Research On Safety And Probiotics Properties Of 5 Isolate Of Lactic Acid Bacteria

Lactic acid bacteria(LAB)are extremely widely distributed in nature and are closely related to human life.They are widely used in agriculture,animal husbandry,food and medicine.Studies have shown that LAB are multi-drug resistant and carry drug-resistant genes.Certain strains can also produce harmful metabolites,which poses certain safety risks.In recent years,researches have been carried out on the regulation of intestinal flora by probiotics.However,the resources of probiotics are still scarce in China.Therefore,research on the safety and probiotic properties of LAB is of great significance to improve the quality and safety of LAB products and develop probiotic resources.In this experiment,five strains of lactic acid bacteria with well-tolerated screened out in the early stage of the laboratory were used as the research objects,using the method of metabolite toxicity test,drug resistance test and animal toxicity test to study its safety.Then,through in vitro probiotic evaluation and dynamic monitoring of mice intestinal flora to detect its probiotic properties,with a view to discovering safe probiotic strains.The main findings are as follows:1.The results of the metabolite toxicity test showed that LAB didn't produce nitrate reductase activity and hemolytic activity.LAB don't carry hemolytic genes cylA,cylB,cylM,virulence genes asal,ace,esp,gelE,tyrosine decarboxylase genes tdc1,tdc2,tdc3 and histidine decarboxylase genes hdcl,hdc2,hdc3.LAB have detected tyrosine decarboxylase activity,and tyramine has been detected in fermented raspberry juice,but the content is very low.2.The results of the drug resistance test showed that LAB have relatively similar drug resistance spectrum.They are all sensitive to cefazolin,cefalotin,ampicillin,trimethoprim and nitrofurantoin.The sensitivity to different ?-lactam antibiotics is different.All 5 strains contained plasmids.Among the 12 resistance genes,four resistance genes(erm,aph,van ?,and aac I)were detected on the plasmids.Six drug resistance genes(aph,erm,van ?,bla ?,aac ?,and aac ?)were detected on genomic DNA.Part of the resistance genes will be expressed under the culture of MRS and MRS with antibiotics,and the resistance genes that are not expressed will not be expressed under the conditions induced by the corresponding antibiotics.3.The results of animal toxicity test showed that the mice were in good behavior and mental condition,and the calculated LD50>10 g/kg.Gavage of different doses of tested strains has a certain promotion effect on the growth of mice.Among them,high and medium doses of Lactobacillus bulgaricus can significantly promote the growth of mice(P<0.05).Compared with the control group,although the blood routine indicators of the experimental groups increased or decreased,there was no significant difference(P>0.05).5 strains have no effect on the serum biochemical indicators of glutamate aminotransferase(ALT)and total bilirubin(T-BIL),direct bilirubin(D-BIL),creatinine(CREA),urea nitrogen(BUN),uric acid(UA),triglycerides(TG),total cholesterol(TC),and blood sugar(GLU)(P>0.05),and no bacterial translocation occurred,indicating that tested strains had no adverse effects on the organs of mice.In general,the tested strain is safe for the organism.4.The results of in vitro probiotic performance evaluation showed that in addition to Lactobacillus rhamnosus,4 strains have the most obvious inhibitory effects on Salmonella,Shigella flexneri,and Escherichia coli.Strains of Lp-2,Lb,and Lc have high hydrophobicity and self-aggregation.Changes of intestinal micro flora and its function is an important index for evaluating the safety of strains and screening strains with probiotic properties.It turned out that,L.bulgaricus increased the number of lactic acid bacteria and Bifidobacterium in the intestinal flora to a certain extent,and decreased the number of Enterobacteria,thus exert the probiotic effect of regulating the intestinal flora.The effective dose of L.bulgaricus for regulating the intestinal flora of mice is 109 CFU/mL,and the adjustment hold about five days after stopping the ingestion.