Comparison of different PCR tests for detecting Shiga toxin-producing Escherichia coli O157 and development of an ELISA-PCR assay for specific identification of the bacteria
| Autor: | Patrick Fach, Joël Grout, Françoise Dilasser, Sylvie Perelle |
|---|---|
| Rok vydání: | 2003 |
| Předmět: |
DNA
Bacterial Microbiology (medical) Enzyme-Linked Immunosorbent Assay Locus (genetics) Escherichia coli O157 Polymerase Chain Reaction Sensitivity and Specificity Microbiology Shiga Toxin law.invention Bacterial genetics fluids and secretions law Chlorocebus aethiops Animals Humans Adhesins Bacterial Vero Cells Molecular Biology Escherichia coli Infections Transaminases Polymerase chain reaction biology Escherichia coli Proteins Shiga toxin Amplicon biology.organism_classification Molecular biology Bacterial adhesin biology.protein bacteria Solution hybridization Carbohydrate Epimerases Carrier Proteins Bacteria Flagellin |
| Zdroj: | Journal of Microbiological Methods. 55:383-392 |
| ISSN: | 0167-7012 |
| Popis: | In an attempt to develop a standard for ELISA-PCR detection of Shiga toxin producing Escherichia coli (STEC) O157, six published PCR tests were tested in a comparative study on a panel of 277 bacterial strains isolated from foods, animals and humans. These tests were based on the detection of the genes rfbE [J. Clin. Microbiol. 36 (1998) 1801] and rfbB [Appl. Environ. Microbiol. 65 (1999) 2954], the 3' end of the eae gene [Epidemiol. Infect. 112 (1994) 449], the region immediately flanking the 5' end of the eae gene [Int. J. Food. Microbiol. 32 (1996) 103], the flicH7 gene [J. Clin. Microbiol. 35 (1997) 656], or a part of the recently described 2634-bp Small Inserted Locus (SILO(157) locus) of STEC O157 [J. Appl. Microbiol. 93 (2002) 250]. Unlike the other PCR assays, those amplifying the rfb sequences were unable to distinguish toxigenic from nontoxigenic O157. These assays were relatively specific to STEC O157, giving essentially a cross reaction with clonally related E. coli O55 and to a lesser extent with E. coli O145, O125, O126. They also detected the Shiga toxin (stx)-negative derivatives of STEC O157. Based on these results, an ELISA-PCR assay consisting of the solution hybridization of amplicons with two probes that ensured the specificity of the amplification was developed. The ELISA-PCR assay, which used an internal control (IC) of inhibition, was able to detect 1 to 10 copies of STEC O157 in the PCR tube. Adaptation of PCR into ELISA-PCR assay format facilitates specific and sensitive detection of PCR amplification products and constitutes a method of choice for screening STEC O157. |
| Databáze: | OpenAIRE |
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