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. 2004 Dec;42(12):5849-53.
doi: 10.1128/JCM.42.12.5849-5853.2004.

Evaluation of multiplex PCRs for diagnosis of infection with diarrheagenic Escherichia coli and Shigella spp

K R S Aranda  1 , U Fagundes-Neto, I C A Scaletsky
Affiliations

Evaluation of multiplex PCRs for diagnosis of infection with diarrheagenic Escherichia coli and Shigella spp

K R S Aranda et al. J Clin Microbiol. 2004 Dec.

Abstract

We have developed two multiplex PCR assays that detect typical and atypical enteropathogenic Escherichia coli (EPEC) isolates, enteroaggregative E. coli (EAEC) isolates, enterotoxigenic E. coli (ETEC) isolates, enteroinvasive E. coli (EIEC) isolates, Shiga toxin-producing E. coli (STEC) isolates, and Shigella spp. The targets selected for each group were eae and bfpA for EPEC isolates, the target of probe CVD432 for EAEC isolates, the genes encoding heat-labile and heat-stable toxins for ETEC isolates, stx(1) and stx(2) for STEC isolates, and ipaH for EIEC isolates and Shigella spp. These PCRs were specific and sensitive for rapid detection of target isolates in stools. Among 150 stool specimens from the acute diarrhea tested, 9 samples (6%) had atypical EPEC, 9 (6%) had typical EPEC, 7 (4.7%) had EAEC, 3 (2%) had EIEC, 3 (2%) had Shigella spp., and 1 (0.7%) had an O26 STEC strain; we also detected mixed infections, 2 (1.3%) with EAEC and Shigella spp., 1 (0.7%) with atypical and typical EPEC strains, and another with atypical EPEC and EAEC strains. One of the multiplex PCRs directly applied to 36 stool specimens correctly identified 100% of EPEC and EAEC isolates.

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Figures

FIG. 1.
FIG. 1.
Multiplex PCR assay 1 of reference strains and clinical and stool samples. Lane 1, EPEC E2348/69 (eae and bfpA) and EAEC 042 (CVD432); lane 2, EPEC HSP43-1 (eae and bfpA); lane 3, atypical EPEC MA343-4 (eae); lane 4, EAEC MA233-1 (CVD432); lane 5, patient sample 5 (eae and bfpA); lane 6, patient sample 6 (eae); lane 7, patient sample 29 (CVD432); lane 8, stool sample 2 (eae and bfpA); lane 9, stool sample 8 (eae and bfpA); lane 10, stool sample 3 (CVD432); lane 11, stool sample 5 (CVD432); lane 12, stool sample 12 (CVD432); lane 13, stool sample 33 (eae); lane 14, stool sample 34 (eae); lane M, DNA molecular size markers (100-bp ladder).
FIG. 2.
FIG. 2.
Multiplex PCR assay 2 of reference strains and clinical samples. Lane 1, ETEC H10407 (LT gene and ST gene), EIEC EDL1285 (ipaH), and STEC EDL931 (stx1 and stx2); lane 2, ETEC 1111-1 (LT gene and ST gene); lane 3, ETEC 2811-1 (LT gene); lane 4, ETEC 2021-1 (ST gene); lane 5, EIEC 733/6 (ipaH); lane 6, S. flexneri 245-5 (ipaH); lane 7, patient sample 48 (ipaH); lane 8, patient sample 75 (ipaH); lane 9, patient sample 127 (stx1); lane M, DNA molecular size markers (100-bp ladder).
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