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Sensitive, specific, and rapid detection of the presence of Shiga toxin-producing organisms such as Escherichia coli O157:H7 and Shigella dysenteriae type 1 in stool
See Laboratory Testing for Infectious Causes of Diarrhea in Special Instructions.
Shiga toxins (also known as Shiga-like toxins, Vero toxins, or Vero-like toxins) are encoded by some strains of Escherichia coli, most notably O157:H7. Shiga toxin can also be produced by other serogroups of enterohemorrhagic Escherichia coli (EHEC), as well as Shigella dysenteriae type 1. Generally, Shiga toxin-producing organisms cause bloody diarrhea, although this is not universal. Unlike some bacterial gastrointestinal infections, antimicrobial therapy is contraindicated, as antimicrobials may exacerbate disease. Treatment is primarily supportive (eg, hydration). A complication of infection by an organism producing Shiga toxin is hemolytic uremic syndrome (HUS). The percentage of people that develop HUS varies among outbreaks of Escherichia coli O157:H7, but generally ranges from 3% to 20%. HUS is characterized by a triad of findings: hemolytic anemia, thrombocytopenia, and kidney failure. Most people recover completely, however, some require permanent dialysis, and some die as a result of complications.
Several diagnostic methods available for the detection of EHEC lack sensitivity, are labor intensive, or have a long turnaround time. There are more than 160 serogroups of EHEC; the first serogroup to be associated with HUS was O157:H7. This is also the serogroup that is most commonly implicated in outbreaks. EHEC O157:H7 is detectable as nonfermenting colonies when cultured on sorbitol MacConkey (SMAC) agar, but the majority of non-O157:H7 Shiga toxin-producing Escherichia coli strains ferment sorbitol and, therefore, are undetectable by this method. The Vero cell line is susceptible to the Shiga toxin, but the assay can take up to 48 hours and is nonspecific. Commercial enzyme-linked immunosorbent assay (ELISA) antigen detection kits have a sensitivity of 90% when compared to culture, but an overnight enrichment step is necessary for adequate sensitivity. PCR detection of stx, the gene encoding Shiga toxin, directly from stool specimens is a sensitive and specific technique, providing same-day results. PCR assay identifies non-O157:H7 Shiga toxin-producing bacteria, extending the utility beyond strains identifiable on SMAC agar.
A positive PCR result indicates the likely presence of Shiga toxin-producing Escherichia coli in the specimen. Although Shigella dysenteriae serotype 1 may produce a positive result, it is extremely rare in the United States.
A negative result indicates the absence of detectable Shiga toxin DNA in the specimen, but does not rule out the presence of Shiga toxin-producing Escherichia coli, as false-negative results may occur due to inhibition of PCR, sequence variability underlying the primers and probes, or the presence of the Shiga toxin gene in quantities less than the limit of detection of the assay. Shiga toxins are encoded on mobile genetic elements and can theoretically be lost by their bacterial host.
Interfering substances in stool may affect the accuracy of this assay; results should always be interpreted in conjunction with clinical and epidemiological findings.
This assay detects stx subtypes stx1, stx2, stx2c, and stx2d. It does not detect stx2e or stx2f, which are seldom associated with human disease.
This assay was prospectively clinically validated using 204 stool specimens submitted for the antigen test (EIA method). In addition, the assay was used to test 85 archived fecal specimens previously tested for either Escherichia coli O157:H7 or Shiga toxin by EIA, with results compared to the prior results. Discordant results on the archived specimens were resolved by submission to the Minnesota Department of Health (MDH) for PCR using different primers. Compared to a combined gold standard (ie, positive by EIA, culture, or MDH PCR) the Mayo PCR assay had 100% sensitivity and specificity; in total, 46 positive and 243 negative specimens were evaluated. No cross-reactivity was observed when tested on a panel of more than 50 organisms commonly found in stool. The analytical sensitivity was 2 targets/mcL.
1. Gould LH, Bopp C: Recommendations for diagnosis of Shiga toxin-producing Escherichia coli infection by clinical laboratories. MMWR Morb Mortal Wkly Rep 2009 Oct;16:v58
2. Grys TE, Sloan LM, Rosenblatt JE, Patel R: Rapid and sensitive detection of Shiga toxin-producing Escherichia coli from nonenriched stool specimens by real-time PCR in comparison to enzyme immunoassay and culture. J Clin Microbiol 2009;47:2008-2012
3. Grys TE, Patel R: Update on Shiga toxin-producing Escherichia coli. Mayo Clinic, Mayo Medical Laboratories Communique
4. Nyre LM, Kiemele DL, Zomok CD, et al: Clinical experience with rapid PCR for detection of Shiga toxin in stool. Abstract of the Annual Meeting of the American Society for Microbiology, 2010 General Meeting, San Diego, CA, May 23-27, 2010