| Vibrio disease is considered to be one of difficulty for shrimp farming,which has greatly reduced the production of shrimp.The loss accounts for about 25%of the total aquaculture loss in the world.In addition,the food safety problems caused by the abuse of antibiotics have become increasingly prominent and the promotion of the national anti-culture policy has made the development of biological feed additives a hot topic.Probiotics have been widely developed as feed bio-additives because of their ability to change the balance of the internal bacterial flora,inhibit the growth of pathogenic bacteria,promote good digestion,reduce the effects of allergens and enhance immune function.Studies have shown that compound addition can usually achieve the effect of supplementing or enhancing the effect,and at the same time,the internal probiotics have a more gentle effect on the host shrimp body and better adaptability than the external probiotics.In this study,26 of lactic acid bacteria and 12 of Bacillus were isolated from the intestines of Litopenaeus vannamei and Macrobrachium rosenbergii.Among them,antibacterial test,adhesion test of lactic acid strains and related hydrolase enzyme activity of Bacillus were measured.The results show that 6-24 shows high antibacterial activity and adhesion at the same time,B9shows good enzyme production capacity,which protease activity is better.The identity of the two strains were determined by 16S r DNA,physiological and biochemical identification.The identification results showed that these two strains were Enterococcus durans 6-24 and Bacillus subtilis B9.6-24 and B9 were made into powder by freeze-drying.Both strains were added to the blood cells,prophenoloxidase system（pro PO）,antioxidant system and lysozyme were determined.The results of the study showed that the total haemocyte count in hemolymph was significantly increased（p<0.05）,the activities of phenol oxidas,superoxide dismutase and lysozyme also increased（p<0.05）after feeding 6-24 and B9,which the number of total blood cells increased by 96.54%,phenol oxidase increased by 48.24%,superoxide dismutase increased by 4.42%,lysozyme increased by 198.00%.The experiment on the influence of digestive enzymes in the intestinal tract of M.rosenbergii showed that the activity of digestive enzymes（amylase,protease,lipase）in shrimp intestinal group with mixed probiotics was significantly increased compared with the control group（p<0.05）.Among them,amylase increased by 48.63%and lipase increased by 44.9%,the activity of protease increased by84.80%which better than other digestive.The study also found that the number of viable bacteria in the intestine of M.rosenbergii decreased by 59.76%（p<0.05）after being fed E.durans 6-24 and B.subtilis B9.Finally,the results of the measurement of the growth status of M.rosenbergii showed that 6-24 and B9 could improve the growth status of M.rosenbergii but the effect was not significantly（p>0.05）.The results of the study showed that E.durans6-24 and B.subtilis B9 can improve the intestinal environment and immune level of M.rosenbergii to a certain extent,and has the potential to develop probiotic bacteria for shrimp feeding.Lysozyme is currently widely distributed in animals,plants and microorganisms,and is an important substance in the innate immune system of organisms.At the same time,lysozyme has the characteristics of non-toxicity and high heat resistance.Therefore,it is widely used as a feed biological additive with broad application prospects in the breeding industry to improve the growth performance and immune status of farmed animals.In addition,most studies have shown that there are gender differences in the immune parameters of many invertebrates.The immune parameters of female individuals will be higher than those of male individuals.This study explored the effect of lysozyme preparations on the correlation coefficient of M.rosenbergii of different genders,adding 270,000 U·kg-1lysozyme to the feed and conducting a 4-week feeding experiment on M.rosenbergii.The growth performance results showed that the weight gain rate of the female group increased by 66.24%（p<0.05）,the specific growth rate increased by 50.03%（p<0.05）,and the feed conversion rate decreased by 40.13%（p<0.05）,but the male population has improved growth compared to the control group but the effect is not significantly（p>0.05）.The study also found that after feeding egg white lysozyme,the immune parameters of both female and male populations were significantly improved（p<0.05）.Among them,the number of total blood cells in the female group increased by 82.57%,the activity of phenol oxidase increased by 90.75%,and the activity of lysozyme increased by 216.77%;the number of total blood cells in the male group increased by 76.92%,the activity of phenol oxidase increased by 90.56%,and the activity of lysozyme increased.126.33%,TF increased by CF.Compared with the control group,the PO activity of TM and TF increased by 90.56%and 90.75%respectively.Interestingly,we found that after feeding egg white lysozyme,the SOD activity level of male population tended to Consistent with the level of SOD activity in female populations,it is speculated that lysozyme can promote SOD expression in male individuals.Finally,the experiment on the effect of digestive enzymes in the intestinal tract of M.rosenbergii showed that egg white lysozyme could significantly improve the activity of lipase in the intestinal tract of M.rosenbergii（p<0.05）,and the female and male groups increased by 111.26%and600.98%respectively.The results show that egg white lysozyme can produce an immune stimulating response to M.rosenbergii in vivo,thereby improving the immune level,growth status and intestinal environment. |
