A novel single-step multiplex polymerase chain reaction assay for the detection of diarrheagenic Escherichia coli

被引:38
作者
Fujioka, Miyuki [1 ]
Otomo, Yoshimitsu [1 ]
Ahsan, Chowdhury Rafiqul [2 ]
机构
[1] Hirosaki Univ, Grad Sch Hlth Sci, Hirosaki, Aomori 0368564, Japan
[2] Univ Dhaka, Dept Microbiol, Dhaka 1000, Bangladesh
关键词
Diarrheagenic Escherichia coli; Rapid diagnostic method; Multiplex PCR; TOXIN TYPE; IDENTIFICATION; STRAINS; GENE; VARIANTS; SEROTYPE; O157-H7; FOOD;
D O I
10.1016/j.mimet.2012.12.010
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Escherichia coli that causes diarrhea in humans is referred to as diarrheagenic E. coli (DEC), and has been categorized into the following 5 groups: shigatoxin-producing E. coli (STEC), enteropathogenic E. coli (EPEC), enteroinvasive E. coli (EIEC), enteroaggregative E. coli (EAggEC), and enterotoxigenic E. coli (ETEC). In this study, we developed a novel one-step multiplex polymerase chain reaction (mPCR) for the rapid detection of 10 pathogenic genes (stx1, stx2, eae, bfpA, invE, aggR, esth, estp, elt, and astA) of DEC. Five categorized strains were used as positive controls for DEC harboring each pathogenic gene, and 828 DEC-like strains, isolated from diarrheal stool samples and assumed to be DEC on the basis of serotyping, were used in the mPCR-based detection of the pathogenic genes. To demonstrate the utility of mPCR, the 828 strains were subjected to our optimized protocol, and the results obtained were compared with those obtained by monoplex PCR. The results showed agreement for all strains. Using mPCR, we also detected 65 DEC and 41 astA-positive E. coli, and 7 of these DEC strains were "O antigen untypable" (OUT). This novel mPCR protocol allowed for rapid, convenient, and economical pathogenicity-based identification of the DEC. (c) 2012 Elsevier B.V. All rights reserved.
引用
收藏
页码:289 / 292
页数:4
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