Evaluation of a technique for identification of Shiga-like toxin-producing Escherichia coli by using polymerase chain reaction and digoxigenin-labeled probes.
A polymerase chain reaction (PCR) technique for the identification of Shiga-like toxin (SLT)-producing Escherichia coli was assessed by using 95 strains of SLT-producing E. coli and 5 Shigella dysenteriae type 1 strains. PCR was used for the amplification of slt gene sequences from whole bacterial colonies. A digoxigenin-labeled DNA probe was used for identification of the PCR products in a spot blot hybridization assay. Modifications were made to adapt this technique for the proper identification of 10 SLT-producing isolates which were refractory to the heat lysis step that was used to liberate whole-cell DNA for PCR and 6 isolates which gave nonspecific amplification products. The sensitivity and specificity of this assay were each 99% when compared with toxin neutralization results by using SLT-specific monoclonal antibodies. These values indicate that this detection technique could be suitable for use in a clinical laboratory.
Begum
D
D
Department of Immunology and Microbiology, Wayne State University School of Medicine, Detroit, Michigan 48201.
Strockbine
N A
NA
Sowers
E G
EG
Jackson
M P
MP
D000911:Antibodies, Monoclonal; D001427:Bacterial Toxins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D015342:DNA Probes; D004269:DNA, Bacterial / Q000235:genetics; D004076:Digoxigenin; D004926:Escherichia coli / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D004927:Escherichia coli Infections / Q000175:diagnosis / Q000382:microbiology; D005069:Evaluation Studies as Topic; D005798:Genes, Bacterial; D006801:Humans; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000706:statistics & numerical data; D012680:Sensitivity and Specificity; D022622:Shiga Toxin 1; D022641:Shiga Toxin 2; D012762:Shigella dysenteriae / Q000235:genetics
D000911:Antibodies, Monoclonal; D001427:Bacterial Toxins; D017931:DNA Primers; D015342:DNA Probes; D004269:DNA, Bacterial; D022622:Shiga Toxin 1; D022641:Shiga Toxin 2; D004076:Digoxigenin