混合乳酸菌对小鼠肠上皮细胞的粘附及抑菌机制
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摘要
目的研究混合乳酸菌粘附及抑菌机制。方法测定混合菌株粘附相关能力及抑菌方式,初步研究菌株粘附相关的表面活性成分。结果混合菌株的表面疏水性、自聚合能力较高,对ICE-6细胞的粘附性亦较高。可抑制G+菌(金黄色葡萄球菌、枯草杆菌、艰难梭菌)和G-菌(大肠埃希菌、沙门菌、假单胞菌)粘附ICE-6细胞,抑制方式效果为排除>竞争>替代;热处理对菌株粘附性影响明显高于胃蛋白酶、胰蛋白酶、LiCl和NaIO_4等处理,表面蛋白(80～140kDa)可使LiCl处理菌株的粘附率提升45%。结论混合乳酸菌主要通过排除方式抑制外源菌的粘附,表面蛋白影响混合乳酸菌的粘附能力。
Objective To study the mechanism of the adhesion-inhibition of mixed Lactic acid bacteria.Methods The adhesion related activity and bacteriostatic way of the mixed strains were detected.The surface active components related to the adhesion was preliminarily researched.Results The mixed strains possessed certain hydrophobicity,aggregation activity and high adhesive activity to the ICE-6 cells.Meanwhile,the adhesion of three Gram-positive bacteria(Staphylococcus aureus,Bacillus subtilis,Clostridium difficile)and three Gram-negative bacteria(Escherichia coli,Salmonella WX29,Pseudomonas brenneri)to the ICE-6 cells were inhibited by the mixed strains.The most effective way of the inhibition was exclusion,followed by competition and replacement.Moreover,thermal treatment affected the adhesion activity of the mixed strains more significantly than pepsin,trypsin,LiCl and NaIO_4 did.Surface protein(80-140 kDa)was extracted and incubated with the treated mixed strains and the adhesion rate was improved by 45%.Conclusion The mixed Lactic acid bacteria mainly used the exclusion mechanism to inhibit the adhesion of extracellular bacteria to to the cells.Surface protein played important roles in the adhesion-inhibition activity of the mixed Lactic acid bacteria.
Objective To study the mechanism of the adhesion-inhibition of mixed Lactic acid bacteria.Methods The adhesion related activity and bacteriostatic way of the mixed strains were detected.The surface active components related to the adhesion was preliminarily researched.Results The mixed strains possessed certain hydrophobicity,aggregation activity and high adhesive activity to the ICE-6 cells.Meanwhile,the adhesion of three Gram-positive bacteria(Staphylococcus aureus,Bacillus subtilis,Clostridium difficile)and three Gram-negative bacteria(Escherichia coli,Salmonella WX29,Pseudomonas brenneri)to the ICE-6 cells were inhibited by the mixed strains.The most effective way of the inhibition was exclusion,followed by competition and replacement.Moreover,thermal treatment affected the adhesion activity of the mixed strains more significantly than pepsin,trypsin,LiCl and NaIO_4 did.Surface protein(80-140 kDa)was extracted and incubated with the treated mixed strains and the adhesion rate was improved by 45%.Conclusion The mixed Lactic acid bacteria mainly used the exclusion mechanism to inhibit the adhesion of extracellular bacteria to to the cells.Surface protein played important roles in the adhesion-inhibition activity of the mixed Lactic acid bacteria.
引文
[1]Bermude BM,Plaza DJ,Munoz QS,et al.Probiotic mechanisms of action[J].Ann Nutr Metab,2012,61(2):160-174.
[2]Schillinger U,Guigas C,Holzapfel WH.In vitro adherence and other properties of Lactobacilli used in probiotic yoghurtlike products[J].Int Dairy J,2005,15(12):1289-1297.
[3]Velez MP,De Keersmaecker SC,Vanderle YJ.Adherence factors of Lactobacillus in the human gastrointestinal tract[J].FEMS Microbiol Lett,2007,276(2):140-148.
[4]Xu H,Jeong HS,Lee HY,et al.Assessment of cell surface properties and adhesion potential of selected probiotic strains[J].Lett App Microbiol,2009,49(4):434-442.
[5]Zhang W,Wang H,Liu J,et al.Adhesive ability means inhibition activities for Lactobacillus against pathogens and S-layer protein plays an important role in adhesion[J].Anaerobe,2013,22:97-103.
[6]CHEN Shuai,ZHOU Fangfang,WANG Yinyu,et al.The adhesive properties of L.casei LC2Wto gastric epithelial cell and its relationship with surface properties[J].Chin J Microecol,2010,22(11):966-969.(in Chinese)陈帅,周方方,王荫榆,等.干酪乳杆菌LC2W对胃上皮细胞的粘附性质及其与菌体表面性质的关系[J].中国微生态学杂志,2010,22(11):966-969.
[7]Logan RP,Robins A,Turner GA,et al.A novel flow cytometric assay for quantitating adherence of Helicobacter pylori to gastric epithelial cells[J].J Immunol Methods,1998,213(1):19-30.
[8]Rojas M,Conwa PL.A dot-blot assay for adhesive components relative to probiotics[J].Method Enzymol,2001,336:389-402.
[9]Santarmaki V,Kourkoutas Y,Zoumpopoulou G,et al.Survival,intestinal mucosa adhesion,and immunomodulatory potential of Lactobacillus plantarumstrains[J].Curr Microbiol,2017,74(9):1061-1067.
[10]Polak BM,Wasko A,Paduch R,et al.The effect of cell surface components on adhesion ability of Lactobacillus rhamnosus[J].Antonie Van Leeuwenhoek,2014,106(4):751-762.
[11]Nikolic M,Jovcic B,Kojic M,et al.Surface properties of Lactobacillus and Leuconostoc isolates from homemade cheeses showing auto-aggregation ability[J].Eur Food Res Tech,2010,231(6):925-931.
[12]Servin AL.Antagonistic activities of Lactobacilli and Bifidobacteriaagainst microbial pathogens[J].FEMS Microbiol Rev,2004,28(4):405-440.
[13]Sanchez B,Urdaci MC.Extracellular proteins from Lactobacillus plantarum BMCM12prevent adhesion of enteropathogens to mucin[J].Curr Microbiol,2012,64(6):592-596.
[14]Neeser JR,Granato D,Rouvet M,et al.Lactobacillus johnsonii La1shares carbohydrate-binding specificities with several enteropathogenic bacteria[J].Glycobiology,2000,10(11):1193-1199.
[15]Mukai T,Asasaka T,Sato E,et al.Inhibition of binding of Helicobacter pylori to the glycolipid receptors by probiotic Lactobacillus reuteri[J].FEMS Immunol Med Microbiol,2002,32(2):105-110.
[16]Lebeer S,Vanderleyden J.Genes and molecules of Lactobacilli supporting probiotic action[J].Microbiol Mol Biol R,2008,72(4):728-764.
[17]Ren D,Li C,Qin Y,et al.Inhibition of Staphylococcus aureus adherence to Caco-2cells by Lactobacilli and cell surface properties that influence attachment[J].Anaerobe,2012,18(5):508-515.
[18]Sara M,Sleytr UB.S-layer proteins[J].J Bacteriol,2000,182(4):859-868.
[19]Silja AJ,Airi P.Lactobacillus surface layers and their applications[J].FEMS Microbiol Rev,2005,29(3):511-529.
[2]Schillinger U,Guigas C,Holzapfel WH.In vitro adherence and other properties of Lactobacilli used in probiotic yoghurtlike products[J].Int Dairy J,2005,15(12):1289-1297.
[3]Velez MP,De Keersmaecker SC,Vanderle YJ.Adherence factors of Lactobacillus in the human gastrointestinal tract[J].FEMS Microbiol Lett,2007,276(2):140-148.
[4]Xu H,Jeong HS,Lee HY,et al.Assessment of cell surface properties and adhesion potential of selected probiotic strains[J].Lett App Microbiol,2009,49(4):434-442.
[5]Zhang W,Wang H,Liu J,et al.Adhesive ability means inhibition activities for Lactobacillus against pathogens and S-layer protein plays an important role in adhesion[J].Anaerobe,2013,22:97-103.
[6]CHEN Shuai,ZHOU Fangfang,WANG Yinyu,et al.The adhesive properties of L.casei LC2Wto gastric epithelial cell and its relationship with surface properties[J].Chin J Microecol,2010,22(11):966-969.(in Chinese)陈帅,周方方,王荫榆,等.干酪乳杆菌LC2W对胃上皮细胞的粘附性质及其与菌体表面性质的关系[J].中国微生态学杂志,2010,22(11):966-969.
[7]Logan RP,Robins A,Turner GA,et al.A novel flow cytometric assay for quantitating adherence of Helicobacter pylori to gastric epithelial cells[J].J Immunol Methods,1998,213(1):19-30.
[8]Rojas M,Conwa PL.A dot-blot assay for adhesive components relative to probiotics[J].Method Enzymol,2001,336:389-402.
[9]Santarmaki V,Kourkoutas Y,Zoumpopoulou G,et al.Survival,intestinal mucosa adhesion,and immunomodulatory potential of Lactobacillus plantarumstrains[J].Curr Microbiol,2017,74(9):1061-1067.
[10]Polak BM,Wasko A,Paduch R,et al.The effect of cell surface components on adhesion ability of Lactobacillus rhamnosus[J].Antonie Van Leeuwenhoek,2014,106(4):751-762.
[11]Nikolic M,Jovcic B,Kojic M,et al.Surface properties of Lactobacillus and Leuconostoc isolates from homemade cheeses showing auto-aggregation ability[J].Eur Food Res Tech,2010,231(6):925-931.
[12]Servin AL.Antagonistic activities of Lactobacilli and Bifidobacteriaagainst microbial pathogens[J].FEMS Microbiol Rev,2004,28(4):405-440.
[13]Sanchez B,Urdaci MC.Extracellular proteins from Lactobacillus plantarum BMCM12prevent adhesion of enteropathogens to mucin[J].Curr Microbiol,2012,64(6):592-596.
[14]Neeser JR,Granato D,Rouvet M,et al.Lactobacillus johnsonii La1shares carbohydrate-binding specificities with several enteropathogenic bacteria[J].Glycobiology,2000,10(11):1193-1199.
[15]Mukai T,Asasaka T,Sato E,et al.Inhibition of binding of Helicobacter pylori to the glycolipid receptors by probiotic Lactobacillus reuteri[J].FEMS Immunol Med Microbiol,2002,32(2):105-110.
[16]Lebeer S,Vanderleyden J.Genes and molecules of Lactobacilli supporting probiotic action[J].Microbiol Mol Biol R,2008,72(4):728-764.
[17]Ren D,Li C,Qin Y,et al.Inhibition of Staphylococcus aureus adherence to Caco-2cells by Lactobacilli and cell surface properties that influence attachment[J].Anaerobe,2012,18(5):508-515.
[18]Sara M,Sleytr UB.S-layer proteins[J].J Bacteriol,2000,182(4):859-868.
[19]Silja AJ,Airi P.Lactobacillus surface layers and their applications[J].FEMS Microbiol Rev,2005,29(3):511-529.