Optimized Preparation Of Nano-selenium Lactic Acid Bacteria And Its Application As Immunopotentiator Of Aluminum Adjuvant Vaccine

Nano-selenium lactic acid bacteria have been proved to have antioxidant,anti-aging and anti-cancer function.However,there is no detailed report on whether nano-selenium lactic acid bacteria can improve the immune effect of vaccines.Therefore,this study optimized the preparation of nano-selenium lactic acid bacteria,explored the potential of different forms of nano-selenium lactic acid bacteria as vaccine immunopotentiator and further analyzed the related mechanisms,so as to provide theoretical basis for clinical use of nano-selenium lactic acid bacteria as immunopotentiator.In order to solve the problem of low efficiency of probiotic lactic acid bacteria transforming inorganic selenium into nano-selenium,this study optimized the preparation conditions of nano-selenium lactic acid bacteria,detected its activity in vitro.In order to find a better application scenario for the optimized preparation of nano-selenium lactic acid bacteria,the immune enhancement effect on four representative vaccines was preliminarily explored.Then according to the test results and site reasons,a mouse model was established for experiment,test group mice oral lactic acid bacteria,nano-selenium lactic acid bacteria,primary purified nano-selenium,and thermally inactivated nano-selenium lactic acid bacteria before vaccination.The changes of antibody levels,attack protection,intestinal pathological section observation,oxidative damage,inflammation and expression of selenoprotein-related factors were measured to determine the effect of aluminum adjuvant inactivated vaccine.In order to more clearly show its immune enhancement effect,we set up a mouse model experiment of nano selenium lactic acid bacteria on the immune effect of 1/2 dose and 1/4 dose vaccine.Finally,the effect of improving vaccine immunity in clinic was further verified in rabbits.The main results are as follows:(1)When 30 μg/mL sodium selenite was added to lactic acid bacteria 23017 from the beginning of the logarithmic phase,the conversion rate of sodium selenite was the highest,reaching 78.4 %,and1 L preparation can realize.Optimized preparation of nano-selenium lactic acid bacteria have stronger hydroxyl radical scavenging ability,DPPH radical scavenging ability,ABTS·+ radical scavenging ability and superoxide anion radical scavenging ability than lactic acid bacteria.And after being stored at 4 ℃ for 7 months,nano-selenium lactic acid bacteria could still reach the standard of viable count of microbial preparations.(2)Nano-selenium lactic acid bacteria can improve the antibody titer of newcastle disease vaccine,Erysipelothrix rhusiopathiae vaccine,porcine circovirus type 2 vaccine and Clostridium perfringens vaccine,and improve the challenge protective effect of inactivated vaccine is better.(3)According to the experimental results and comprehensive consideration,the aluminum adjuvant inactivated vaccine represented by Clostridium perfringens Type A inactivated vaccine was selected.The model of lactic acid bacteria,nano-selenium lactic acid bacteria,heat-inactivated nano-selenium lactic acid bacteria and crude nano-selenium to enhance the immune effect of aluminum adjuvant inactivated vaccine in mice was established.The main results were as follows:Nano-selenium lactic acid bacteria,heat-inactivated nano-selenium lactic acid bacteria and crude nano-selenium can improve antibody level after vaccination with aluminum adjuvant inactivated vaccine,and the effect is better than lactic acid bacteria;Adding lactic acid bacteria,nano-selenium bacteria,heat-inactivated nano-selenium lactic acid bacteria and crude nano-selenium can significantly alleviate the clinical symptoms of immunized mice after challenge;Body weight of immunized mice was significantly reduced by challenge.Lactic acid bacteria,nano-selenium bacteria,heat-inactivated nano-selenium lactic acid bacteria and crude nano-selenium could effectively slow down the weight loss;Pathological sections showed that lactic acid bacteria,nano-selenium bacteria,heat-inactivated nano-selenium lactic acid bacteria and crude nano-selenium can effectively alleviate the intestinal injury caused by challenge in immune mice.(4)Nano-selenium lactic acid bacteria can up-regulate the mRNA expression levels of selenoprotein related functional genes(GPx1,GPx2,GPx3,GPx4,SelH,SelI,SelW,SelT,SelO,Sephs2,Sepp1,Sep15,DIO1);Lactic acid bacteria,nano-selenium lactic acid bacteria,heat-inactivated nano-selenium lactic acid bacteria and crude nano-selenium is more conducive to the recovery of oxidation-antioxidant system balance in mice after immunization and challenge;Lactic acid bacteria,nano-selenium lactic acid bacteria,heat-inactivated nano-selenium lactic acid bacteria and crude nano-selenium can regulate the imbalance of inflammation-related factors after vaccine immunization and challenge.(5)Heat-inactivated nano-selenium lactic acid bacteria can increase the serum specific IgG antibody level of mice vaccination with 1/4 doses,1/2 doses of vaccine and reduce the mortality of1/2 dose vaccine immunized mice after challenge.(6)The results of rabbit test showed that nano-selenium lactic acid bacteria could improve the serum specific IgG antibody level of immunized rabbits and the neutralizing antibody level of immunized rabbits.Significantly decreasing the DAI score can effectively reduce the intestinal injury caused by immunized rabbits attacking the alpha-toxin of clostridium perfringens Type A.The above results showed that the inorganic selenium conversion rate of lactic acid bacteria was further improved by optimizing the preparation of nano-selenium lactic acid bacteria.Nano-selenium lactobacillus can significantly improve the immune effect of aluminum adjuvant inactivated vaccine in mice,which may be related to the regulation of oxidative stress level and the influence on the expression level of selenium protein-related functional gene.At the same time,nano-selenium lactic acid bacteria can significantly improve the effect of aluminum adjuvant inactivated vaccine in rabbits,and provide new ideas for better application of nano-selenium lactic acid bacteria.