. — Amino acid composition of raw freeze-dried whole ground fish. The samples were prepared after fish were held in ice for periods up to 11 days. increase in methionine sulfoxide. We do not know whether this compound was present in the hake that we studied. Fish Protein Concentrate The same methods were used as with the standard reference sample. That is, the amino acids were determined by the method of Spack- man, Stein, and Moore (1958). Table 5 shows the concentration of amino acids in the FPC's processed from the fish held in ice. The data indicate that about 1009c of the amino acids were recovered. The essential amino acids constituted 47% of the total amino acids in the FPC made from fish stored 2 days, but the concentration of these amino acids dropped to 43 Sr after the fish had been stored 11 days. Individual amino acids decreased in concentration. Of these amino acids, leucine and isoleucine decreased slightly, whereas lysine and histidine decreased markedly after the 8th day of storage. The total concen- tration of lysine was about 11 '^r less in the FPC made after the fish had been stored for 11 days than in the FPC produced after they had been stored for 2 days. The concentration of histidine Table 5. — Amino acid composition of FPC prepared from raw fish held in ice for periods up to 11 days. Concentration of the Standard Reference So given omino acid in the Tiples after they were held: Amino acid Concentration of the samples extracted o given amino acid in the fter they were held for: 2 doys 1 6 days 8 doys 11 doys 2 days | 6 doys 8 days 11 days Perirnl 0/ Iht protein (N X 6.25 j Percent oi the prot ein (N X 6.25) Lysine 7.63 7.44 7.72 7.56 Lysine 867 8.14 8.38 7.72 Histidine 1.92 1.77 1.92 1.76 Histidine 2.06 1.88 2.00 1.74 Ammonia 1.68 1.57 1.58 1.59 Ammonia 1.47 1-57 1 39 1.44 Arginins 5.96 5.82 6.05 5.76 Arginine 7.12 698 7.24 6.92 Aspartic acid 9.50 9.41 9.45 953 Aspartic acid 10.36 10.36 10.17 10.00 Threonine 4.07 4.16 4.08 4.14 Threonine 4.45 4.48 4.51 4.44 Serine 4.08 4.22 4.11 4.11 Serine 4.60 4.59 4.70 4.75 Glutamic acid 14.05 14.32 14.27 14.23 Glutamic acid 15.47 15.57 15.34 15.27 Proline 4.57 4.79 4.69 4.76 Proline 5.01 5.64 559 6.47 Glycine 7.70 8 23 768 7.52 Glycine 8.04 9.20 912 10.23 Alanine 6.50 6.56 6.36 6.41 Alanine 6.78 7.10 6.96 7.23 Valine 4.88 4.78 4.63 4.98 Valine 5.14 5.24 4.95 4.90 Methionine 3,02 3.00 2.91 3.05 Methionine 3.32 3.32 3.46 3.29 Isoleucine 4.21 4.17 4.13 4.32 Isoleucine 4.52 4.46 4.37 4.24 Leucine 7.07 7.03 7 06 7.21 Leucine 7.70 7.54 7.44 7.17 Tyrosine 3.01 2.90 2.88 2.99 Tyrosine 3.39 3.31 3.28 3.14 Phenylalanine 3.82 3.98 3.98 3.94 Phenylolanine 4.12 4.05 4.07 3.91 Total amino acid recovery 91.99 92.58 91.92 92.27 Total amino acid recovery 100.75 101.86 101.58 101.44 Percent essential omino acids 46.29 45.51 44.21 46.30 Percent essential amino acids 47.00 45.25 45.70 43.71 148 DUBROW ET AL. : EFFECT OF ICE STORAGE decreased about 15.5'^f within the same period of time. Both glycine and proline increased in percentage of the total amino acid concentra- tion. This increase could possibly be due to the lack of enzymatic breakdown of the fish collagens, thereby increasing the percentage of these amino acids as compared with that of the amino acids of the myofibrillar proteins. In retrospect, an analysis of the raw, unpro- cessed fish for free amino acids or total non- protein nitrogen would have made the interpre- tation of these results more certain. No marked differences in the amino acid pattern of the standard reference sample could be detected after storing the whole fish in ice for periods up to 11 days. The amino acid pat- tern of the FPC's produced from the same batch of fish as was the standard reference sample, did, however, show changes, which were more pronounced in the FPC processed from 11-day-old fish. These changes appeared to be the result of alcohol extraction of solubles that were apparently formed during ice storage and not leached out by the melt water from the ice. PROTEIN QUALITY Table 6. — Mean weight gained, food consumed, and ad- justed protein efficiency ratio of groups of eight rats fed freeze-dried whole hake prepared from fish stored in ice, compared with casein. Storage time Meon werghr gained Mean weight of food consumed Adjusted protein efficiency ratio^ Day: Grami Grams 2 158.6 ± 3,16 390 ± 5.7 3.46 ± .05 6 150,8 ± 3,08 385 ± 5,9 3.35 ± .08 B 155.6 ± 5,28 381 ± 7.7 3.49 ± .07 11 148.6 ± 3.35 400 ± 3.9 3.18 ± .07 Casein 113.5 ± 5.65 323 ± 3.9 3,00 ± .00 ^ The protein efficiency ratios were odjusted ta a protein efficiency ratio of 3,00 for casein. protein quality of the standard reference sample taken on the 11th day was similar to that of casein and therefore was lower than that of the three samples taken earlier. Proximate composition and concentrations of amino acid do not account for the difference ob- tained in the quality of the protein in the sample of fish held in ice for 11 days. Because the fish were from the same lot and were chosen randomly, we can only speculate either that the utilization (digestibility) of the protein (amino acids) was decreased or that compounds de- pressing growth were formed during storage. STANDARD REFERENCE SAMPLE Protein efficiency ratios were determined by the method of Campbell (1960). Diets of the standard reference samples and of FPC pre- pared from raw fish stored in ice were fed ad libitum to male albino rats (Charles River strain), which were randomly allotted to groups of 10 animals. The samples were added to a basal diet at a 10 ""r level of crude protein. Gain in weight and consumption of food were re- corded each week for 4 weeks, and the protein efficiency ratio was calculated as (weight gain)/' (weight of protein consumed) . A diet in which casein was the source of protein was used as a reference. Table 6 shows the data obtained from the animal-feeding studies comparing the quality of the protein of the various samples. Except for the sample jjrepared from fish held 11 days, the protein quality of the standard refei'ence samples was better than that of casein. The FISH PROTEIN CONCENTRATE The same methods were used to determine protein quality as were used with the freeze- dried fish. Table 7 shows the data obtained from the feeding tests made on FPC's produced from the fish held in iced storage. All the FPC's gave a greater gain in weight and a higher pro- Table 7. — Mean weight gained, food consumed, and pro- tein efficiency ratio of groups of eight rats fed diets of FPC prepared from raw fish stored in ice for periods up to 11 days compared with casein. Storage time Mean weight gained Mean weight of food consumed Adjusted protein efficiency ratio! Days 2 6 8 11 Grams 154.0 ± 8,63 155.1 ± 8.12 154.4 ± 4.95 145.4 ± 4.80 Grams 363 ± 12.0 362 ± 12.3 368 ± 7.6 358 ± 10.3 3.62 ± .05 3.65 ± .09 3.59 ± .10 3.47 ± .10 Casein 113.5 ± 5.65 323 ± 3.9 3.00 ± .00 1 The protein efficiency ratios were adjusted to a protein efficiency ratio of 3.00 for casein. 149 FISHERY BLXLETIN: VOL. 69, NO. I tein efficiency ratio than did the casein. Diets containing FPC made from fish stored for 2, 6, and 8 days in ice resulted in protein efficiency ratios ranging between 3.59 and 3.65. The diet containing FPC from the U-day-old fish yielded a slightly lower gain in weight and a protein efficiency ratio of 3.47. These results agree with those obtained with the standard reference samples made from the same fish. The nutritive quality of the 1 1-day standard reference samples, however, was poorer than that of the FPC sample. This anomalous result suggests either an improved utilization of protein as a result of ex- traction with isopropyl alcohol or the removal of some factor that may have depressed growth. Freeze-dried fish produced from whole red hake stored in ice 2 to 8 days did not differ in protein quality. Freeze-dried fish produced from whole red hake stored for 11 days, how- ever, was lower in protein quality but still had a protein efficiency ratio equal to that of casein. FPC produced from whole fish stored for 2 to 11 days showed no diff'erences in protein quality. All the FPC's had protein efficiency ratios high- er than that of casein. SUMMARY AND CONCLUSIONS Whole red hake were stored, in ice for 2, 6, 8, and 11 days. The fish were organoleptically evaluated for freshness at each storage period and were then processed by freeze-drying to form a reference sample or by solvent extraction with isopropyl alcohol to form a fish protein concentrate. These products were then analyzed for proximate composition and amino acid con- centration and for protein quality. The results of the subjective evaluation for freshness indicated that the fish stored up to 8 days were still acceptable for food but that those stored for 11 days were not acceptable. The proximate composition and the amino acid concentration of the freeze-dried whole samples of fish showed very little change as a result of the storage of the raw fish in ice. Rat- feeding tests indicated a loss in protein quality of the freeze-dried sample prepared from 11- day-old raw fish. Protein efficiency ratio values ranged from 3.35 to 3.49 for fish stored up to 8 days, whereas the llth-day sample resulted in a protein efficiency ratio of 3.18. All protein efficiency ratio values, however, were equal to the value for casein or were higher. The proximate composition of FPC's pro- duced from fish stored up to 8 days in ice re- mained relatively constant. The crude protein in the concentrate produced from fish stored for 1 1 days decreased about 2.5 9f The concentra- tion of amino acids also followed this pattern with a resultant lowering in the concentration of lysine and a slight increase in that of proline and glycine. The protein quality of the FPC processed from the 11-day-old fish was also slightly lower than that of FPC processed from fresher fish. All FPC's, however, had protein efficiency ratios higher than that of casein. We conclude that storage of whole hake in ice up to 8 days is a satisfactory means of hold- ing them prior to extracting the ground hake with isopropyl alcohol to produce FPC. LITERATURE CITED Brown, Norman L., and Harry Miller, Jr. 1969. Experimental production of fish protein con- centrate (FPC) from Mediterranean sardines. Commer. Fish. Rev. 31(10): 30-33. Campbell, J. A. 1960. Evaluation of protein in foods for regulatory purposes. J. Agr. Food Chem. 8(4): 323-327. Cohen, Edward H., and John A. Peters. 1963. Effect of storage in refrigerated sea water on amino acids and other components of whiting (Merluccius bilinearis) . Fish. Ind. Res. 2(2): 5-11. DuBROw, David, and Olivia Hammerle. 1969. Holding raw fish (red hake) in isopropyl alcohol for FPC production. Food Technol. 23(2) : 254-256. HoRWiTZ, William (chairman and editor). 1965. Official methods of analysis of the Associa- tion of Official Agricultural Chemists. 10th ed. Association of Official Agricultural Chemists, Washington, D.C., xx + 957 pp. Smith, Preston, Jr., Mary E. Ambrose, and George N. Knobl, Jr. 1964. Improved rapid method for determining total lipids in fi.sh meal. Commer. Fish. Rev. 26(7) : 1-5. Spackman, Barrel H., William H. Stein, and Stanford Moore. 1958. Automatic recording apparatus for use in the chromatography of amino acids. Anal. Chem. 30(7): 1190-1206. 150 LABORATORY REARING OF THE DESERT PUPFISH, Cyprinodon macularius David Crear' and Irwin Haydock^ ABSTRACT The desert pupfish, Cyprinodon macularius, may ba reared in the laboratory for use in the study of embryology, genetics, physiology, and behavior. It is euryhaline (0-70 %c) and eurythermal (8°-44.6° C) and may be useful as a bioassay for either freshwater or marine pollutants. In the Salton Sea area of California, the recent introduction of exotic species and the encroachment of civilization have drastically reduced the formerly abundant pupfish populations. Laboratory rearing eliminates the need for continuous exploitation of a rapidly contracting natural population and could supply adequate stocks for sanctuaries, thereby preserving the species from extinction. Laboratory apparatus and conditions are described for maintaining larval and adult pupfish. Parasites and diseases encountered are dis- cussed and successful treatments described. Methods for spawning and rearing the desert pupfish in the laboratory are detailed. These methods may also be applicable to many other species of pupfish that are in danger of extinction. The desert pupfish, Cyprinodon macularius Baird and Girard, is a killifish (Cyprinodonti- dae) native to the Lower Colorado River Basin from southern Arizona to southern California and the Sonoyta River of northern Sonora, Mex- ico (Miller, 1948). It thrives under the harsh conditions of the desert environment. It lives in fresh water as well as highly saline pools that few other vertebrates can tolerate. Its ability to survive in such environments, plus other important biological characteristics listed in Table 1, renders it an exceptionally hardy laboratory animal potentially valuable for re- search in many fields. POTENTIAL FOR RESEARCH Desert pupfish has many characteristics fa- vorable for embryological research. It can be spawned with relative ease and can be main- tained in the laboratory throughout the year to supply large eggs (approximately 2 mm in di- ameter) , suitable for vital marking and grafting ' Formerly, California Department of Fish and Game, Inland Fisheries Branch, Sacramento, Calif.; present address: School of Public Health, 1890 East-West Road, University of Hawaii, Honolulu, Hawaii 96822. ' Formerly, California Department of Fish and Game, Inland Fisheries Branch, Sacramento, Calif.; present address: Southern California Coastal Water Research Project, 10845 Lindbrook Drive, Los Angeles, Calif. 90024. Manus and reached a high of 12 % in mid-May (Fig. 1); at this time, individual females were caught with GSI's of more than 17 %. Peak spawning in the Salton Sea was observed in May and early June in 1969. Le.ss frequent sampling and obser- vations confirmed a similar pattern of events in 1970. In the years 19.5.5, 1956, and 19.57, Whitney (1961) found that the peak abundance of croaker eggs and larvae in the Salton Sea fell in middle and late May. Laboratory Cycle and Effects of Photoperiod, Water Temperature, and Food Immature fish. — Young-of-the-year croakers Salton Sea Bairdiella 1968- A M J MONTH Figure 1. — Seasonal change in GSI of female Bairdiella icistia captured in the Salton Sea. Hori7.ontal line in- dicates the mean GSI value; vertical line indicates range; on either side of the mean, open bar indicates the standard deviation, and closed bar two standard er- rors of the mean of each sample. The number of fish sampled is given in parentheses. Horizontal dashed line indicates 5 % GSI for comparison w-ith laboratory fish (Fig. 2). 164 HAYDOCK: GONAD MATURATION OF GULF CROAKER Table 2. — GSI values of female Bairdiella icistia cap- tured in Salton Sea compared with captured fish matured in the laboratory. Group Dots of sample Weight (mean ± SD) GSI (mean ± SD) 1 23- X -<59i 10.47 ± 3.56 096 It 0.28 9 S-Xll-69 34.30 ^ 5.15 0.93 ±L 0.15 11 19-X:i-69 35.40 :+; 4.78 1.07 ± 0.46 10 16-11 -70 60.14 ^ 11.84 5.46 ± 3.32 5 2 23- X -691 10.47 ± 3,56 0,96 ± 0.28 9 5-X1I-69 31.90 ^ 8.04 0,92 ± 0.14 10 19.x 11-69 33.10 zt 4.94 0-80 ± 0.09 9 16-11 -70 39.40 ± 7.16 1.28 ± 0.22 5 4- IV -70 49.30 ± 5,01 9.69 ± 3,46 5 3 14- V.70» 32.38 ± 8.32 5.90 ± 3.84 25 4 I4-V-701 170.4 ± 84.1 7.93 ± 1.35 24 5 19-111-69 161.6 =t 22.8 10.2 ± 1.94 23 Treatments Group 1- Laboratory stock fish sampled irregularly. All fish kept on 16L:8D photoperiod at 22° C. Group 2. Laboratory stock seporated from group 1 and maintained on 8L:160 photoperiod at 14° C until 16.11-70, when they were switchea step-wise (15 min/day) to 16L:8D and 22° C. Group 3: Young fish captured during their first breeding season in Salton Sea. Group A: Fish more than 1 year old, captured during the breeding season in Salton Sea. Group 5: Fish more than I year old, captured following the breeding season in Salton Sea ond matured early under laboratory conditions of 15L:9D and 14° to 16° C. 1 At capture. before their normal breeding season, reached maturity in the laboratory sometime prior to being sampled in mid-March 1969 (Table 2, group 5). These fish experienced 15L:9D and ambient La Jolla seawater temperature (14°- 16° C) during 4 months in the laboratory. Cold water may slow down the maturation process, but it is evidently not as important as the stim- ulation of long days. A second sample of adult fish was captured in mid-May (at the peak of the breeding season), subjected to various experimental laboratory conditions and sampled every 2 weeks to deter- mine the status of their GSI (Fig. 2). Fish maintained on long days (16L:8D) at 14° C and 22° C showed a slow decline in GSI from a high of 12 9^ at capture to below 5 % by late-August and September. The GSI values at 22° C were more variable and, in general, showed a more rapid decline than those at 14° C. Similar fish given 10L:14D at high and low temperatures showed a similar but much more rapid decline in GSI, and their GSI also declined to a lower overall level (1-2 '/c by mid-August) than that of fish which never experienced short-day con- ditions. Both groups of short-day fish subsequently showed a slow but steady increase in their GSI in response to having the photoperiod increased 15 min/day from lOL to 16L. Although this increase was not followed through a complete cycle, it was evident that exposure to long days for 2 to 3 months would have been required to bring the fish up to the GSI level necessary for spawning (about 5 % — see below). It is likely that adult fish brought into the laboratory just prior to the normal increase in GSI observed in the Salton Sea would respond rather quickly, I estimate within 1 month, if they were given adequate light, temperature, and food. It may also be possible to mature fish rapidly after they have gone through their na- tural GSI decrease (Fig. 1), but this was not tested. Effects of Hormones on Maturation of Fish Maintained in the Laboratory Groups of fish maintained on various light and temperature regimes were subjected to hormonal treatments to enhance gonad maturation. Adult fish captured prior to the breeding season were not available for these experiments, which were conducted after the spawning season on fish undergoing a decline in GSI, as in Fig. 2. None- theless, the results obtained probably indicate the extent to which maturation can be influenced by hormone treatment. In croakers, the tech- nique of hormone-induced maturation is of rel- atively little practical importance, since fish can be matured by appropriate manipulation of photoperiod, temperature, and feeding sched- ules. The results are, therefore, reported for their possible application to other species in which maturation proves more intractable. Since the treatments were carried out on fish being used for photoperiod and temperature ex- periments, the results can only be evaluated in relation to the GSI value of the population under each set of conditions. In some cases, sham-in- jected controls were used while in others unin- jected fish sampled for the light- temperature 165 FISHERY BULLETIN: VOL. 69. NO. I 15n LaboratoTLj Bairdiella 1969 5- 15r 10 - r a c _ IGL-.SD 22C — -1 lOL*-^ 16L r I i 22c - \ " \ - J- X~~-i;— 2; — + J 1 1 1 1 - \ 1 1 1 1 1 1 leUSD 14C i^F I I I I I I I I L_ I I J J A MONTH N lOL*^ 16L 14C l\l-.-I' -rn I I I I J L. J J A MONTH N Figure 2. — GSI of female Bairdiella icistia captured during the spawning season and maintained under the following laboratory conditions: (a) 16L:8D, 22° C; (b) 16L:8D, 14° C; (c) 10L:14D, 22° C for 2 months; then, photoperiod was increased 15 min/day to 16L:8D; (d) 10L.14D, 14° C for 2 months; then, photope- riod was increased 15 min/day to 16L:8D. Mean GSI of all fish at capture (May 13, 1969) was 12.2 %. Dashed value in (d) indicates GSI of five fish injected with 1 mg of salmon pituitary 1 day prior to last sampling per- iod. Horizontal line at 5 % GSI indicates approximate minimum level necessary for successful hormone-induced spawning. studies served as controls when they were sched- uled to be sampled at the same time as the in- jected fish. In all cases the gonad index responded positively to hormone treatment. Differences seen in the tabular data (Tables 3, 4, and 5) were due to the number of injections, strength and type of hormone injected, the water temperature and the GSI value of the fish at the beginning of the experiment. In Tables 3, 4, and 5 the symbols (S, E, + , — ) record the initial (2 days after the first in- jection) and the maximal (at some point during the series of injections) response noted for each group of fish. Two factors, water temperature and GSI, were found to have an im])ortant bear- ing on the results observed following hormone injection. Fish kept in 14° C water showed the most consistent and largest positive response to long-term hormone injections which were giv- Table 3. — GSI of Bairdiella icistia given five injections, every other day for 9 days (17-26-IX-1969). Photope- riod was 16L:8D and temperature 14° C during this e.\- periment. The response of the fish to injection was as- sessed every day and the maximal response is indicated by S (= spawned viable eggs), E (= obtained non- viable eggs), -f (= swollen papillae observed, no eggs obtained), or — (^ no observed response). The sec- ond column under each heading indicates the reproductive status of the fish 2 days after the first injection. The third column (in parentheses) indicates the maximum reaction noted during the experiment. Treatment 1 mg salmon pituitary extract 100 lU HCG 5 mg OOCA Uninjected control GSI 12.4 -1- (E) 12.1 - (E) 11.4 - (E) 4.1 (-) 11.6 + (E) 1 0.0 — (E) 6.1 - (E) 3.6 (-) 11.1 + (E) 8.8 - (-H 5.2 - (E) 3.2 (-) 10.9 - !-) 6.6 - (+) 4.6 - (-H 2.7 (-) 6.0 - (-1-) S.9 - (-) malo 2.7 (-) X * SD 10.4 ± 2.6 8.7 ±: 2.5 6.8 ± 2.7 3.2 i: 0.6 166 HAYDOCK; GONAD MATLRATION OF GULF CROAKER Table 4. — GSI of Bairdiella leistia given six injections over a 15-day period (18-VIII-69 — 2-IX-69). Each in- jection consisted of 1 mg salmon pituitary extract. Fish in groups 1 and 2 experienced a 16L:8D photoperiod; groups 3 and 4 experienced 10L:14D for 2 months (to 28-VII-69) followed by day length increases to 16L:8D (by 22-VIII-69). Groups 1 and 3 were maintained at 22° C, groups 2 and 4 at 14° C throughout the experiments. Symbols (S, E, +, — ) are the same as in Table 3. Group 1 (16U 22° C) Gro (16L, up 2 14° C) Group 3 (10L-16L, 22° C) Group 4 (10L-16L, 14° C) Injected Control Injected Control Injected Control Injected Control 6.5 E (E) 10.2 (-) 5.0 E (E) 3.2 (-) 4.2+ (E) 1.9 (-) 4.2 + (+) 1.3 (-) 3.6 - (-) 0.9 (-) 11.3 + (E) 9.4 + (E) 9.1 + (E) 9.1 + (E) 7.6 + (E) 10.2 (-) 6.0 (-) 5.5 (-) 5.2 (-) 2.3 (-) 5.6 - (E) 3.6 (-) 4.9- (E) 1.6 (-) 1.8 - (+) 1.4 (-) 1.6 - (-) 1.4 (-) 1.2 - (-) 1.3 (-) 7.3 - (E) 2.4 (-) 3.3 - (+) 2.0 (-) 1.9 - (-) 1.9 (-) 1.7 - (-) 1.6 (-) 1.2 - (-) 1.4 (-) X ± SD 4.7 ±1.1 3.5 + 3.8 9.4 ± 1.3 5.8 ± 2.8 3.2 ± 2.0 1.9 ± 1.0 3.1 ± 2.5 1.9 ± 0.4 Table 5.— GSI of Bairdiella icistia given three injections, one every other day for 7 days (16-IX— 23-IX-1969). Photoperiod was 16L:8D and temperature 22° C during this experiment. Symbols (S, E, +, — ) are the same as in Table 3. 1 mg salmon 1 mg salmon 1 mg carp 0.1 mg salmon Control sesame oil Holtfreter's sesame oil sesame oil sesame oil 5.5 S (S) 5.0 S (S) 8.1 +(E) 6.0 -(E) 4.9 (-) 4.9 S (S) 5.0 S (S) 6.3 + (E) 3.3 - (+) 1.4 (-) 4.7 -(E) 4.6 -(E) 2.7 -(E) 3.0 - (+) 1.4 (-) 4.1 -(E) 3-6 -(E) 2.7-(+) 2.5 - (+) 1.1 (-) 1.8-(-) I.4-(-) 2.5 - (+) 1 .5 - (-) 1.0 (-) X± SD 4.2 ± 1 .4 3.9 ± 1 .5 4.5 ± 2.6 3.2 ±1.7 2.0 ± 1.6 en every other (3ay for 1 to 2 weeks (Table 3; Table 4, groups 2 and 4). Fish in 22° C water also showed a positive response (Table 4, groups 1 and 3; Table 5) ; however, this response is less clear because warm-water fish occasionally shed their eggs prior to sampling and this re- duced the observed GSI. In general, fish with GSI values below 5 % did not respond to the first injection ( — ), re- sponded with a weak swelling in the genital area ( + ) , or gave nonviable eggs ( E ) only after several injections. This indicated that threshold values of GSI and temperature exist below which "growth" and above which hydration and ovulation occur in response to hormone in- jections. These values will be further discussed in the section on ovulation. Here it will suffice to point out that hormone treatment does give rise to increases in gonad size which can perhaps be considered the equivalent of gonad growth. The tabular data indicate that salmon pitui- tary caused the greatest increase in GSI of fish kept in cold water. Salmon was followed by HCG and then DOC A (Table 3) . Salmon pitui- tary produced smaller increases in warm water than in cold (Tables 3, 4, and 5), and the re- sponse tended to vary in proportion to the dosage used (Table 5). The relatively small response at 22° C in Table 5 was probably due in part to the lesser number of injections (3) in this batch and in part to the fact that three of the fish spawned relatively large quantities of eggs (see qualitative responses in Table 5) , thus reducing their GSI values, which were measured after testing for the presence of viable eggs. How- ever, the results of the longest series of injections (Table 4) suggest that there is a general pat- tern of greater increase in GSI in cold water, except in the case of fish with a very low initial GSI. In warm water (Table 4, group 1; Table 5), fish produced viable eggs (S) or nonviable eggs (E) which could be forced out the day after the first injection. Cold-water fish (Table 3; Table 4, groups 2 and 4) required several injections to produce a response and never spawned viable eggs on stripping. In a further experiment, young-of-the-year fish collected in October were injected shortly thereafter with various concentrations of salmon pituitary to assess usefulness of immatures as a bioassay in testing dose-response relationships. Each fish received five injections over a 10-day period. At each dose level a large number of fish (20) was injected, but, because of the difficulty in identifying the sex of these immature fish, the number of females actually injected was some- what smaller. Also, the gonads were quite small, and the overall GSI response was slight. This made any meaningful analysis difficult. How- 167 FISHERY BULLETIN: VOL. 69, NO. 1 Table 6. — GSI of young female Bairdiella icistia after five injections of various concentrations of salmon pitui- tary extract over a 10-day experimental period (4-XI- 14-XI-69). Dosa Weight (s) mean ± SD GSI mean ± SD N Control sesome oil 26.07 ± 5.78 79 ± 0.08 13 0.1 mg salmon 23.90 ± 6.23 0.82 ± 0.22 10 0.5 mg salmon 28.40 ± 4.46 88 ± 0.18 13 1.0 mg salmon 26.97 ± 6.88 0.99 ±0.19 17 ever, the results (Table 6) do indicate a general increase in GSI corresponding to dose. That these fish were not too small to respond to treat- ment is indicated by the high GSI observed in similar sized fish captured in Salton Sea during the breeding season (Table 2, group 3). It is possible that the rather small response observed was due in part to the fact that the fish were handled frequently and did not feed readily dur- ing the course of the experiment. This test would probably be more successful if carried out with 30- to 50-g fish kept under short-day conditions at 14° C. GONAD HYDRATION Water Uptake Following Hormone Injections Weight gain on various hormones. — Short term changes in body weight occurred 1 to 2 days following the injection of various hormones. This weight change was recorded as a percent of the initial total body weight (Table 7) and was found to vary from slightly negative values to positive values of over 13 Sr of the body weight. Single injections were given, and fish were weighed prior to, and 30 hr after, injection. With few exceptions, the fish showed little or Table 7. — Effects of hormones of GSI of Bairdiella icistia at 30 hr post-injection. or ovulate.) (1 = did, = did not hydrate Initial Weight GSI (% final weight) Initial Weight GSI Preparation, dose, and dote body weight (l) change (% initial weight) Hydrote Ovulate Preparation, dose, ond date body weight (s) change (% initial weight) (% final weight) Hydrate Ovulate Carp pituitary lOmg Salmon pituitary 5 mg-Con. lO-VI-70 51.07 -1-10.99 9.34 1 91.7 -1-10.09 25.94 1 1 (poor 70.46 -H5.42 19.80 1 62.3 -1-8.06 26.64 I %%' 72.79 -f 11.72 25.50 1 PMS 100 lU eggs) 57.22 -f 12.82 28.60 1 27-IV-70 86,8 — 21.00 0(?) 1 Carp pituitary 5 mg 1 2-V-70 88.7 -HIS — 0(?) I 1 1 16-VI-70 71 40 54.22 - 5.42 8.14 7-V-70 PMS 50 lU 173.6 -1-11.98 24.02 1 Salmon pituitary 9-VI-70 80 38 —3.06 2.90 0.1 mg, 5-VI-70 48.33 -0.93 2.38 54.89 -M.IO 14.60 I I 55.90 -2.45 3.60 69.90 -f-7.77 18.50 1 I 49.40 -f2.39 10.97 (1) 62.18 -1-12.06 24.10 I 1 66.14 -1-0.06 12.41 HCG 100 lU Salmon pituitary 28-IV-70 68.5 28.76 1 0.5 mg, 2-VI-70 65.18 -2.59 3.83 14-IV-70 87,5 29.37 1 62.77 -1.96 5.02 15-IV.70 84.5 30.29 1 74.83 -0.64 5.18 56.22 -1-8.75 21.76 1 1 HCG 50 lU 16-VI-70 78.51 -2.09 2.69 Salmon pituitary 1 mg 57.03 -1.95 3.65 28-IV-70 96.12 -1-9.40 15,90 1 1 67.91 — 1.29 3.68 15-V-70 65 50 -fl3.40 -_ 1 1 80.41 -f7.15 19.80 1 1 18-V-70 79.90 -1-13.10 __ 1 1 15-V-70 131.20 -(-600 1 1 Oxytocin 20 lU ie-vi-70 59.12 -0.70 1.60 Salmon pituitary 2 mg 65.66 -1.02 3.37 lO-lV-70 93 30 __ 24.47 I t 72.11 -0.11 3.95 23-111-70 68.92 __ 25.67 1 I 60.08 4-2.39 11.97 1 IO-lV-70 100.03 - 28.08 1 1 DOCA 5 mg Salmon pituitary 5 mg 15- VI -70 54.78 -0.51 4.07 28-V-70 66.1 -f7.35 20.79 1 (few 53.70 -3.07 4.28 65.4 +6.)\ 21.23 I !Pew 55.33 -0.55 7.98 eggs) 73.10 -1.20 9.78 168 HAYDOCK : GONAD MATURATION OF GULF CROAKER no response to 0.1 and 0.5 mg salmon pituitary, 5 mg carp pituitary, 50 lU HCG, 20 lU oxy- tocin, and 5 mg DOCA. On the other hand, 1 to 5 mg salmon, 10 mg carp, and 100 lU HCG, gave uniformly positive results, all fish showing weight gains of 5 to 13 %. Variable results were observed with 50 and 100 lU PMS. Al- though most PMS fish which were weighed showed some gain in weight, this was in general less than that observed with carp, salmon, and HCG. In fact, it was noted that two fish in- jected with 100 lU PMS spawned freely without ever appearing grossly bloated, a characteristic of all fish which were spawned with other prep- arations. The time scale of weight gain. — A comparison was made of the weight gained by fish given one injection of 5 mg salmon, 10 mg carp, and 50 III PMS (Table 8). The time span of hy- dration was arbitrarily divided into the weight gained between 7 and 23 hr post-injection and the total weight gained, including that added between 23 and 30 hr. At 30 hr ovulated eggs, if present, were stripped from the fish. Gen- erally, all fish lost weight in the first 7 hr, pro- bably because of handling and lack of feeding during the experiment. The weight gains are due mostly to water uptake and movement of water into the gonad. Table 8. — Effects of hormones on time-course of weight gain. fleets some fundamental difference in the way these preparations afl!"ect the physiological mech- anism causing hydration. The time-course of hydration (Table 8) may be important in de- termining the condition of eggs at ovulation (Table 7). It should be noted that among the three groups tested for the time-course of hy- dration, viable eggs were obtained only from the PMS-injected fish (Table 7) ; unfortunately, there is no comparable data on the time-course of weight gain in fish given 1 mg salmon, which also produces viable eggs. Factors Affecting Hydration GSI. — It is apparent from Table 7 that the gonad must be close to 5 % of the body weight to respond to an otherwise adequate dose of hormone. Although GSI could not be measured prior to injection, almost all fish which failed to respond had final GSI's below 5 %. Table 9 presents further confirmation of this. These fish, injected with 1 mg salmon, came from a stock which had shown a general decline in GSI, because of being kept on a long photoperiod for an extended time. Of four injected fish, three hydrated and one of these subsequently spawned. The fish that neither hydrated nor spawned had a final GSI of just under 3 %. Only 1 of 11 un- infected fish from this same stock showed a GSI above 5 %, while 3 more were above 4 %. Table 9. — GSI of Bairdiella icistia measured on 26-VI-70. Initial fish weight Percent weight gain Preparation A. Fish (1 injected = did, with 1 mg salmon = did not hydrate pituitary, after or ovulate). GSI hod declined ond dose 7-23 hr post-injection 7-30 hr post-injection Initial Weight change GSI G % % body weight (% initial (% final Hydrate Ovulate Salmon pituitary 5 mg 66 1 4.58 7.35 weight) weight) 91.7 7.72 10.09 53.10 -1-14.73 24.42 1 I 65,4 4.59 6.11 65.38 -1-7.40 10.55 1 62.3 4.41 8.46 60.14 -H4.21 6.94 ) Carp pituitary 10 mg 57.22 7.38 13.13 79.89 -2.21 2.77 51.07 4.17 10.44 B. Unin iected fish from same tank. 72.79 6.76 13.11 Total weight GSI PMS 50 lU 54.89 0.13 5.25 62.18 4.86 13.11 67.65 1.63 69.9 2.88 9.69 53.20 93.28 1.79 0.05 58.86 4.38 The results sho w that t he weight increase 63.48 1.27 in the final 7 hr p rior to sp awning is less than 61.90 78.60 4.57 1.70 50 % of the total increase with sail non pitui- 74.70 2.08 tary, more than 50 ^f with P MS, and a bout equal 53.24 69.29 2.59 7.86 when carp pituitai -y is used This ev idently re- 59.46 4.80 169 FISHERY BULLETIN: VOL. 69, NO. I Hormone dosage. — In general, there appears to be a threshold response to dosage. Fish with high GSI values and between 50- and 100-g body weight hydrated after one injection of 10 mg carp but not 5 mg (Table 7) . With salmon, 1 to 5 mg were adequate doses for 50- to 350-g fish while 0.1 and 0.5 mg were inadequate except in one case. In the case of HCG, 100 lU caused hydration while 50 lU did not except in a single case. Note, however, the low GSI value meas- ured for three of the fish given 50 lU HCG. It must be noted that the highest dosages used were adequate for hydration but inhibited spawning (see section following on ovulation). This was true for 10 mg carp, 5 mg salmon and, especially, 100 lU HCG where the fish continued to gain weight and eventually died in the tank without ovulating a single egg. These results would suggest that it is important to determine the lowest possible dosage which will consistently bring about hydration. Temperahire. — A temperature threshold un- derlies the entire spawning process. Between 14° and 17° C the fish did not hydrate in re- sponse to an otherwise adequate dosage of 1 mg salmon pituitary. Two days later these same fish spawned within 30 hr when given a second 1-mg-salmon injection 24 hr after being trans- ferred to 22° C water. OVULATION Ovulation and Hydration as Separate Events Some early results with 100-IU-PMS and 100- lU-HCG injections led to speculation that the two hormones were acting on different physiological processes. PMS brought about ovulation with- out gross hydration while HCG hydrated fish to the point of death without ovulation (Table 7). This result was not confirmed with 50-IU doses, but, in general, the impression gained was that PMS produced high quality eggs with less hydration than either HCG or salmon pituitary. In sharp contrast to the PMS results, HCG caused uniform hydration but, with one excep- tion, failed to bring about ovulation. In a pre- liminary test, it was found that oxytocin (20 lU) or salmon (1 mg) caused some eggs to be ovu- lated when the injection was given 24 hr after the fish were injected with HCG. Oxytocin and salmon pituitary had the same effect on carp- injected fish which otherwise did not ovulate. Ovulation without apparent hydration was also achieved by using multiple, subthreshold doses of 0.1 and 0.5 mg salmon pituitary, but the time of ovulation could not be accurately pre- dicted, and therefore the eggs obtained usually were not viable. (The importance of obtaining eggs just at ovulation is discussed in the section on fertilization.) In summary, carp pituitary, HCG, DOCA, and oxytocin were uniformly inadequate for bring- ing on ovulation except in the case of one fish treated with HCG. Both carp- and HCG-in- jected fish hydrated, some becoming grossly distorted. On the other hand, salmon pituitary and PMS regularly brought about ovulation. Dosage appeared to be critical in the case of salmon pituitary, as nonviable eggs resulted from injections of 5 mg and no spawn could be obtained with a single dose of 0.1 and 0.5 mg. A 1-mg-salmon dosage seems to be optimal for fish of 50- to 100-g total weight. Both 50- and 100-IU dosages of PMS gave good results with remarkably clear eggs obtained from all fish. In one case, a 50-IU dose of HCG was adequate for spawning, but 100 lU appeared to be inhib- itory to ovulation. The Time Scale of Ovulation The combined results from 28 fish which pro- duced viable eggs following injection with a single dose of salmon (1-3 mg) showed that the average time elapsed from injection to spawning was 30.4 hr, with a standard deviation of 3.3 hr and a range of 24.5 to 35.5 hr (Fig. 3). This 30-hr latent period following injection gener- ally held regardless of the type of hormone used, its dosage, or the time of day the injection was made. In the case of a few large fish (100 to 300 g) given 1 mg salmon, a second 1-mg in- jection was given 24 hr later, but this did not affect the time of spawning. Five fish given 5 mg salmon spawTied 28 hr after injection, but their eggs were not viable. In one experiment. 170 HAYDCXTK: GONAD MATURATION OF GULF CROAKER I I I I I I I I I I I I I I I I I 060O 1200 TIME OF INJECTION tPST) Figure 3. — Time of injection (Pacific Standard Time) and time of spawning (hours post-injection) in relation to photoperiod. All injections were 1-3 mg salmon pitui- tary extract and all fish produced viable eggs. Solid black bar indicates dark period; dashed line indicates mean time of spawning. five fish were injected with 1 mg salmon at a time corresponding with the beginning of the laboratory dark cycle. All these fish spawned 24 hr later, indicating a possible enhancement by the normal diurnal cycle of glandular activity. Natural spawning in the Salton Sea is I'elated to the normal diurnal light cycle, with most spawning occurring in the early evening. Factors Affecting Ovulation It has already been shown that GSI level, hormone dosage and type of hormone are criti- cal interacting factors which must be considered in any spawning eff'ort. Injection of high levels of salmon (5 mg) may possibly assure a more uniform hydration response (see GSI of Table 7), but the nonviable eggs which result speak against using more than the minimal dose found to give consistent results. The eflfect of temperature on ovulation per se was not studied. Hydration is effectively blocked at temperatures lower than 17° C, but this effect was reversed after 24 hr acclimation at 22° C. In the cases in which this transfer was carried out, a second injection was given 24 hr after transfer, and spawning took place approximately 30 hr later. As a matter of practical interest, it was found that fish could be injected and spawned twice (tried, successfully, with two fish) or three times (one fish) with a period between spawnings of 3 to 4 weeks. This is in contrast to the much longer time required for maturation after fish had slowly resorbed their gonads in photoperiod experiments (Fig. 2). Apparently the rapid emptying of the gonad consequent upon hor- monal injection quickly leads to a renewed cycle of egg maturation. The direct and indirect effects of various in- jections on the gonad were assessed by biopsy following spawning or the lack of spawning. These qualitative observations are listed in Table 10; no attempt is made to interpret these results, except to point out that fish injected with salmon pituitary extract had gonads most closely re- sembling those of naturally spawning fish. Table 10. — Appearance of mature Bairdiella icistia ovaries during natural spawning and 30 hr after various hormone injections, and color reaction of fish to injections. 1. Sollon Sea fish at spawning Gonad color white, light yellow or red-orange. Consistency of ■nature gonad is granular with patches of tronsporent eggs which are close to being ovulated or are lying free in the ovarian lumen. 2. Salmon pituitary extroct Gonad color and consistency very close to naturally spawning fish. Mast eggs ovulated and free in lumen. Fish blanch on injection. 3. Corp pituitary extract Gonad color red-orange; few eggs ovulated. In fish given 5 mg dose, blood clots oppeared to be blocking oviducts near vent. Fish blanch on injection. 4. PMS Gonad whitish, translucent; strikingly different from other prepa- rations. Most eggs ovulated and free in lumen. Possibly, greater degree of ovulotion with less hydration makes gonad appear lighter in color. Fish do not blanch on injection. 5- HCG Eggs either not ovulated or partially ovulated; those not ovulated appear as white patches in the ovary. Many vacuoles and dis- persed oil drops appear in eggs. Fish do not blanch on injection. 6. Oxytocin Ovary was very bloody. Eggs white (not hydroted); different sized eggs (mostly large) apparent in ovarian folds. Fish do not blanch on injection. 7. DOCA Fish showed no observable reaction. FERTILIZATION Relationship ot Egg Viability to the Time of Ovulation Shortly before ovulation, eggs could be squeezed from females by applying strong pres- sure to the abdomen, but eggs obtained in this way still had an investiture of blood vessels and ovarian tissue and could not be fertilized. An analysis of viability in relation to the time after 171 FISHERY BULLETIN: VOL. 69, NO. 1 Table 11. — Fertilization and viability of Bairdiella icistia eggs tested over a 4-hr period following first ovulation. Fish was injected with 1 mg salmon pituitary extract at 0930 l-V-70; first eggs expressed with some difficulty at 1530 2-V-70. Time of fertilization Development to early tailbud Hatching 1530 hr (not quite running ripe) % 100 % 91 % 84 1630 hr (running ripe) 100 91 83 1730 hr 100 78 72 1830 hr 90 62 44 1930 hr (eggs spotty, opaque) 5 52 34 ovulation was made in the case of one fish which remained ripe for 4 hr (Table 11). To check for viability, eggs were test-fertilized at hourly intervals following the first sign of ovulation, taken as the earliest time when normal eggs could easily be expressed from the fish by gentle pressure applied to the abdomen. Fertilization and early cleavage remained above 90 '^f up to 3 hr post-ovulation. By 4 hr the eggs looked crinkled, opaque, and spotty, and less than 5 % could be fertilized. A further measure of via- bility was made by culturing 100 early cleaving eggs from each batch until hatching. A decrease in hatching success was noted in the eggs ob- tained 2 hr after the initial ovulation, and hatch- ing decreased still further in the 3- and 4-hr post-ovulatory samples. It appears that the maximum grace period for egg-taking is about 2 hr. In another experiment I studied eggs from a larger sample of 10 fish determining fertil- ization success as a function of time after ovu- lation; the optimum time for taking eggs was 1 hr after the fish first showed signs of running ripeness and gave viable eggs. Although eggs rapidly deteriorated when kept in the ovary following hormone-induced ovula- tion, it was found that they retained their ability to be fertilized up to several hours after they were placed in a moist storage chamber. Eggs placed in seawater remained fertile for several minutes; in one case, a few cleaving eggs re- sulted from fertilization carried out after the eggs had been in seawater for 30 min. Viability of Sperm Although eggs kept in seawater remained viable for several minutes, sperm were no longer able to fertilize eggs 30 sec after the sperm mass had been introduced into seawater. It is thus readily apparent that croaker sperm and eggs should be mixed immediately after the sperm is obtained, in order to achieve maximum fertil- ization. Microscopic examination showed that sperm were immediately activated by addition of water and retained motility for a period of 1-5 min. In some tests it was apparent that water from the Salton Sea caused greater ac- tivity for a longer time than water taken from the ocean at La Jolla, Calif., but there was great variability between males, and a proper tech- nique of quantifying this relationship awaits further studies. Number of Eggs Obtained by Hormone Treatment The number of eggs obtained by hormone in- jection varied between 700 and 1,000 per gram of fish wet weight (Table 12). This provided 50,000 to 100,000 eggs for experiments from each fish of 50 to 100 g used in this study. Table 12. — Number of eggs obtained from hormone-induced spawning of Bairdiella icistia. Wet weight GSI Ripe eggs Actuol count (1 g eggs) Eggs/g fpsh weight Approximate Total fish gonad Body weight Gonad weight total eggs/ fish G C % % 82.0 17.7 21.6 16.0 74.0 4,700 841 69,000 86.6 17.6 20.3 15.5 78.1 ■793 '69,000 91.7 26.0 25.9 197 76.1 5,590 1.101 101,000 342.7 62.1 __ _- 750 ■699 ■240,000 441.5 129.7 -- 17.1 - - ■878 ■388,000 Indicates volues colculoted from meosured parameter and mean number of eggs per gram counted. 172 HAYDOCK: GONAD MATURATION OF GULF CROAKER DISCUSSION MATURATION Many experiments have shown that gonad maturation can result from hormone therapy (for reviews see Pickford and Atz, 1957; Ahsan and Hoar, 1963; Atz and Pickford, 1964; and Hoar, 1969), but in most cases this is a long and tedious approach and has proved of prac- tical use only on a short-term basis for eluci- dating mechanisms of hormone action. A re- cently described catheter implant technique (Frogner and Hendrickson, 1970) , allowing fre- quent or continuous administration of hormones, has been used with partial success to mature mullet, Mugil cephalus, with a minimum of dam- age from excessive handling (Shehadeh, person- al communication ) . A mass of tangled catheters is envisioned if this technique were applied to commercial fish production, but the ease of this method may have considerable merit for exper- imental situations. Implanted pituitary glands might also be used to enhance maturation, and this could easily be tested in croakers. In the present study, a slight increase in GSI, possibly reflecting enhanced gonad "growth," followed 1 to 2 weeks of hormone injections giv- en every other day to fish held in 22° C water. Even greater "growth" enhancement was ob- served in 14° C water, and these fish could have been spawned using techniques which were fully developed later in the study. However, for prac- tical purposes, it proved simpler to mature croakers in large groups using appropriate schedules of long days, warm water, and abun- dant feeding. The fact that fish kept in cold water respond to hormones by gonad enlargement without sub- sequent hydration or ovulation indicates that different temperature thresholds exist for these various processes. It is possible that the rate of absorption of hormone is considerably slowed in cold water, as some fish do develop a slight reaction following several days of injection at 14° C. The general relationship of light and temper- ature to gonad maturation is well known (see Harrington, 1959; Henderson, 1963; Wiebe, 1968; and Hoar, 1969, for reviews) and requires no lengthy discussion here. It is sufficient to note that long-day photoperiod (16L:8D) and high temperature water (22° C) induce gonad maturation in croakers several months prior to the normal breeding season observed in the Salton Sea. Also, a combination of long days and low temperature (14° C) will retard the normal GSI decline when the fish have been cap- tured at the peak of breeding. This technique may prove useful for maintaining fish in a ma- ture state for prolonged periods; such fish may be subsequently spawned following transfer to warm water (22° C) for a period of 1 day. Studies on the relationship between maturity and spawning indicate the existence of a GSI threshold value of about 5 %, below which hor- mone injections are ineffective. Also, fish brought to maturity with photoperiod and tem- perature control eventually resorb gonadal tis- sues if they are not subsequently spawned. This resorption process requires several weeks, and the gonad will not grow in response to photo- period and temperature during this time. Fish which are spawned with hormones do not show a refractory state and can be respawned within a few weeks. The practical implication of these findings is that the GSI of maturing fish should be frequently checked so that spawning can be- gin soon after the 5 'r GSI threshold is reached and the fish should be spawned before they reach the maximum GSI value and begin gonad re- sorption. A useful approach would be to hold stock supplies of fish on short days at low tem- perature and mature separate groups as needed for experiments. Samples of croakers taken throughout the year from Salton Sea showed that maturation is quite rapid, the GSI increasing from 2 to 10 % in a little over 1 month. It is probable that the increased light, temperature, and food stimu- lation available in the laboratory could bring about even more rapid maturation, but the pro- per fish (early spring) to test this were not ob- tained during this study. HYDRATION Hormone-induced gonadal hydration is a rel- atively rapid phenomenon which is completed 173 FISHERY BULLETIN: VOL. 69, NO, 1 in a little over 1 day following injection under laboratory conditions. In croakers, the total water uptake is reflected primarily in increased gonad weight and may amount to more than a 10 % increase in total body weight. A detailed study of the gonadal hydration of carp, Cyprinus carpio, and goldfish, Carasshis aurahis, following injections of carp pituitary extracts showed a similar pattern of water movement into the go- nad with respect to time (Clemens and Grant, 1964). These authors measured the increased water content of the gonad following injection and found that, in the case of males, the peak of seminal fluidity was 24 hr after a single ip injection. A similar response was observed with im injection. Goldfish females injected with 10 mg/g carp pituitary extract showed sim- ilar responses, increasing gonad water by up to 7.2 % over carrier-injected controls. Un- fortunately, the changes they describe are in the relative water content of gonads and various other tissues including blood, and no mention is made of any increase in total body weight re- sulting from water taken up from the external medium. Hydration under laboratory conditions results in a grossly distorted appearance in females, the abdominal cavity becoming bloated several hours prior to spawning. In the Salton Sea, fe- males appear plump but never grossly enlarged at spawning. It is possible that naturally spawning females hydrate and ovulate fre- quently but in small amounts over the course of the breeding season and that the laboratory fish show the maximum hydration and ovulatory response because of unnatural overstimulation with the injection of salmon pituitary. Use of 10 mg carp pituitary and 100 lU HCG caused an equally strong hydration response, but gen- erally this did not culminate in ovulation when these preparations were used alone. A dose threshold for response was indicated by the in- ability of 1 to 5 mg of carp pituitary to cause hydration. With 100 lU HCG, continued hy- dration without ovulation evidently overstressed the fish and led to their eventual death. On the contrary, PMS gave somewhat variable re- sults, but appeared to have less effect on gonad hydration while, at the same time, proving to be a potent ovulating agent. Subthreshold doses of salmon pituitary do not appear to cause hy- dration, but a sequence of injections given at daily intervals will eventually lead to ovulation of small quantities of eggs. This may reflect the response of exceptionally ripe eggs which are able to hydrate and ovulate. Thresholds of GSI (above 5 %), water tem- perature (between 17° and 22° C), and hormone dose (e.g., 1 mg or more of salmon pituitary for 50- to 100-g fish) exist, and if any one of these factors is below its threshold, hydration does not occur. OVULATION Ovulation in croakers is a rapid process, taking 1 to 2 hr for completion when induced with hormones in the laboratory. The period between injection and spawning includes the hydration phase and culminates in ovulation at about 30 hr post-injection. Stevens (1966) found a similar 30-hr latent period for fully mature striped bass, Roccus saxatilis. Clemens and Sneed (1962) found a shorter latent period of 15 hr for goldfish. Fontenele (1955) gave injections to several Brazilian fish species at 6- hr intervals. He stated that spawning usually occurred just prior to the 5th injection (i.e., close to 30 hr after the first injection), although in most cases the fish were allowed to spawn naturally in ponds and were not tested by strip- ping. Indian carp are also allowed to spawn naturally in ponds after injection. Chaudhuri (1960) states that spawning may come 6 to 8 hr after the first injection of very mature fish; if a second injection is necessary, the total elapsed time may be 14 to 18 hr. It would ap- pear that in most recorded cases (see above and Pickford and Atz, 1957, Table 46) hormone injection will bring about final maturation and spawning within 1 to 2 days if the gonads are fully mature. In only a few cases (e.g., Joseph and Saksena, 1966) have longer series of in- jections been successfully used to produce viable eggs. A constant time period for spawning latency was found to hold for croakers used in this study. When GSI, water temperature, and hor- 174 HAYDOCK: GONAD MATURATION OF GULF CROAKER mone dosage all exceed certain threshold levels, the fish spawned viable eggs an average of 30 hr after the first injection. Hormone dosages above threshold and the type of preparation had no apparent effect on this result, which exhibited only small variability. Subthreshold doses of salmon pituitary did delay ovulation, but this de- lay could not be accurately predicted, and there- fore the spawn obtained was never viable. Sub- threshold doses might theoretically be useful if females were to spawn naturally in captivity, but croakers never exhibited complete spawning behavior in tanks following hormonal injections. Injection of oxytocin in hydrated females and in males appeared to cause heightened pre- spawning behavior, with males following and touching the vent of females, but actual spawn- ing was not observed. Hydrated females event- ually expelled their eggs into the tanks if they were not hand-stripped shortly after ovulation. Actually, the relatively constant latency and the fact that fish must be hand-stripped are highly advantageous to scheduling laboratory oper- ations. At Salton Sea, eggs of gulf croakers sampled from the plankton and staged at various times during the day and night showed that there is a diurnal pattern of spawning, most early cleav- age stages appearing in the early evening (Whit- ney, 1961). The same diurnal pattern was found in a closely related species from the East Coast, B. chrysura (Kuntz, 1914). When the effect of this diurnal pattern on laboratory spawning was tested by injecting fish at a time corresponding to the beginning of the labora- tory dark cycle, all five injected fish spawned just at "dusk," 24 hr after injection rather than the usual 30 hr. However, such enhancement was not found in any subsequent spawning attempts carried out at many diff"erent times of day and night. Clemens and Sneed (1962) found no change in latency in goldfish, groups of which were injected at 2-hr intervals over a period of 12 hr. Evidently, the injection of hormones usually overrides any effect of diurnal spawning patterns. Clemens and Sneed (1962) found that the latent period decreased with increasing temper- ature, doubling from 12 hr at 30° C to 25 hr at 10° C. Croakers spawn above 20° C in the Salton Sea, and eggs develop to hatching between 20° and 30° C in the laboratory (Robert C. May, Scripps Institution of Oceanography, personal communication). However, all laboratory spawning was accomplished at 20° to 22° C, and no tests were run to determine if higher tem- peratures would decrease the latent period. At temperatures below 17° C, the croaker does not hydrate or ovulate in response to hormone in- jection. For 50- to 100-g croakers, with GSI levels above 4 to 5 %, a single injection of 50 lU PMS, 50 lU HCG, or 1 mg salmon pituitary proved adequate to induce spawning. A dosage of 100 lU PMS and 2-3 mg salmon also produced viable eggs, but 5 mg salmon produced nonviable eggs in the four fish tested. A single injection of 10 mg (not 5 mg) carp pituitary or 100 III HCG was adequate for hydration but not for ovulation. An injection of 20 lU oxytocin or 5 mg DOCA apparently had little or no effect on hydration, although DOCA may have caused some slight change in the GSI. Oxytocin may affect spawning directly. Liley (1969) reviews evidence that the spawning re- flex is controlled by behavioral stimulation of the CNS which releases oxytocin. Oxytocin is used up during the reproductive season of fishes, e.g., Fjindulics and Oncorhynchus (Perks, 1969). The possibility that a second hormone, acting (in concert or independently) directly on ovula- tion, was absent from HCG or in too low a con- centration in carp pituitary was evaluated in a preliminary way by injecting 10 mg carp or 100 lU HCG fish with 20 lU oxytocin at 30 hr or an otherwise inadequate dose (0.1 mg) of salmon 24 hr after the initial injection. Evidence was obtained for ovulation shortly after injection (oxytocin) or at 30 hr (salmon), although the eggs usually were not viable. These experiences indicated that hydration alone was not sufficient to initiate ovulation and that the latter may be a separately controlled process. The apparent contrast observed with respect to the different abilities of HCG and PMS to hydrate and ovulate fish may possibly be 175 FISHERY BULLETIN: VOL. 69, NO. I interpreted as additional evidence tiiat a two- hormone system exists for reproductive control in fishes similar to the FSH-LH system of birds and mammals (see Ahsan and Hoar, 1963; Hy- der, 1970, for details). HCG is LH-like while PMS is FSH-like; fish pituitary extracts show strong LH and slight FSH activity in mammals,, but there is a great deal of conflicting evidence and interpretation (Sundararaj and Goswami, 1966; Hoar, 1969). Other evidence indicates that PMS acts like a combination of FSH and LH when tested in mammals (Ball, 1960), but this effect can be modified by the dosage used. Hoar (1969) pi-esently considers it likely that teleost pituitaries contain only a single gona- dotropin. Sundararaj and Goswami (1966) demonstrate how wide the range of conflicting results can be, when they report that hypophysectomized cat- fish, HeteropnenMes fossilis, spawned ripe eggs after injection with appropriate concentrations of LH, HCG, PMS, and DOCA, while FSH brought about ripening but no spawning (LH contamination was possible) . PMS did cause ovulation in striped bass (Stevens, 1966) . HCG has been used successfully in other fish spawn- ing studies (e.g., Sneed and Clemens, 19.59; Stevens, 1966). The fact that both PMS and HCG can lead to successful spawning and, yet, reflect basically antagonistic systems in mam- mals should make these hormones prime targets in future experiments. It is quite evident that considerable work re- mains to be done to untangle the connections be- tween hydration and ovulation, which are cer- tainly i-elated events, but may be controlled by different hormones acting at different threshold levels. The puzzling fact that one out of four .50 lU HCG fish hydrated and spawned while three out of three 100 I U fish hydrated Ijut never spawned, might be explained by postulating that a "criti- cal dose" exists, with doses above or below this level being unable to induce the complete se- quence of spawning events. The three 50 lU fish which did not hydrate would perhaps have spawned if their GSI had been above 5 Sf . A "critical dose" phenomenon might also be in- volved in the observed difference in hydration and the comjilete lack of spawning obtained with 5 mg and 10 mg carp pituitary, as both groups showed GSI values above 5 % ; in this case the "critical dose" might lie between 5 mg and 10 mg. Carp is considered a universal donor by Cle- mens and Sneed (1962) and was successfully used to spawn several species of freshwater fishes. Bioassay with goldfish showed 100 lU HCG to be equivalent to 0.5 mg acetone-di'ied carp pituitary (Sneed and Clemens, 1959), and ovulation was obtained with 100-1600 lU HCG and 0.5-3.0 mg carp. Most other workers also report no inhibition of spawning from very large doses, but they all point out the critical nature of exceeding some lower threshold dose to initiate ovulation. The strength of pituitary extracts for spawning is assumed to be related to the reproductive status of the donors, a datum not given by the company selling the carp pitui- taries used in the present experiments. Salmon pituitaries, however, were removed only from fish graded at the hatchery for optimal ripeness and the glands were taken within 15 min of death. Nonetheless, from the results of this study 1 mg of salmon pituitary appears to be 5 to 10 times more potent than 1 mg carp and about equal to 50 lU HCG or PMS, although definite qualitative differences in response exist. A truly valid comparison of the strength of var- ious fish pituitary preparations can of course be made only by standardized bioassay (for reviews of methods see Clemens and Sneed, 1962; Das and Kahn, 1962; and Yamazaki and Donaldson, 1968a and 1968b). It is clear that the effects of hormones vary with the GSI level of the experimental fish. Most of these spawning experiments were carried out over a 2-month period beginning in mid-Ajn-il and ending in mid-.June, while the GSI was gradually decreasing in the stock of fish used for the experiments. Thus, it is difficult to directly compare the effects of 50 lU and 100 lU HCG, as they were tested almost 2 months apart and the average GSI values of the experimental fish may have been somewhat different. The effect of the population's declining GSI is clear in the 176 HAYDOCK: GONAD MATURATION OK OLLF CROAKER case of a standard 1 mg dose of salmon pitui- tary, which caused hydration, but produced eggs from only one fish in the last test (late June 1970) carried out with the same stock of fish which had been spawned regularly with the same dose over the prior 2 months. FERTILIZATION Several early attempts to fertilize gulf croaker eggs all ended in failure. These eggs were ob- tained from hormone-induced spawning, and they appeared normal in all respects; however, the sperm mass was dispersed in the water some time prior to adding the eggs. Later studies showed that no fertilization resulted when the sperm and eggs were mixed more than 30 sec after sperm had been placed in water, while eggs retained their ability to be fei-tilized for several minutes when kept in water and for several hours when stored in moist chambers. The early failures to fertilize eggs thus resulted from not utilizing diluted sperm quickly enough. It is well known that sperm may be stored for long periods of time if it is maintained in concen- trated form or is not activated by the diluent. The rapid decrease in the viability of sperm in water is probably important for maintaining the genetic integrity of the spawners; its signifi- cance for practical laboratory work is that sperm should be added to the eggs and not vice versa. Sperm tended to be more active and to re- main motile longer in water from the Salton Sea than in ordinary seawater from La Jolla, Calif. Moreover, developing eggs always floated in Salton Sea water (salinity in 1970, about ;57 /i. on the basis of total dissolved solids) , while they sank in La Jolla seawater (33.5 ',,,). These ob- servations may have important implications for the salinity tolerance and adaptability of Salton Sea fishes (transplanted originally from the Gulf of California), matters of crucial interest in the initiation of this study of fish reproductive physi- ology. Several batches of eggs obtained from hor- mone-induced spawning were allowed to develop to hatching, and a few of the resulting larvae were reared to metamorphosis in the laboratory on a diet of rotifiers, Bianchionus plicatilis,io\- lowed by brine shrimp, Artemia sali)ia, nauplii. Thus the entire life history of the gulf croaker can probably be completed under controlled lab- oratory conditions. This opens up the jjossibility of using this species for many other biological studies where large numbers (50,000-100,000) of pelagic eggs are desired from a marine spe- cies of known genetic history. Some of these studies are now in progress (Robert C. May, Scripps Institution of Oceanography, personal communication) . It is hoped that future studies will include comparative work on this species, especially with respect to the possible adapta- tions of Salton Sea croakers since their sepa- ration from the Gulf of California population. SUMMARY 1. Adult and immature gulf croakers captured by beach seining in the Salton Sea were trans- ported to the Fishery-Oceanography Center lab- oratory in La Jolla, Calif., and used in labora- tory studies on gonad maturation and hormone- induced spawning. 2. A bacterial disease which invariably devel- oped on recently captured or frequently handled fish was successfully treated with Fui'acin anti- liiotic. 3. Long photoperiods (16 hr of light ijer 24 hr) and warm water (22° C) , along with optimal feeding, accelerated the gonadal maturation of females captured prior to their natural cycle of gonadal maturation. These fish were ready to spawn in the laboratory 1 to 3 months prior to the spawning season observed in the Salton Sea. Male fish became ripe under all combinations of laboratory conditions and remained ripe throughout the study. 4. Concomitant field studies confirmed earlier work showing that female croakers ripened in April, while day length was increasing, and spawned when the water temperature reached about 20° C; peak spawning occurred in May of 1969 and 1970. 5. Mature fish, captured during the spawning season at the Salton Sea, quickly resorbed their gonads when held under short photoperiods (10 hr of light) in the laboratory, but similar fish 177 FISHERY BULLETIN: VOL. 69. NO. 1 maintained on long photoperiods (16 hr of light) remained in spawning condition for 2 months (at 22° C) or 3 months (at 14° C) beyond the normal season. 6. At 14° C, injection of mature fish with salmon pituitary, carp pituitary, chorionic go- nadotropin from human pregnancy urine (HCG), and deoxycorticosterone acetate (DO- CA) caused increases in gonad size over sham- injected or uninjected fish. 7. A single injection of 1 mg (acetone dried) salmon pituitary, 50 lU of gonadotropin from pregnant mare serum (PMS) or 50 lU of HCG induced spawning in mature croakers (50-100 g) with gonad index values about 5 "^.'f . Fish with gonad index values below about 5 ^r did not re- spond to otherwise adequate hormone doses. Hormone-spawned fish could be spawned a sec- ond or third time at 1- to 2-month intervals. 8. Eggs could be stripped from the fish an average of 30.4 hr following injection. This latent period consisted of a slow hydration phase of water uptake followed by a rapid ovulation phase which released eggs from the follicles in- to the ovarian lumen. 9. The eggs remained viable only for 1 to 2 hr following ovulation, unless they were stripped from the fish and stored in moist chambers. Each female produced 700 to 1000 eggs per gram of wet body weight. 10. Sperm are viable for less than 30 sec after dispersion in water. 11. Low dosages (0.1 mg) of salmon pituitary were insufficient to cause hydration, while very high dosages (5 mg) caused hydration but, evi- dently, inhibited ovulation. High dosages (100 lU) of HCG caused fish to overhydrate and eventually die without having ovulated. 12. Carp pituitary caused hydration but was inadequate for ovulation. Deoxycorticosterone acetate and oxytocin, given alone, had little or no effect on the fish. 13. Fish did not respond to single hormone in- jections if the water temperature was at or be- low 17° C. One day of acclimation to a higher temperature was sufficient to prepare fish from cold water for spawning. 14. A few larvae hatched from eggs obtained by hormone-induced spawning were reared through metamorphosis; thus, the entire life cycle of the gulf croaker can be completed under laboratory conditions. ACKNOWLEDGMENTS Many people contributed to the total success of this project, a cooperative effort made pos- sible by Alex Calhoun, Chief of Inland Fisher- ies, California Department of Fish and Game, and Reuben Lasker, Program Director for Be- havior and Physiologj', National Marine Fish- eries Service Fishery-Oceanography Center, La Jolla. Their interest made possible the funds and facilities for this study, supported by Fed- eral Aid to Fish Restoration Funds, Dingell- Johnson Project California F-24-R, "Salton Sea Investigations." Robert F. Elwell, Senior Research Supervisor, IFB, Sacramento, saved me from many admin- istrative details and provided aid and encour- agement at critical points in time. Fred Murin, Salton City, was my able field assistant and com- panion; and David Crear, now at the University of Hawaii, was of inestimable value in our lab- oratory ventures. Robert C. May. Scripps In- stitution of Oceanography, La Jolla, contributed much of his time to the successful completion of the study. Technical assistance was provided by Charles F. Wright and Bert D. Kitchens, National Ma- rine Fisheries Service Fishery-Oceanography Center, La Jolla, who built the fish-holding facil- ities and maintained the critical seawater sup- ply. W. H. Jochimsen and D. R. Von Allmen, Nimbus Fish Hatchery, Rancho Cordova, ar- ranged for me to collect salmon pituitaries. Da- vid Powell, Curator of Fishes, Sea World-San Diego, taught me how to transport and care for fish, and Vince Catania, California Deiiart- ment of Fish and Game at Antioch, showed me how to capture Salton Sea fishes. To each of these persons I extend my gratitude for their help, interest, and ideas. I also thank the many other members of the California Fish and Game Department and in- terested Salton Sea sportsmen and merchants 178 HAYDOCK: GONAD MATURATION OF GULF CROAKER that provided aid, comfort, and fellowship which made the sometimes arduous field work a pleasant and rewarding experience. LITERATURE CITED Ahsan, S. Nazar, and William S. Hoar. 1963. Some effects of gonadotropic hormones on the threespine stickleback, Gasterosteus acvle- (itua. Can. J. Zool. 41(6): 1045-1053. Atz, James W., and Grace E. Pickford. 1959. The use of pituitary hormones in fish culture. Endeavour 18(71): 125-129. 1964. The pituitary gland and its relation to the reproduction of fishes in nature and in captivity. An annotated bibliography for the years 1956- 1963. FAO (Food Agr. Organ. U.N.) Fish. Biol. Tech. Pap. 37, 61 p. Ball, J. N. 1960. Reproduction in female bony fishes. Symp. Zool. Soc. London 1: 105-135. Ball, Robert C, a.nd Edward H. Bacon. 1954. Use of pituitary material in the propagation of minnows. Progr. Fish-Cult. 16(3): 108-113. Bardach, John E. 1968. The status and potential of aquaculture. Vol. 2, particularly fish culture. American In- stitute of Biological Sciences, Washington, D.C. Available Clearinghouse for Federal Scientific and Technical Information, Springfield, Va., as PB 177 768, 225 p. Breder. Charles M., Jr., and Donn Eric Rosen. 1966. Modes of reproduction in fishes. Natural History Press, Garden City, N.Y. .w + 941 p. Carpelan, Lars H. 1961. Physical and chemical characteristics. In Boyd W. Walker (editor). The ecology of the Salton Sea, California, in relation to the sport- fishery, p. 17-32. Calif. Dep. Fish Game, Fish Bull. 113. Chaudhuri, Hiralal. 1960. Experiments on induced spawning of Indian carps with pituitary injections. Indian J. Fish. 7(1) : 20-48. Clemens, Howard P., and F. Blake Grant. 1964. Gonadal hydration of carp (Cyprimts cnrpio) and goldfish {Carassius auratus) after injections of pituitary extracts. Zoologica (New York) 49(4) : 193-210. Clemens, Howard P., and Kermit E. Sneed. 1962. Bioassay and use of pituitary materials to spawn warm-water fishes. U.S. Fish Wildl. Serv., Res. Rep. 61, iv + 30 p. Das, S. M., and H. A. Khan. 1962. The pituitary and pisciculture in India, with an account of the pituitary of some Indian fishes and a review of techniques and literature on the subject. Ichthyologica 1(1-2): 43-58. Dodd, J. M. 1955. The hormones of sex and reproduction and their effects in fish and lower chordates. Mem. Soc. Endocrinol. 4: 166-187. Finucane, John H. 1970. Pompano mariculture in Florida. Amer. Fish Farmer 1(4) : 5-10. Fontenele, Osmar. 1955. Injecting pituitary (hypophyseal) hormones into fish to induce spawTiing. Progr. Fish-Cult. 17(2): 71-75. Frogner, Karl J., and John R. Hendrikson. 1970. A technique for coelomic administration of drugs to fish without handling. Progr. Fish- Cult. 32(3) : 142-146. Gunter, Gordon. 1967. Vertebrates in hypersaline waters. Contrib. Mar. Sci. 12: 230-241. Harada, T. 1970. The present status of marine fish cultivation research in Japan. Helgolaender wiss. Meere- sunters. 20: 594-601. Harrington, Robert Whiting, Jr. 1959. Photoperiodism in fishes in relation to the annual sexual cycle. In Robert B. Withrow (editor), Photoperiodism and related phenomena in plants and animals, p. 651-667. AAAS (Amer. Ass. Advan. Sci.) Publ. 55. Hedgpeth, J. W. 1959. Some preliminary considerations of the bi- ology of inland mineral waters. Arch. Oceanogr. Limnol. 11 (Suppl.) : 111-139. Henderson, Nancy E. 1963. Influence of light and temperature on the reproductive cycle of eastern brook trout, Sal- velinut; fontinaUs (Mitchill). J. Fish. Res. Bd. Can. 20(4) : 859-897. Hickling, C. F. 1962. Fish culture. Faber and Faber, London. 259 p. Hoar, William S. 1969. Reproduction. In W. S. Hoar and D. J. Randall (editors), Fish physiology. Vol. 3, p. 1- 72. Academic Press, New York. Hoar, W. S., and D. J. Randall (editors). 1969, 1970. Fish physiology. Academic Press, New York. 5 vol. Hora, S. L., and T. V. R. PiLLAY. 1962. Handbook on fish culture in the Indo-Pa- cific region. FAO (Food Agr. Organ. U.N.) Fish. Biol. Tech. Pap. 14, vii -1- 204 p. Hyder, Mohamed. 1970. Histological studies on the testes of pond specimens of Tilapia nigra (Gunther) (Pisces: Cichlidae) and their implications of the pituitary- testis relationship. Gen. Comp. Endocrinol. 14(1) : 198-211. 179 FISHERY BULLETIN: VOL. 69. NO. 1 Joseph, Edwin B., and Vishnu P. Saksena. 1966. Determination of salinity tolerances in mum- michog {Fii7idtdiis heterocUtua) larvae obtained from hormone-induced spawning. Chesapeake Sci. 7(4) : 193-197. KuNTZ, Albert. 1914. The embryology and larval development of Bairdiella chrysura and Anchovia mitchilli. U.S. Bur. Fish., Bull. 33: 3-19. Lasker, Reuben, and Lillian L. Vlymen. 1969. Experimental sea-water aquarium. U.S. Fish Wildl. Serv., Circ. 334, iv + 14 p. Leitritz, Eabl. 1959. Trout and salmon culture (hatchery meth- ods). Calif. Dep. Fish Game, Fish Bull. 107, 169 p. LiLEY-, N. R. 1969. Hormones and reproductive behavior in fish- es. In W. S. Hoar and D. J. Randall (editors), Fish physiology. Vol. 3, p. 73-116. Academic Press, New York. Malven, p. V. 1970. Interaction between endocrine and nervous systems. Bioscience 20(10) : 595-601. McInerney, John E., a.nd David 0. Evans. 1970. Action spectrum of the photoperiod mech- anism controlling sexual maturation in the three- spine stickleback, Gasterosteus aculeatus. J. Fish Res. Bd. Can. 27(4): 749-763. New, D. a. T. 1966. The culture of vertebrate embryos. Logos Press, Academic Press. London, x ^- 245 p. Perks, A. M. 1969. The neurohypophysis. In W. S. Hoar and D. J. Randall (editors), Fish physiology. Vol. 2, p. 111-205. Academic Press, New York. Pickford, Grace E., and James W. Atz. 1957. The physiology of the pituitary gland of fishes. New York Zoological Society, New York, xxiii + 613 p. Shelbourne, J. E. 1964. The artificial propagation of marine fish. In F. S. Russell (editor), Advances in marine biology. Vol. 2, p. 1-83. Academic Press, London. 1970. Marine fish cultivation: Priorities and progress in Britain. In William J. McNeil (edi- tor). Marine aquiculture, p. 15-36. Oregon State University Press, Corvallis. Sneed, Kermit E., and Howard P. Clemens. 1959. The u.se of human chorionic gonadotrophin to spawn warm-water fishes. Progr. Fish-Cult. 21(3) : 117-120. Stevens, Robert E. 1966. Hormone-induced spawning of striped bass for re.servoir stocking. Progr. Fish-Cult. 28(1): 19-28. Sundararaj, Bangalore I., and Shashi V. Goswaml 1966. Effects of mammalian hypophysial hormones, placental gonadotrophins, gonadal hormones, and adrenal corticosteroids on ovulation and spawning in hypophysectomized catfish, Hetcropneiistes fos- silis (Bloch). J. Exp. Zool. 161(2): 287-295. 1968. Some aspects of induced spawning in the hypophysectomized catfish, Heteropneustes fos- silis. In Symposium on comparative endocrinol- ogy, p. 189-193. Nat. Inst. Sci. India, New Delhi. Tsu-iTiKi, H., p. J. Schmidt, and M. Smith. 1964. A convenient technique for obtaining pitui- tary glands from fish. J. Fish. Res. Bd. Can. 21(3) : 635-637. Walker, Bo^t) W. 1952. A guide to the grunion. Calif. Fish Game 38(3) : 409-420. Walker, Boyd W. (editor). 1961. The ecology of the Salton Sea, California, in relation to the sportfishery. Calif. Dep. Fish Game, Fish Bull. 113, 204 p. Whitney, Richard R. 1961. The bairdiella, BairdieUa icistius (Jordan and Gilbert). In Boyd W. Walker (editor). The ecology of the Salton Sea, California, in relation to the sportfishery, p. 105-151. Calif. Dep. Fish Game, Fish Bull. 113. 1967. Introduction of commercially important spe- cies into inland mineral waters, a review. Con- trib. Mar. Sci. 12: 262-280. Wickler, W. 1966. Breeding aquarium fish; an introduction to the biology of their reproduction [in German]. Transl. by D. W. Tucker. Van Nostrand, Prince- ton, N.J. 112 p. Wiebe, John P. 1968. The effects of temperature and daylength on the reproductive physiology of the viviparous seaperch, Cymatngnster aggrcgata Gibbons. Can. J. Zool. 46(6): 1207-1219. Yamazaki, Fumio, and Edward M. Donaldson. 1968a. The .spermiation of goldfish (Cnrasshis aaratus) as a bioassay for salmon {Oncorlnpichus tahawytscha) gonadotropin. Gen. Comp. Endo- crinol. 10(3) : 383-391. 19fi8b. The effects of partially purified salmon pituitary gonadotropin on spermatogenesis, vitel- logenesis, and ovulation in hypophysectomized goldfish (Carassiits anratus) , Gen. Comp. En- docrinol. 11(2) : 292-299. 180 HARMONIC FUNCTIONS FOR SEA-SURFACE TEMPERATURES AND SALINITIES, KOKO HEAD, OAHU, 1956-69, AND SEA-SURFACE TEMPERATURES, CHRISTMAS ISLAND, 1954-69 GUNTHER K. SECKEL' AND MARIAN Y. Y. YONG'' ABSTRACT Harmonic functions have been fitted to time-series, sea-surface temperatures and salinities in order to facilitate studies of the oceanographic climate near Hawaii and Christmas Island. The manner in which Fourier analysis has been adapted to this application has been described. The standard errors of estimate for Koko Head temperatures and salinities are less than 0.26° C and less than 0.05/^», respectively. The standard errors of estimate for Christmas Island temperatures are approximately 60 % above those for the Koko Head temperature. The expected values of the Koko Head tem- perature and salinity functions have an uncertainty of ±0.1° C and ±0.015^fr, respectively, when samples are obtained twice weekly. Error terms of the Christmas Island temperatures, with daily sampling, are on average 0.07° C. Harmonic analysis spanning the entire sampling duration shows that long-term variations in the Christmas Island temperature and Koko Head salinity are larger than the seasonal variations. Seasonal variations in the Koko Head temperatures are dominant and longer term variations small. The results of the harmonic analyses are presented in the appendixes: (1) a listing of coefficients that define the Koko Head temperature and salinity functions for each year and the Christmas Island temperature functions for each quarter of each year, (2) graphs of the fitted curves together with the observed values for each year. In this paper harmonic functions are presented of sea-surface temperatures and salinities that have been regularly measured near Koko Head, Oahu (lat. 21°16' N., long. 157°41' W.) since 1956 and at Christmas Island (lat. 1°51' N., long. 157°23' W.) since 1954 (Fig. 1). Sea-surface temperatures and salinities change in response to, and therefore reflect, sea-air interaction processes (heat exchange, evaporation minus precipitation) and ocean- ographic processes (advection, diff'usion). For example, the mean sea-surface temperature for a month at Koko Head provides a measure of the mean heat content of the water near the surface. Thus, if the mean temperature for March is above that for February, then meteor- ological and oceanographic processes must have taken place to raise the mean heat content of Figure 1. — Location of Koko Head, Oahu and Christmas Island. ' National Marine Fisheries Service Environmental and Fishery Forecasting Center, Monterey, Calif. 93940; formerly National Marine Fisheries Service Hawaii Area Fishery Research Center, Honolulu, Hawaii. ' National Marine Fisheries Service Hawaii Area Fishery Research Center, Honolulu, Hawaii 96812. 165° 160' 155' 150" W {{ / _ KOKO HEAD OAHU t^ N. c^.HAWAI 1 ¥\ . 10* 10* ^CHRIST* AS 1 165- 160' I55' ISC Manuscript received September 1970. FISHERY BULLETIN. VOL. 69, NO. I, 1970. 181 FISHERY BULLETIN: VOL. 69. NO. I the surface water in March above that in Feb- ruary. This concept was used in studies of the Hawaiian oceanographic climate (Seckel, 1962, 1969) and has been ajipHed to Hawaiian fishery problems (Seckel and Waldron, I960; Seckel, 1963). Rigorously, the theory of distribution of pro- perties in the sea states that the change of sea- surface temperature during a time interval, say from the first day of one month to the first day of the next month, is equal to the integral of all meteorological and oceanographic processes affecting the temperature during the time in- terval: 1^ 8 h — S II = f (all processes) J/. e I, is the temperature at the beginning and di, is the temperature at the end of the interval. In application, the choice of e „ and 0/, presents the following problems: The difference in the observed temperatures at times a and b also reflects the effect of short-term variability ("noise") that is not of interest in monitoring the large-scale events. If one uses monthly mean temperatures in the heat budget equation that include observations made 15 days before and after times a and b, then the change of temperature incorporates the effect of processes that lie outside the interval of interest. Al- though mean values usually provide an adequate measure of the temperature change during given time intervals, the true change of temperature can be obscured. One can overcome the problems caused by the two unsatisfactory methods of obtaining measures of the temjierature change by finding suitable functions that filter out un- desirable short-term variability without obscur- ing the basic temperature and salinity trends. Techniques that can be used in the smoothing of time series data have been reviewed by Hol- loway (1958) and usually involve moving aver- ages of the data to which weighting factors have been assigned. Curve fitting provides another method of aji- proach. A useful technique that has been used in this report, is to obtain an analytic expression for the temperature and salinity as a function of time by Fourier analysis. The Fourier series is efficiently, and therefore inexpensively, de- rived by computer. Efficiency is furthered in that graphs can be produced by automatic plot- ter. The Fourier series provides a least-squares fit of the observed values. It permits filtering of undesired variability, facilitates statistical evaluation of the data, and — within limits — pro- vides insight into the properties of the distri- bution. These advantages will become apparent in the following sections of this report. The results of the analyses for each year of observation are presented in the appendi.x in both tabular and graphical form. THE FOURIER METHOD Fourier series are well known, widely applied, and adequately described in texts of advanced calculus. A good description can be found in Sokolnikoff (1939) where the derivation of the Fourier coefficients by least-squares method is also presented. The temperature or salinity is expressed as a function of time, t, in the Fourier series: .S„ (/) = h L (/l„cos/;a)' + B,is\nnu>i), where w 2tt 1,2,3, , and T is the fundamental period. For example, if harmonic analysis is to be performed on data collected for a dura- tion of 1 year, T would be 365 days. The Fourier series contains the coefficients .4(1, A„, and B„ that are given by the Fourier integrals 2 An = and «,, = i=T T ^0 F{!)cos{nui)ili. n 0.1,2, F(thin{nui)dr. n = 1,2.3, The coefficient Aq is the special case of A^i with n — 0. In our application F(i) is the temper- ature or salinity at the time t. Of course, the functional relationship between temperature and time or salinity and time is not known so that 182 SECKEL and YONG : HARMONIC FUNCTIONS F{t) is the observed temperature or salinity at the time t. Furthermore, F{t) is known only at finite intervals of time so that the above Fourier integrals must be obtained by numer- ical integration. This integration, approxi- mating the area under the curves F{t) cos (nU) and F{t) sin (?i<.>f), is performed by summing areas of rectangles with height G(t) cos (wuO or G{t) sin (no>t), and with width Af, the sampling interval. The finite difference form of the Fourier in- tegrals is A„ = and X G{l)iCos{nut)Ati. n = 0,1,2, ...k. B„ = — X G(l)iSm(nut)M,. n = 1,2,3, ...k. • i = \ The number of samples in the interval ( io t = T \s m + 1, and G(t)i = ViiFU,) 4- F(/,. i)], / = 1,2,3, ...m. The time used to evaluate the geometric factor is 1/2 (^' + i'-\)- Other schemes of obtaining the best estimate of G(t) cos {riud) during the interval Ai can be used but would not signifi- cantly affect the results in our application (see Kaplan, 1953: p. 168-172). Library programs for the evaluation of Four- ier coefficients by computer usually require that the sampling interval, A?, be constant. Since this condition is not necessarily met in our ap- plication, a more flexible computer program was written to evaluate the coefficients. In this program the sampling interval may vary, and the number of samples for the basic period of analysis need not be the same in each application. The Fourier coefficients evaluated in the above manner enable us to describe anal.vtically the temperature or salinity as a function of time. If we wish to go further and gain insight into the properties of the temperature or salinity distribution, it is more useful to express the Fourier series as a sum of cosines: S„ (!) = — + X Cicos u(ni — a„), 2 " n = 1,2,3 k. The transformation is accomplished by the use of the trigonometric indentities A„ = C„ cos wa„, B,, = C, sin uia.i. and C„=±{Al+ B^y\ B„ ua„ — arctan - A, In the application described in this report the fundamental period in the Fourier series is the sampling duration or any portion of this duration that may be arbitrarily chosen; the amplitudes and phase angles do not necessarily coincide with natural variations in temperature or salinity; and the harmonic functions have no predictive value. In some cases, such as the Koko Head tem- peratures with a well-defined annual cycle, the fundamental period of the Fourier series de- rived for each year approximates the annual cycle. At Christmas Island, however, an annual temperature cycle is not always clearly apparent. Despite the fact that choice of the fundamental period may be arbitrary and may not coincide with a naturally occurring period, the spectrum is resolved beyond the first few harmonics. For example, if the fundamental period, n — 1, is 12 months then the period of the first harmonic, n = 2, is 6 months. A naturally occurring 9 months cycle in the observations would in this case not be resolved. As n increases, however, resolution improves to 4, 3, 2.4, 2, etc., months. The highest harmonic, or ?i-value, to which harmonic analysis can be carried, is limited by the number of observations. In the ideal case and when samples are equally spaced in time, there must be at least 2n observations, i.e., at least two samples per cycle. In nature, where we are dealing with noncyclical variations and unequal spacing of samples a sinusoidal curve cannot be resolved with only two samples, and 183 FISHKRY BLXLETIN: VOL. 69. NO. 1 a minimum of four or, better, six samples is required to achieve good resolution. For ex- ample, sea-surface temperatures are to be mon- itored and the fundamental period of observa- tions is to be 12 months. Resolution of a 1- month cycle (?i = 12) , requires four samples per month, or sampling once per week. APPLICATION OF THE FOURIER METHOD In practice, the Fourier method described above must be adapted to each specific applica- tion. In addition to the minimum number of samples necessary in order to attain a desired resolution another restriction applies to vari- ations in the sampling interval. Although the computer program used to obtain the results of this paper allows a varying sampling interval, thus accepting a sequence with missing obser- vations, the sampling interval can be allowed to vary only within limits. For example, at least four samples per month are necessary to resolve a monthly cycle. This cycle will, how- ever, not be resolved if the samples are taken on four consecutive days, rather than being evenly distributed throughout the month. It is also possible to aid the hai-monic analysis in rapid convergence to its best fit with the ob- served values by adjusting the fundamental period of analysis and by performing some pre- liminary operations which are described below. APPLICATION TO KOKO HEAD SEA- SURFACE TEMPERATURES AND SALINITIES The sampling station is located near Koko Head at the exposed, eastern shore of Oahu so that the sea-surface temperatures and salinities measured there reflect open-ocean conditions. The salinities appear to be affected by runofl' only on rare occasions of heavy rainfall. Both the tempei'atures and salinities are based on bucket samples. The salinity is determined in the Hawaii Area Fishery Research Center, Honolulu. Before 1961 samples were collected at weekly intervals and subsequently twice weekly, usually on Tuesday and Friday mornings. Occasionally sampling has been missed. The computer pro- gram must therefore accept data with an ir- regular sampling interval. The basic period for analysis has been chosen to be 1 year. Harmonic analysis began with the first sample and ended with the last sample of the year. The sampling time, in days and months, was converted to days of the year be- ginning with the first of the year. Owing to a longer term trend, the value of a property at the beginning is not necessarily the same as at the end of an annual cycle. In the case of Koko Head salinities and Christmas Island temperatures, it will be seen later that an annual cycle is, in fact, not always apparent. The noncyclic trend during the analysis period can be obtained by linear approximation. Rapid convergence to the best fitting function can then be achieved by performing the harmonic anal- ysis on the residuals of the observed values from a linear fit. In our application the first observed value, F(to), and the last observed value, F(ti), for the period were used to obtain the linear equation S' = F(?o) + bi FUi) - FUo) where b = • '/ - '0 The residuals, /?„, = F(t,n) — [F(to) + bt^}, m — 0.1,2, ... /, were used to obtain the Fourier coefficients. The Koko Head temperatures and salinities for each year are then expressed by the function k S = K + bt + ^ C„ coK 1 2 3 '. 5 6 7 8 9 10 11 12 13 llib 0.21 0.19 0.17 0.16 3.16 0.1'. 0.1'. 0. I', n.i3 0.13 0.12 0.12 0. U 1957 t). 36 0,29 0.2'. 0.2* 0.23 0.23 0.23 0.21 0.20 0.20 0.19 0.19 0.18 1S53 0.31 0. 29 0.2'. 0.24 0.22 0.22 0.22 0.22 0.22 0.20 0.20 0.18 0.17 1<)59 0.9 O.-^S 0.38 0.36 0.3'i 0.30 0.28 0.27 0.27 0.27 0.27 0.26 0.26 1966 0.<.3 U.32 0.32 0. 30 0.30 0.27 0.27 0.27 0.26 0.26 0.26 0.26 0.26 196 7 0.45a 0.06 9 0.066 0.059 0,059 0.056 19 59 0.174 0.099 0.076 0,075 0.074 1960 0.090 0.083 0.081 0,078 0.077 1961 0.06 4 0.061 0.051 0,047 0.04 3 196 2 0.049 0.046 0. 046 0.044 0.043 1963 0.054 J. 53 0.052 0.046 0.045 196 4 0.086 0.078 0.069 0.063 0.061 1965 0.094 0.085 0.078 0.072 0.072 1966 0.044 0.043 0.042 0.037 0.037 1967 0.079 0.07S 0.07'. 0.072 0.068 196 3 0.060 0.057 0.051 0.046 0.042 0.030 0.030 0.029 0.028 0.025 0.041 0.039 0.037 0,034 0,034 0.053 0.053 0.052 0.049 0.048 0.073 0.069 0.064 0.060 0.058 0.075 0.069 0.063 0.356 0.053 0.038 0.037 0.036 0,033 0,030 0,041 0.037 0.037 0.036 0.035 0.045 0.043 0.041 0.037 0.037 0.059 0.052 0,052 0.051 0.044 0.071 0.066 0.064 0.058 0.056 0.034 0.U34 0.033 0.033 0.030 0,061 0.055 0.054 0.052 0,051 0.040 0.038 0.038 0.035 0.C35 0,025 0.028 0.030 0,032 0,030 0.030 0,041 0,036 0.036 0.054 0.053 0.054 0.050 0.042 0.038 0.027 0.028 0.02 8 0.034 0.034 0.034 0.034 0.033 0.033 0.044 0.04 3 0.0 38 0.054 0.048 0.045 0.030 0.029 0.028 0.050 0.046 0.044 0.033 0.033 0,033 187 FISHERY BULLETIN: VOL. 69, NO. 1 Table 3.— Standard error of estimate (° C) for each quarterly temperature function at Christmas Island, 1954-68, with harmonic analysis carried out in sequence to n =r 1, 2, 3, . . . and 7. N -VALlJf S YEAR QUARTER 1 2 3 4 5 6 7 1954 0.-.4 0.41 0.41 0.41 0. 32 0.30 0,29 0.36 0.35 0.33 0.33 0.37 0.30 0.30 0.51 0.50 0.50 0.50 0. 47 0.45 0.44 J. 39 U.33 0.37 0.33 0. 30 0. 30 0.29 19 65 0.3 0.2R 0.27 0. 26 0.26 0.26 0.25 0.29 0.29 0.28 0.28 0.28 0.27 0.26 0.3<. 0.33 0. 33 0. 33 0.32 0.32 0.32 0.46 0.45 0.43 0.42 0. 40 0.39 0.39 1956 0.40 0. 38 0. 38 0.37 0.36 0.35 0.35 0.52 0.50 0.4 8 0.48 0.46 0.45 0.45 0.43 0. 47 0.45 0.44 0.43 0.41 0.41 0.38 0.39 0. 36 0.3s 0.36 0.32 0.32 1957 0.48 0. 46 0.45 0.44 0.43 0.43 0.43 0.61 0.54 0.54 0.54 0.53 0.51 0.51 0.44 0.44 0.43 0.43 0.40 0.39 0.38 0.40 0.39 0.35 0.34 0.33 0.30 0.28 1958 0.26 0.25 0.24 0.24 0.24 0.23 0.23 0.33 0.33 0.33 0.32 0.3 1 0.30 0.29 0.37 0.35 0.30 0. 29 0.28 0.28 0.28 0.32 0.31 0.28 0.27 0.26 0.25 0.25 19 59 0.41 0.34 0.30 0. 28 0.28 0.27 0.27 J. 40 0.38 0.36 0.35 0. 34 0.34 0.33 0.48 0.39 0. 36 0.34 0.37 0.32 0.32 0.43 0.39 0.36 0.32 0. 31 .29 0.29 1960 C.30 0.29 0.27 0.2b 0.26 0.25 0.25 0.35 0. 33 0.32 0.31 0.31 0.31 0.31 0.32 0. 31 0.30 0.27 0.26 0.26 0. 25 T.39 0.32 0.29 0.26 .2 6 0.24 0.23 1961 0. 36 0.34 0.34 0.32 0.31 0. 30 0.27 0.34 0.34 0.31 0.27 0.26 0.26 0.25 0. 36 0.29 0.29 0.28 0.27 0.25 0.25 0.2 6 0.24 0.22 0.22 0.20 C. 19 0. 18 1962 .39 0.34 0.30 0. 30 0.30 0.2 7 0.27 0.38 (1.33 0.31 0.30 0.29 0.27 3.25 0.26 0.74 0.71 0. 20 0.70 0.1 9 0.19 0. 30 0.26 0.25 0.24 0.24 3.24 0.24 196 3 0.36 0.36 0.34 T.34 0.32 0.32 0.28 0. 46 0.38 0.31 0.3 .78 0.27 0.71 0.36 0.29 0.29 0.77 0.26 0.25 0. 24 0.30 0.29 0.28 0.26 0. 26 O.Z' n.25 1964 0. 32 0.32 0.31 0.30 0.79 0.29 0.28 0.37 0.31 0.30 0.29 0.28 0.28 0.27 0.34 0.31 0.79 0.29 0.28 0.27 0.26 0.28 0.27 0.26 0. 73 0.22 0.21 0.21 1965 0. 30 0.29 0.28 0.27 0. 26 0.75 0.25 0.36 0.32 n. 79 J. 29 0.29 0.29 0.29 0.53 0.52 0.45 0.42 0. 39 0.38 0.37 0.37 0.32 0.31 0.30 0.30 0.30 0.29 1966 0.42 0.39 0.35 0. 34 0.33 0.30 0.29 0.42 0.35 0.35 0.34 0.34 0.31 0.30 0.60 0.51 0.48 0. 46 0.43 0.42 0.40 0.68 0.68 0.53 0.52 0.51 0.51 0.47 1967 0.42 0.40 0.34 0.34 0.34 0.32 0.32 0.40 0.39 0.36 0.36 0.36 .36 0.35 0.39 0.38 0.37 0.36 0.34 0.33 0. 33 0.32 0.30 0.28 0.27 0. 27 0.27 0.26 1968 0.42 0.37 0.36 0.33 0.33 0.33 0.32 0.37 0.35 0.31 0. 11 0.31 0.30 0.30 0.29 0.29 0.29 0.30 O.'l 0, 78 0.28 0.28 0.27 0.26 0.24 0.23 0.21 0.21 188 SECKEL and VONG; HARMONIC FUNCTIONS in 1959 and 1965 when it is 0.054/rr, and 0.045%f, respectively. At Christmas Island (Table 3), the average standard error of estimate at n = 4 (resolution of 1 month) is near 0.33° C and therefore about 60 % higher than that for the Koko Head temperatures. As previously mentioned, high temperature variability is to be expected at the Christmas Island sampling site. A standard error of estimate based on all samples used to obtain a function obscures the month-to-month changes in variability that may have occurred. At Koko Head the month-to- month changes in temperature variability as re- flected by the standard error of estimate for each month ranges from 0.05° to 0.45° C, the same values for the Koko Head salinities range from 0.006%<. to 0.136%f , and those for Christmas Island temperatures range from 0.17° to 0.66° C. Assuming that sampling error remains constant, the range of variability reflects changes in oceanographic conditions. The standard error of estimate computed from the temperature and salinity observations of each month also reflects sampling quality in that low values indicate the residual variability in the ocean plus sampling error. For the Koko Head temperature, low values of the monthly standard error of estimate are near 0.1° C and for the Koko Head salinity they are near 0.02%c. The sampling error is therefore with- in ±0.1° C for the temperature and ±0.02^0 for the salinity. These are the limits to be ex- pected when bucket sampling of the temperature and salinity is carefully done. Finally, how is the quality of fit affected by sampling frequency and how reliable are the expected values that may be obtained from the harmonic functions? The constraint imposed by the sampling frequency on the resolution that may be attained by harmonic analysis has al- ready been discussed. The present question con- cerns improvement of fit when the sampling frequency is increased above the minimum re- quirements. At Koko Head the sampling frequency was increased from once to twice weekly in 1961. No significant change can be seen in the stand- ard errors of estimate listed in Tables 1 and 2 as a result of doubling the sampling frequen- cy. This observation is consistent with results obtained from oceanographic data collected at Ocean Weather Station "P" in the Gulf of Alaska. Tabata (1964: Table 8) lists the monthly mean value and the standard deviation of the temperature at 10-m depth based on data obtained twice daily, data obtained every second, third, fourth, fifth, sixth, and seventh day of July 1959 and May 1961. For July 1959 the mean temperatures range from 10.70° to 10.81° C and the standard deviations range from 0.60° to 0.76° C. For May 1961 the mean temper- atures range from 5.84° to 5.90° C and the standard deviations range from 0.39° to 0.46° C. In May 1961 Koko Head temperatures and salinities were sampled on 25 days. The mean of all temperature observations was 24.67° C with standard deviation 0.27° C. The mean of temperatures taken every fifth day was 24.58° C with standard deviation 0.39° C. The mean of all salinity observations was 34.759%o with standard deviation 0.051%r. The mean of salinities taken every fifth day was 34.772%o with standard deviation 0.058%^. The temper- ature results from Koko Head are comparable to those from Ocean Weather Station "P" in that mean values and standard deviations based on diff'erent sampling frequencies fall within ap- proximately the same range. The standard er- rors of estimate for the May 1961 Koko Head temperatures and salinities, based on the har- monic functions with resolution of 1 month, are lower than the standard deviations, namely, 0.25° C and 0.02T/,c, respectively. The stand- ard errors of estimate as well as the standard deviations do not change significantly when the sampling frequency is increased above the re- quired minimum to attain a desired resolution by harmonic analysis. Increasing the sampling frequency does, how- ever, improve the confidence limits of a mean value or the expected value of a harmonic func- tion. A good measure of the confidence limits of a mean value is the standard error of the mean (the standard deviation divided by the square root of the number of samples) . Return- 189 FISHERY BULLETIN: VOL. 69. NO, 1 ing to Tabata's table the standard error of the mean for July 1959 is for twice daily sampling every day 0.086° C, and for twice daily sampling every seventh day 0.253° C. For the same sampling frequencies in May 1961 the standard errors of the mean are 0.053° and 0.15° C, re- spectively. For the May 1961 Koko Head tem- peratures the standard error of the mean is 0.055° C with 25 samples and 0.16° C with sampling every fifth day. The standard error of the mean for the May 1961 Koko Head sa- linities is 0.010'/,, with 25 samples and 0.024',, with sampling every fifth day. On the basis of these considerations, the expected values ob- tained from the temperature functions have an uncertainty of ±0.10° C, and those from the sa- linity functions have an uncertainty of ±0.015'/(c when samples are obtained twice weekly. At Christmas Island temperatures are sampled daily rather than twice weekly as at Koko Head. In consequence, despite the larger variability, expected values obtained from the harmonic functions have approximately the same uncer- tainty as those obtained from the Koko Head harmonic functions. This statement is con- firmed by considering the error terms that can be obtained by taking the difference of the ex- pected values at the midpoint of the 30-day over- lap portion of the Christmas Island temperature functions (see appendix D). On average this er- ror term is 0.07° C and ranges from to 0.26° C. SOME PROPERTIES OF THE TEMPERATURE AND SALINITY DISTRIBUTIONS Although the harmonic functions are merely analytic expressions of the temperature and sa- linity as a function of time, they do provide, to some extent, insight into the nature of the dis- tributions. For instance, the monthly standard error of estimate, mentioned in the previous section, provides a measure of the month-to- month changes in variability. At Koko Head there is no seasonal pattern in this variability of the temperature; however, there is a seasonal pattern in the variability of the salinity. The monthly standard errors of estimate of the sa- linity function with harmonic analysis carried out to n = l;i, are listed in Table 4. Table 4. — Standard error of estimate (/(c) for each month, 1956-68, of the Koko Head salinity. Harmonic analysis is carried out to n = 13. MONTH YEAR 1 2 3 4 5 6 7 8 9 in 11 12 iq^o O.OIO 0,017 0,02 7 0.04H 0,064 0.052 0.02 4 0,008 0.012 0.014 0.014 0.015 195 7 o.c^ 0.013 0,021 0.015 0,030 0.034 0.02 9 0.017 0.036 0.031 O.OIH 0.034 1958 O.OCo 0.041 0,052 0.049 0,059 0.02 6 0.02 8 0.028 0.013 0.02 3 0.044 0.022 1959 0.0 'i9 0.035 0,044 0.136 0,040 0,036 0.054 0.02 3 0.047 0.032 0.023 0.035 1960 CO'.? 0.032 0.0 18 0.019 0,056 0.043 0.075 0.035 0.033 0.014 0.014 0.024 1961 0,036 0.019 0.017 0.0 19 0.027 0.054 0.01 1 n.020 0.025 0.021 0.023 0.023 1962 0,054 0.0 40 0,064 0.021 0.013 0.J23 0,02 5 0,033 0.031 0.031 O.niB 0.02 7 1963 0.CZ9 0.026 O.Oltl 0.073 0.045 0.036 0.02 1 0.025 0.0 20 0.022 0.032 0.036 1964 0,031 0.033 0.031 0.030 0.0 29 0.019 0.035 0.050 0.053 0.052 0.024 0.036 1965 CO** 0.053 0.059 0.092 0.03? 0,043 0.034 0.016 0.018 0,033 0.019 0.019 1966 ,026 0.016 0.011 0.014 0.072 0,021 0.011 0.012 0.016 0.03? 0.065 0.036 1967 0,026 0.0 29 0.097 0.055 0.050 0.015 0.019 0.021 0.017 0.031 0.0 29 0.056 1968 0.034 0.024 0.057 0.041 0.040 0.019 0.035 0.026 0.018 0.021 0.016 0.038 190 SECKEL and YONG; HARMONIC FUNCTIONS In each year excepting 1957, 1964, and 1966, highest variability occurred during the first 7 months of the year. In 1957 a seasonal pattern was not clearly apparent and in 1964 and 1966 highest variability occurred during the last 5 months of the year. Although the seasonal pat- tern of variability has not been examined in de- tail, it is consistent with the results of previous studies (Seckel, 1962, 1969). First, Hawaii is located in the vicinity of a relatively high sa- linity gradient that delineates the boundary of the North Pacific Central Water. Thus, the salinity measured at the Koko Head sampling station is sensitive to variations in the location of this water type boundary. Secondly, north- ward displacement of water (warm advection) tends to occur during the first 7 months of the year. In consequence the water tyi^e boundary that generally lies south of the Koko Head sampling station during autumn and winter is brought to within the vicinity of the sampling station. The months with higher variability tend to be associated with declines in the Koko Head salinity. Insight into the nature of the distributions is also obtained by examining the spectra of the harmonic functions. It is evident from the fig- ures in appendix B, that considei-able temper- ature and salinity variability at Koko Head occurs with timespans of 35 to 60 days. Rather than showing the amplitudes for each harmonic of every function, the 13-year mean of the ab- solute magnitude of amplitudes for each har- monic of the Koko Head temperature and sa- linity functions is presented in Figure 2. For both the temperature and the salinity, the amplitude of the annual cycle (n = 1) is largest. The am]ilitudes then decline rapidly with in- creasing harmonics ton =5. In the case of the temperature, a slight increase in amplitude oc- curs at M = 6 and )i = 9. Similar small in- creases in amplitudes occur in the case of the salinity at w = 7 and n = 9. The increased amplitudes at « = 6 and w = 7, resolving 60- and 52-day periods, reflect the climatic signals described by Seckel (1962, 1969). The in- creased amplitude at w = 9, resolving a 41-day period, reflects shorter term variability that may be due to large geostrophic eddies with dimen- I 1 I 1 1 I I T -r 1 1 1 1 11 I 1 r Mill llllllii I 2 3 4 5 6 7 8 9 10 II 12 13 365 182 122 91 73 61 52 46 40 36 33 30 28 PERIOD IN DAYS Figure 2. — Mean magnitude of amplitudes for each harmonic of the Koko Head temperature and salinity functions, 1956-69. sions near 200 km (Wyrtki, 1967) or eddying flow near the Hawaiian Islands. LONG-TERM HARMONIC FUNCTIONS Long-term harmonic functions with the fun- damental period spanning the entire duration of observations, can be obtained by the method described before in this paper. Temperatures and salinities were used as computed for the 191 FISHERY BULLETIN: VOL. 69. NO. I 1st and 16th of each month from the harmonic functions whose phase angles and coefficients are tabulated in appendixes A and C. Harmonic analysis was carried to w = 42 for the Koko Head temperature and salinity, and n = 48 for the Christmas Island temperature, giving in each case a 4 months' resolution. The fitted curves resulting from this analysis are shown in Figure 3, together with the values that were used as input data. Clearly the annual cycle forms the dominant signal in the Koko Head temperature curve. In the Koko Head salinity and Christmas Island temperature curves longer term changes are more pronounced than the an- nual cycle. The relatively large deviations of the input data from the long-term function are to be ex- pected. The figures of appendixes B and D show that variations with a duration of less than 4 months can be relatively large and are not re- solved by the long-term analyses made. The spectra of the long-term harmonic func- tions for the Koko Head temperatures and sa- linities and the Christmas Island temperatures are shown in Figure 4. As is also apparent from Figure 3, the spec- trum of the Koko Head temperature function is distinct fi'om those of the Koko Head salinity and Christmas Island temperature functions. In the former the 12-month period has the most pronounced amplitude, but in the latter two, al- though the annual period has a large amplitude, the amplitudes of longer period changes are large and for some periods exceed those of the annual period. CONCLUSION The results of this paper show that sea-surface temperatures and salinities regularly monitored at island sampling stations can be expressed by harmonic functions of time. Advantages of an- alytic expressions for the temperature and salin- ity were cited in the introduction. Important applications will be in climatic oceanography where one may wish to filter out undesired "back- ground noise." At Christmas Island, for ex- ample, the short-term variability with a dura- tion of 1 month or less can be filtered out by using only the harmonic terms to n = 3 in the quarterly functions. At Koko Head, the vari- ability with duration of less than 50 days, that may be due to large geostrophic or island-in- duced eddies, can be filtered out by using only the harmonic terms to w = 7 in the annual functions. We mentioned in the introduction that the rates of change of temperature reflect the cli- matic processes of change and that distortions or aliasing may occur when monthly mean temperatures are used to compute the change of a property. Consider, for example, the Christ- mas Island temperatures from March to May 1968 (appendix D, days 61 to 152). In Table 5 are listed the monthly mean observed temper- atures, the month-to-month changes of mean temperature, the expected temperatures from the harmonic functions for the 16th of each month (computed with harmonic terms up to w = 4), and the month-to-month changes of expected temperatures. It is clear from this illustration that the use of mean values would result in an underestimate of the rise in temperature from March to April, and would obscure the decline in temperature from April to May. The ex- ample is not isolated and other instances can be found in both the Koko Head and the Christ- mas Island data. Table 5. — Month-to-month temperature differences using mean observed temperatures and expected tempera- tures from the harmonic function, Christmas Island, March to May 1968. Mean temperature Change of mean temperature Expected temperature Change of expected temperature March 1968 April 1968 May 1968 ° C 25.1 26.0 26.2 ' C 09 0.2 ° C 25.1 26.3 26.0 ' c 1.2 -0.3 The results also aid in the choice of an opti- mum sampling frequency. Both the desired confidence limit and the desired resolution must be considered. If the harmonic functions are to be used in monitoring the oceanographic climate as is the case of those presented in this paper, then the limits of about ±0.1° C for the expected temperature value and ±0.02%ppp'^CD'^O'O00^O(i)rOO'^int\JO00U)^tvJ — CJ>0DN.inVfOrj— o S^towioo)^— odiDtn^cNjcvi — oo>(T>(Ijcy^oa>Ki0vc\JO'D>fi"O''iw— oiCDKiomvKiw PERIOD IN MONTHS Figure 4. — Spectra of the long-term harmonic functions for Koko Head temperatures, 1956-69, Koko Head salinities, 1956-69, and Christmas Island temperatures, 1954-69. 194 SECKEL and YONG : HARMONIC FUNCTIONS suming that temperature and salinity samples are of Koko Head quality, then for a resolution of 1 month, weekly sampling is sufficient. Oc- casionally, however, a scheduled sample is not taken or an erroneous value must be eliminated. In such cases sampling gaps would become too large for the desired resolution. Undesirable sampling gaps can be avoided by doubling the minimum sampling frequency. The simplicity and economy of deriving har- monic functions by computer are of practical value, particularly in the analysis of data sampled automatically. By this method large quantities of data can be brought into useful form rapidly. The results of this paper, based on manual sampling, are useful in the investigations of changes with a duration of more than 1 month. Automated sampling would broaden the spec- trum and permit analyses of shorter term var- iations such as diurnal changes, changes of tidal period, and other changes with durations of less than 1 month. Automated sampling would also improve the quality of data since instruments can be placed in locations where undesirable variability is min- imized and where manual sampling is difficult. At Koko Head, for example, samples are ob- tained from an exposed rock ledge where the island effects on the temperature and salinity are small. At Christmas Island, however, the sampling site is convenient and the best obtain- able for manual sampling, but it is not the best in terms of monitoring open-ocean temperatures. This shortcoming is often also the case when temperatures and salinities are measured at tide stations located in protected bays or harbors. The value of regularly monitoring the sea- surface temperatures and salinities has been demonstrated in many instances. For example, empirical relations between Koko Head temper- atures and salinities and the availability of skip- jack tuna to the Hawaiian fishery have been demonstrated (Seckel, 1963). Bjerknes (1969) has shown the relationship between anomalously high equatorial sea-surface temperatures using primarily Canton Island observations, and the intensification of the North Pacific westerlies and trades. This relationship must, in turn, affect temperatures and salinities in the North Pacific. In view of these factors, serious consideration should be given to the establishment of auto- mated sampling stations at selected islands in the Pacific. The derivation of harmonic func- tions, as demonstrated in this paper, would make reduction of data into usable form simple and economical and so facilitate the study of pro- cesses which govern the climate in both ocean and atmosphere. LITERATURE CITED Bjerknes, J. 1969. Atmospheric teleconnections from the equa- torial Pacific. Mon. Weather Rev. 97(3) : 163-172. HOLLOWAY, J. LeITH, JR. 1958. Smoothing and filtering of time series and space fields. Advances in Geophysics 4: 351-389. Academic Press, New York. Kaplan, Wilfrbh). 1953. Advanced calculus. Addison-Wesley Pub- ishing, Cambridge, Mass., 679 pp. Seckel, Gunter R. 1962. Atlas of the oceanographic climate of the Hawaiian Islands region. U.S. Fish Wildl. Serv., Fish. Bull. 61: 371-427. 1963. Climatic parameters and the Hawaiian skip- jack fishery. In H. Rosa, Jr. (editor), Proc. World Sci. Meet. Biol. Tunas Related Species. FAO Fish. Rep. 6, 3: 1201-1208. 1969. The Hawaiian oceanographic climate, July 1963-June 1965. Bull. Jap. Soc. Fish. Oceanogr., Spec. No. (Prof. Uda's Commem. Pap.), pp. 105- 114. Seckel, Gunter R., and Kenneth D. Waldron. 1960. Oceanography and the Hawaiian skipjack fishery. Pac. Fisherman 58(3): 11-13. Sokolnikoff, Ivan S. 1939. Advanced calculus. McGraw-Hill Book Co., New York and London, 446 pp. Tabata, S. 1964. A study of the main physical factors gov- erning the oceanographic conditions of station P in the northeast Pacific Ocean. D. So. thesis, Univ. Tokyo, 264 pp., 55 figs., 17 tables, 41 pp. appendixes. Wyktki, Klaus. 1967. The spectrum of ocean turbulence over dist- ances between 40 and 1000 kilometers. Deut. Hydrog. Z. 20(4) : 176-186. 195 FISHERY BULLETIN: VOL. 69, NO. ! a, CO a I a s e o u SB 0) o u fl CS bo j5 X s C5 3 j3 C3 O cS M Sh 7 0) 3 -S a '3) 0) J3 to >. cS T3 S '■0 T3 bo c Oh IT o IT 00 in o> CD a: 4- c X CD (N; nT in - a- *0 1 (*> o m. (NJ f\j in (N) in CO C -0 C in (D in IN. in rsj CC in (T CO c IT o o in o 00 g rvj in ■c INJ 1 (NJ in m 1 •* in r«-i (Nl in (T in 1 cr 1 in 1 CT' IN; fNJ 'C in CD ^ 4" cr 0- CC 0- 1 o 1 (M 1 -4- in CD 00 (T (NI •0 in 03 in 4- (NJ cr CN cn 00 1 IT CO fNJ O -0 CD o o CD (\J O (T J- cr (M 0- CD o 0- in in 1 ■c (\j (T o 1 CD o in in in (N P»- (-1- 4- pr. (NJ ^ 00 in CD in (T ir> - (NJ in {\J O (NI (NJ CD •J- (Nl O IT 1 4- 1 o 1 O 00 CC 0 CD * in in IT CD CD o in in 1 o 1 00 in t in o O ■^ 03 CD CT fSJ (NJ o o 0- (NI in o 4- (NJ •o (NI o CO (M eo IT 1 CD m 1 in 1 in 1 O ^ 1 (T in CD O 4- CD "• (NJ ? >f CD in in c CD ^ * - 4- 00 tD o o cr in (VJ m 1 in - 4- fSl oc h- (N) o (Nl ■a •c CT 1 't in 1 CO (Nl 1 in in *0 -0 f- %D 1 PM 00 CD IT ^ cr in * (^ in CD (Nl CO CO (^ m in in ■0 -0 (NJ o in CO in .0 in r- tn 0^ o ■NJ c o m o •J- o o in ■a o m in o in CO o o o .0 -0 c J- o in o (NJ O D O 1 o 1 o o o o ( o o o o o 1 o O O ■n CT- D O o o o o in (M CD O -J- PVJ o in o o in o ^0 in o in CO in o in m •4- O Z) O o o O o o o O o O o o 1 O ( O ?• CD D - •J- CO m <0 CD O in o 0- o m •4- O in o o in in o O pg o 1 1 o o 1 o 1 O 1 o o 1 o O 1 o 1 o 1 o o 1 o 1 J- o ^ o -i m o in m o in O CO St o CD in o -t o in CD o CO IT. m o CO o o 1 o O 1 1 O O O t o 1 o o 1 o o O o _. o o 00 in o o o 0* in in fM CO ro in m o ■*• o in — (M 1 ) 1 1 Y 1 1 1 4- a -< m o o o o o o CD o o o CD O o o o o o O o O O 00 o o o m o o o o o O O o 00 IN O o o o o o o o o o 1 o o o o o o o 1 o 1 o o o o o in CD CD ir m m •4- (M m fNI m in 00 -r 1^ o o OD -r o o o in CD in o <» cr IM in fNJ in fM •J- 0 cc CL- (M in fM 'J- CD cc o •4- m m -J- r^ r^ in CO 1 -a- fM fM o r^ -0 o 1 in o oc fM O fM cc cr O cr IN in -0 cr cr fM m fM CD fM fM 00 1 •J- r^ 'M ir 1 fM 00 o cr o r- 1 -T 00 ■J- ^ f- in o f\J cr o cr r*- in in *n r- fM ft) cr CO fM O oo f\l o fM 1 00 1 CD ro 1 tM fM in 1 I f^ in 1 in CO cr 1 a- o o •J- CT CT (M cc 00 IT, h- -r (M fM (Nl in O O cr o cr f^ in cr fSJ 1 o O 0^ o in 'J- fM in ■^ fvj 1 1 1 IT o ao CD in fM O cr in o fM m cr rg fM cr cr •C CO - 1 00 in 1 (M in 1 00 in in cr fM o cr 1 cr <£ ro o o nC fM in cr fM f*1 o fM ■4- in 0" IN cr fM 0- cr cr IT in 1 CD CO 1 in 1 rf\ fM f*^ 1 1 cr 1 1 00 f^ 1 m I a o CD in rg fM O O CJ* cr cr o o in in ■C o o cr o in o •r »n 0* in -t (M fM m m in f*> o 1 1 t fM CD o o ^ •* fM OO r- cr o cr cr cc cr <33 cr <0 O 't fSJ o in r^ fM i in fM 1 fM 1 ^ fn I 1 •O f^ - 0* in o fM in o o in cc (T O in cr O (X) ^0 rg 1 <0 in 00 (D in "* 1 fM CO •r r cr in in O CD o fM o 00 X o o cr (T" in cr in •O CD O rg CD 1 ^ 1 00 in C3 > Sh o O o E 3 C < 3. 3UniVd3dN31 V3S 3. 3UniVU3dtM31 V3S 206 SECKEL and YONG: HARMONIC FUNCTIONS o \ 1 CM O s S a 3 rt ;-i D IV X> a g £ a, P o o 0. 3UniVd3dW31 V3S 3. 3UniVd3dlN31 V3S 207 FISHERY BULLETIN: VOL. 69, NO. 1 P. Q 3. 3UniVtJ3dN3i V3S O. 3»niVd3dW31 V3S 208 SECKEL and YONG: HARMONIC FUNCTIONS 0, 3UniVd3dN3i V3S 0. 3aniva3dW3i V3S 209 FISHERY BULLETIN: VOL. 69, NO. I to B j3 2 5 o a z a. p- < Q ea hnrengns). J. Fish. Res. Bd. Can. 26(7) : 1919-1926. Dassow, John A., Max Patashnik, and Barbara J. KOURY. 1970. Characteristics of Pacific hake (Merluccius productuts) that effect its suitability for food. In Pacific hake, p. 127-136. U.S. Fish Wildl. Serv., Circ. 332. DuBRow, David L., Norman L. Brovi'N, E. R. Pariser, Harry Miller, Jr., V. D. Sidwell, and Mary E. Ambrose. 1971. Efl'ect of ice storage on the chemical and nutritive properties of solvent-extracted whole fish — red hake, Urophycis chuss. Fish. Bull. 69 (1) : 145-150. DuBROw, David, and Olivia Hammerle. 1969. Holding raw fish (red hake) in isopropyl alcohol for FPC production. Food Technol. 23 (2) : 254-256. Knobl, George M., Jr. 1967. The fish protein concentrate story. Food Technol. 21(8) : 1108-1111. LowRY, Oliver H., Nira J. Rosebrough, A. Lewis Fakr, AND Rose J. Randall. 1951. Protein measurement with the folin phenol reagent. J. Biol. Chem. 193(1): 265-275. Sen, D. p., T. S. Sayanarayana Rao, S. B. Kadkol, M. A. Krishnaswamy, S. Venkata Rao, an-d N. L. Lahiry. 1969. Fish protein concentrate from Bombay-duck (Harpoden nehereus) fish: Effect of processing variables on the nutritional and organoleptic qualities. Food Technol. 23(5): 683-688. SlEBERT, G. 1962. Enzymes of marine fish muscle and their role in fish spoilage. In Eirek Heen and Rudolf Kreuzer, Fish in nutrition, p. 80-82. Fishing News (Books) Ltd., London. Ting, Chao-titin, M. W. Montgomery, and A. F. Anglemeier. 1968. Partial purification of salmon muscle cath- epsins. J. Food Sci. 33(6): 617-620. Whaley, Wilson M. 1966. Production of fish proteins. U.S. Patent No. 3,252,962. 246 NOTES EQUIPMENT FOR HOLDING AND RELEASING PENAEID SHRIMP DURING MARKING EXPERIMENTS' Personnel of the National Marine Fisheries Service Biological Laboratory at Galveston, Texas, have conducted numerous mark-recap- ture ex]3eriments to obtain information on the movement, growth, and mortality of penaeid shrimp. These experiments were carried out under a variety of conditions at sea and in coastal bays. Several types of specialized equipment were developed to overcome problems of holding, handling, and releasing shrimp during the mark- ing phase of these experiments. Some of this equipment has been described previously by Costello (1964). Holding tanks, a cooling unit, and two devices used to transport shrimp to the sea floor are described here. Holding Facilities A number of factors were considered in the design of tanks for holding shrimp. Construc- tion materials had to be relatively light in weight, require little maintenance, and be nontoxic to shrimp. Provisions also were needed to permit lapid water exchange, minimize water turbu- lence within tanks, and control water temper- ature. The tank design in Figure 1 meets these needs and has proved successful for both sea- and land-based operations. It is constructed of light gray fiberglass with wood reinforcement and weighs about 114 kg (250 lb.) . Advantages of the light color are that it reflects heat and makes shrim]) easily visible in the tank. To permit rapid drainage or water exchange, a polyvinyl chloride (PVC) pipe, 7.6 cm (3 inch- es) in diameter, is molded into each end of the tank near the bottom. Filter .screens, used to ' Contribution No. 304, National Marine Fisheries Service Biological Laboratory, Galveston, Texas. prevent loss of shrimp in outflowing water, have a large surface area to minimize clogging. A one-quarter section of PVC pipe, 7.6 cm (3 inches) in diameter, is molded to the top of the tank at each end as a splash rail to reduce spillage. Five sets of guides in the tank support baffles that reduce water turbulence at sea and are used to separate groups of shrimp in a tank (Fig. 1). The baffles have a frame of aluminum flashing covered with sheets of patterned aluminum 0.063 cm (0.025 inch) thick. During field use, a series of two to four tanks are linked to provide either recirculating water or continually flowing new water. The pump used depends on the volume of water required. Normally, we use a cast-iron pump powered by a 0.5-hp electric motor (110-220 v) that dis- charged 114 to 132 liters (30 to 35 gal) per min. As the water is discharged into the tanks, it passes through siphon filler-drain nozzles (Cos- tello, 1964) which draw air into the circulation system and aerate the water. The aeration unit (Fig. 2), made of 1.9-cm (0.75-inch) pipe, may be attached temporarily at any convenient place on the tank. The amount of air that enters the water is regulated by valves in each air line. Because it is difficult to keep shrimp alive when water temperatures exceed about 27° C (80° F), cooling units are used to lower and maintain temperatures in holding tanks. A cooling unit of our own design is shown in Fig- ures 3 and 4. The casing consists of a PVC pipe, 25.4 cm (10 inches) inside diameter, 45.7 cm (18 inches) long, and 0.9 cm (0.37 inch), thick, and top and bottom pieces of PVC flat stock, 30.5 by 30.5 by 1.3 cm (12 by 12 inches by 0.5 inch) with circular grooves 0.6 cm (0.25 inch) deep. 0-ring gaskets that fit the grooves prevent leakage of water. The refrigerant coil is made from 0.9-cm (0.37-inch) diameter stain- less steel tubing, 9 m (30 ft) long. A thei-mo- stat sensor receptacle, inserted through the top 247 Figure 1. — The holding tank, baffles, and filters used in shrimp marking experiments. Figure 2. — Aeration unit and siphon filler-drain nozzle through which water enters tank. plate of the cooling tank, consists of a piece of 0.6-cm (0.3-inch) diameter stainless steel tubing, 20.3 cm (8 inches) long, and is sealed at one end. The top of the chilling tank is reinforced by a 30.5 by 30.5 cm (12 by 12 inches) frame made from angle aluminum stock 3.8 by 3.8 by 0.6 cm (1.5 by 1.5 inches by 0.25 inch). Two experiments were completed to deter- mine the cooling capability of the unit. Water was recirculated through the chilling unit at rates of 114 to 132 liters (30 to 35 gal) per min, and thermographs recorded water and air temperatures (Fig. 5). The temperature at- tained after 24 hr was about 15.6° C (60° F) and was lower than that required for shrimp- marking procedures. Field observations have indicated that water temperatures can be main- tained within 2° C (4° F) of desired levels, ir- respective of fluctuations in air temperatures. A table top with plastic pans 33.0 cm (13 inch- es) long, 38.1 cm (15 inches) wide, and 13.97 cm (5.5 inches) deep equipped for continuous water circulation (Fig. 6) slides over the lip of the holding tank and extends about 5 cm (2 inches) beyond the ends of the tank. When in 248 ^ «r 'L^. NUT ' WASHER - 5 Figure 3. — One-hp single-pliase cunipressor and con- densing unit (12,000 BTU factory rated) attached to a PVC chilling tank and mounted on angle iron stand. A. vibration joint; B. expansion valve; C. thermostat control; D. compressor; E. sight glass; F. dryer; H. condensing unit; J. chilling tank; K. angle iron stand. place, the table tojj serves as a work area for staining and tagging shrimp which are held in the pans. Equipment for Releasing Shrimp Three types of release devices have been de- veloped to protect shrimp from exposure to pre- dation during their return to the sea floor. Cos- tello (1964) described a release box that is low- ered to the bottom with a winch and opened by Figure 5. — Reduction of water temperatures in a 1,892- liter (500-gal) tank compared to surrounding air tem- peratures during trials with the chilling unit. £ ALUMINUM SUPPORT COIL INLET STAINLESS STEEL ROD- 2S4 CM PVC PIPE— - (10 IN I PVC TOP PLATE STAINLESS STEEL ROO PVC BOTTOM PLATE / FlGiniE 4. — Details of the chilling tank assembly. — 1 — I — I — r- 249 PLASTIC PAN 330>3e II 13 9 CM (13115x550 IN ) I 9 CM PLVWOOO (0 75 IN ) 3 CM PLASTIC CONNECTION (0 12 IN ) TUBING CLAMP PLASTIC "T" Figure 6. — Removable table top and holding pans equipped for continuous water circulation. a messenger dropped down the cable. Use of this device is restricted to large vessels equipped with a winch, and requires that the vessel be stopped when shrimp are released. To circum- vent these requirements, we designed an ex- pendable release canister that can be put over- board while a vessel is underway and a release tube for use in shallow water. The canister (Fig. 7) is constructed of high- impact styrene plastic formed into a hollow cyl- inder. Tabs on each of the styrene plastic end pieces have holes to accommodate retaining rods used in assembling the canister. Assembly and loading are accomplished in a cradle attached to the inner wall of a holding tank so that shrimp will remain submerged until the canister is ready to be put overboard. Slots in the canister allow it to fill with water. A salt block, a rubber band, and a paper clip constitute the release mechanism. This mech- anism is set by folding together the two ends of the styrene plastic sheet (thus forming a cyl- inder) and securing them with a rubber band. When ends A and B (see canister. Fig. 7) are folded, the paper clip is inside the canister with the attached rubber band inserted through a hole (end A) and a slot (end B). The salt block is then inserted in the loop formed by the rubber band, and the retaining rods are removed. A cement weight (1.1 kg or 2.5 lb.) is attached and the canister is lowered to the water surface and released. When the salt block dissolves, tension in the canister wall pulls the rubber band from the slot (end B) and the canister disas- sembles, releasing the shrimp. Although never obsen-ed during the actual release of shrimp in offshore waters, this release devise was tested in shallow estuarine waters and in the labora- tory. In all tests it performed as expected. The canister accommodates up to 100 shrimp that CANISTER ENDS r76CM (3IN ) r*4^ (0 03 IN) -008 CM Thick ASSEMBLY OF CANISTER B SALT BLOCK, 06 CM (0 25 IN) ^^ 11.4 CM (4 SO IN j 81.3 CM (32 IN) CANISTER SIDE ASSEMBLED CANISTER WEIGHT Figure 7. — Disposable release canister showing release mechanism, loading cradle, and method of assembly. 250 are released on the sea bottom within 5 to 15 min from the time the unit enters the water, depending on the size of the salt block. The release tube (Fig. 8), intended for use in shallow water, consists of two telescoping aluminum pipes, each about 3 m (10 ft) long. To release shrimp, the outer pipe is lowered to the bottom and shrimp are poured from a pail into the funnel. After each pail of shrimp is poured into the unit, the apparatus is flushed with several pails of water to insure that shrimp do not remain in the tube. The pouring and flushing of one pail of shrimp usually take about 1 min. The new equipment described herein and the improved techniques for staining and tagging described by Neal (1969) enabled us to hold. (lOIN ) FUNNEL TELESCOPING ALUMINUM PIPES LINE FOR LOWERING ' 12 7 CM 5 InT 3 1 M (10 FT) mark, and release large numbers of shrimp. We can now process between 1,500 and 3,000 shrimp per day, depending on the type of mark used. Literature Cited COSTELLO, T. J. 1964. Field techniques for staining-recapture ex- periments with commercial shrimp. U.S. Fish Wildl. Serv., Spec. Sci. Rep. Fish. 484, 13 p. Neal, Richard A. 1969. Methods of marking shrimp. FAO (Food Agr. Organ. U.N.) Fish. Rep. 57, 3: 1149-1165. Dennis A. Emiliani National Marine Fisheries Service Biological Laboratory Galveston, Texas 77550 AN ADULT BLUEFIN TUNA, Thunnus thynnus, FROM A FLORIDA WEST COAST URBAN WATERWAY' The bluefin tuna, Thunnus thymius (Linnaeus), is a wide-ranging pelagic species occurring in most tropical and temperate seas (Gibbs and Collette, 1966: 119). In the Gulf of Mexico exploratory and commercial catches have been limited to the northern, western, and central parts, from waters beyond the continental shelf. The collection of a large adult from the Florida west coast represents a new record for the Flor- ida shelf. The specimen, a female, was captured by local fishermen with harpoons in a waterway at Hudson, Fla., (lat 28°21'24" N, long 82°42'42" W) on 10 May 1970. It weighed 239 kg (525 lb.), was 244 cm (96 inches) in fork length and 168 cm (66 inches) in girth, and appeared to be in healthy but lean condition, characteristic of post-spawning fish in May on the Bahama Banks (Rivas, 1955: 139). Histological examination of gonadal tissue sectioned at 6 ,u, and stained with Harris hema- toxylin and Eosin Y showed early and late atretic Figure 8. — Release tube used to place marked shrimp on the bottom. ' Contribution No. 154. 251 body formations, indicating recent spawTiing. The stomach contained a mussel, Brachidontis recurv2is (Rafinesque), a piece of marl (no doubt ingested accidentally), and a digenetic trematode ; gills and other tissues were free of parasites. The fish had entei'ed the waterway through a shallow navigation channel from the adjacent grass flats. The waterway, a network of chan- nels cut through marl, consists of several 161 m by 15 m "fingers" branching from a 1370 m by 22 m central channel, with depths averaging less than 5 m at low tide. Surface salinity in the waterway, tested at a sub.^equent low tide, was 27.0 '/i,; surface temperature in the adjacent Gulf was 26° C. This occurrence, although admittedly irreg- ular, may help to fill a gap in our emerging picture of the origin and distribution of Gulf of Mexico bluefin tuna stocks. Bluefin tuna are taken from the Greater Antilles in spring, with substantial numbers of large adults being reported from the Windward Passage in April (Bullis, 195.5: 6) . During May they begin their dramatic migration through the Straits of Florida toward the summer feed- ing grounds (Rivas, 1954, 1955). The occurrence of large bluefins at Grand Cayman and east of Cozumel in April (Bullis and Mather, 1956: 9) suggests that at least a component of these Cariljbean stocks may undertake a similar northward movement through the Yucatan Straits and into the Gulf of Mexico. The occurrence of ripe or nearly ripe females in the Gulf in May and of small juveniles (less than 8 cm) in the northern Gulf in late May and early June (Mather, 1962: 5) implies that these stocks sjiawn in the Yucatan Straits or in the Gulf of Mexico proper. Our spent female on the Florida shelf could be from the Caribbean stock or from a stock wintering in the Gulf (Bullis, 1955: 13; Wathne, 1959: 16). We are indebted to Gordon D. Marston, St. Petefsburrj Times, for informing us of the in- cident, to Bud and Marvin Mattix and others for allowing us to examine the fish and viscera, to Richard B. Roe for providing information on specimens collected during exploratory fishing by U.S. Fish and Wildlife Service vessels in the Gulf of Mexico and Caribbean, to Alice Gennette for preparing histological sections, and to Frank J. Mather, III for critically reviewing the man- uscript. Literature Cited Bullis, Harvey R., Jr. 1955. Preliminary report on exploratory long-line fishing for tuna in tlie Gulf of Mexico and the Caribbean Sea. Part I - Exploratory fishing by the Oregon. Commer. Fish. Rev. 17(10) : 1-15. Bullis, Harvey R., Jr., and F. J. Mather, III. 1956. Tunas of the genus Thunnus of the northern Caribbean. Amer. Mus. Nov. 1765, 12 p. GiBBS, Robert H., Jr., and Bruce B. Collette. 1966. Comparative anatomy and systematics of the tunas, genus Thunnus. U.S. Fish Wihil. Serv., Fish. Bull. 66(1) : 65-130. Mather, Frank J., III. 1962. Distribution and migration of North Atlantic bluefin tuna. Proc. 7th Int. Game Fish Conf. Pap. 6, 7 p. Rivas, Luis Rene. 1954. A preliminary report on the spawning of the western north Atlantic bluefin tuna (Thunnus thynmis) in the Straits of Florida. Bull. Mar. Sci. Gulf Carib. 4(4): 302-322. 1955. .A comparison between giant bluefin tuna (Thunnus thynnus) from the Straits of Florida and the Gulf of Maine, with reference to migra- tion and population identity. Proc. Gulf Carib. Fish Inst., 7th Annu. Sess., p. 133-149. Wathne, Fredrick. 1959. Summary report of exploratory long-line fishing for tuna in the Gulf of Mexico and Car- ibbean Sea, 1954-1957. Commer. Fish. Rev. 21 (4) : 1-26. Robert W. Topp and Frank H. Hoff Florida Department of Natural Resources Marine Re.iearch Laboratory Sf. Petrrshurg, Fla. 337.31 252 SWIMMING SPEED, TAIL BEAT FREQUENCY, TAIL BEAT AMPLTrUDE, AND SIZE IN JACK MACKEREL, Trachurns symmetric^ AND OTHER FISHES John R. Hunter and James R. Zweifel' ABSTRACT The tail beat frequency and tail beat amplitude of jack mackerel, Trachnrus symmetricus, 4.5 to 27.7 cm were measured at speeds of 15 to 212 cm/sec. Tail beat amplitude was a constant proportion of length at all speeds but tail beat frequency changed with speed; thus speed depended only on fre- quency of the tail beat an° 5- 15 10 Leuciscus • 1 .-•O'- • 1- • .-. <■■ 1 1 15 10 Scomber FISHERY BULLETIN; VOL. 69, NO. 2 5 Corossius 10 . .■ •■*•■ 15 20 10 15 20 10 15 20 15 10 Solmo OL-^ 10 15 20 F-Fn Figure 5. — Relationship between swimming speed corrected for minimum s|)eed over lenprtli and tail beat fre- quency corrected for minimum frequency for Trdrhiiriis and Scomber from the present data, and for Leuciscus, Salmo, and Carassius from Bainbridge (1958). Graph at lower right shows individual regression lines for all above species, equations for linos are given in Table 6. 262 HUNTER and ZWEIFEL: SWIMMING SPEED AND TAIL BEAT FREQUENCY Table 5. — Slopes for the velocity-frequency relationship for individual Carassius studied by Bainbridge (1958) when the general relationship is slope ^= 0.68L; esti- mated minimum speed when Vq = O.SIL-/^; observed minimum swimming speed (I'obs) > the tail beat fre- quency Fq estimated by substitution of Vg into the cor- rected slope equation ; and the lowest observed tail beat frequency (F„f,J. Length (cm) * = 0.66L ^0 = = 0.81i2/3 *'obs ^0 = 2.22i" -1/3 '^obs 4.6 3.04 2.25 3.50 1.34 1.30 7.0 4.62 2.98 9.30 1.17 1.52 9.5 6.27 3.66 11.60 1.04 2.03 15.2 10.03 5.01 33.70 0.90 3.38 22.5 14.85 6.52 13.70 0.78 1.63 1 Data from Bainbridge (1958). Table 6. — Minimum speed (Vq), minimum tail beat fre- quency (Fq), the coefficient K in equation V — F„ = L(KF — Fq) arranged in order of K. Species N >'o "o K Triakis kenlei 6 0.15L^/^ 1.66L" -1/3 0.93 TrackuTus svmmetricus 176 ■o.aor''' 3.98i" -1/3 0.83 Scomber japonicus 261 1.3U^^^ 3.51i" 1/3 0.82 Leuciscui leuciscus^ 149 0.67i2/3 3.04/." -1/3 0.74 Carassius auratus^ 111 lo.eii"''^ 2.22i" -1/3 0.66 Salmo gairdneri^ 109 0.52Z,2/^ 2.81i" 1/3 0.64 Sardinops sagax 9 2.23i^^^ 3.48/." -1/3 0.50 1 y theoretical estimote based on equation of Magnuson (1970). 3 One deviant fisli omitted; if fish included. A' = 74, K = 0.66. » Original dota from Bainbridge (1958). 4 beats/sec on the abscissa were from this single deviant fish. If the deviant fisli is included K = 0.66, but if not, K = 0.82. We are inclined to use K = 0.82 because the values for the four fish were very similar and the protocol indicated that the deviant fish may have been overly fa- tigued when tested. Triakis appears to have a relatively high coefficient but not too much significance can be attached to the exact value for Triakis or for Sardi^iops because these were based on so few measurements. In sum, the speed-tail beat equation (Case II) — Table 6 — was biologically as well as statisti- cally relevant, was sensitive to specific differ- ences in swimming behavior, provided an un- biased correction for length, and made possible a more accurate estimation of swimming speed from tail beat frequency than heretofore has been possible. TAIL BEAT AMPLITUDE We pointed out previously that tail beat ampli- tude was a constant and was directly propoi-- tional to length and consequently the size coeffi- cients for amplitude are probably the same as those for length. Thus amplitude (A) in centi- meters can be substituted for length in the ori- ginal Case II equation V = a^vl^/s -j_ ^ L * F. When this was done for Trachurus using all individual amplitude values (iV = 176) , we ob- tained the equation; V = —6.5767/12/3 + 3.5637/1 * F. The amplitude coefficient may be also estimated by substitution of the ampli- tude-length relationship for Trachurus (A = 0.23177L) , into the Case II equation. The tail beat amplitude data collected by Bainbridge (1958) were insufficient for specific estimates of an amplitude coefficient. The mean amplitudes for each of the fish we studied and for each of those studied by Bainbridge were nearly the same, when adjusted for body length. Variation within a species was as great as that between species (Figure 6). The relationship UJ Q 3 Q. s < Z < UJ S 3- CorosBius (FROM BfllNeRlOGE, 19581 Enq roulis • Leuciscus (FROM BfilNBRIOGE, 19581 D Solmo (FROM BaiNBRlDGE, 1958) Sofdino ps * Scomber o Trachurus ^ Tnokis DO o i 10 15 20 25 TOTAL LENGTH (cm) 30 35 Figure 6. — Relationship between mean tail beat ampli- tude and length for every fish we studied and all those studied by Bainbridge (1958), A = 0.21L. between mean tail beat amplitude and length for all species combined was A = 0.21L. 263 FISHERY BULLETIN: VOL. 69. NO. DISCUSSION In all previous studies speed was divided by length then related to tail beat frequency. In our data when speed was converted to body lengths per second the relationship between speed and frequency was nearly identical to that given by Bainbridge (1958) for Carasslus, Leuciscus, and Salmo The confidence intervals for the slopes in the speed-frequency relation- ship in Eiifhynntis and Thmnivs (Yuen, 1966) and in Sarda (Magnuson and Prescott, 1966) overlap the slope in the Bainbridge equation and the ones for Trachurus and Scomber when the body length conversion is used. Thus, when speed is in body lengths per second, the relation- ship between it and frequency is about the same in all fish studied to date from goldfish to mack- erel and is adequately described by the Bain- bridge equation V/L = bf. Thus the Bainbridge equation provides a description of the average relationship for fish in general but little sig- nificance can be attached to specific differences in slope. If more than a rough estimate of speed is required or if specific difterences are important, or if estimates are needed near the minimum swimming speed it would be neces- sary to use the equation developed in this study. Bainbridge (1958) concluded from his data that the frequency-speed relationship was curvi- linear below a frequency of about 5 beats sec because fish modulated their tail beat amplitude. His evidence for this conclusion was that in some fish amplitude appeared to decrease at low- er frequencies, and that the distance per beat, calculated by dividing speed by frequency, de- clined at frequencies below 5 beats sec but was constant above that frequency. His evidence for amplitude modulation at low speeds was weak. In the three Salmo studied no trend ex- isted; in Cara,'isius he suggested there might be a decrease in amplitude in one of the two fish studied, and in one of the two Leuciscus studied a trend existed slightly stronger than the one in Camssius. In sum, the evidence for a de- cline in amplitude measurements was based on possible trends in two of the seven fish studied. Two fish could easily give a misleading picture of the general trend in the data, especially when the variability in ami)litude measurements are considered. In our .studies we measured the tail beat amplitude in every fish at all possible speed levels and no evidence existed for a consistent change in amj^litude with speed. In Bainbridge's data the departure of distance traveled per beat from a constant at low fre- quencies was caused by the division of speed liy frequency. Had the line relating frequency to speed passed through the origin, no bias would have existed but because the line inter- sected the abscissa at about 1 beat/sec division by frequency produced an artificial curvilinear trend at lower frequencies. We produced the same trend in distance per beat in our data by dividing speed by frequency but the trend was eliminated when a correction for the in- tercept was used. Thus the curvilinear trend in distance per beat in Bainbridge's data was an artifact caused by the method of calculation and consequently distance per beat was a con- stant at all frequencies. In addition, the appar- ent nonlinearity below 5 beats/sec in his graphs relating speed divided by length to frequency was also the result of the same intercept problem. Therefore, no evidence exists for consistent amplitude modulation at any speed range and speed appears to be related only/ to tail beat frequency and length in the species studied by Bainbridge (1958) as well as in the ones we studied. We concluded that during steady swim- ming at any speed, tail beat amplitude is a constant proportion of body length of the order of 0.21 L. That the mean amplitude during steady swim- ming was constant does not mean that amplitude is not modulated under certain conditions. It is widely known that fish modulate tail beat amplitude when they accelerate (Gray. 1968). Further, we had the impi'ession that some of the variability in the speed-frequency relation- shi]) was caused by diflJ'erences in amplitude. These diff'erences were infrequent and irregular in occurrence and consequently we were not able to evaluate them statistically. We are inclined to believe, however, that fish occasionally made minor adjustments in amplitude and frequency over the entire range of sjieeds, but these adjust- 264 HUNTER and ZWEIFEL: SWIMMING SPEED AND TAIL BEAT FREQUENCY ments were merely individual deviations from the general relationship we have described. We do not wish to detract from the original and important contribution of Bainbridge (1958), by emphasis on the differences between his and our conclusions. His basic conclusions and equations were not greatly different from our own. We were able to examine more closely the form of the relationships he described be- cause of a larger sample size made possible by the availability of automatic film analysis equip- ment and because of the existence of his data in the literature. The question of species-specific size effects re- mains unresolved. In our general model a good fit was obtained in seven species when the min- imum stalling speed was proportional to L~^^, the frequency at this minimum speed was propor- tional to L-'^^ and the slope coefficient was pro- portional to W. A com])arative study on speed- related size effects in fishes would certainly be of value. It also remains to be resolved whether or not it was appropriate to apply the minimum swim- ming speed equation developed by Magnuson (1970) for Euthijnnus affinis, a fish that lacks a swim bladder, to such a broad assortment of species. The equation implies a functional re- lationship between minimum speed and hydro- static equilibrium and implies existence of neg- ative buoyancy at minimum speeds. We do not know if these relationships exist in all species; nevertheless his equation did provide a reason- able estimate for minimum speed and it func- tioned well in our equation. The relationship between swimming speed and tail beat frequency we have described could be used in any application where it is necessary to measure swimming speeds of fish. For ex- ample, a sonic internal tag could be developed that telemetered tail beat frequency and thus the speed of free-swimming fish could be mon- itored continuously over extended periods. The tail beat frequency-speed relationship could be used for size or species identification using Continuous Transmission Frequency Mod- ulated sonar as suggested by Hester (1967). The increase of speed with frequency (our K value) varied from species to species and thus might be used for identification. If size were known, the minimum observed velocity would provide additional information for identification. Alternatively, if the species were known, min- imum (or maximum) speed would provide an indication of size. The equation could also be used to estimate size from tail beat amplitude, but caution should be exercised because in our study amplitude was not modulated and conse- quently, we do not know whether or not speed and tail beat amplitude are linearly related within an individual. ACKNOWLEDGMENTS We wish to thank David Holts and George W. Rommel of the NMFS Fishery-Oceanography Center, La Jolla, Calif., who assisted in con- ducting the experiments and in calibration of the apparatus. Albert Good wrote the program for digitizing the photographic data, and John J. Magnuson, Department of Zoology, University of Wisconsin, Madison, Wis., reviewed the man- uscript. LITERATURE CITED Alexander, R. M. 1959a. The densities of Cyprinidae. J. Exp. Biol. 36: 333-340. 1959b. The physical properties of the swimblad- ders of fish other than Cypriniformes. J. Exp. Biol. 36: 347-355. Bainbridge, R. 1958. The speed of swimming of fish as related to size and to the frequency and amplitude of the tail beat. J. Exp. Biol. 35: 109-133. 1960. Speed and stamina in three fish. J. Exp. Biol. 37: 129-153. Beamish, F. W. H. 1966. Swimming endurance of some northwest At- lantic fishes. J. Fish. Res. Bd. Can. 23: 341-347. Conway, G. R., N. R. Glass, and J C. Wilcox. 1970. Fitting nonlinear models to biological data by Marquardt's algorithm. Ecology 51 : 503-507. FlERSTINE, H. L., and V. WALTERS. 1968. studies in locomotion and anatomy of scom- broid fishes. Mem. S. Calif. Acad. Sci. 6: 1-29. Gray, J. 1968. Animal locomotion. Weidenfeld, London, 479 p. 265 FISHERY BULLETIN: VOL. 69. NO. 2 Hester, F. J. 1967. Identification of biological sonar targets from body-motion Doppler shifts. Mar. Bio-Acoustics 2: 59-74. Hunter, J. R. 1966. Procedure for analysis of schooling behavior. J. Fish. Res. Bd. Can. 23: 547-562. Magnuson, J. J. 1970. Hydrostatic equilibrium of Eutbynnus af- finis, a pelagic teleost without a gas bladder. Copeia 1970: 56-85. Magnuson, J. J., and J. H. Prescott. 1966. Courtship, locomotion, feeding, and miscel- laneous behaviour of Pacific bonito (Sarda chil- iensis). Anim. Behav. 14: 54-67. Olson, J. R. 1967. Flowmeters in shallow-water oceanography. Nav. Undersea Warf. Center, San Diego, Calif. TP 5, 44 p. Tranter, D. J., and P. E. Smith. 1968. Filtration performance. In D. J. Tranter (editor), Reviews on zooplankton sampling meth- ods, p. 27-56. UNESCO (U.N. Educ. Sci. Cult. Organ.) Monogr. Oceanogr. Method. 2, Part 1. Yuen, H. S. H. 1966. Swimming speeds of yellowfin and skipjack tuna. Trans. Amer. Fish. Soc. 95: 203-209. 266 SUSTAINED SPEED OF JACK MACKEREL, Trachurus symmetricus John R. Hunter' ABSTRACT Jack mackerel, Trachurus symmetricus, were forced to swim for up to 6 hr at various speeds in an activity chamber. The probit estimate for the swimming speed at which 50% of Trachurus would fa- tigue during 6 hr was 93.4 cm/sec (8.4 L/sec) for fish 10.0 to 11.9 cm and was 22.4 L^'^/sec for fish 9.0 to 17.6 cm where L is the total length of the fish in centimeters. At higher speeds, Trachurus, 15 cm, swam for 3 min at 160 cm/sec or 10 L/sec. The swimming speed at which 50% fatigued declined exponentially with time for about the first 22 min of swimming and thereafter declined linearly with time. The possible significance of the time-speed relationship for Trachurus is discussed. Although a substantial literature on the swim- ming speed of fishes exists (see Bainbridge, 1958; Gray, 1968), few reliable estimates of maximum sustained speed exist. Much of the literature on swimming speed of fishes is con- cerned with estimates of maximum speed or burst speed, that is, speeds that can be main- tained for only a few minutes or less. A sus- tained speed implies, on the other hand, that the animal is capable of swimming at that speed for hours. For example, Brett (1967) recom- mended a minimum of 200 min for a fixed sus- tained speed test. Fairly wide agreement exists that 2 to 3 L/sec can be maintained for an hour or more and salmonids and herring seem capa- ble of sustaining 3 to 4 L/sec for such periods (Blaxter, 1969) . These conclusions were drawn primarily from studies of freshwater fish and salmon; no estimates of maximum sustained speeds have been made for fast-swimming pe- lagic marine forms. The object of this study was to determine the sustained speed thresh- hold of jack mackerel, Trachurus symmetricus, a pelagic marine fish of commercial importance. The body form and musculature of Trachurus appear to be designed for greater hydrodynamic efficiency at high speeds than other species here- tofore studied. In Trachurus, lateral muscula- ture is concentrated in the anterior portion of the trunk, and inserts by tendons on a small deeply forked caudal fin. ' National Marine Fisheries Service Fishery-Ocean- ography Center, La JoUa, Calif. 92037. Manuscript received January 1971. FISHERY BULLETIN: VOL. 69, NO. 2, 1971. In addition to the interest in comparing the sustained speed capabilities of Trachurus with that of fish with other body forms, sustained speed data have significance in prediction of migratory capabilities and physiological limits. APPARATUS AND METHODS The apparatus used in the experiments was an activity chamber provided with a water cur- rent of various calibrated speeds. The appa- ratus was the same as the one described and figured by Hunter and Zweifel (1971) in this issue except that a port was provided in the transparent hatch of the swimming chamber so that fatigued fish could be removed by hand from the downstream screen without reducing the flow in the chamber. The error in estimating the water speed in the swimming chamber did not exceed 10% and it was assumed that the fish were swimming at the estimated speed. The experimental design was essentially the same as that used by Brett (1967) for deter- mining the sustained speed threshhold for sock- eye salmon, Oncorhynchus nerka. Fifty-five groups of five Trachurus (9.0 to 17.6 cm total length, mean = 12.43 ± 0.11 cm) were subjected to a fixed speed of 38 to 160 cm/sec for 360 min or longer after an introductory period of about 30 min at a low speed. A time-lapse camera photographed the fish at 1-min intervals and the time to fatigue for each fish was determined from the photographs. The temperature of the water in the activity chamber and in the holding 267 FISHERY BLXLETIN: VOL. 69, NO. 2 tank (a plastic swimming pool 15 ft in diameter) was maintained by a temperature regulation system at about 18.5° C. The mean test temper- ature was 18.48 ± 0.03° C. The fish were cap- tured near Santa Catalina Island, Calif., on 12 September 1969. Tests began 2 weeks later and ended on 21 November 1969. Fish were fed an abundant ration of chopped squid, anchovies, and frozen brine shrimp. Probit analysis, a sta- tistical technique first applied to sustained speed data by Brett (1967), was used to estimate sus- tained speed threshholds. Variability in length posed a problem in the analysis. Although all fish were from the same school, differences in length existed; also the fish grew in the course of the study. These dif- ferences were insufficient, however, to determine the form of the relationship between length and sustained speed. In general, the relationship between length and sustained speed for other species (Bainbridge, 1962; Brett, 1965), theo- retical considerations (Gray, 1968; Fry and Cox, 1970), and relationships between length and other swimming capabilities (Magnuson, 1970), indicate that speed is proportional to a fractional power of length equal to about U"^ - "'. In addition, the minimum swimming speed of Trachurus was proportional to L"^ when esti- mated from Magnuson's equation (Hunter and Zweifel, 1971). In light of the above evidence it seemed preferable to use 0.6 as the coefficient of length, although unity has been commonly employed in cases where length coeflficients were unknown. As an alternative to this procedure I also estimated the percent fatigued at different speeds in centimeters per second and in body lengths per second for a narrow length range (10.0 to 11.9 cm total length) where the effect of diflFerences in length would be negligible. RESULTS Within a few minutes after Trachurus were placed in the swimming compartment they be- came quiescent, swam steadily, and remained in about the same position in the compartment throughout the test or until they became fa- tigued and fell against the rear screen. This was in contrast to some other species which did not swim steadily, but swerved and oscillated from side to side. The relationship between water speed and percent fatigue had the normal sigmoid form of a dosage response curve (Finney, 1952). Probit estimate of the applied water speed at which 50^; fatigue occurred in 360 min of swim- ming and the 95 5r confidence limits were 94.40 ± 5.15 cm/sec for Trachurus 10.0 to 11.9 cm total length, N = 127 (Figure 1, Table 1). Thus, Table 1. — Swimming endurance of Trachurus sym- metricus in cm/sec and in L^Vsec. Length 10.0-11.9 cm Length 9.0-17.6 cm Speed S Percent fatigued ler gth Speed' i0.6/sec N Percent cm/sec Mean SO fatigued 71 8 10.69 0.61 15.9 3 78 16 13 10.71 0.47 16.9 10 85 16 31 10.84 067 17.8 14 92 14 50 10 97 0.54 18.7 17 6 99 21 62 11.23 0.43 19.7 18 17 106 21 76 10.85 0.56 20.6 23 39 113 15 100 11.41 0.42 21.5 39 31 120 8 100 11.27 0.45 22.5 42 67 138 8 100 11.20 0.51 23.4 19 58 Tolol 127 24.3 25.3 26.2 27.2 28.1 29.0 30.0 30.9 31.8 32.8 33.7 Total 20 29 19 9 4 3 1 3 7 10 4 294 60 72 100 100 100 100 100 100 100 100 100 ' Totol speed range divided into 20 equal intervals; speeds listed are midpoints of those intervals. 50 ''r of Trachurus in this length range could be expected to sustain a speed of about 8.4 L/sec or 22.1 L" Vsec for 360 min. For all Trachurus (N = 294) the water speed at which 50 ':r fa- tigue occurred after 360 min of continuous swimming and the 95 ''r confidence intervals were 22.4 ± 1.2 L" Vsec. The first estimate, based on a narrow length range, and the second one, based on all data, were reasonably close. On the other hand, when all data were in the form V L'" the 50^r threshold was 9.34 L sec which is higher than the preceding estimates. Inspection of these data, however, showed that the coefficient for length clearly was less than one and that use of unity biased the estimate. 268 HUNTER: SPEED OF JACK MACKEREL 98 r Figure 1. — Probit lines for sustained speed threshold for 6 hr of forced swimming at 18.5° C in juvenile Trachurus symmetrieus. Upper panel, range fish length 10.0 to 11.9 cm iV = 127, probit = 0.077A' - 2.238; lower panel, range fish length 9.0 to 17.6 cm, N =; 294, speed expressed in L" ^/sec where L is the total length of the fish, probit = 0.355:? — 2.958. To determine the form of the relationship between the duration of the swimming period and the ability to maintain a certain speed, probit estimates of speed for five levels of fa- tigue were made for swimming periods varying from 10 to 360 min. The form of the relation- ship was about the same for all fatigue levels; speed estimates declined exponentially with time for short swimming periods and linearly with time for longer ones (Figure 2). The point 40r u 30 UJ 99% 100 200 300 MINUTES Figure 2. — Relation between speed in L'^'^/sec and the time it can be sustained for 1, 25, 50, 75, and 99 per- cent fatigue levels in Trachelitis symmetrieus. Estimates of speed for each fatigue level made at 10-min intervals of cumulated time. of inflection from the exponential to the linear relationship was examined in detail for the 50% fatigue level. Probit estimates of the speed at which 50% of the fish fatigued were made for 2-min intervals of swimming cumulated over the first 100 min of observation. The data were plotted on semilog paper and a line fit by eye to the exponential function. The point of in- flection appears to occur at about 22 min (Fig- ure 3) . Thus, speed at which 50% fatigued and the duration of the swimming period were ex- ponentially related for durations up to about 22 min and were linearly related for longer pe- riods of swimming. The performance of juvenile Tmchimis at high speed was of interest. Fifteen fish 14.6 cm mean total length (range = 13.4 to 16.6 cm) swam at the highest speed used in the study (160 cm/sec) for 2 to 6 min, mean time 3.4 min. Thus, Trachurus 15 cm total length were able to swim for about 3 min at about 10 L/sec or about 32 L" Vsec. A slightly higher level of 269 FISHERY BULLETIN: VOL. 69, NO. 2 \ 1 1 1 1 1 1 1 1 T- - \ - \ - •n. - 1. i-jt-lA 1 1 I 1 lihl 20 30 MINUTES 100 Figure 3. — Relation between speed at which 50% Trach- unis fatigued and the duration of the swimming period. Duration of swimming period in minutes plotted on log scale to show exponential trend ; line fit by eye. Speed estimates made at 2-min intervals of cumulated time over the first 100 min of swimming. performance in length per second is obtained if we consider smaller fish. For example, fish of mean length 11.2 cm (length range 10.4 to 11.9 cm, A'^ = 8) swam for 3 to 5 min (mean = 4.5 min) at 139 cm/sec or about 12 L/sec. This difference between large and small fish becomes negligible if 0.6 is used as a coefl^cient of length instead of unity because, as was pointed out previously, the length coeflicient for Trachurus appears to be less than 1. DISCUSSION The exponential decline in swimming speed with time in fish is well documented; see for example Bainbridge (1960), Brett (1967), and Blaxter (1969). The general form of the re- lation between time and swimming speed in other fish resembles that for Trachurus al- though the speeds and endurance times are dif- ferent in Trachurus. The physiological mech- anisms responsible for the exponential relation- ship between swimming speed and endurance are generally believed to be the limited enery stores in the muscle, the rate these stores can be replaced and the rate catabolites are removed from the muscle (Bainbridge, 1960). A study by Pritchard, Hunter, and Lasker (1971) in this issue has provided an explanation for the form of the speed-time relationship in Trach- urus. Pritchard et al. found that at speeds where an exponential relationship exists be- tween time and speed the principal cause of failure of Trachurus was most likely the de- pletion of glycogen in the white muscle. On the othei- hand, fish that failed at speeds near the 6-hr 50% threshold, where a linear relation- ship exists between speed and time, had de- pleted not only the glycogen in the white muscle but that in the red muscle and liver as well. Thus, in Trachurus the form of the time-speed relationship could be explained on the basis of the extent of glycogen reserves available for locomotion and the time required to mobilize them from sites other than the white muscle. An exponential relationship between speed and time could be produced when the speeds are so high that the glycogen supply would be limited almost entirely to the white muscle because the supply in the white muscle would be used up and the fish would fail before significant amounts of glycogen could be mobilized from other sources. A linear relationship could exist where swim- ming speeds are sufficiently low that reserves in the white muscle could not be depleted before other sources in the red muscle and the liver are mobilized. We have, on one hand, a high rate of consumption using a more limited supply of fuel which could lead to an exponential relation- ship between speed and time and, on the other hand, a much lower rate of consumption using a relatively much larger fuel supply which could produce a linear relationship with time. An exponential relationship between energy con- sumption and swimming speed would enhance these effects. Let us now consider the significance of the 6-hr sustained speed threshold determined for Trachurus. When compared with other deter- minations, this threshold appears to be unique because of different physiological mechanisms and because it is higher than those estimated for other fish. Trachurus at threshold speed appeared to use glycogen as fuel, white muscle for locomotion and maintained a high lactic acid 270 HUNTER: SPEED OF JACK MACKEREL level in the muscle (Pritchard et al., 1971). These results are inconsistent with the conclu- sion that at sustained cruising speeds, fish use lipid metabolism to drive red muscle (Bone, 1966; Gordon, 1968; Blaxter, 1969) and that no oxygen debt is incurred (Brett, 1963). Re- liance on glycogen as the principal fuel pro- bably severely limits the time a speed can be maintained as compared with one where lipid metabolism is used exclusively. Thus the bio- chemical evidence indicates that the 6-hr speed threshold for Trachurus probably could be main- tained only for a period of hours or perhaps days but certainly not weeks as one would ex- pect if fat were used as fuel. The 6-hr thresh- old was also considerably above sustained speed thresholds for other fish where presumably fat may be employed as fuel. Brett (1967), in a study directly comparable with the current one, found the 50% fatigue time for sockeye salmon was 4 L/sec (about 11.3 U"^) whereas for comp- arable size jack mackerel it would be about 7.6 L/sec or 22.0 L"^. Other less comparable data give sustained or cruising speeds in the range of 3 to 4 L/sec (Blaxter, 1969). Thus, Trach- urus has special physiological and structural adaptations that permit swimming for periods of hours at elevated speeds and it was the thresh- old for this swimming behavior that was meas- ured. Other fishes, especially the scombroid fishes, may have similar abilities. For example, skipjack tuna can swim at 8 knots, or about 43 L"^, for over an hour (Commercial Fisheries Review, 1969) and yellowfin tuna and skipjack tuna have higher levels of white muscle gly- cogen than many other species of fish (Barrett and Connor, 1964). It seems possible another speed threshold may exist for Trachurus below the present one where fat is the principal fuel, only red muscle is used for locomotion, and swimming can be main- tained almost indefinitely. It would not be sur- prising if this lower threshold were closer to those determined for other fishes. LITERATURE CITED Bainbridge, R. 1958. The speed of swimming of fish as related to size and to the frequency and amplitude of the tail beat. J. E.xp. Biol. 35: 109-133. 1960. Speed and stamina in three fish. J. Exp. Biol. 37: 129-153. 1962. Training, speed and stamina in trout. J. Exp. Biol. 39: 537-555. Barrett, I., and A. R. Connor. 1964. Muscle glycogen and blood lactate in yellow- fin tuna, Thunnus albacares, and skipjack, Katsu- wonus pelamis, following capture and tagging. Inter-Amer. Trop. Tuna Comm., Bull. 9: 219-268. Blaxter, J. H. S. 1969. Swimming speeds of fish. FAO (Food Agr. Organ. U.N.) Fish. Rep. 62: 69-100. Bone, Q. 1966. On the function of the two types of myotomal muscle fibre in elasmobranch fish. J. Mar. Biol. Ass. U.K. 46: 321-349. Brett, J. R. 1963. The energy required for swimming by young sockeye salmon with a comparison of the drag force on a dead fish. Trans. Roy. Soc. Can., Ser. 4, Vol. 1, Sect. 3: 441-457. 1965. The relation of size to rate of oxygen con- sumption and sustained swimming speed of sock- eye salmon {Oncorhynchus nerka) . J. Fish. Res. Bd. Can. 22: 1491-1501. 1967. Swimming performance of sockeye salmon (Oncorhynchus nerka) in relation to fatigue time and temperature. J. Fish. Res. Bd. Can. 24: 1731-1741. Commercial Fisheries Review. 1969. Underwater tuna school tracked by sonar. Commer. Fish. Rev. 31(11) : 9-10. Finney, D. J. 1952. Probit analysis. 2d ed. Cambridge Univ. Press, Cambridge, Engl., 318 p. Fry, F. E. J., and E. T. Cox. 1970. A relation of size to swimming speed in rainbow trout. J. Fish. Res. Bd. Can. 27: 976-978. Gordon, M. S. 1968. Oxygen consumption of red and white mus- cles from tuna fishes. Science 159: 87-90. Gray, J. 1968. Animal locomotion. Weidenfeld, London, 479 p. Hunter, J. R., and J. R. Zweifel. 1971. Swimming speed, tail beat frequency, tail beat amplitude, and size in jack mackerel, Trach- urus symmetriciis, and other fishes. Fish. Bull. 69: 253-266. Magnuson, J. J. 1970. Hydrostatic equilibrium of Euthynnus af fin- is, a pelagic teleost without a gas bladder. Copeia 1970: 56-85. Pritchard, A. W., J. R. Hunter, and R. Lasker. 1971. The relation between exercise and biochem- ical changes in red and white muscle and liver in the jack mackerel, Trachurus symmetrictts. Fish. Bull. 69: 379-386. 271 THE TRANSPLANTING AND SURVIVAL OF TURTLE GRASS, Thalassia testudinum, IN BOCA CIEGA BAY, FLORIDA' John A. Kelly, Jr., Charles M. Fuss, Jr., and John R. Hall= ABSTRACT Turtle grass was transplanted to an unvegetated, dredged canal and a hand-cleared portion of a flour- ishing grass bed. Complete or partial success was attained in 7 of 14 methods used. The best method, in which short-shoots (rhizomes removed) were dipped in a solution of plant hormone (Naphthalene Acetic Acid) and attached to construction rods for transplanting, was 100% successful and may be suitable for general application. Turtle grass, Thalassia testudinum, and other marine grasses are an invaluable asset to the marine ecosystem. They are primary producers and form an essential ecological niche in which a great number and variety of species find food and shelter. They are also important agents in the control of substrate erosion and the de- positions of sediments (Stephens, 1966). Uncontrolled dredging and filling of sub- merged lands have destroyed many turtle grass beds and their dependent fauna, some of which are economically important. An immediate need exists not only for sharply restricting further destruction of sea grass beds but also for re- placing lost beds. One method of replacing them may be by transplanting sea grasses to areas that are suitable for their growth or to areas that are made favorable by soundly planned en- gineering (Phillips, 1960; Strawn, 1961). Areas surrounding spoil banks and finger-fill canals (dredged canals between filled land mass- es) would be suitable if they were constructed to supply zones of optimum depth for growth of marine grasses. Unsuccessful earlier attempts to transplant turtle grass in Tampa Bay showed that the main problem was erosion by tidal currents. Turtle grass is buoyant, and new transplants tend to ' Contribution No. 64 from the National Marine Fish- eries Service Biological Laboratory, St. Petersburg Beach, Fla. 3.3706. ' National Marine Fisheries Service Biological Lab- oratory, St. Petersburg Beach, Fla. 33706. work free of the sediments and float to the sur- face when disturbed by water movement (Phil- lips, personal communication)." Another ma- rine plant, eelgrass (Zostera marina), was transplanted successfully on the coast of Wash- ington by Phillips (1967) and in the Aleutian Islands by Jones* and McRoy' (personal com- munication), but details on methods are not yet published. Successful growth of turtle grass under artificial conditions (Fuss and Kelly, 1969) led us to attempt transplanting it from one field location to another as described in the present paper. Turtle grass spreads vegetatively by creeping rhizomes (long-shoots) buried in the substrate (Figure 1). Work by Tomlinson and Vargo (1966) showed that this growth is dependent entirely upon the vigorous activity of meriste- matic tissue in the apexes of rhizomes. The apex is also the only source of short-shoots (erect lateral branches) that develop from buds at this site. In the Miami area (Phillips, 1960) and tropical parts of its range, the plants also re- produce by flowering. Tampa Bay, however, is near the northern limit of the flowering capa- bihty of Thalassia (Phillips, 1960); thus, we Manuscript received January 1971. FISHERY BULLETIN: VOL. 69. NO. 2, 1971. ' Phillips, Ronald C, Department of Botany, Seattle Pacific College, Seattle, Wash. 98119. ' Jones, R. D., Jr., Range Manager, Bureau of Sport Fisheries and Wildlife, Aleutian Islands National Wild- life Refuge, Cold Bay, Alaska 99571. ° McRoy, C. P., Institute of Marine Science Univer- sity of Alaska, College, Alaska 99701. 273 FISHERY BULLETIN: VOL. 69, NO. 2 m ^"O"'- M RHIZOME , iW-SHOOlJf APEX (lONG-SHOOl) i IT // ^^ Figure 1. — External features of Thalassia testudinum. confined our restoration studies to the trans- plantation of adult plants. This paper describes the procedures for and results of transplantation of turtle grass into modified environments. trol site was 95.5% sand (>62.5/x) and 4.5% silt and clay (<62.5/Lt) on a dry weight basis. At the planted areas of the finger-fill canals, sediments averaged 98.6% sand and 1.4% silt and clay. No analysis of the carbonate fraction was made for these samples; however, all sites had shell fragments, which appeared to be more abundant in the canals than at the control site. MATERIAL AND METHODS The work was divided into two phases: Phase I extended from July 1966 through August 1967 and phase II from April through October 1967. In phase I, methods of deflecting and reducing the force of tidal currents and waves in the vi- cinity of transplants and of anchoring new trans- plants in the substrate were tested. Concrete building blocks were laid in parallel rows at both transplant sites to form enclosed areas for sheltering new transplants against the forces of moving water (Figure 3). Plugs of grass approximately 8 inches square (20 X 20 cm) DESCRIPTION OF TRANSPLANT SITES All experiments took place in the southern end of Boca Ciega Bay, Fla., an elongate coastal lagoon joined to Tampa Bay and separated from the Gulf of Mexico by a line of barrier islands (Figure 2). The area it encompasses is a par- amount example of grass bed destruction by hydraulic engineering (Hutton et al., 1956; Phillips, 1960; Taylor and Saloman, 1968). A rectangular area 8.2 by 21 m (27 by 7 ft) in a large turtle grass bed was cleared by hand to serve as the control site. Two other trans- plant areas of the same size were in two adjacent finger-fill canals in a large land-fill development. Construction of houses had not begun along the canals selected, and none was built during the experiments. Boating in the canals was light, and during periodic inspections we saw no disturbance of the plants directly attributable to man. Sediments from transplant sites were an- alyzed by particle size. A sample from the con- FlGURE 2. — Locations of experiments (phases I and II, and control area). 274 KELLY, FUSS, md HALL: TRANSPLANTING TURTLE GRASS and containing four to five short-shoots were dug from natural beds adjacent to the control site. Three methods of transporting and anch- oring the plugs were tried: (1) placing them in tin cans, (2) balling them in burlap, and (3) temporarily bagging the roots and rhizomes in polyethylene, which was removed just before planting. A total of 120 plugs was transplanted — 60 at the control site and 60 at the finger-fill canal. At both locations, 30 were placed inside and 30 outside of enclosures. Each group of 30 plugs was planted three ways: 10 in cans, 10 in bur- lap, and 10 unanchored (Figure 3). Phase II consisted of testing additional an- choring devices to hold individual sprigs of turtle grass in the substrate of the finger-fill canal. The devices were cast iron, 2-inch (5.1-cm) pipe, brick, and construction rod. Sprigs used in this study were single short-shoots with leaves, many roots, and with or without a por- tion of the parent rhizome. Also tested in phase II was the plant hormone, NAPH (Naphtha- lene Acetic Acid),° which is used for rooting grass stolons and plant cuttings. Sixty sprigs, obtained from the same natural bed as the plugs in phase I, were washed and prepared for the experiment by breaking entire rhizomes from some, breaking only the apexes of rhizomes from others, and leaving the rhi- zomes attached and entire on others. Half of the sprigs were placed in a 10% solution of NAPH in seawater for 1 hr. The other half were left untreated. The sprigs were planted in groups to test various combinations of treatment and non treatment with NAPH, presence and ab- sence of apexes of rhizomes, presence and ab- sence of rhizomes, and types of anchors ( Figure 4). Sprigs anchored with construction rod had no rhizomes. Sprigs anchored with pipe had rhizomes that were buried in hand-dug holes; whereas, sprigs anchored with brick were simply placed on the surface of the substrate and their POLYETHYLENE ' Manufactured by Nutri-Sol Chemical Company, Tampa, Fla. 33609. References to trade names in this publication do not imply endorsement of commercial products. Figure 3. — Details of concrete-block wave and current barriers, tin-can anchors for plugs, and placement of transplants at planting sites. rhizomes held in contact with the sediment by the weight of the brick. RESULTS Transplants were considered successful if they established themselves in the new envi- ronment and exhibited new rhizome growth (Figure 5) . Individual sprigs met these criteria if short-shoots appeared healthy, had new roots, and had either given rise to a new rhizome or were still part of an old long-shoot with an active apex. Plug transplants (phase I) were con- sidered successful if only one of the short-shoots met the above criteria. 275 FISHERY BULLETIN: VOL. 69, NO. 2 CONSmUCTJON ROD APEXES o oo w o oo w o oo w o o oo w o o o NR o o NR o (TTT] W Em Em W - WITH [ O - WITMOUf NR NO RHIZOME OlD SHORI-SHOOT NEW SHORI-SHOOI APEX NEW RHIZOME (LONG-SHOOI) NEW ROOTS Figure 5. — New rhizome, root, and short-root growth on a sprig of turtle grass transplanted without an intact rhizome apex. made in the finger-fill canal in phases I and II, respectively, and 40 Sr of the transplants made in the control bed (Tables 1 and 2). Transplant attempts made with burlap in phase I were not included in the above percentages because all failed within 1 month. FiGUKE 4. — Details and treatment of sprigs anchored by pipe, brick, and construction rod; and placement of transplants at planting sites. Plugs in phase I planted July 1966 were re- moved from the sites late in August 1967, ap- proximately 13 months after planting. Six of the 40 transplants at the canal area and 16 of the 40 at the control area were successful (Table 1). Planting individual sprigs of turtle grass in phase II yielded similar results. Of the 60 sprigs planted in the second canal in April 1967 and removed in mid-October 1967 (about 6 months after they were transplanted), 11 were successful (Table 2). Successful new growth of rhizomes repre- sented 15 and 18'; of the number of transplants EROSION CONTROL Results of planting plugs within concrete- block enclosures were completely different in the finger-fill canal and grass bed locations (Table 1). In the canal the only successful transplants grew within the protection of the block enclosures; none planted without this pro- tection survived, and most of the latter failed within the first 6 months of the experiment. Most of the successful plugs placed in the con- trol area were planted outside the concrete blocks and, throughout the study, appeared to be in better condition than those inside the en- closures. The enclosures fulfilled their purpose in the canal but ai)i)eared detrimental in the control area. In the latter region, surrounding grass beds apparently provided suflicient pro- tection from water movement. Enclosures in 276 KEXLY. FUSS, and HALL: TRANSPLANTING TURTLE GRASS Table 1. — Surviving transplants, successful transplants, and seasonal mortality of transplant- ed plugs of Thalassia in the finger-fill canal and the control site, phase I, July 1966 through August 1967. Method of protecting and anchoring plants Surviving plants Mortality Winter Spring Summer Inside concrete block enclosures; Anchored in cons Unanchored Totol Outside concrete block enclosures: Anchored in cons Unanchored Total Grand total No. % No. Finger-fill canal site No. No. No. 10 10 3 3 30 30 3 2 4 4 I 20 6 30 5 8 1 10 10 9 10 1 20 19 I Inside concrete block enclosures: Anchored in cans Unanchored Total Outside concrete block enclosures: Anchored in cans Unanchored Total Grand total 10 10 1 2 3 30 5 4 1 1 2 1 20 3 3 15 9 2 2 I 10 10 1 2 7 6 70 60 1 1 1 1 20 3 13 65 1 2 1 40 10 1 Transplants survived but did not exhibit new rhizome growth. 3 Transplants exhibited new rhizome growth. the control area often filled with a heavy accumu- lation of light-robbing algae, dead grass, and other detritus, which quickly resulted in burial and death of the entire transplant. ANCHORING METHODS FOR PLUGS Of the plugs anchored with tin cans, 50% were successful at the control site, but only 15% in the canal (Table 1) . None of the plugs plant- ed in cans outside of the concrete-block enclo- sures survived. Cans were thus ineffective against currents unless used in conjunction with the concrete-block current barrier. Plugs transported in polyethylene bags and then directly transplanted served to evaluate the effect of tin cans. We noted no adverse effects from the metal in the cans. The ratio of suc- cesses of unanchored to anchored transplants was 3:5. The reasons for the rapid failure of plugs with roots and rhizomes wrapped in burlap is unknown. Possibly decomposition products of the decaying burlap, such as H2S, or toxic chem- icals in the material caused the plants to die. ANCHORING METHODS FOR SPRIGS In phase II sprigs were planted with added anchoring devices but without the aid of the wave and current barriers. Construction rod was the most effective device used to anchor sprigs. It was the easiest to handle because all sprigs fixed to it were trans- planted without rhizomes and were simply fas- tened to the rod with plastic-coated wire and inserted into hand-dug holes in the substrate. Of the 12 sprigs anchored with rod, only the 6 that had been treated with the hormone NAPH became established (Table 2). Sprigs that did not survive failed early in the experiment and simply disappeared, probably because they were dislodged by water movement in the canal be- fore roots were developed. 277 FISHERY BULLETIN: VOL. 69, NO. 2 Table 2. — Surviving transplants, successful transplants, and monthly mortality of transplanted sprigs of Thalassia in a finger-fill canal, phase II, April through October 1967. Method of anchoring and treoting Trons- plants Surviving plants Mortality! Unsuccessful* Successful^ May July Aug. Sept. Pipe* With apexes, with NAPH With opexes, without NAPH Without opexes, with NAPH Without apexes, without NAPH Total Bricks With opexes, with NAPH With apexes, without NAPH Without opexes, with NAPH Without opexes, without NAPH Total Construction rod^ With NAPH Without NAPH Total Grand total 60 n 33.3 16.7 25 24 7 3 12.5 6 6 2 6 4 2 33 3 6 6 6 6 6 3 3 24 4 2 8.3 3 15 6 6 100.0 6 4 2 12 6 50.0 4 2 2 * Mortality not observed in June. ' Transplants survived but did not exhibit new rhizome growth. * Tronsplants exhibited new rhizome growth. * Rhizomes were buried; two sprigs per anchor. ^ Rhizomes were not buried; three sprigs per anchor. « Rhizomes were removed before planting; tv/o sprigs per anchor. Pipe and brick were poor anchors. The sprigs anchored with pipe were transplanted with their rhizomes and special care was required in bury- ing them to avoid breakage. Almost half of the 24 sprigs held with pipe lived to the end of the experiment, but only 3 exhibited new rhizome growth (Table 2) . Sprigs secured to the bottom with brick were not buried but were simply laid on the bottom and the substrate was scooped over them by hand. They were also transplanted with their rhizomes but were difficult to handle because of their tendency to slip out from under- neath the brick before they were finally set in place. Six of the 24 sprigs lived for awhile, but only 2 were successful. Sprigs that were anchored with brick and failed did so shortly after they were planted. Water movement prob- ably eroded away enough sediment to allow the buoyant sprigs to float from under the brick. TREATMENT OF TRANSPLANTS IN PHASE II The effect of NAPH on marine grasses is ap- parently similar to its effect on terrestrial plants, primarily inducing rapid and heavy rooting. Ten of the 11 sprigs producing new rhizomic growth were treated with it (Table 2) . Because of the small number of transplants attempted and successes achieved, we cannot definitely establish the significance of NAPH in such ex- periments. Our results indicate to us, however, that NAPH was one of the main factors con- tributing to transplant success. Particular care was taken to avoid damaging rhizomes and rhizome apexes of sprigs before and during transplanting. No apparent advant- age was gained from this care ; invariably old rhizomes withered away and were replaced by new ones developing from the bases of the short- shoots. MORTALITY OF TRANSPLANTS Visual checks made throughout the year showed that the most critical period for the survival of turtle grass was during the first 3 months after transplanting. In phase I, mor- tality of plugs planted in the canal was 60% through the third month (October), 22.5% through the sixth month (January), zero 278 KELLY. FUSS, and HALL; TRANSPLANTING TURTLE GRASS through the 3-month period February-April, and 2.5Cf during the remainder of the study. Losses in the control area for the same time intervals were 25, 10, 5, and 5%, respectively. Mortality experienced during phase II was also high. Over half (BTSr ) of the sprigs trans- planted in April failed before the end of the third month (July) and T;} from August to Oc- tober. Additional failures within this phase might have occurred had the experiment con- tinued through the winter. CONCLUSIONS AND RECOMMENDATIONS Our experiments resulted in the first success- ful field transplantation of turtle grass. All new short-shoots produced by transplants were from the new rhizome apexes (Figure 5). This finding supports the observations of Phillips (1960) and Tomlinson and Vargo (1966) that buds on the rhizome apex are the only source of short-shoots. It is also in agreement with findings in the tank culture of Thalassia (Fuss and Kelly, 1969). Continuous growth of turtle grass depends on the activity of vigorous rhi- zome apexes, but the apexes do not contain the only meristematic tissue in the plant. New rhizomes can be produced from residual meris- tematic tissue present in the old short-shoot. Phillips (1960) observed such branching in the field and stated that it could account for the continued growth of turtle grass if the apex of the rhizome were damaged or lost, but be- lieved that the frequency of this branching was small. Tomlinson and Vargo (1966) also re- ported that vegetative branching in short-shoots occurs and indicated that it is rare. Undamaged leaves may not be required for sprig transplanting. Further studies are needed to determine, for example, if the leaves could be cut back to reduce the surface area and buoy- ancy of the sprig. Results of investigations in Boca Ciega Bay by Prest, Saloman, and Taylor' show that turtle grass leaves clipped as much as 50% of their original height (about 26 cm) would regrow as much as 3 to 4 cm (1.2 to 1.6 inches) per week. It would thus appear that physical damage to leaves is quickly overcome by regrowth of the plant. We have shown that turtle grass can be trans- planted in the field and that it will grow in an area denuded by coastal dredging. A simple transplant method using only the short-shoots of this grass, the hormone NAPH, and con- struction rod was 100% successful (six trans- plants) in a land-fill finger canal (Table 2). This method has value for use in restoring Thalassia to estuarine environments when conditions fa- vorable for plant growth exist or can be arti- ficially created. We must emphasize however, that no large-scale transplant program has been attempted. Moreover, recent observations (No- vember 1970)' of vegetative growth into our original control site indicate that turtle grass spreads at an annual rate of only 20 cm (8 inches) or less. LITERATURE CITED Fuss, C. M., Jr., and J. A. Kelly, Jr. 1969. Survival and growth of sea grasses trans- planted under artificial conditions. Bull. Mar. Sci. 19: 351-365. HuTTON, R. F., B. Eldred, K. D. Woodburn, and R. M. Ingle. 1956. The ecology of Boca Ciega Bay with special reference to dredging and filling operations. Part I. Fla. State Bd. Conserv., Tech. Ser. 17, 87 p. Phillips, R. C. 1960. Observations on the ecology and distribution of the Florida seagrasses. Fla. State Bd. Con- serv., Mar. Lab. Prof. Pap. Ser. 2, 72 p. 1967. On species of the seagrass, Halodule, in Florida. Bull. Mar. Sci. 17: 672-676. Stephens, W. M. 1966. Life in the turtle grass. Sea Frontiers 12: 264-275. Strawn, K. 1961. Factors influencing the zonation of sub- merged monocotyledons at Cedar Key, Florida. J. Wildl. Manage. 25: 178-189. ' Unpublished data on file National Marine Fisheries Service Biological Laboratory, St. Petersburg Beach, Fla. 33706. ' Unpublished data on file National Marine Fisheries Service Biological Laboratory, St. Petersburg Beach, Fla. 33706. 279 FISHERY BULLETIN: VOL. 69. NO. 2 Taylor, J. L., and C. H. Saloman. Tomlinson, P. B., and G. A. Vargo. 1968. Some effects of hydraulic dredging and 1966. On the morphology and anatomy of turtle coastal development in Boca Ciega Bay, Florida. grass, Thatassia testudinum (Hydrocharitaceae). U.S. Fish Wildl. Serv., Fish. Bull. 67: 213-241. '^^ Vegetative morphology. Bull. Mar. Sci. 16. 748-761. 280 EFFECT OF DIETARY FISH OIL ON THE FATTY ACID COMPOSITION AND PALATABILITY OF PIG TISSUES' Robert R. Kifer,° Preston Smith, Jr..'' and Edgar P. Young" ABSTRACT Basically, this report deals with the problem of a "fishy" flavor in the meat of pigs, which sometimes results when pigs are fed fishery products, such as fish meal, above a certain concentration in the diet. In this study, pigs were fed diets containing fish oil to investigate specifically: (1) the effect, on the taste of the meat, of feeding pigs fish oil, (2) the effect, on the taste of the meat, of withdrawing the oil from the diet at given times, (3) the fatty acid composition of the various body tissues of the pigs, and (4) the relation of composition to the taste of the meat. The principal findings of the study were: (1) The amount of the fish oil co3 fatty acids fed and de- posited was significantly positively correlated with the weighted organoleptic score' when the pigs were fed the oil containing diets to a market weight of 90.9 kg. (2) Removal of the fish oil from the pigs' diets when the pigs obtained body weight (of either 68.0 or 79.5 kg) resulted in a loss of the signifi- cant positive correlation above. (.3) Differences in the degree of unsaturation and in fatty acid comp- osition were found among the oils in the tissues examined. (4) A signifiant positive correlation was obtained between the quantity of the characteristic fatty acids (&>3) of fish oil fed and the quantity de- posited in three of the four tissues examined, the exception being the longissimtis dorsi tissue. Both the processors of fishery industrial prod- ucts and the feed manufacturers who use the products are sometimes confronted with the problem of a fishy flavor in the carcasses of animals fed diets in which these products are included. Fish oil fed directly to the animals or fed as a residual component of fish meal or of fish solubles has been shown to pi-oduce an off-flavor under certain conditions (Banks and Hilditch, 1932; Hilditch and Williams, 1964). Through practical research, the problem has been partly solved by reducing the quantity (that is, the percentage) of fish oil in the diet or by eliminating the oil during an interval of time before the animals are marketed (Frazer, Stot- hart, and Gutteridge, 1934). This latter tech- nique is not always effective, especially when fairly high (8.25'^r) levels of fish oil have been fed (Anglemier and Oldfield, 1957). ' Contribution number 4304 Maryland Agriculture Experiment Station, Department of Animal Science, Project number C33-Scientific Article A-1586. ' National Marine Fisheries Service, Washington, D.C. 20235. " Department of Animal Science, University of Mary- land, College Park, Md. 20740. ' Note the organoleptic score increased with greater unacceptability. Manuscript received January 1971. FISHERY BULLETIN: VOL. 69. NO. 2, 1971. Investigations to relate more specifically the causal agents of the oflF flavor resulting from the use of fish oil have led to the hypothesis that the long-chain polyunsaturated fatty acids of the C20-22 series commonly found in fish oil are pi-ecursors of the flavor-producing compo- nents (Banks and Hilditch, 1932; Marion and Woodroof , 1963 ; Miller, Gruger, Leong, and Knobl, 1967). Investigations by the Animal Nutrition Unit of the Bureau of Commercial Fisheries (now the National Marine Fisheries Service) Technological Laboratory, College Park, Md., using chickens, have indicated that a further partitioning of the C2(i 22 fatty acid series results in a positive correlation between individual fatty acids of these series deposited and the detection of the off-flavor (Miller et al., 1967). In a continuation of this line of investigation, the work reported here was divided into four experiments. Their purposes were to determine the following information: 1. The relation between the menhaden-oil fat- ty acid fed and the fatty acid pattern of tissue samples (namely, those of the outer and the in- 281 FISHERY BULLETIN: \'0L 61. NO 2 ner backfat, the longissimus dorsi muscle, and the liver) of pigs fed various diets with and without menhaden oil and for various intervals of time before they are marketed. 2. The organoleptic effect of the different di- etary levels of menhaden oil on the meat of the pigs and the retention or disap])earance of the off-flavor by removal of menhaden oil from the diet of the pigs when they reach a body weight of 68.0 or 79.5 kg and are subsequently marketed when they reach a weight of 90.9 kg. 3. The relation, if any, between the detection of off-fiavor and the pattern of fatty acid de- liosition in the tissue samples. 4. The metabolic interrelation of fatty acids of the various fatty acids of the omega families (coS, 0)6. 0)9). RELATION BETWEEN MENHADEN OIL FATTY ACIDS FED TO PIGS AND DEPOSITIONAL PATTERNS OF THESE FATTY ACIDS IN THE PIG TISSUES Callow (1935, 1938) indicated that the rate of deposition of fat in pigs is correlated with the iodine number of the fat and that slower growing pigs deposit a more unsaturated fat. Accordingly, we felt that our exiierimental pigs should be handled so that they would develop uniformly, thus minimizing the variation in the composition of depot fat resulting from differ- ential rates of growth. The first part of this experiment was a gen- eral study to monitor the uniformity of growth of the pigs and of the development of their car- casses. That is. we wanted to determine wheth- er the diets fed and our treatment of the pigs would I'esult in any abnormalities that might invalidate the specific findings in this first ex- periment and in the other three experiments to follow. UNIFORMITY OF GROWTH OF PIGS AND OF DEVELOPMENT OF CARCASSES Uniformity of Growth Described here are the diets, the allotment and management of the pigs, and the statistical analyses used. The diets were balanced on an equal-protein and equal-calorie basis and were fortified to sup- ply all the known nutrients required by pigs. Crude menhaden fish oil that had been stabilized with butylated hydroxy toluene' was added at levels of 0.4 Sr to 1.4 '^r. The oil replaced var- ious proportions of cerelose and Solka Flox' to give isocaloric and isonitrogenous diets (Table 1). The diets were mixed in a ribbon-type mix- er and were pelleted weekly through a 12-mm die. Steam was not used in the pelleting pro- cess. Table 2 shows the gas-liijuid chromato- graphic analyses of the oil and of the diets fed. " Level of aildition is trade secret. " Trade names are used merely to simplify descrip- tions; no endorsement is implied. Table 1. — Diet formulation used in experiment to determine the dietary level of menhaden oil that will impart off- flavors to the meat of pigs. Concentration of the g ven ingredients in the diet when th e percer tage ingredients of menhaden oil in the diet was: 0.4 0.6 0.8 I.O 1.2 1 '-^ % % % % Te % % fixed basal ingredients: Corn US #2 67.0 67,0 67.0 67.0 670 67.0 67.0 Soybean oil meal 203 20.3 20.3 20.3 20.3 20.3 20.3 Alfalfa leaf meal 3.0 3.0 3.0 3.0 3 3.0 3.0 Dicolcium phosphate 2.0 2.0 2.0 2.0 20 20 2.0 Salt {trace mineroD* .6 .6 .6 .6 .6 .6 .6 Vitamin mix^ .2 .2 .2 .2 .2 .2 J2 Variable ingredients; Cerelose 6.9 5.5 4.8 4 1 3.4 2.7 2.0 Cellulose 1.0 1. 5 20 2.5 3.0 3.5 Menhaden oil — ,4 .6 .8 1.0 1.2 1.4 Sufficient (race minerals and vitamins were present to meet the reciuiremenrs of the Notional Reseorch Council. 282 KIFER, SMITH, and YOUNG: EFFECT OF DIETARY FlSil OIL Table 2. — Gas chromatographic analysis of methyl esters of the fatty acid components of the menhaden oil and of the diets fed to pigs. Folly QCid Concenlrolion of Ihe given folly acid in menhaden oil Concenlrolion of Ihe given folly acid in the diet when Ihe percentage of menhoden oil in the diet wos: 0.4 1 0.6 1 0.8 1.0 1.2 1.4 % % % % % % % % M4:0 5.96 0.17 1.03 1.42 1.62 1.98 2.05 2.51 Mil .. 0.05 0.05 006 0.06 0.08 0.08 0,10 15:0 0.34 0.13 0.05 15 0.21 0.06 0.06 0.07 ? air 0.06 0.17 0.05 0.06 0.27 0.27 0.31 15:1 0,09 __ tr Ir tr Ir tr tr 16:0 13,10 11.77 13.15 13 59 13.26 14.10 13.46 14.85 = 16:1 a'7 10.36 0.31 1.31 1.59 1.86 2,31 2.34 2.85 17:0 0.64 0.16 0.21 0.24 026 0,26 0,27 0.31 7 __ 0.11 tr tr tr Ir tr tr 1(5:2 __ _^ .^ __ __ 17:1 1.06 0.10 0.23 0,29 0.32 0,36 0.39 0.4S ? 0.10 0.04 0.04 0,03 0.05 0.05 18:0 4.36 2.70 2.86 2.95 3.17 3-16 3.26 3.17 18:1 019 27.59 27.45 24.97 24.06 23.57 22,90 22.63 20.44 19:0 1,45 0.10 0.27 0.35 0.42 0,43 0.48 0,58 18:2 u6 1.57 51.22 44.60 41.86 40.96 39.14 38.22 35.22 ? 0,32 tr tr tr tr tr tr tr ? 024 on 0.13 0.14 0.18 0.12 0.16 0.17 20:0 0,31 0.78 0.78 0.75 0.81 0,65 0.68 0.69 18 3 o'3 0.94 3.31 2.60 2.89 2.75 2,62 2.49 2.97 20 1 1.32 0.40 0.68 0.73 0.76 0-75 0.79 0.86 18 4o-3 289 0.50 0.65 0.76 0,84 0.94 1.11 7 0.37 0.07 0.17 0.19 0.24 0,21 0.27 0.30 20:2 u9 0.18 0.07 O.II 0.12 0.13 0,07 0.13 0.14 20:2 0)6 0.06 tr tr tr ir Ir Ir 0.06 20:3 o9 0.13 tr 0.08 0.08 009 0,05 0.10 0.11 22:3 ^6 0.05 Ir tr 0,04 ir Ir tr tr 20:4 o;6 0.69 0.22 0.37 042 0.42 0,37 0.45 0.53 22:1 oj2 0.30 tr 0.12 0,09 0.12 0,07 0.13 0.16 20:4 o;3 1.25 0.08 0.29 0,34 0.41 0,42 0.52 0.59 20:5 0.3 12.95 0.11 2.19 2,95 3.35 3,96 4.28 5.01 7 __ 0.23 0.40 0,26 0.34 0,15 0.28 0.40 24:0 09 __ tr tr tr Ir tr tr 22:4 w6 059 0.29 0.53 0.47 0.48 0,46 0.54 0.57 22:5 oj6 0,43 .. tr 0.24 Ir 0,26 0.46 0.45 22:5 oj3 1.63 0.40 0.50 0.63 0,65 0.77 0.86 22:6 0)3 8.47 — 1.71 2.49 2.76 3,29 3.44 4.14 "14:0" meons that the fatly acid has 14 carbon atoms per molecule ond no unsoluroted bond. "16:1 oj7" means thot in the folly acid Ihe unsaturated bond occurs at Ihe seventh bond from the terminal methyl group. "Ir" means trace. Seven Yorkshire gilts each weighing about 27.3 kg were allotted to each of the seven treat- ment groups. Two of the seven pigs of each menhaden-oil group were fed the appropriate oil-containing diet until they attained a body weight of 68.0 kg and then were fed the control diet until they attained a body weight of 90.9 kg. Similarly, two additional pigs of each men- haden-oil group were fed the appropriate oil- containing diet to a body weight of 79.5 kg and then also were fed the control diet to a body weight of 90.9 kg. The remaining three pigs were continuously fed the various test diets con- taining menhaden oil until they each also at- tained a body weight of 90.9 kg. Feed was offered twice a day (for a maximum of 1 hr per feeding) to the pigs in individual crate- type pens. This interval of time was considered to be adequate to permit the pigs to eat the same total amount of food that they would have eaten ad lib. Data on rates of gain and consumption of feed were recorded weekly. Data obtained on rates of gain and utiliza- tion of feed were subjected to an analysis of variance (Snedecor, 1956). Table 3 presents the rates of gain, utilization of feed, and quantity of oil consumed by the pigs fed diets containing the various percentages of menhaden oil. Results of the analyses of variance for each 283 FISHERY BULLETIN' : \ OL. 09. NO- I Table -Rates of gain, utilization of feed, and quantity of oil consumed by pigs fed diets containing various percentages of menhaden oil. Relative amount of nienhaden Average doily gain Ratio of feed to gain Mean quontity of oil consumed by pigs fo o body weight of: oil in diet Mean SD Mean SD 68.0 kg 1 79.5 kg 90.9 kg ■/o kv. H *? H i:% 63 065 3.45 0.136 0.4 .64 065 3.26 .105 52 0.62 0.85 0.6 .60 ,047 3.34 .093 0.85 0.96 1.30 0.8 .64 .045 3.47 095 1.03 1 32 1.90 1.0 .64 .025 3.26 .080 1.20 1.48 2.16 1.2 .66 .068 3.28 .100 1.53 1.78 2,70 1.4 .64 .044 3.25 .084 1.61 2.12 3.22 criterion of evaluation indicate that tlie.se cri- teria did not differ significantly. Development of Carcasses The .yield of lean cuts was obtained as an accumulative value for the four commercial lean cuts — namely, hams, loins, shoulders (picnics), and Boston butts. Ci'os.s-sectional measure- ments of the longlsi^hnus dorsi muscle of the loin were obtained by cutting- the loin at the 10th rib, tracing- the muscle area onto pajjer, and measuring the perimeter of the area by means of a ])lanimeter to convert the encom- passed area to square centimeters. The thick- ness of the backfat was based on an average of three measurements taken at positions opposite the first rib, the last rib. and the last lumbar vertebra. Table 4 presents the data on the dressing per- centage, lean-cut percentage, loiu/lssiiniis dorsi area, and backfat thickness obtained from pigs fed the various diets containing menhaden oil. The analyses of variance for each criterion of evaluation indicate that no significant differ- ences occurred among these factors that reveal the growth reaction of the pigs to their diet. Thus the pigs develojied uniformly during the feeding trials. Consequently any differences that may be found in the fatty acid composition of the tissues should be related to the oil in the diet rather than to markedly different growth of the pigs. RELATION OF DEPOSITIONAL PATTERNS TO FATTY ACIDS IN OIL FED TO PIGS In this section, we are concei-ned with the fol- lowing three sulyects: (1) the differences found in the degree of saturation both within and among tissues, (2) the fatty acids identified, and (3) the relations of the quantity of fatty acids fed to the quantity deposited in the var- ious tissues. Differences Found in Degree of Saturation Both Within and Among Tissues Described here are (1) the tissue samjiles used, (2) the extraction of lipids, (.3) the prep- aration of methyl esters, and (4) the quantita- tive gas-liquid-chromatographic technique. Samples were taken from the outer and the inner backfat tissue, the lo)igissi>nus dorsi, and the liver in the following manner. From each animal, a sample of backfat was obtained dor- sally to the 10th to 12th ribs. This sample was then divided into the "outer" and "inner" fat layers. Samples of the muscle were taken from the eye of the Io)u/issimiis dorsi at the 10th rib. Samples of the liver were taken from the right central lobe. All samples were i)!aced in vials, Ijrotected with nitrogen, and held at —20° C until the lipids were extracted from them. The lipids were extracted from the samples by the homogenization of the tissue in a mechan- ical blender with a 2: 1 mixture of chloroform and methanol for 2 min. The solvent mixture was added in the proportion of 5 ml of mixture to 1 g of sami^le. The slurry was filtered through a Buchner funnel, and the filter paper and the nonfilteral)le portion were re-extracted for an- other 2-min period. The filtrate was evaporated in a rotary vacuum evaijorator over a 60° C wa- ter bath. The dried sample was redissolved in 284 KIFER. SMITH, and YOUNG; EFFECT OF DIETARY FISH OIL Table 4. — Dressing percentage, lean-cut percentage, longistiimns dorsi area, and backfat thickness obtained from pigs fed various diets containing menliaden oil. Relative Relative yield of; Lonsiilimul doni area Backfar amount of menhaden Dressing Lean c jtsi thickness oil in diet Mean SD Mean SD Meon SD Mean SD % % % % % cm^ rm" cm cm 83 8 ±2.36 38.9 ±2.45 32.39 ±5.78 3.56 ±062 0.4 83.4 ±1.23 40.2 ±1.68 33.68 ±4.83 3.30 ± .45 0.6 82.0 ±1.89 39.6 ±1.07 31.87 ±3.68 3.61 ± .19 0.8 83.1 ±1.06 39.4 ±1.32 30.78 ±5.08 3.61 ± ,29 I.O 82.7 ±1.62 40.1 ±0.88 32.78 ±4.39 3.30 ± .27 1.2 82.0 ±2.21 38.3 ±2.14 31.74 ±4.00 3.53 ± ,62 1.4 82.1 ±1.86 38.1 ±1.61 30.91 ±5.19 3.65 ± .40 petroleum ether (30° to 60° C boiling point), poured into a separatory funnel, and washed twice with a 20^^ solution of NaCl. The layer of petroleum ether was evaporated in the rotary evaporator, and the e.xtracted fat was trans- ferred to containers in which it was protected by nitrogen and was stored at — 20° C until methyl esters were prepared from it for analysis. The methyl esters of the fatty acids were pre- pared as follows: Five ml of anhydrous methanol and about 50 mg of freshly cut and shiny sodium were placed into a small test tube. After the sodium had reacted, six to eight drops of the extracted oil were added and heated to reflux on a steam bath for 2 min with agitation. The end point of the reaction was signaled when the solution became clear. The reaction solution was quenched with 5 ml of distilled water and was transferred to a sep- aratory funnel. The mixture was extracted with two 10-ml portions of petroleum ether (30° to 60" C boiling point). The final water layer was discarded, and the two petroleum ether extracts were combined. The petroleum ether solution was washed with 10 ml of 5 ""r aqueous HCl so- lution. The acid wash was followed by succes- sive washes with 15-ml and 10-ml aliquots of 20''r NaCl solution. The washing was com- pleted when pH paper tested neutral. The ethereal solution of methyl esters was dried over 3 g of anhydrous Na2S04, filtered, and evaporated over a 60° C water bath, using a vacuum rotary evaporator. To check for purity, we made a thin-layer chromatogram of the ester solution using silicic acid paper. Methyl myristate was used as the control. A solution of 90 parts petroleum ether, 10 parts ethyl ether, and 1 part formic acid was used to elute the esters. The chromatogram was develo])ed in iodine vapor. Methyl esters of pure fatty acids were used as reference standards for the C14-24 saturated acids, Cii; -:24 monoenoic acids, plus linoleic, lino- lenic, ai'achidonic, eicosapentaenoic, and docosa- hexaenoic acids. Also concentrates of 16: 2, 16:3, 16:4, and 18:4 methyl esters that were obtained by fractional distillation and urea-inclusion com- pound fractionalization were used as reference standards." As a secondary reference mixture, methyl esters from whole menhaden oil were also analyzed. From a plot of the logarithms of the retention times (relative to stearate) versus the number of carbon atoms, nearly linear relations were observed for homologous series (Farquhar, Insull, Rosen, Stoff'el, and Ahrens, 1959). Iden- tifications were further verified by applying the graphical method of James (1960) for analyses on columns jiacked with diethylene glycol succi- nate polyester and Apiezon L. These plots pro- vided the necessary reference data for identifi- cation of the various tissue lipids analyzed." ' Tlie staff of the National Marine Fisheries Service Technological Laboratory, Seattle, Wash., made the fractional distillations and urea-inclusion compound frac- tionations. ' The fatty acids of the oil fed and of the animal tissues were identified initially in collaboration with the staff of the National Marine Fisheries Service Techno- logical Laboratory, Seattle, Wash. 285 FISHERY BULLETIN: VOL. 69, NO. Methyl esters of fatty acids taken from the various tissues were analyzed with an F&M Bio- medical Model 400 gas chromatograph. The in- strument was equipped with a hydrogen flame detector. The column used was comi)osed of 4.0- mm ID by 243.8-cm Pyrex glass containing 5.0'^ r (by weight) of diethylene glycol succinate poly- ester (DEGS from Wilkens Instrument and Re- search Inc.) supported on 80- to 90-mesh acid- base washed and siliconized flux-calcinated diatomaceous eai-th (Anakron ABS). The op- erating conditions were as follows: column temperature, 165° C; flash-heater temperature, 285° C; detector temperature, 200° C; and in- itial attenuation that corresponds to 10 to 14 amp full-scale deflection. The inlet pressure of the column measured 40 psi of helium, the flow measured 53 ml per min at the outlet of the column. The size of the injected sample was about 0.12 juliter. The area-percent method was used to deter- mine the corresponding peak areas of the curves obtained from the gas-liquid chromatographic recorder. The fatty acid composition (in aver- age percentage) of each sample was calculated by multiplying peak height by retention time and then multiplying this product by 100 and dividing by the total area. Certain diff'erences were obtained in the total degree of saturation and quantity of specific Table 5. — Summary of gas-liquid cliromatographic analyses indicating comparative degree of unsaturation and quantity of selected fatty acids within and among the tissues obtained from pigs fed either 0% or 1.47c dietary menhaden oil. Type of fatty octd Concentration of the vorious fatty acids in the vorious tissues when the relative amount of menhaden oil in the diets was: 0% Backfat Inner tissue Outer tissue Longijiimul dorsi tissue Liver tissue 1 .4% Backfat Inner tissue Outer tissue Longiisimus dorti tissue Liver tissue % % % % % % % % Saturated fatty ac ids 34.56 28.37 33.74 34,63 35.42 28.77 33,05 34.66 Unsaturated fatty acids 65.44 71.63 62,82 65,37 64.58 71.23 64,62 65.34 Unsaturoted bond in the fatty acids: I 47.68 49.03 39,38 17,53 47.17 46.16 45,96 15.44 2 16,35 20.49 14,40 16.85 17.39 20.75 11.46 17.43 3 0.93 1.40 1.37 2.35 1.40 1.73 1.15 1.85 4 0.40 0.72 4.95 19.08 0.84 0.88 3.27 12.80 5 0.19 0.27 2.03 2.96 0.80 1,53 2.17 8.52 6 of 0.03 0.05 0.69 2.26 0.40 0,50 0.62 8,11 Equivalent degree unsoturotion in the fatty acids: 1 47,68 49.03 39.38 17.53 47.17 46,16 45.96 15.44 2 32.70 40.98 28.80 33.70 34.78 41,50 22.92 34.86 3 2.79 4.20 4.11 7.05 4.20 5,19 3.45 5,55 4 1.60 2.88 19,80 76.32 3.36 3,52 13.08 51.20 5 095 1.35 10,15 14.E0 4.00 7,65 10.85 42.60 6 ed 0.18 0.30 4,14 13.56 2.40 3,00 3.75 48.66 Total 85.90 98.74 106,38 162.96 95.91 107,02 99.98 198.31 Individually select fatty acids: 16:0 19.90 18.49 21,78 13.04 20.39 18,64 19.62 12.80 18:0 12.18 7.78 9,24 19.37 12.49 8,08 10,74 18.76 18:1 u9 43.67 44.94 34,33 15 86 40.52 42,04 41,16 13.57 18:2 (..-d 15.07 18,99 13,24 15.27 16.18 19.40 10.47 15.87 18:3 li3 0.74 1.18 0,59 0.44 1.02 1,37 0.52 0.57 18:4 u6 0.34 0.49 2,44 18.29 0.32 0,41 I.6I 12.15 20:4 0.-3 0.06 0.08 2.42 0.49 0.20 0,31 1.61 0.54 20:5 k3 0.09 0.13 0.98 0.51 0.27 0,52 1.12 4.18 22:5 f3 0.10 0.14 1.05 2.45 0.53 l.OI 1.04 4.34 22:4 k3 0.03 0,05 0.69 2.26 0.40 0.50 0.62 8.11 Note: The equivalent degree of unsaturation In the fatty adds was obtained by multiplying the number of double bonds by the quantity of fatty acid. 286 KIKER. SMITH, and VOLXG: EFFECT OF DIETARY FISH OIL fatty acids found within and among the tissues examined. All the fatty acids that were identi- fied will be discussed in the next section. For illustrative purposes. Table 5 presents selected results obtained with the various tissues. The outer backfat had the lowest total concentration of saturated fatty acids of all the tissues, regard- less of whether the diet contained menhaden oil or did not contain it. The remaining- tissues (inner backfat. liver, and lonpissimus doisi) were all higher than the outer backfat and did not differ markedly from each other in the total concentration of saturated fatty acids. The difference in degree of saturation when confined to comparisons between the inner and outer backfat is in agreement with rei^orts l\v Banks and Hilditch (1932) and Sink, Watkins, Ziegler, and Miller (1964). The simple ratio of the total quantity of saturated to unsaturated fatty acids, however, does not describe the true character of the unsaturated fatty acids found within the tissues or among them. An examination of the quantity of unsatura- tion on the basis of the number of double bonds and the relative quantities of the corresponding fatty acid groups indicates marked differences among the tissues. Both the longissimns do)-si tissue and the liver tissue contain markedly less fatty acids with one unsaturated bond than do either of the back- fat tissues, regardless of the dietary treatment. This difference no doubt is reflected by the 18:1 co9 content. The concentration of fatty acids with two un- saturated bonds in the outer backfat tissue is higher than that in the remaining tissues and apparently indicates a differential concentration of 18:2 w6. The difference most evident among the tis- sues with respect to the fatty acids with three unsaturated bonds is the higher concentration found in the liver tissue. Both the longissimus dorsi and the liver tissue contained considerably more of the four-unsatu- rated-bond fatty acids than did the backfat tis- sues. The liver, in turn, contained about four times the concentration found in the Inngissimus dorsi. Incorporating menhaden oil into the diet lowered the magnitude of these differences among the tissues. The relative differences among the tissues in the case of the longissimns dorsi tissue reflect about equal quantities of the isomeric fatty acids 20:4 ojG and 20:4 coo. The concentration of the fatty acids with four unsaturated bonds in the liver tissue is due jirimarily to the 20:4 w6 iso- mer; only small concentrations of the 20:4 w3 isomer were found. Similarly, the concentration of fatty acids with five and six unsaturated bonds in the lon- gissimns dorsi and liver tissues was markedly higher than in the l)ackfat tissues. The incor- lioration of menhaden oil into the diet resulted in increased concentrations of these fatty acids in all tissues, although the differences among tissues were of the same magnitude as the dif- ferences occurring in the absence of the men- haden oil. The variable concentrations of the fatty acids with five and six unsaturated bonds, owing to treatment differences, reflect differ- ences in the quantities of 20:5 coo. 22:. 5 coo, and 22:6 w3 fatty acids. On the basis of the equivalent degree of un- saturation obtained by the multiplication of the number of unsaturated bonds by the quantity of fatty acids of that category, the relative degree of unsaturation of the four tissues is: inner backfat, 8-5.9; outer backfat, 98.7; longissimus dorsi. 106.4; and liver, 164.0. The incorporation of menhaden oil did not change the relative dif- ferences among tissues, but it did result in a treatment difference. The relative degree of unsaturation among the treatments was of the magnitude of 10 to 30 units greater for all tis- sues exce])t the loiigissimus dorsi. Thus, these results generally conform with those previously reported that various tissues differ in fatty acid comjiosition (Brown and Deck, 1930: Banks and Hilditch, 1932; Sink et al, 1964) and that dietary oils alter this fatty acid jjattern and degree of unsaturation of the animal tissues of monogastric animals (Ellis and Isbell, 1926a, 1926b; Ellis and Zeller, 1930; Ellis, Rothwell, and Pool, 1931; Bhattacharya and Hilditch, 1931; Hilditch and Pedelty, 1940). 287 FISHERY BULLETIN: VOL. 69, NO. 2 Fatty Acids Identified Table 6 reports the fatty acids identified by the method of gas-liquid chromatographic anal- ysis described in the preceding section. Table 6. — Fatty acids identified in pig tissues. Presence or absence of the fotty acid in: Fatty acid Backfat Longisjimui dorii tissue Liver Inner tissue Ou ter tissue tissue 22:6 a-3 + -1- -1- + 22:5 a-3 + + + + 20:5 a..3 + + -f- + 20:4 u3 + + -t- + 18:4 <.-3 + + + + 18:3 c;3 + -t- -t- + 22:5 ic6 _ Irace trace + 22:4 ^6 — + + + 20:4 u6 -1- -1- + + 20:2 ..■6 — — + + 18:2 ^•6 + + + + 21:1 u9 _ — + + 20:2 u9 + + + + 20:1 10-9 + + + + 18:1 1^9 + + + + 22:1 (?) — — trace + 20:3 (?) + + + + 16:2 + + + + 16:1 u7 + + + + 15:1 + + trace + 14:1 + + + + 20:0 + -t- -1- + 19:0 + — + + 18:0 + -t- -t- + 17:0 + + + + 16.0 + + + + 15:0 + + -t- + 14:0 + + + 4- Twenty-eight fatty acids were identified in the liver tissue, whereas a lesser number was identified in the three other tissues (inner and outer backfat and Io)n;issim us doisi) . The fatty acids identified included those reported by Sink et al. (1964) plus unsaturated 18, 20, 22 carbon fatty acids of three of the fatty acid families — co3, 0)6, and aj9 — according to current classi- fication (Mohrhauer and Holman, 1963a). With respect to the two backfat tissues, the acids found in addition to those reported by Sink et al. (1964) are as follows: 15:1, 16:2. 20:1 aj9, 18:4 co3, 20:2 aj9. 20:3, 20:4 0)3, 20:. 5 0)3, 22:4 o)6, and 22:5 oj3. The liver and lonf/isshmis dorsi tissue also contained 20:2 oj6, 21:1 w9, 22: 1, 22:5 w6, and 22:6 w3. Ilill (1966) reported, however, the presence of most of these fatty acids in various tissues of miniature pigs with the exception of 20:4 oj3, which we found in our pigs. All of these fatty acids, except 20:4 o)3, have also been noted in rat tissue (Mohrhauer and Holman, 1963a, 1963b, 1963c) , and all of them including 20: 4 oj3, have also been noted in chick tissue (Miller et al., 1967), in fish tissues and in seal tissue (Ackman, Burgher, and Jangaard, 1963; Ack- man, Jangaard, Hoyle, and Brockerhoff', 1964). The relation of the fatty acids fed (A') to those deposited in the various tissues (Y) was established by correlation and polynomial re- gression analyses. A polynomial regression computer program prepared by the Biomedical Division of the University of California, Los Angeles, was used. The extent of analysis of the data was limited to the fourth polynomial degree. Regression coefficients, standard errors of regression, correlation coefficients, analyses of variance, and data plots (predicted and ob- served) were obtained. Correlation and polynomial regression anal- yses of the gas-liquid chromatographic data presented in Tables 7 to 10 indicate that the marine-type polyunsaturated fatty acids of the linolenic acid (oj3) family were deposited in all four tissues examined. In general, a significant positive correlation was obtained between the quantity of the o>3 fatty acids fed and the quantity deposited in the various tissues. This relation was not ob- tained, however, with the longissimus dorsi tis- sue. The only explanation we have is that the reaction caused by difficulties in the extraction of the fatty acids and their subsequent sepa- ration masked any pattern. Definite relations between the amounts of most of the o)3 fatty acids fed to pigs and the amounts deposited were found in the liver tis- sues and in the inner backfat tissues and the outer ones. Specifically, the quantity of two of the men- haden oil fatty acids (22:5 w3, and 22:6 o)3) found in the liver was positively correlated (0.01 'r ) with the quantity of oil fed to the pigs until they were of market weight (90.9 kg) . The correlation for 20:5 coS approached significance. 288 KIFER, SMITH, and YOUNG: Table 7. — Liver tissue: EFFECT OF DIETARY HSU OIL concentration of fatty acids found in liver tissue and correlation to quantity of various fatty acids fed for various time intervals. Fatly acid 22:6 i^3 20:4- a.-3 18:4 u3 20:4 u6 18:2 i^6 weight group Concentrotion of fotty acid in liver tissue when the percentage of menhaden oil in the diet wos; Correlotion coefficient «g — — — — — — — — .Irea ptTCent 0/ fatty acid — — — 90.9 2 25 5.70 7.88 7.72 840 8.90 7.49 0.69** Quadratic 79.5 2 25 4.65 4.98 5.11 6.25 5.82 7.43 0.70" Linear 68.0 2.25 4.30 4.70 4.33 5.69 5.54 5.12 0.65* Linear 90.9 2.55 3.66 4.42 4.48 5.12 5.17 4.85 0.78** Quodratic 79.5 2.55 2.94 3.59 3.47 4.00 3.73 4.05 0.56* Linear 68.0 2.55 2.70 4.64 3.75 3.70 3.93 4.11 0.59* Linear 90.9 0.56 3.55 6.23 6.52 8.11 8.46 8.14 0.89** Quadratic 79.5 056 1.57 2.07 1.96 3.33 2.30 2.20 0.50 680 0.56 1.00 1.25 1.65 2.12 3.49 2.20 0.67* Linear 909 039 0.71 054 0.50 0.56 0.43 044 -0.21 79.5 0.39 1.45 0.75 0.50 0.60 069 0.29 -0.42 68.0 039 0.58 0.11 067 0.71 0.54 0.89 0.39 — 90.9 008 0.11 0.05 08 0.09 0.12 0.07 0.00 79.5 008 0.26 0.05 07 0.04 0.06 0.06 -0.29 68.0 0.08 0.07 0.05 0.08 0.06 10 0.09 0.33 — 90.9 0.47 0.67 0.59 0.66 0.73 0.94 0.66 0.26 79.5 0.47 0.77 0.89 0.54 0.56 051 0.48 -0.39 __ 68 0.47 0.52 0.45 0.57 0.50 0.39 0.59 0.12 — 90.9 0.24 0.05 0.06 0.05 12 Oil 0.03 -0.27 79.5 0.24 0.23 0.10 0.05 0.17 0.16 0.00 -0.60* Linear 68.0 0.24 0.17 0.10 0.10 009 014 0.03 -0.83** Linear 90.9 1 21 0.43 0.28 033 0.29 0.33 0.21 -0.65** Cubic 79.5 1.21 0.53 0.34 0.45 0.47 0.48 0.40 -0.63* Cubic 68.0 1 21 0.74 0.74 0.64 058 0.53 0.63 -0.69* Linear 90.9 18.58 12.37 11.53 10.50 882 8.84 9.69 -0.70" Quadratic 79.5 18.58 15.23 14.48 14.65 15.12 14.99 12.84 -0.19 ._ 68.0 18.58 17.93 16.95 15.98 16.06 16.66 13.92 -0.74** Lineor 90.9 0.52 0.19 0.18 020 14 0.17 0.17 -0.52* Cubic 79.5 0.52 0.43 0.50 0.46 0.26 0.40 36 -0.34 68.0 052 0.20 0.40 0.23 0.40 0.48 0.38 0.13 _ 90.9 15.87 17.03 16.17 16.32 15.95 15.93 15 79 -0.33 .. 79.5 15 87 14.28 15.69 16.00 17.05 15.93 15.40 0.16 68.0 15.87 16.78 16.27 16.10 16.39 14.48 16.41 -0.12 — 90.9 27 0.31 0.13 0.15 15 0.15 0.14 -0.59** Linear 795 0.27 0.52 0.45 0.30 0.13 0.25 020 -0.40 .. 68.0 027 0-42 0.22 0.28 0.29 0.27 0.18 -0.47 — 90.9 0.82 054 0.60 0.54 0.59 0.58 0.54 -0.12 .. 79.5 0.82 1.29 0.78 0.68 0.44 0.63 0.52 -0.48 68.0 0.82 42 0.59 0.56 1.25 0.59 1.05 0.52 — 90.9 0.28 0.24 0.19 0.30 0.20 0.23 0.17 -0.22 „ 79.5 0.28 0.44 0.41 0.26 0.20 0.21 0.22 -0.38 ._ 68.0 0.28 0.22 0.27 0.25 0.22 0.23 0.25 -0.28 ~ 90.9 16.61 15.28 13.16 14.80 13.56 14.91 14.17 -0.09 .. 79.5 16.61 16.49 18.55 16.18 12.67 13.33 13.56 -0.49 68.0 16.61 15.64 15.55 16.09 14.59 9.68 1299 -0.54 _. P <.os P < 01 Polynomial regression analyses of these data in- dicate that the incorporation pattern of the fatty acids (20:5 co3, 22:5 co3, 22:6 aj3) was quad- ratic, with the rate of deposition being greater at the lower levels in the diet. Removal of men- haden oil from the diet of the pigs at the two body weights, 68.0 or 79.5 kg, did not alter this pattern markedly. The principal changes were a reduction in the relative degree of significant response (O.Ofr to 0.059r) and an alteration 289 FISHERY BULLETIN: VOL. 69, NO. 2 Table 8. — Inner backfat tissue: concentration of fatty acids found in backfat tissue and correlation to quantity of various fatty acids fed for various time intervals. Fatty acid Pig weight group Concentration of fotty acid in inner fat tissue wtien the percentage of me oil in the diet was: back- nhoden Correlation coefficient Kind of regression 0.4 0.6 _L 0.8 1.0 1.2 1.4 90.9 79.5 68.0 . percent ol jatty acid 0.29 0.54 0.51 0.37 0.20 1 .06 0.49 0.31 0.28 0.48 0.40 0.31 0.76" 0.56* 0.49 22:6 u;3 0.13 0.19 0.09 0.28 0.21 15 Linear Linear 22:5 u;3 90,9 79.5 68.0 10 0.10 0.10 0.35 0.25 0.18 0.48 0.46 0.20 0.54 0.28 0.39 0.96 0.60 0.24 0.73 0.23 0.54 0.88 0.19 0.51 0.77" -0.01 0.71" Linear Linear 20:5 ^3 90.9 79.5 680 07 0.07 007 0.12 0.08 04 0.18 0.17 Oil 0.29 0.20 0.12 0.44 0.27 0.12 041 0,16 0.33 0.39 0.22 0.19 0.71" 0.55* 0.71*> Linear Linear Linear 20:4 US 90.9 79.5 68 06 0.06 0.06 015 Oil 0.06 0.17 0.14 0.10 0.24 0.23 0.19 0.34 0.34 0.11 0.26 0.19 0.14 0.29 0.17 0.14 0.72" 0.43 0.89'* Linear Linear 18:4 u3 90.9 79.5 68.0 06 0.06 0.06 0.10 0.10 0.06 0.13 0.11 0.04 0.13 0.11 0.08 0.18 0.17 0.10 0.16 0.06 0.15 0.18 0.12 0.30 0.70* • 0.15 0.54 Linear 18:3 u3 90,9 79.5 68 0.74 0.74 0-74 0.82 0.72 86 0.89 0,88 080 0.94 0.92 0.77 1.04 1.08 1.03 1.02 0.77 0.71 1.10 0.82 1.15 0.52' 0.17 0.52 Linear 22:5 u6 90,9 79.5 68.0 Not identified - -- 22:4 U!6 909 79.5 68.0 Not identified - -- 20:4 m6 90.9 79.5 68.0 34 034 0.34 0,33 028 0.36 0.33 0.35 0.30 0.29 0.38 0.34 0.39 0.34 0.34 0.44 0.32 0.32 0.33 0.33 0.28 0.03 0.09 -0.60- Linear 20:2 w6 90.9 79.5 68.0 Not identified - - 18:2 u6 90.9 79.5 68.0 15.07 15.07 15.07 1553 13.15 17.45 1545 15.43 16.70 15.25 16.46 14.69 17.51 17.97 17.48 16 30 15.16 13.12 17,38 14.06 17.09 0.41 0.10 -0.01 — P <.05 P <.01 in the patterns of incorporation of these fatty acids (quadratic to linear). No statistical relation was found for the re- maining three fatty acids (18:3 w3, 18:4 c <.01 Kg 0.81 .83 .91 1 26 1,29 1,36 1.77 1.86 2.07 2,00 2.15 2,35 2,55 2.74 2.74 2.82 3.20 3.25 .l! % ol Jill 0.4 1.0 1.2 23 21 27 22 22 25 31 26 32 30 34 35 29 36 28 37 32 296 KIFER. SMITH, and YOUNG: EFFECT OF DIETARY FISH OIL Table 21.— Panel test Trial II— weighted organoleptic scores obtained with inner backfat of pigs fed various levels of menhaden oil in the diet until the pigs attained a body weight of 79.5 kg. Quantity of oil consumed (X) Weighted organoleptic score (}') Kg As % of diet 21 0.55 .64 0.4 25 27 .94 .99 .99 0.6 20 21 la 1.30 1.34 0.8 20 20 1.36 1.48 1.0 35 23 1.77 1.2 34 2.11 1.4 32 Table 22. — Panel test Trial II — weighted organoleptic scores obtained with inner backfat of pigs fed various levels of menhaden oil in the diet until the pigs attained a body weight of 68.0 kg. Quantity of oil consumed (A') W 'ighted organoleptic score ()') Kg .is % of dut 21 0.45 .54 0.4 23 20 .72 0.6 21 .98 1.09 0.8 23 22 1.21 1.21 1.0 22 25 1.54 1.2 26 1.61 1.61 1.4 23 17 Table 23. — Correlation and polynomial regression anal- yses of quantity of menhaden oil consumed (.Y) to weighted organoleptic score {Y) when the oil was fed until the pigs attained a body weight of 90.9, 79.5, or 68.0 kg. Oil fed to Correlotion coefficient Regression coefficient Standard error of regression Lost degree of polynomial significant Degree Kg 90.9 79.S 68.0 0.82" .49 .21 2.169 2.375 0.617 0,371 1.325 0.967 329.58" 3.21 0.41 P <.01 loss of the significant positive correlation be- tween the variables, although the correlation coefficient obtained for die group weighing 79.5 kg approached significance. These results of organoleptic tests are in agreement with reports of Miller et al. (1967), which indicate that co3 family fatty acicds, when fed and subsequently deposited, are positively correlated with organoleptic scores obtained with broiler flesh. The results are in partial agreement with the hypothesis of Banks and Hilditch (1932), who suggested that the fatty acids of the C20-22 series are associated with an off- (fishy) flavor. Both the results reported here and those reported by Miller et al. (1967) indicate that fatty acids of the coS family con- taining 18 to 22 carbon atoms are positively cor- related with the incidence and degree of off"- flavor in pig or broiler flesh. These fatty acids may be causal agents for the oflf-flavor, or they may not be. In fact, they probably are the precursors of the compound producing the off- flavor. In these experiments, the inclusion of the men- haden oil in the diet of the pigs resulted in no physiological abnormalities other than the pro- duction of off-flavor and an alteration in the pat- tern of fatty acids in the tissues. This result was not unexpected, because previous work at the National Marine Fisheries Service Techno- logical Laboratory at College Park had indi- cated that levels of menhaden oil in excess of 10% of the diet are necessary to produce the physiological abnormalities of e.Kudative diath- esis and muscular dystrophy exiierimentally. Adding various antioxidants (vitamin E, sele- nium, and ethoxyquin) to the diet at compen- satory levels prevented the development of these abnormalities (exudative diathesis and muscu- lar dystrophy) in chicks fed menhaden oil at high concentrations (Miller, Leong, Knobl, and Gruger, 1965). METABOLIC INTERACTIONS OF FATTY ACIDS OF THE OMEGA FAMILY ( (o3, u6, w9) Mohrhauer and Holman (1963a), Rahm and Holman (1964), Tinsley (1964), and Lowry and Tinsley (1966) have demonstrated that feeding 297 FISHERY BULLETIN: VOL, 69, NO. 2 rats increasingly higher concentrations of lin- olenicacid (18:3co3) increases the concentration of the fatty acids of the &;3 family in the liver and that the proportion of the fatty acids of the oleic (18:1 w9) and linoleic (18:2 coG) fam- ilies are concomitantly reduced. They hypothe- size that this interaction is due to the compe- tition for enzymes necessary for elongation and desaturation within the individual families of fatty acids. Since our pig e.xperiment included an in- creasing quantity of 18:3 coS in the diet, the question arose as to whether this hypothesized competitive interaction actually occurred. Trial II results were analyzed by correlation analysis and polynomial regression analysis as previously described. The quantity of men- haden oil consumed constituted the X axis, and the quantity of the 17:1 o)9 or 18:2 co6 family fatty acid in question the Y axis. The 0)3 family fatty acids incorporated into the diet of the pigs as menhaden oil and sub- sequently ingested resulted in a significantly depressed deposition of the quantity of certain members of the w6 and c