Test Id : MTBT

When non-fixed specimen is available, a mycobacterial culture on that specimen must always be performed in addition to this test. If your facility is unable to perform mycobacterial culture, order CTB / Mycobacteria and Nocardia Culture, Varies concurrently with this test.

Specimen source is 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 Mycobacterium tuberculosis DNA is unlikely.

Preferred Paraffin-embedded tissue block:

Supplies: Tissue Block Container (T553)

Specimen Type: Formalin-fixed, paraffin-embedded tissue block (FFPE)

Sources: Body tissue

Container/Tube: Tissue block

Collection Instructions: Submit a FFPE tissue block to be cut and returned.

Acceptable Paraffin-embedded tissue block:

Specimen Type: Formalin-fixed, paraffin-embedded tissue block (FFPE)

Sources: Body tissue

Container/Tube: Sterile container for each individual cut section (scroll).

Collection Instructions: Perform microtomy and prepare five separate 10-micron sections. Each section (scroll) must be placed in a separate sterile container for submission.

See Specimen Required.

Preferred method for rapid detection of Mycobacterium tuberculosis complex DNA in formalin-fixed, paraffin-embedded tissue specimens

Detecting M tuberculosis complex

This test is not intended for the detection of latent tuberculosis and must not be used as a substitute for tests intended for detection of latent tuberculosis such as the tuberculin skin test or an interferon gamma release assay.

Each year, Mycobacterium tuberculosis accounts for more than a million deaths and is responsible for millions of newly diagnosed cases of tuberculosis worldwide. M tuberculosis is spread from person-to-person via respiratory transmission and has the potential to become resistant to many or all antibiotics currently used if antimycobacterial treatment is not promptly initiated. Therefore, rapid and accurate detection of M tuberculosis in patient specimens is of clinical and public health importance.

Conventional culture methods can generally detect M tuberculosis in 2 to 3 weeks, although up to 8 weeks of incubation may be required in some instances. Developed at Mayo Clinic, this rapid polymerase chain reaction (PCR) assay detects M tuberculosis complex DNA directly from specimens without waiting for growth in culture and, therefore, the results are available rapidly after receipt in the laboratory. A mycobacterial culture must always be performed in addition to the PCR assay. The PCR assay is rapid, but culture has increased sensitivity over the PCR assay. The PCR assay targets a unique sequence within the katG gene, which is present in members of the M tuberculosis complex. In addition, the assay can detect genotypic resistance to isoniazid mediated by mutations in the katG target, when present.

Not applicable

A positive result indicates the presence of Mycobacterium tuberculosis complex DNA. Members of the M tuberculosis complex detected by this assay include M tuberculosis, Mycobacterium bovis, Mycobacterium bovis bacillus Calmette-Guerin (BCG), Mycobacterium africanum, Mycobacterium canettii, and Mycobacterium microti. The other species within the M tuberculosis complex (eg, Mycobacterium caprae, Mycobacterium pinnipedii, and Mycobacterium mungi) should, in theory, be detected using the primer and probe sequences in this assay, but they have not been tested at this time. This assay method does not distinguish between the species of the M tuberculosis complex.

A negative result indicates the absence of detectable M tuberculosis complex DNA.

Isoniazid (INH) resistance mediated through a katG variant will be reported when observed but lack of a katG variant does not imply that the isolate is susceptible to INH. There are other genetic loci in addition to katG that can contribute to resistance for this drug.

An inhibition result indicates that inhibitors are present in the specimen that could prevent the detection of M tuberculosis DNA. A new specimen can be resubmitted under a new order, if desired.

This rapid polymerase chain reaction (PCR) assay detects Mycobacterium tuberculosis complex nucleic acid and, therefore, does not distinguish between viable, disease-related organisms and nucleic acid persisting from prior infection. Test results should be correlated with patient symptoms and clinical presentation before a definitive diagnosis is made.

A negative result does not rule out the presence of M tuberculosis complex or active disease because the organism may be present at levels below the limit of detection for this assay.

This test has not been studied for use with specimens from patients being treated with antituberculous agents and, therefore, should not be used to determine bacteriologic cure or to monitor response to therapy. It is not known how long the PCR assay can remain positive following treatment for M tuberculosis.

The sensitivity of this test from formalin-fixed, paraffin-embedded tissue is approximately 63%; therefore, testing of additional specimens should be considered if the result from the first specimen is negative.

A mycobacterial culture on a non-formalin-fixed specimen must always be performed in addition to the PCR test when non-fixed specimen is available. If your facility is unable to perform a mycobacterial culture, CTB / Mycobacteria and Nocardia Culture, Varies  should be ordered.

The sensitivity of the PCR assay from acid-fast smear positive specimens is approximately 96% compared to mycobacterial culture, but sensitivity of the PCR from a smear negative specimen is lower, and a negative result does not rule out M tuberculosis complex.

1. Iseman MD: A Clinician's Guide to Tuberculosis. 2nd ed. Lippincott Williams and Wilkins; 2013

2. American Thoracic Society; Centers for Disease Control and Prevention; Infectious Diseases Society of America. Treatment of tuberculosis. MMWR Recomm Rep. 2003 Jun 20;52(RR-11):1-77. Erratum in: MMWR Recomm Rep. 2005 Jan 7;53(51):1203

3. Ortiz-Brizuela E, Menzies D, Behr Mad: Testing and treating Mycobacterium tuberculosis infection. Med Clin North Am. 2022 Nov;106(6):929-947. Doi: 10.1016/j.mcna.2022.08.001

Following specimen processing, genomic DNA is extracted, and the purified genomic DNA is placed on the LightCycler instrument, which amplifies and monitors, by fluorescence, the development of target nucleotide sequences after each polymerase chain reaction (PCR) cycle. A specific target sequence from a portion of the katG gene from Mycobacterium tuberculosis complex is amplified and the resulting segment is detected by melt-curve analysis using sequence-specific fluorescence resonance energy transfer hybridization probes. The LightCycler PCR assay is a closed PCR system that greatly reduces the potential for false-positive results due to specimen cross-contamination as compared with traditional open-system PCR or other amplification methods like transcription-mediated amplification.(Buckwalter SP, Connelly BJ, Louison LK, et al: Description, validation, and review of a decade of experience with a laboratory-developed PCR test for detection of Mycobacterium tuberculosis complex in pulmonary and extrapulmonary specimens. J Clin Tuberc Other Mycobact Dis. 2022 Nov 12;29:100340. doi: 10.1016/j.jctube.2022.100340)

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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.

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