Aquaculture Europe 2014

October 14-17, 2014

Donostia–San Sebastián, Spain


I.B. Falk-Petersen1, S. Peruzzi1 and V. Puvanendran2
1Faculty of Biosciences, Fisheries and Economics, University of Tromsø, Breivika, N-9037 Tromsø, Norway, 2Nofima AS, N-9291 Tromsø, Norway

Fish egg and larval development, growth  and mortality are generally highly influenced by temperature  (Blaxter 1991; Geffen et al. 2006; Puvanendran et al. 2013) as well as feed quality (Busch et al. 2010). The national cod-developmental project CODE (RCN-199482) was established in order to explain the mechanisms behind early life history success or failure. The present investigation aims at identifying possible developmental differences between larvae raised under different temperature regimes.  
Materials and methods
Cod broodstock
Sperm from 8 males and eggs from 18 females from cultured Atlantic cod  (2007 and 2008 year class at the National Cod breeding Centre (NCBC) in Tromsø, North Norway), were hand-stripped, gametes were pooled and fertilization carried out according to Hansen & Puvanendran (2010).  
Experimental setup
Fertilized eggs were transferred to 16 upwelling incubators (375 ml eggs in each) and 8 of the incubators were kept at 4±0.2 °C until hatch while 8 were kept at 9.5±0.2 °C (T increased in 32 hours, ~0.5 °C every 3 hours ). At 100% hatch, larvae were transferred to 190 L circular experimental tanks (stocking density 26 000 larvae). Larvae from eggs incubated at 4 °C were transferred to 8 tanks at 4 °C while those from 9.5 °C were transferred to 8 tanks at 9.5 °C. The temperature of 4 tanks at 4 °C was gradually increased to 9.5 °C (LH) while the other 4 tanks were kept at 4 °C (LL). The temperature of 4 tanks at 9.5 °C was gradually decreased to 4 °C (HL) while the other 4 tanks were kept at 9.5 °C (HH). Thus, 4 different test groups were established in order to study temperature effects on development and growth during the larval and early juvenile stages.
Larval rearing
Microalgae (Nannochloropsis) were supplied during the first 10 days and the larvae were fed with Phosphonorse and Micronorse enriched rotifers (4000 individuals L-1every four hours until 29 dph). During the last 4 days of rotifer feeding, Artemia was added and fed to the larvae until 46 dph. During the weaning period after 32 days, dry feed (Algonorse coldwater) was used (pellet size 200-300 µm initially and gradually increased to 300-500 µm at 64 dph).
Larval sampling
Larval samplings were conducted at 5 predetermined stages (based on von Herbing et al. 1996) from all 4 treatments (100 % hatch, stage 4 (after first-feeding), stage 8 (start metamorphosis), stage 11(mid-metamorphosis) and stage 12 (end metamorphosis).
Ten larvae were sampled randomly from each replicate, anaesthetized with MS-222 and transferred to small glass tubes with 4 % phosphate-buffered formaldehyde and kept in the fridge before being embedded in Technovit and/or Paraplast for comparative histological studies of organ and tissue development.   
Sections of hatched larvae (100 % hatch) from the high and low egg incubation temperature regimes did not reveal obvious differences with regard to organ or tissue histomorphology and larvae sectioned at stage 4 also appeared similar. Thyroid-follicles were prominent in all larvae at stage 4, the stomach was apparent in stage 11 larvae and hepatocytes became vacuolized at this stage.The HL-group suffered high mortality after stage 8 and the LL group after stage 11, but the larvae studied until then had apparently normal histomorphology. At stages 8 and 11 the swimbladder was generally more expanded in the LH-group, but similar in LH- and HH-group at stage 12. The digestive tract was more differentiated in many individuals from the LL and LH-regime at stages 8 and 11. At stage 12 the digestive tract of the HH-group was more differentiated than that of the LH-group.
Discussion and conclusion
Higher temperatures appeared to support growth and survival of cod larvae in this experiment. Sectioned larvae  showed similar histomorphology at the selected developmental stages. Even if the HL-regime was detrimental to the larvae and mortality  also surprisingly high in the LL-regime, no significant abnormalities were detected in organs and tissues compared to the LH and HH groups. The overall histomorphology reflected individual growth variations in and between groups and was most advanced in the HH-larvae at stage 12 (end metamorphosis).   
Blaxter, J.H.S., 1991. The effects of temperatur eon larval fishes. Netherlands journal of                    Zoology, 42:336-357.      
Busch, K.E.T., Peruzzi, S., Tonning, F., Falk-Petersen, I.B., 2011. Effect of prey type and size on the growth, survival and pigmentation of cod (Gadus morhua  L.) larvae fed rotifers (Brachionus plicatilis) or natural zooplankton.. Aquaculture Nutrition, 17:595-603 .
Geffen, A.J., Fox, C.J., Nash, R.D.M. 2006. Temperature-dependent developmental rates
of cod Gadus morhua eggs. Journal of Fish Biology, 69:1060-1080.
Hansen, Ø.J. & Puvanendran, V., 2010. Fertilization rate and blastomere morphology as predictors of egg and juvenile quality for domesticated Atlantic cod, Gadus morhua, broodstock. Aquaculture research, 41:213-216.  
Hunt von Herbing, I.,R.G. Boutilier, T. Miyake and B.K. Hall. Effects of temperature on morphological landmarks critical to growth and survival in larval Atlantic cod (Gadus morhua). Marine Biology, 124:593-606
Puvanendran, V., Falk-Petersen, I.B., Lysne, H., Tveiten, H., Toften, H., Peruzzi, S., 2013. Effects of different temperature increment regimes during egg incubation of Atlantic cod on egg viability and larval quality. Aquaculture Research, 1-10.