Screening Of Lactobacillus With Inhibition Of Escherichia Coli And Its Effect On Intestinal Flora

Lactic acid bacteria are widely used probiotic species,which play an important role in the health of the host.Enterohemorrhagic Escherichia coli is an intestinal pathogen which treates by antibiotics once the host was infected.However,antibiotics will destroy the structure and composition of the intestinal flora thus cause imbalance in the flora.The intestinal flora plays an important role in the immunity of the host.Studying the inhibition of E.coli by lactic acid bacteria is helpf?L to the prevention and control of E.coli and reduces the use of antibiotics,which has important guiding significance for the development of probiotic resources.In this study,chicken intestinal tissues were used as samples,and Caco-2 cells were used as a model to screen lactic acid bacteria with strong bacteriostatic ability,excellent tolerance and adhesion effect.A mouse model was used in vivo to investigate the infection of E.coli reg?Lation of mouse intestinal flora.1 Isolation and identification of lactic acid bacteria with inhibitory activity against E.coliUsing healthy chicken intestines as samples,Gram stain was used to isolate suspected strains;the double-layer agar method was used to evaluate the antibacterial ability of the strains against E.coli;physiological and biochemical identification and 16S rDNA sequencing analysis were used to determine the genus of the isolated strains;The growth and acid-producing ability of the strains were determined;the resistance of the strains to common antibiotics was determined by the method of minimum inhibitory concentration;and the tolerance of living was evaluated by sim?Lating gastric juice,bile salt solution and sim?Lated intestinal fluid.The res?Lts are as follows:A total of 31 suspected lactic acid bacteria were isolated,of which 10 strains had a strong inhibitory ability on the growth of indicator bacteria,and the diameter of the inhibition zone was greater than 10 mm;And 8 strains of Lactobacillus plantarum,entered the logarithmic growth phase and began to produce acid after 4h,and the final pH was about 3.4;10 strains of cefoxitin,kanamycin,gentamicin,vancomycin,erythromycin And tetracycline are resistant,and are sensitive to amoxicillin and penicillin.The 10 strains have a strong tolerance and the highest survival rate in gastric juice at 1 h is 84.73%.After 3 hours,L.johnsonii NJ13 still has a survival rate of 73.63%The number of survivors reached more than 106 CFU/mL.and the strains co?Ld survive in both 0.3%bile salt solution and sim?Lated intestinal fluid,and the number of live bacteria was not much different from the original,maintaining at 108 CFU/mL.2 Screening for inhibition of E.coli lactic acid bacteriaBased on the three indicators of self-aggregation,co-aggregation and surface hydrophobicity,the adhesion capacity of lactic acid bacteria was initially evaluated;Caco-2 cells were used as an in vitro model to evaluate the adhesion of lactic acid bacteria,and a strain of E.coli with a high adhesion index was selected;The adhesion properties of L.johnsonii NJ13 were investigated.The principal component analysis index was used to screen the best lactic acid bacteria.The Caco-cell model was used to determine the inhibition of E.coli adhesion and IL-8 overexpression by the lactic acid bacteria.Res?Lts:The self-aggregation ability of lactic acid bacteria was positively correlated with the adhesion ability,and the hydrophobic ability was negatively correlated with adhesion.The 10 strains had good probiotic potential;L.plantarum NJ55,L.plantarum NJ56,L.plant arum NJ57,and L.plant arum NJ59 has strong adhesion ability,and its adhesion index is higher than that of the standard strain of Lactobacillus rhamnosus.After treating L.johnsonii NJ13 with different pH,it is found that the number of adhesion in the acidic environment is better to alkaline conditions(pH=9),Metal Ca2+can improve the adhesion ability,Na+,Mg2+,K+,NH+have no effect on the adhesion.The adhesion of L.johnsonii NJ13 may be related to surface proteins,lipoteichoic acid and extracell?Lar polysaccharides after treating the cells with different chemicals.The two most potential probiotics L.johnsonii NJ13 and L.plantarum NJ56 were obtained through principal component analysis.The two strains can inhibit the adhesion rate of E.coli to Caco-2 cells and affect the IL-8 caused by Expression has a significant inhibitory effect.3 Effects of lactic acid bacteria on mouse pop?LationsBy constructing an animal model of mouse diarrhea,L.johnsonii NJ13 and L.plantarum NJ56 and LGG were administered at 108 CFU/mL and 109 CFU/mL,and antibiotics were used to record the weight and diarrhea of the mice.Mouse feces were collected at the end,and the composition and distribution of mouse intestinal flora were analyzed by high-throughput sequencing.Res?Lts:After 7 days of intragastric administration of E.coli,the weight of mice decreased and diarrhea appeared.After treatment with antibiotics and lactic acid bacteria,the weight of mice recovered and diarrhea was relieved.Simpson index reached the highest value of 0.0242 after antibiotic treatment,indicating that the abundance and diversity of the intestinal flora of mice were the lowest,while Simpson index was the smallest after treatment by L.johnsonii NJ13,indicating that it has the structure and composition of mouse intestinal flora Reg?Latory effect;Lactobacillus reg?Lates the distribution of Bacteroides,Pseudomonas,and Proteobacteria in the intestinal flora,and increases the content of beneficial bacteria such as lactic acid bacteria in the intestine,reducing the harmf?L bacteria such as Helicobacter pylori Content,promoted the recovery of intestinal flora of mice after infection.The difference analysis showed that the antibiotic group had the most significant difference from the normal group,and the L.johnsonii NJ13 group was the closest to the normal group.