| Lactic acid bacteria(LAB)strains can enhance the growth performance,immune response and disease resistance for aquatic animals and have become one of the alternative substance for antibiotics.Dross vinegar is a unique cuisine in the chino town of Hainan province,which has a long history of eating and contains large amounts of lactic acid bacteria according to the previous research.In this study,a variety of Dross vinegar samples were collected from Hainan chino town and the potential probiotic lactic acid bacteria strains were screened from them.Their haemolytic activity,antagonistic activity,auto-aggregation and hydrophobic activity were also tested.The screened lactic acid bacteria were mixed in the artificial feeds and were fed to the brown-marbled grouper(Epinephelus fuscoguttatus)and white shrimp(Litopenaeus vannamei),and its effect on growth performance,innate immune parameters were determined.The main results of this study were showed as follows.(1)In this study,125 strains of acid-producing bacteria were screened from 11 kinds of Dross vinegar using MRS + CaCO3 solid medium.Twenty-three lactic acid bacteria strains were screened by exclusion of acid and hydrogen peroxide experiments and protease hydrolysis experiments.Then the investigated probiotic properties include Antagonistic effect of aquatic animal pathogens,Haemolytic activity,Adhesion capacity(auto-aggregation,cell surface hydrophobicity)were evaluated.The effects of 16S rDNA to the 23 strains of lactic acid bacteria were identified.Based on the above studies,two lactic acid bacteria were screened,and the strains were named LAB041 and LAB 1036,respectivey.The results of the study on the characteristics of LAB041 showed that LAB041 is.Weissella confusa.It exhibited type a-hemolytic activity and had strong adhesion capability.Its cell surface hydrophobicity in Toluene,Ethylacetate,Chloroform were 23.4%,58.58%and 66.73%,respectively.It also exhibited a significant auto-aggregation property(12.03%-27.50%)after 3h incubation,but its antagonistic effect of aquatic animal pathogens was relatively weak.Their inhibition zones against Vibro.harveyi,V.anguillarum,V.Parahaemolyticus and V.alginolyticus were 8.33mm,8.49mm,7.74mm,9.52mm,respectively.LAB 1036 is Lactobacillus plantarum strain,which exhibited typical y-hemolytic activity.Its antimicrobial activity was relatively strong and their inhibition zones against Vibro.harveyi,V.Parahaemolyticus and V.alginolyticus were 12.38mm,12.77mm,11.58mm,respectively.Its adhesion capacity was soso.Its cell surface hydrophobicity in Toluene,Ethylacetate and Chloroform were 47.40%,18.57%and 60.31%,respectively.Its auto-aggregation property were 18.10%-30.95%after 3h incubation.(2)By adding Lactobacillus plantarum strain1036 and Weissella confusa strain041 in the brown-marbled grouper feed made by myself,the effect of the dietary administration of two Lactobacillus strains on the growth performance and innate immune of the brown-marbled grouper were evaluated.Experient divided into four groups:SBW group,SBP group,SBC group,each group has three replicates,each replicates has thirty brown-marbled grouper.The SBW group were fed with LAB 041 diet 1×108CFU/g),the SBP group were fed with LAB1036 diet(1×108CFU/g),the SBC group were fed with control diet.The growth performance and innate immune parameters were examined at 1,2,4 and 8 weeks of post-feeding.Results showed that the brown-marbled grouper fed a diet containing LAB041 demonstrated increased percent weight(PWG),specific growth rate(SGR)compared to the control group,but there was no significant difference(P>0.05).The brown-marbled grouper fed with lactic acid bacteria,the number of non-specific immune indexes have been enhanced.There were significant differences in the non-specific immune indexes between the SBP group and the SBW group,and the non-specific immune indexes were significantly different from those of the control group.For example,The respiratory burst activity of macrophages in SBW group was significantly lower than that in control group at 4 weeks(P<0.05);The activity of catalase in SBP group and SBW group was significantly lower than that in control group at 4 week(P<0.05);The serum alkaline phosphatase activity of SBW group was significantly higher than that in control group(P<0.05),SBP group was opposite,and there was significant difference between SBW group and SBP group(P<0.05)at the second week;The total complement content of SBP group was significantly higher than that of the control group in the second week,while the SBW group was significantly lower than the control group,and SBP group and SBW group were significantly different(P<0.05).However,there was no significant difference in the serum superoxide dismutase,liver-body ratio and spleen-body ratio between the experimental group and the control group(P>0.05).The challenge test which is separated from 1,2,4,8 week showed that supplementation of both strains can enhance the protection against Vibrio harveyi 385 infection,and with the feeding time prolonged,the protection rate gradually increased.Fish fed with 8 weeks,the relative percent survival(RPS%)of SBP group was 36.4%,while the relative percent survivalof SBW group was 27.26%.(3)The effects of two lactic acid bacteria strains on the growth,immunity and stress resistance of Litopenaeus vannamei were studied by adding Lactobacillus plantarum strain1036 and Weissella confusa strain041 in Litopenaeus vannamei feed.Experient divided into three groups:DXW group,DXP group,DXC group,each group has two replicates,each replicates has four thousand Litopenaeus varmamei.The DXWgroup were fed with LAB 041 diet(1×108CFU/g),the DXP group were fed with LAB1036 diet(1×108CFU/g),the DXC group were fed with control diet.The growth performance and innate immune parameters were examined after 8 weeks of post-feeding.The results showed that the survival rate of shrimp in the lactic acid bacteria group was higher than that in the control group after eight weeks,the survival rate of shrimp in the DXW group was 73.63%,the DXP group was 91.25%;Body length,final wet weight,weight gain rate(WGR),specific growth rate(SGR)were significantly higher than those in the control group(P<0.05).The DXP group was significantly higher than the DXW group(P<0.05);The feed conversion rate of lactic acid bacteria group was significantly lower than that of control group(P<0.05).The shrimp fed with lactic acid bacteria,the number of non-specific immune indexes have been enhanced,such as serum total antioxidant capacity of lactic acid bacteria group(T-AOC),serum alkaline phosphatase acitivty(AKP),acid phosphatase acitivty(ACP),polyphenol oxidase acitivty(PLO)were higher than the control group,but only the serum total antioxidant capacity(T-AOC)acid phosphatase(ACP)activity of DXW group and increased significantly compared to the control group(P<0.05).In conclusion,Lactobacillus plantarum strain1036 and Weissella confusa strain 041 could inhibit the antibacterial activity of Aquatic Pathogenic Bacteria in vitro,producebacteriocin,and has a certain adhesion.The oral administration of two Lactobacillus could improve immunity and enhance the disease resistance of the brown-marbled grouper.In the same way,two strains of lactobacillus were added to the shrimp feed could also enhance the growth performance and improve immunity. |
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