Optimization Of Fermentation Conditions For Lactobacillus Reuteri GL005 Strain And Evaluation Of Its Antidiarrheal Effect

Lactobacillus reuteri,as a food-grade probiotic approved by the Ministry of Health of the People’s Republic of China,has important probiotic functions and has been widely used in the food and fermentation industries.However,insufficient research and attention have been paid to the immunomodulatory effects and mechanisms of animal gut health.Therefore,in this experiment,the probiotic properties of Lactobacillus reuteri were tested and domesticated,and then the initial p H and medium components of the domesticated Lactobacillus reuteri GL005were optimized,and the Lactobacillus reuteri GL005 was further observed.The effect of fermentation broth on the rabbit model of diarrhea caused by enterotoxigenic Escherichia coli aims to provide data support and theoretical basis for the application of Lactobacillus reuteri in veterinary clinics.This experiment is divided into three parts:(1)Probiotic testing and domestication of Lactobacillus reuteri to study its probiotic properties;(2)The optimum p H and medium composition of Lactobacillus reuteri GL005 were optimized by single factor test,orthogonal test and response surface test,and the optimal culture conditions of Lactobacillus reuteri GL005 were studied;(3)The rabbits were divided into four groups:control group,model group,antibiotic group and probiotic group(n=4 in each group);the enterotoxigenic E.coli was injected into the model group,probiotic group,and antibiotic group by intraperitoneal injection at 0.7 m L/kg to establish a diarrhea model,and the control group was intraperitoneally injected with the same amount of normal saline.After successful modeling,the probiotic group was given 2.5 m L/kg Lactobacillus reuteri GL005,the antibiotic group was given 0.04 g/kg norfloxacin,and the model group and control group were given the same amount of normal saline for 10 days.In the experiment,the therapeutic effect of the fermentation broth of Lactobacillus reuteri GL005 on diarrhea of young rabbits was studied.The results showed:(1)The Lactobacillus reuteri provided was extremely poor in tolerance to 0.3%bile salt,so the bile salt acclimation test was carried out,and the domesticated Lactobacillus reuteri GL005 had strong tolerance in simulated digestive juice,the survival rate of cultured 4h remained above 97%.The supernatant of Lactobacillus reuteri GL005 had good inhibitory effect on four pathogens,among which the inhibitory effect on Salmonella was the strongest(18±0.41)mm.Lactobacillus reuteri GL005 can quickly enter the logarithmic growth phase,reaching the highest growth point at 16 h,and the p H value is also stable at about 4.27 after 16 h.(2)The p H optimization results showed that the optimum p H of Lactobacillus reuteri GL005 was 5.8;with MRS as the basal medium,the optimized growth medium was:brown sugar 20.25 g/L,glucose 10.13 g/L,maltose 5.06 g/L,soy peptone114.67 g/L,yeast extract 38.22 g/L,diammonium hydrogen citrate 2 g/L,sodium acetate 7.5g/m L,dipotassium hydrogen phosphate 2 g/L,magnesium sulfate 0.098 g/L,manganese sulfate 0.03 g/L,Tween 80 1 g/L,statically cultured in this medium for 16 hours,the number of viable bacteria in the culture medium was 9.8×10~9 CFU/m L,which was 7.54 times higher than that in the basal medium.,successfully achieved high-density culture of Lactobacillus reuteri GL005.(3)The results of animal experiments showed that the immune organ index of rabbits in the probiotic group was higher than that in the control group,indicating that Lactobacillus reuteri GL005 could positively promote the development of immune organs in rabbits;Lactobacillus reuteri GL005 significantly reduced inflammatory factors IL-1β,IL-6and TNF-α,and the level of reduction was better than that of the antibiotic norfloxacin;The repair effect of Lactobacillus reuteri GL005 on ileal villi was better than that of the antibiotic group,and the value of ileal villus height/crypt depth(V/C)of the rabbits in the probiotic group was the largest among the four experimental groups.The test results showed that the domesticated Lactobacillus reuteri GL005 has the characteristics of potential good probiotics,and the number of viable bacteria cultured in the optimized optimal conditions of Lactobacillus reuteri GL005 increased by 7.54 times compared with that before optimization;Lactobacillus reuteri GL005 has a good health care effect on diarrhea in young rabbits.This experiment lays the foundation for the development of rabbit-derived probiotics,and provides data support and theoretical basis for the application of Lactobacillus reuteri GL005 in veterinary clinics.