Effects Of Complex Probiotics In The Seed Breeding Of Litopenaeus Vannamei

With the rapid development of economy,people’s demand for aquatic products has been increasing.The scale of aquaculture and the density of fish farming is expanding,which lead to a threat to the ecological balance of the watery environment.The negative effects of antibiotics and chemicals are obvious.Now the aquaculture industry urgently needs to find a healthy and sustainable development path.Probiotics can not only improve the watery environment,but also inhibit or kill the pathogens in the water.It can also promote the growth and the immunity of aquatic animals.Litopenaeus vannamei,as one of the major aquaculture animals in China,has a high economic value.The result of shrimp culture of L.vannamei is closely related to the quality of its seedlings.During the larval stage of L.vannamei,several metamorphic processes are experienced,with changes in morphology,life habits,food habits and the formation of the immune system.In this study,we screened and separated functional strains from water samples,then used with the strains which our laboratory preserved in cultivation process of Litopenaeus vannamei seedlings.We analyzed the effects of probiotics on the structure of microbial community of shrimp larvae and evaluated application potential of compound probiotics in the cultivation of shrimp prawn.The main contents of the study are as follows:1.Isolation and identification of nitrifying bacteria and acidogenic bacteria in seawaterDifferent probiotics have different functions.With the rapid rising of nitrogen concentration in the rearing water,the self-purification ability of the water is seriously reduced.Moreover,ammonia nitrogen and nitrite have certain toxicity to aquatic animals and affect the health and growth of aquatic animals.The nitrifying bacteria can reduce the ammonia nitrogen in the culture water.Lactobacillus is a microorganism that can ferment carbohydrates to lactic acid.Lactobacillus colonize in the intestine of animals and produce lactic acid,which can reduce the number of pathogenic bacteria.In this study,through the selective bacteria,two different strains were screened from the water samples.One has nitrification,the other can produce the lactic acid.It was identified by the 16s rRNA method and phylogenetic trees were constructed.The results showed that BC109 was Bacillus and BC702 was Enterococcus.The results of blood plates showed that the two strains did not have hemolytic activity when the number of bacteria reached 1×10~6 CFU,which indicated that the two strains could be used as probiotics in aquatic production.2.The screening of compound probiotics formula.In order to control the diseases caused by vibrio,such as Acute hepatopancreas necrosis syndrome(AHPNS),more and more probiotics have been applied to the cultivation of the shrimp.In this study,we screened compound probiotic formulas and methods of addition.Five probiotic strains and a V.parahaemolyticus have been selected.First,we tested the antagonistic effects between the five probiotics,and the results showed that the Pseudomonas CDM8 has antagonistic effects on the other four strains.Probiotics were combined and added in different ways.After the addition of probiotic bacteria,the total bacterium counts(TBC)and vibrio bacterium counts(VBC)in the water were monitored.We found two ways adding Pseudomonas CDM8 and compound probiotics alternately both can maintain the ratio of VBC/TBC from 10%to 16%during the experiment and reduce the vibrio content in water at the same time.This experiment lays the foundation for the application of microecological agents in the future.3.Effects of probiotics combination on breeding of Litopenaeus vannamei under zero-water exchange conditionIn order to explore a breeding technology with the application of probiotics combinations for Litopenaeus vannamei under zero-water exchange conditions,four probiotic bacteria with different ecological functions were formulated to two combinations(i.e.,PBE and PEN)in this study,which were applied in the breeding experiment from nauplius larval phase V(N5)to postlarvae phase V(PL5).Each experimental group contained six parallels and the control group was set at the same time.Culturing water was not exchanged throughout the experiment,and probiotics combinations were continuously applied to the water in the PBE and PBN groups.The larval metamorphosis rate,total culturable bacterial count(TBC)and total presumptive Vibrio bacteria(TPVB)in larva and water were counted at the phases of zoea III(ZIII),mysis III(MIII)and PL5.At the end of the experiment,the survival rate of postlarva in each experimental group was determined,and the microbial communities from postlarva were sequenced using an Illumina platform.The results showed that there was no disease outbreak in each group,and that probiotics combinations significantly decreased(P<0.05)relative abundances of TPVB in postlarva and culturing water in the groups of PBE and PBN compared to that in the control group,as well as significantly increased(P<0.05)the larval metamorphosis rate(~15%)and survival rate(~15%)at the phases of ZIII and PL5,respectively.In addition,there was no striking difference of microbial diversities and structures of the core populations in the microbial communities from postlarva in the three groups,and every microbial community was dominated by Rhodobacteriaceae with the relative abundance ranged from 40%to 86%.Taking together,probiotics combinations can effectively inhibit the overabundance of Vibrio in rearing-water and postlarva and improve the survival rate of postlarva,thereby achieving healthy breeding of L.vannamei under zero-water exchange conditions.