Molecular Differentiation of M. tuberculosis complex (MTBC)

Differentiation of MTBC members through sequence analysis of a combination of genes. MTBC constitutes a genetically related group that encompasses M. tuberculosis (including 'M. tuberculosis subsp canettii'), M. bovis, M. bovis BCG, M. africanum, M. caprae, M. microti and M. pinnipedii.

Solid or liquid media growth. For solid media growth, isolated colonies on plated media are preferred. For liquid culture, a minimum volume of 4 mL of actively growing culture is preferred. Mycobacterium tuberculosis complex species must show visible growth and be no more than 6 weeks old.

N/A

Ship all cultures at room temperature (DO NOT freeze) for overnight delivery, and prior to Wednesday each week to ensure receipt by Friday.

Shipping of specimens shall be done by a TDG certified individual in accordance with TDG regulations. For additional information regarding classification of specimens for the purposes of shipping, consult either Part 2 Appendix 3 of the TDG Regulations or section 3.6.2 of the IATA Dangerous Goods Regulations as applicable.

None

Requisitions must be faxed prior to shipping isolates to NRCM - attention Joyce Wolfe, Program Manager, NRCM/NML at 204-789-2036. Requisition for the NRCM must be completed and signed off by the supervisor/designate of the submitting laboratory and include the source of specimen, patient gender, date of birth, clinical history, submitting laboratory identifier and submitter information. Also include isolate characteristics: microscopy, pigmentation, culture characteristics, growth rate/temperature, and identification. Requested testing for urgent submissions should be accompanied with justification.

Submitted cultures will be rejected and a resubmission requested if there is inadequate growth, or there is contamination. Cultures will also be rejected if appropriate documentation and justification is incomplete or missing.

Species identification will be accomplished through sequence analysis of genetic targets as needed. The identification will be determined by comparison of sequence data to an in-house quality controlled databases.

Urgent submissions: 4 calendar days from the date of specimen receipt. Routine testing from culture: 10 calendar days from the date of specimen receipt.

1. Niemann S, Harmsen D, Rusch-Gerdes S, and Richter E. (2000) Differentiation of Clinical Mycobacterium tuberculosis Complex Isolates by gyrB DNA Sequence Polymorphism Analysis. J. Clin. Microbiol. 38:3231-3234.

2. Huard RC, de Oliveira Lazzarini LC, Butler WR, van Soolingen D, and Ho JL. (2003) PCR-Based Method To Differentiate the Subspecies of the Mycobacterium tuberculosis Complex on the Basis of Genomic Deletions. J. Clin. Microbiol. 41:1637.

3. Frothingham, R. (1995). Differentiation of Strains in Mycobacterium tuberculosis Complex by DNA Sequence Polymorphisms, Including Rapid Identification of M. bovis BCG. J. Clin. Micrbiol. 33:840-844.

4. Akos Somoskovi, Jillian Dormandy, Linda M Parsons, Michel Kaswa, Khye Seng Goh, Nalin Rastogi, Max Salfinger. Sequencing of the pncA gene in members of the Mycobacterium tuberculosis complex has important diagnostic applications: Identification of a species-specific pncA Mutation in Mycobacterium canettii, and the Reliable and Rapid Predictor of Pyrazinamide Resistance. J Clin Microbiol. 2006 Nov 29; : 17135430 (P,S,E,B)

5. Kamerbeek J, Schouls L, Kolk A, van-Agterveld M, van-Soolingen D, Kuijper S, Bunschoten A, Molhuizen H, Shaw R, Goyal M, and van-Embden J. (1997) Simultaneous detection and strain differentiation of Mycobacterium tuberculosis for diagnosis and epidemiology. J.Clin.Microbiol. 35:907-914.