Screening Of Lactobacilli With Antagonistic Activity Against Enteroinvasive Escherichia Coli

Enteroinvasive Escherichia coli （EIEC） is one of the main pathogenic Escherichia coli, andpeople have paid great much attention to it for its rapid reproductive speed and invasive property.At present, the widely used treatment of diseases caused by EIEC is the antibiotic treatment, butthe method could cause problems such as coliform disorders, double infection and antibioticsresidue. The approach of using probiotic strains with antagonistic activity to treat EIEC diseaseis now a new direction to avoid the abuse of antibiotics. Therefore, in the present study lacticacid bacteria （LAB） strains isolated from Chinese traditional food were screened for theantagonistic activity against EIEC, and these isolated strains were investigated for their abilitiesto inhibit EIEC adhesion and invasion to epithelial cell lines HT-29and the probiotic properties.In the present study,187LAB strains were selected from fermented dough, pickles, saltedmeat, baby feces and fermentation dairy products by MRS, M17and TPY media. By measuringthe inhibition zone of the lactobacilli spent culture supernatant （SCS） on EIEC growth, wescreened24LAB strains which had strong inhibition ability on EIEC. Various treatments to theSCS of different starins were conducted to compare the residual inhibition effects of the samplesof protease-treated, heat-treated （100℃,1h） and pH-treated （adjusted LAB-SCS to neutral）against EIEC. After heating and treatments with catalase, trypsin and proteinase K, theseLAB-SCS did not lose their antagonistic activities against EIEC, but when the SCS of these LABstrains were adjusted to pH7.0, they lost their antagonistic activities against EIEC growth.The probiotic properties of24selected LAB strains were studied including the adhesionindices to the HT-29cell lines, and tolerance to low acid and high bile concentration. Based onthe results two potential probiotics were screened out, Lactobacillus plantarum CCFM233andCCFM231. Their inhibition zone diameters on EIEC were15.50±0.28mm and15.00±0.40mmseparately. It appeared that part of their SCS inhibitory may result from proteinaceous substances,and their antimicrobial activity required an acidic environment. They all had very high adhesionrates to HT-29cells, and their separate adhesion indices were45±6/cell and29±2/cell. In thecondition of pH2for1h, the residual strain counts were2.5×10⁶and1.6×10⁷CFU ml^-1. After16h incubation with2‰bile, the two strains L. plantarum CCFM233and L. plantarumCCFM231, still had more than3.2×10⁵CFU ml^-1viable bacteria in MRS.In vitro assay showed that L. plantarum CCFM233and L. plantarum CCFM231couldstrongly antagonize the adhesion and invasion of EIEC to the HT-29cell lines. In the protectionof the two strains, the adhesion index of EIEC was reduced about66%and50%,correspondingly, and the invasion rate of EIEC was decreased about31.2%and34.7%,respectively. Furthermore, the prebiotic utilizing ability of L. plantarum CCFM233and L.plantarum CCFM231was studied. The experimental results showed that L. plantarumCCFM233and CCFM231had good abilities of utilizing galacto-oligosaccharides,fructo-oligosaccharide, xylo-oligosaccharide, malto-oligosaccharideslow.Lactobacillus plantarum CCFM233and Lactobacillus plantarum CCFM231were shownnot only to inhibit the growth of EIEC significantly, but also to have high adhesion to HT-29celllines and good tolerance to low acid and high bile. Furthermore, the two strains could stronglyantagonize the adhesion and invasion of EIEC to the HT-29cell lines. This study suggests that Lactobacillus plantarum CCFM233and Lactobacillus plantarum CCFM231could be used aspotential probiotics against EIEC.