Studies On Dietary Protein Requirement And Optimal Protein To Energy Ratio Of Lateolabrax Maculatus Under Different Water Temperatures

This study investigated the protein requirement and optimal protein to energy ratios（P/E）of spotted seabass（Lateolabrax maculatus）under different water temperatures（27℃,33℃）.Moreover,we assessed the hepatic and spleen transcriptome of spotted seabass under different water temperatures to explore the differences in nutrient metabolism and immune response of the spotted seabass.The main research results are as follows:（1）Studies on the dietary protein requirement of spotted seabass under different temperaturesSix diets were formulated containing 35,38,41,44,47 or 50%crude protein（P35,P38,P44,P47,P50）,and each diet was used to feed fish（initial weight:2.00±0.01 g）at a low temperature（27℃;LT）and high temperature（33℃;HT）.So there are 12 treatment groups（6protein levels×2 temperature levels）.The quadruplicate groups of fish were fed the test diets for8 weeks.During the feeding period,fish were fed three times daily to satiation.In the end of experiment,samples are collected for analysis.The results showed that growth and feed utilization were significantly（P<0.05）influenced by both dietary protein level and water temperatures but no significant interaction effect（P>0.05）.The highest weight gain was recorded in the P47LT group which was significantly higher than those of P35LT,P38LT,P41LT and all HT groups.Moreover,feed conversion ratio decreased by increasing the dietary protein level up to 47%at both water temperatures.Intestinal protease activity was increased by increasing the dietary protein level at both water temperatures,and HT groups exhibited higher activity than their respective LT groups（P<0.05）.Hepatic citrate synthase and succinate dehydrogenase activities were significantly（P<0.05）influenced by protein level and water temperature and their highest value was detected in the P47LT group.Higher water temperature led to a significant enhancement of serum triglyceride concentration（P<0.05）.Also,we assessed the hepatic transcriptome and compared the genes associated with metabolism of fish reared at LT or HT.The KEGG enrichment analysis showed that most of the DEGs were involved in the metabolism processes of amino acids,fatty acids and glucose and some signaling pathways.The transcriptomic results indicated that high temperature could induce changes in three major nutrient metabolism and heat stress in spotted seabass.Based on the results of this experiment,the protein requirements of spotted seabass reared at 27℃ and 33℃ are 46.8%and 44.9%,respectively.The minimum dietary protein requirements of spotted seabass are different when reared at two water temperatures and this may be attributed to alterations of liver metabolism.（2）Studies on the dietary optimal protein to energy ratios（P/E）of spotted seabass under different water temperaturesSix diets were formulated with different P/E:22.31,23.27,23.47,24.05,24.35,25.55 g MJ^-1,using two dietary protein levels（44%and 47%）and three dietary lipid levels（8%,11%and 14%）.Each diet was used to feed fish（initial weight:2.67±0.01 g）at a low temperature（27℃;LT）and high temperature（33℃;HT）.So there are 12 treatment groups（6 protein to energy ratios×2 temperature levels）.The triple groups of fish were fed the test diets for 8 weeks.During the feeding period,fish were fed three times daily to satiation.In the end of experiment,samples are collected for analysis.The results showed that growth was significantly（P<0.05）influenced by P/E and water temperatures but no significant interaction effect（P>0.05）.Feed conversion ratio（FCR）significantly（P<0.05）influenced by P/E,and fish fed 22.31P/E of LT exhibited the lowest FCR.At this time,the protein level was 44%and the fat level was 14%.The activities of intestinal protease,amylase and lipase was neither affected by P/E or water temperature（P>0.05）.Hepatic citrate synthase and succinate dehydrogenase activities were significantly（P<0.05）influenced by water temperature and the higher value was detected in LT group.The analysis based on weight gain showed that the optimal P/E ratio of spotted seabass under different water temperatures are 24.35 g MJ^-11 and 23.47 g MJ^-1,respectively（3）Effect of water temperature and P/E on non-specific immunity of spotted seabassThe design of this experiment is same as Experiment 2.To explore effect of water temperature and P/E on the immunity,the non-specific immune activity of spotted seabass with different protein to energy ratios under different water temperatures were studied.Moreover,we analyzed the non-specific immune activity and spleen transcriptome at high and low temperatures（27℃,33℃）.The results showed that protein to energy ratios had no significant effect on the non-specific immunity of fish（P>0.05）,but significantly affected by water temperature（P<0.05）.High temperatures would reduce the activity or content of lysozyme,complement 3,complement 4 and immunoglobulin M.To clarify the molecular mechanism,spleen transcriptome of spotted seabass under different temperatures was analyzed.The KEGG enrichment analysis showed that most of differentially expressed genes were involved in immune network for IgT production and phagosome pathways,and the high temperature group related genes are down-regulated.The results show that water temperature can inhibits the non-specific immunity of spotted seabass,and analysis of spleen transcriptomics results suggest that immunosuppression may be due to genes down regulation in the"IgT production in the intestinal immune network" and "phagosome" pathways.