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

作者：Fujioka Miyuki^*; Otomo Yoshimitsu; Ahsan Chowdhury Rafiqul

来源：Journal of Microbiological Methods, 2013, 92(3): 289-292.

DOI：10.1016/j.mimet.2012.12.010

• Diarrheagenic Escherichia coli
• Rapid diagnostic method
• Multiplex PCR

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 %26quot;O antigen untypable%26quot; (OUT). This novel mPCR protocol allowed for rapid, convenient, and economical pathogenicity-based identification of the DEC.

• 出版日期2013-3