Study On The Mechanism Of Antagonism To Helicobacter Pylori By Two Potential Probiotics And Preliminary Study On Its Dairy Technology

Helicobecter pylori(H.pylori),which caused different kinds of digestive system diseases,including peptic and even gastric cancer,was classified by IARC(International Agency for Research on Cancer,part of the WHO)as a class 1carcinogen.In 1996,the standard therapy for H.pylori eradication,clarithromycin based triple therapies,had been created and applied widely.However,serious adverse effects increasing antibiotic resistance and failure rates for H.pylori eradication had been caused.Plenty of clinical researches confirmed that probiotics had functions of regulating gastrointestinal envrionment and reducing antibiotic-induced adverse effects.Research on lactic acid bacteria,one of the most popular probiotics,was highly foucused by academia and industry.In this dissertation,two strains analysed by probiotic assesment,Lactobacillus plantarum ZDY 2013(L.plantarum ZDY2013)and Bifidobacterium bifidum WBIN03(B.bifidum WBIN03),had been used to investigate the antagonism mechanism of H.pylori and application in dairy production as probiotics food.The main results were shown as follows:The inhibition effect of L.plantarum ZDY2013 and B.bifidum WBIN03 was investigated by using microbial physiological and morphological technology.Both of the two strains and their mixure could reduce the urease viability of H.pylori SS1(P < 0.001),with inhibition rate of 40.42%?39.75%?38.96%.In well diffusion assay,diameters of inhibition zones of the two strains of potential probiotics were19.17 ± 0.58 cm?21.17 ± 2.52 cm,respectively.It was observed that morphology and integrity of H.pylori SS1 cells had been influenced after treated with supernatant of two strains of potential probiotics via transmission electron microscopy analysis.Time-kill assay showed that it were organic acids in potential probiotics supernatants that had antibacterial ability.The acid category and yield were investigated by high performance of liquid chromatography(HPLC),L.plantarum ZDY 2013 and B.bifidum WBIN 03 could inhibit the growth of H.pylori SS1 through producing lactic acid and acetic acid.To explore L.plantarum ZDY2013 and B.bifidum WBIN03 antagonistic mechanism of H.pylori SS1 adhesion,non-specific and specific adhesion from two perspectives had been tested.Moreover,effects of the two strains on the transcription of cytokines induced by H.pylori infection were analyzed.The results of non-specific adhesion evaluation showed that the autoaggregation ability of the the two strains of potential probiotics was good,self-polymerization percentages of which reached 36.20% and 43.78% at 4 h,respectively.The hydrophobic percentage of L.plantarum ZDY2013 reached 40.62%,which was significantly higher than that of B.bifidum WBIN03(P < 0.001).Specific adhesion assay on AGS cells showed that the adherence rate of L.plantarum ZDY2013 was 42.53%,and that of B.bifidum WBIN 03 was 1.52 times higher,indicating that both of them could adhere to AGS Cells well,while the binding force was stronger with the latter.Both two strains could effectively inhibit the adhesion of H.pylori SS1 via three modes of replacement,exclusion and competition,and the competitive effect was the best.Under this model,the inhibitory rates of L.plantarum ZDY2013 and B.bifidum WBIN03 were up to 54.46% and 74.26%,respectively.In addition,q-PCR analysis showed that after treatment with B.bifidum WBIN03,L.plantarum ZDY2013 and mixture of the two strains,the relative expression level of TNF-? was significantly lower than that of the H.pylori treatment group(P < 0.001);among them,the relative expression levels of TNF-? in the H.pylori treated group were 1.67,7.95,21.73,7.94,12.37 and 16.78 times of the potential probiotic treatment group and the pretreatment group,respectively.In conclusion,L.plantarum ZDY2013 and B.bifidum WBIN03 could effectively antagonize the adhesion of H.pylori SS1 to human gastric adenocarcinoma cells and regulate the transcription level of related cytokines induced by H.pylori infection.L.plantarum ZDY2013 fermented milk product was produced.The optimum fermentation conditions were: inoculation amount of 1.5%,milk powder concentration of 15%,fermentation temperature at 42?,fermentation time of 11 h.