Screening Of Lipid-Lowering Probiotics And Their Effects On Growth,Immunity And Anti Stress Of Epinephelus Lanceolatus♂×Epinephelus Fuscoguttatus♀

Fat is an essential nutrient for the growth of fish,it is also an important source of energy and essential fatty acids.While appropriate amounts of fatty acids and energy is essential for the growth of fish,excessive fat in the feed will lead to over accumulation of fat in the liver of hybrid grouper,causing fatty liver disease.The resulting metabolic disorder and decline of anti-stress ability and immunity often lead to repetitive disease break outs and huge economic losses.In order to adjust the metabolism and absorption of juvenile hybrid grouper lipids,reduce the accumulation of fat in the liver and abdomen,this study aimed at screening lipid-lowering probiotics from the fish intestine bacteria.Firstly,multiple probiotic characteristics,including lipid-lowering capacity,pathogen antagonism activity,gastrointestinal environmental tolerance,safety,etc.,were evaluated in vitro,and the obtained potential lipid-lowing probiotics were identified.Then the in vivo effects of these potential lipid-lowering probiotics on the growth,non-specific immunity,lipid metabolism and anti-ammonia nitrogen stress ability of juvenile hybrid grouper were studied by feeding trials.The results were as follows:1.Isolation,screening and identification of in vitro lipid-lowering probioticsIn this study,52 lactic acid bacteria isolates were obtained from the intestines of Siganus fuscescens and Golden pompano using MRS agar medium.By measuring the hemolysis,lipid-lowering ability,pathogen antagonistic ability,artificial gastrointestinal fluid tolerance and adhesion ability of all isolates,3 potential lipid-lowing probiotics were screened.Then the lipid-lowing capacity of these isolates and some laboratory stored probiotics were reexamined,and finally 3 isolates(LZ8,58 and F2)with strong in vitro lipid-lowing capacity were obtained.16 S r RNA sequencing identified LZ8,58 and F2,as Lactoccus Lactis,Lactobacillus plantarum and Enterococcus faecalis,respectively.2.In vivo lipid-lowering effect of lipid-lowering probiotics in juvenile hybrid grouperLactoccus Lactis LZ8,Enterococcus faalice F2 and Lactobacillus plantarum 58 were added to basal diet and high-fat diet respectively.The growth performance and lipid-lowering effect of different probiotics in juvenile hybrid grouper were studied through 8-week feeding experiment.The results showed that L.plantarum 58 could significantly reduce the body fatness,viscera ratio,serum T-CHO,TG,and LDL-C content(P< 0.05)of high-fat diet hybrid grouper.The liver H.E staining and oil red O staining lipid droplet area results indicated that L.plantarum 58 can alleviate liver steatosis and prevent the occurrence of fatty liver disease.Strains L.lactis LZ8 and E.faecalis F2 also showed certain lipid-lowering effects,but L.plantarum 58 displayed the best serum lipid-lowing effect in juvenile hybrid grouper.3.Effect of lipid-lowering probiotics on the ammonia nitrogen stress tolerance of juvenile hybrid grouper.The eight groups of juvenile hybrid grouper were challenged with ammonia nitrogen stress for 24 h before samples were taken to measure the liver and intestine sections,serum non-specific immunity,antioxidant and stress indicators of juvenile hybrid grouper.The results showed that there was no significant differences in non-specific immunity indexes(LZM,GOT,GPT and ALB)before and after ammonia nitrogen stress in the HF group.The LZM activity of H8 group before ammonia nitrogen stress was not significantly different from that after ammonia nitrogen stress(P> 0.05).In C8 group and CF group,the antioxidant indexes(GSH-Px,SOD and T-AOC)were significantly increased,and MDA was significantly decreased(P<0.05).The content of lactic acid(stress index)of C8 group,CF group,H8 group and HF group increased significantly(P< 0.05).The above results indicated that L.Lactis LZ8 and E.faecalis F2 exhibited antioxidant,anti-stress and immunity-boosting effects on juvenile hybrid grouper.The mortality rate after 96 h ammonia nitrogen stress was lowest in F2 supplemented group,therefore,E.faecalis F2 had the best protective effect against ammonia nitrogen stress in juvenile hybrid grouper.