Screening Of Psychrophile Candidate Probiotic Strain From The Rainbow Trout In Vitro And Evaluation Of Antiviral

The study is based on natural waters Benxi in Liaoning Province backyard get healthy rainbow trout as experimental animals, immediately under laboratory sterile conditions forced to kill, obtain the gut and other parts of the low temperature separation strains; Rainbow trout psychrotrophic strains were isolated temperature gradient screening; by the method of molecular biology psychrotrophic strains were identified, remove pathogens and opportunistic pathogens; Rainbow trout psychrophilic probiotic strain probiotic properties in vitro screening to determine psychrotrophic strains of probiotic strains of the candidate; addicted to rainbow trout psychrotrophic probiotic strains preliminary screening for antiviral properties; using TNF-α activitated HT29-cells and PBMCs System isolates the second tier of candidates for antiviral screening; through the multiple screened 6 Rainbow trout psychrophilic probiotic strains and 1 ‰ infectious hematopoietic necrosis virus(IHNV) of the EPC cells were co-cultured in the culture supernatant was measured viral titers to determine the candidate strains in vitro antiviral potency; for 6 Rainbow trout psychrophilic probiotic strains physiological and biochemical tests to determine the optimal medium The optimum temperature and optimum growth pH; candidate strains screened out environmental stress tolerance test, high bile acid test, resistance to low p H tests; explore strains of microencapsulation methods and optimization; to develop feeding program Animal tests to evaluate the safety of candidate strains; continuous feeding 7 days after the attack eight times the LD50 were IHNV(Sn1203 strain) of the 6 candidates for in vivo anti-virus capabilities to verify and evaluate the effect of feeding a candidate strain immunity, Determination of serum ACH50, SOD, LYS, Ig M and IL-2 content in the testing process; serum antibody titers.In this study, low temperature separation from Rainbow trout and other parts of the intestine get 83 Rainbow isolates; by strict psychrotrophic temperature gradient screen, 18 strains has been screening. By psychrotrophic strain probiotic properties in vitro screening, the triple antiviral properties in vitro screening method to obtain the 6 Rainbow trout having higher antiviral potency in vitro psychrotrophic strains of probiotics; the use of microencapsulation methods coated 6 Rainbow trout body psychrophilic candidate strain probiotic strains continuous feeding of animal tests to verify its good probiotic characteristics and safety; then dorsal injection attack drug, verify multiple vitro antiviral effective screening method, candidate strains have favorable in vivo anti-viral properties, 32#(Lactococcus lactis subsp.HL1) strain, 10#(Arthrobacter sp.HL4) strain, 46#(Psychrobacter glacincola HL6) and 58#(Bacillus sp.HL5) strain has favorable probiotic properties and anti-viral properties, can stimulate the body immune response, for IHNV infection have a protective effect, The relative percents of survival(RPS), respectively, 22.5% and 22.5%, 15% and 7.5%. 32# Lactococcus lactis subsp.HL1 in 0.5% of strains capable of bile acid and high-acid p H4.0 normal growth medium, the candidate strain safety and protective efficacy evaluation have shown immunity excellent probiotic properties and anti-viral properties, the serum total complement(ACH50), superoxide dismutase(SOD), lysozyme(LYS), IgM and IL-2 content with the control group was significantly(P <0.05), and higher than other test strains. 32# Lactococcus lactis subsp. HL1 has highest anti-virus capabilities, through mucosal immune function, can effectively stimulate the body to produce an immune response can be further used as an oral live vector vaccine candidate strains.