Isolation And Idetification Of Liseria Monocytogenes And The Inhibition Or Degradation Of Its Biofilm Formation

Listeria monocytogenes, one of important food-borne pathogens, cause theoutbreak of food-borne disease repeatedly. Some of the listeria strains can formbiofilms and stick to food processing and retail facilities’ surface, making them moreresistant to negative environment. And they are not easy to remove and will harm thefood safety. In this study, Listeria monocytogene was selectively isolated from foodsamples collected from local markets, and identified according to SNT0184.1-2005combined with physiological and biochemical identification method. This thesis alsostudies the inhibition and degradation effect of iron complex of pyridine carboxylicacid and poisson gold ester series on biofilm formation by ListeriamonocytogenesATCC19111using the96microplates assay. The results are asfollowing:（1）According to standard SN/T0184.1-2005of China entry-exit inspection andquarantine industry,18suspected listeria monocytogenes strain were isolated byculturing in selective medium Fraser broth and on PALCAM agar plate from30foodsamples collected in local markets. The multiple PCR amplification methodcombined with the rhamnose, xylose fermentation and Gram staintest were used toidentify those strains. There are13strains identified preliminarily to be listeriamonocytogenes. They are from eight different samples, and the detection rate is about26.7%.（2）Biofilm formation of the13identified strains were studied by culturing in LB,TSB and BHI separately.11strains out of the13strains were found to be able toformbiofilm obviously, which are T12、T13、O11、F11、H14、Q11、S11、R12、M12、F13、R11. They are from8different samples, including dumpling of two brands,raw beef, beef meat balls, chicken, chicken wings, dumplings from wet market, andpork. All of themform most biofilms in LB culture medium with OD value over0.7.（3）Poisson Gold ester series and their sodium salts (0.006μmol/ml,0.013μmol/ml,0.025μmol/ml) were added into the Listeria monocytogenes ATCC19111culture at0h,and incubated for20hours. Bacteria growth and biofilm formation were examined by 96microplate method. In general the sodium salts of Poisson Gold ester series havegood inhibition effect on growth of Listeria monocytogenes ATCC19111and certaindegradation effect to the biofilms.(4) Poisson Gold ester series (0.05μmol/ml,0.1μmol/ml,0.15μmol/ml) were addedinto the Listeria monocytogenes ATCC19111culture at0h,4h,18h,24h. The bacteriagrowth and biofilm formation after incubated for20hours were examined.It wasfound that the three Poisson Gold ester which were added into the Listeriamonocytogenes ATCC19111culture at0h and4h can inhibit the growth of thebacteria effectively. However, when they were added at8h or later, the inhibitioneffect is reduced, showing that poisson gold ester series show better inhibition effecton listeria culture medium of low concentration. For biofilm formation, esters addedat different time period and different concentration displayed an effectiveinhibition.Within the selected concentration range, it is not found that highconcentrations of paraben ester have stronger antibacterial and inhibit effect onbiofilm formation.（5）Aqueous solution of iron complexes of pyridine carboxylic acid (0.1μmol/ml，0,2μmol/ml，0.3μmol/ml) were added to the Listeria monocytogenesATCC19111culture at0h,4h,8h,18h and24h respectively, and incubated for20hours. Bacteriagrowth and biofilm formation were examined by96microplate method. Resultsindicated that every time iron complexes of pyridine carboxylic acid shown obviousinhibition and degradation effect on the biofilm fromation of Listeria monocytogenes.And the best concentration for inhibition effect is0.1μmol/ml. However, it can notsignificantly inhibit the growth of the bacteria.