24 



ONTOGENY AND SYSTEMATICS OF FISHES-AHLSTROM SYMPOSIUM 



the basis for ontogenetic studies of fishes. These are essential 

 for the identification of ichthyoplankton collections, and also 

 present characters for systematic analysis. Data provided in 



these descriptions have proved useful in studies of the physi- 

 ology, behavior and ecology of the early stages of fishes. 



National Marine Fisheries Service, Southwest Fisheries 

 Center, P.O. Box 271, La Jolla, California 92038. 



Synopsis of Culture Methods for Marine Fish Larvae 

 J. R. Hunter 



THE objective of this paper is to provide a synopsis of present 

 technology for small-scale laboratory culture of marine fish 

 larvae. The technology of marine fish culture is relevant to this 

 book because it is one of the best ways to obtain a taxonomic 

 series. "Ahlie" Ahlstrom was a strong proponent of this ap- 

 proach and I lectured on the subject at his request for his courses 

 on larval fish systematics. Marine fish culture has often been 

 reviewed (May, 1970-, Houde, 1972a; Houde and Taniguchi, 

 1979; Shelboume, 1964; Kinne, 1977) and many additional 

 references may be found in the previous reviews. The key feature 

 of my review is that it is a condensed practical guide and key 

 to the literature for beginners interested in small-scale laboratory 

 culture of marine fish larvae; culture of freshwater fishes is not 

 considered. 



Eggs 



Sources. — Pelagic fish eggs can be obtained from plankton tows, 

 by catching ripe fish and fertilizing the eggs, and by induction 

 of spawning of laboratory brood stock. 



Let eggs taken in plankton tows stand in quart bottles for 0.5 

 h, then remove plankton from bottom of jar and add fresh sea 

 water (a second decanting may be required). Jars are stored on 

 their sides in an insulated ice box with a refrigerant for 24 h or 

 longer with the temperature kept within spawning range. 



Virtually all marine clupeoid fishes (Blaxter and Hunter, 1982) 

 and probably most other pelagic marine fishes spawn at night, 

 hence running ripe fish are more common at night or just before 

 sunset (final egg maturation or hydration occurs just before 

 spawning). After an egg is spawned in sea water its fertility 

 decreases but the maximum time for it to become infertile is 

 highly variable among species, varying from 6 minutes to over 

 3 hours (Ginzburg, 1972). Sperm in sea water may remain fertile 

 for days (Ginzburg, 1972) although fertility periods as short as 

 30 seconds have been observed (Haydock, 1971). Owing to the 

 great variation in the time eggs and sperm remain fertile it is 

 preferable that sperm and eggs be mixed immediately after they 

 are obtained. 



Storage of gametes may be helpful since mature males and 

 females are not always available simultaneously and crosses 

 between subpopulations may be desired. It is well known that 

 sperm can be stored for extended periods ( 10 or more hours) if 

 kept cool and maintained in the concentrated form and not 

 activated by sea water (Ginzburg, 1972; Erdahl and Graham, 

 1980). Fertilization of Clupea harengus eggs may be obtained 



after 6-7 days dry storage at 4° C but a high hatching rate is 

 expected only after periods less than 36 h (Blaxter and Holli- 

 day, 1963). It is now possible to extend the life of fish sperm 

 for much longer periods using cryopreservation techniques 

 (- 196°C) (Erdahl and Graham, 1980). Various cryoprotective 

 agents have been used to freeze sperm of marine fishes including 

 glycerol (Blaxter and Holliday, 1963), glucose, NaCI, Ringer's 

 solution and fish serum (Hara et al., 1982). 



The stress of capture causes female Katsiiwonus pelamis to 

 ovulate and spawn within 24 h after capture but eggs are often 

 not viable (Kaya et al., 1982), Maturing marine fish in the lab- 

 oratory and spawning them by hormone injections has become 

 routine in recent years and is preferable to stress techniques. 

 Examples include Engraulis mordax (Leong, 1971), Scomber 

 japonicus (Leong, 1977), Chanos chanos (Liao et al., 1979), 

 Bairdiella icistia (Haydock, 1971), Paralichthys denial us and 

 Pseudopleuronectes americanus (Smigielski, 1975a, b) and oth- 

 ers (see review of Lam, 1982). Induction of spawning in the 

 laboratory may require an open sea water system, large holding 

 tanks (e.g., -3 m dia. or larger), temperature and light control. 



Handling and stocking.— To count eggs without damaging them 

 we recommend a polished wide bore (~3 mm) pipette; count 

 30-50 late stage eggs at a time in a depression slide under a 

 dissection microscope, and wash eggs off the slide by immersion 

 of the entire slide in sea water. Counting eggs is critical because 

 higher mortalities and slower growth result from excess stocking 

 densities (Houde, 1975 and 1977). As a rule stocking densities 

 in rearing tanks of 8 eggs/I or less seems preferable and most 

 rearing successes have occurred when stocking did not exceed 

 20 eggs/1 (Houde, 1975). Similarly, the mortality of Mugil ceph- 

 a/(« larvae seems to remain constant (2-3% loss/day) at stocking 

 densities of 1-30 larvae/1 (Kraul, 1983). 



Apparatus 



Containers and lighting. — Larvae appear to grow faster and show 

 fewer signs of starvation when reared in large containers (100 

 1) rather than in smaller ones (10 1) (Theilacker, 1980b). Opti- 

 mum container size doubtless varies with species but 40 1 con- 

 tainers are probably the minimum size that should be used and 

 I prefer 100-400 1 containers. We use cylindrical black fiberglass 

 containers although excellent results are obtained using ordinary 

 rectangular glass aquaria (Houde, 1975). 



It is traditional to provide a daily cycle of illumination to 



