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传统方法和分子方法在鉴定生羊肉中大肠杆菌O157:H7时检测方法的影响

Influence of Detection Methods in Characterizing Escherichia coli O157:H7 in Raw Goat Meat Using Conventional and Molecular Methods.

作者信息

Tabashsum Zajeba, Nazneen Mafruha, Ahsan C R, Bari M L, Yasmin M

机构信息

Department of Microbiology, University of Dhaka.

出版信息

Biocontrol Sci. 2016;21(4):261-264. doi: 10.4265/bio.21.261.

Abstract

Presence of Escherichia coli O157:H7 on fresh goat meat samples (n= 40) of Dhaka city was analyzed using conventional and molecular methods. A total of 86 presumptive E. coli O157:H7 colonies were isolated from 60% of the samples using selective agar plating method. After conventional biochemical assay followed by API 20E assay, only 11 isolates were found to be E. coli O157:H7. Further serological test identified only four isolates that has strong agglutination reaction against anti-H7 sensitized latex. The biochemically and serologically confirmed isolates were then screened for major virulence factors include eaeA, rfbE, fliC, stx1 and stx2 genes by PCR. PCR analysis of positive isolates showed, 10 isolates were eaeA and rfbE genes positive but fliC gene was only in six, indicating that these isolates were H7 positive with flagellum antigens which might not expressed or detected in serotyping tests. Multiplex PCR against eaeA, stx1 and stx2 genes of the isolates showed similar results as when done individually. These results revealed that only 7% of the primary presumptive E. coli O157:H7 was found to be stx producing E. coli O157:H7 and thus greatly influenced the detection of the pathogen in meat samples.

摘要

采用传统方法和分子方法分析了达卡市新鲜山羊肉样品(n = 40)中大肠杆菌O157:H7的存在情况。使用选择性琼脂平板培养法,从60%的样品中总共分离出86个疑似大肠杆菌O157:H7菌落。经过传统生化检测,随后进行API 20E检测,仅发现11株分离株为大肠杆菌O157:H7。进一步的血清学检测仅鉴定出4株对抗H7致敏乳胶有强烈凝集反应的分离株。然后通过PCR对经生化和血清学确认的分离株进行主要毒力因子筛查,包括eaeA、rfbE、fliC、stx1和stx2基因。对阳性分离株的PCR分析显示,10株分离株eaeA和rfbE基因呈阳性,但fliC基因仅在6株中存在,这表明这些分离株H7呈阳性且带有鞭毛抗原,可能在血清分型检测中未表达或未检测到。针对分离株的eaeA、stx1和stx2基因进行的多重PCR结果与单独检测时相似。这些结果表明,最初疑似的大肠杆菌O157:H7中只有7%被发现是产志贺毒素大肠杆菌O157:H7,因此极大地影响了肉类样品中病原体的检测。

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