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Test ID STFRP Shiga Toxin, Molecular Detection, PCR, Feces

Reporting Name

Shiga Toxin PCR, F

Useful For

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

 

This test is not recommended as a test of cure.

Specimen Type

Fecal


Additional Testing Requirements


In some cases, there may be local public health requirements that impact Mayo Clinic Laboratories (MCL) clients and require additional testing on specimens with positive results for this test. MCL recommends that clients retain an aliquot of each specimen submitted for testing to perform such additional testing, if needed. Alternatively (not preferred), clients who want their specimen returned from MCL should call MCL as soon as possible, at the latest within 96 hours of specimen collection, to request that MCL return an aliquot of the submitted specimen to them. Clients will be responsible for submitting their specimens to appropriate public health departments.



Necessary Information


Specimen source is required.



Specimen Required


The high sensitivity of amplification by polymerase chain reaction requires the specimen to be processed in an environment in which contamination of the specimen by shiga toxin DNA is unlikely.

 

Submit only 1 of the following specimens:

 

Preferred:

Specimen Type: Preserved feces

Supplies: Culture and Sensitivity Stool Transport Vial (T058)

Container/Tube: Commercially available transport system specific for recovery of enteric pathogens from fecal specimens (15 mL of nonnutritive transport medium containing phenol red as a pH indicator, either Cary-Blair or Para-Pak C and S)

Specimen Volume: Representative portion of feces; 5 mL

Collection Instructions:

1. Collect fresh fecal specimen and submit in container with transport medium.

2. Place feces in preservative within 2 hours of collection.

Specimen Stability Information: Ambient (preferred) <7 days/Refrigerated <7 days

 

Acceptable:

Specimen Type: Unpreserved feces

Supplies:

-Stool container, Small (Random), 4 oz Random (T288)

-Stool Collection Kit, Random (T635)

Container/Tube: Fecal container

Specimen Volume: Representative portion of feces

Collection Instructions: Collect fresh fecal specimen and submit representative sample in fecal container.

Specimen Stability Information: Refrigerated (preferred) <7 days/Frozen <7 days


Specimen Minimum Volume

1 mL

Specimen Stability Information

Specimen Type Temperature Time Special Container
Fecal Varies 7 days

Reference Values

Not applicable

Day(s) Performed

Monday through Sunday

Test Classification

This test was developed and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. It has not been cleared or approved by the US Food and Drug Administration.

CPT Code Information

87798

Clinical Information

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 E 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 E 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 due to complications.

 

Several diagnostic methods are available for the detection of EHEC but 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 frequently implicated in outbreaks. EHEC O157:H7 is detectable as non-fermenting colonies when cultured on sorbitol MacConkey (SMAC) agar, but the majority of non-O157:H7 Shiga toxin-producing E 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. Polymerase chain reaction (PCR) detection of stx, the gene encoding Shiga toxin, directly from fecal specimens is a sensitive and specific technique, providing same-day results. The PCR assay identifies non-O157:H7 Shiga toxin-producing bacteria, extending the utility beyond strains identifiable on SMAC agar.

Interpretation

A positive polymerase chain reaction (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 E coli and may occur due to inhibition of PCR, sequence variability underlying primers or probes, or the presence of Shiga toxin DNA 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.

Cautions

Interfering substances in the fecal specimen 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.

 

Repeat testing should not be performed on specimens collected less than 7 days apart.

Clinical Reference

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. 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

3. Procop GW, Church DL, Hall GS, et al: The Enterobacteriaceae. In: Koneman's Color Atlas and Textbook of Diagnostic Microbiology. 7th ed. Wolters Kluwer; 2017:213-315

Method Description

This method employs a target-specific detection system, including polymerase chain reaction (PCR) primers and fluorescent resonance energy transfer (FRET) hybridization probes designed for the stx1 and stx2 genes. The LightCycler instrument amplifies and monitors target nucleic acid sequences by fluorescence during PCR cycling. This is an automated PCR system that can rapidly detect amplified product development. The detection of amplified products is based on the FRET principle. For FRET product detection, a hybridization probe with a donor fluorophore, fluorescein, on the 3' end is excited by an external light source, which emits light that is absorbed by a second hybridization probe with an acceptor fluorophore, LC-Red 640, at the 5' end. The acceptor fluorophore then emits a light of a different wavelength that is measured with a signal that is proportional to the amount of specific PCR product. The process is completed in a closed tube system.(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)

Report Available

1 to 2 days

Reject Due To

Formed feces or feces in gel transport medium
EcoFix preservative; formalin or PVA fixative
Preserved feces received frozen		 Reject

NY State Approved

Yes

Method Name

Real-Time Polymerase Chain Reaction (PCR) using Fluorescent Resonance Energy Transfer (FRET)

Testing Algorithm

For more information see Laboratory Testing for Infectious Causes of Diarrhea.

Forms

If not ordering electronically, complete, print, and send 1 of the following forms with the specimen:

-Microbiology Test Request (T244)

-Gastroenterology and Hepatology Test Request (T728)

-Renal Diagnostics Test Request (T830)

-Coagulation Test Request (T753)