Molecular confirmation of drug resistance in MTBC

Molecular confirmation of resistance to the first line anti- tuberculosis antimicrobials isoniazid, rifampin, ethambutol, and pyrazinamide. Portions of the following genes/genetic regions will be sequenced: inhA and katG (isoniazid), rpoB (rifampin), embB (ethambutol), pncA (pyrazinamide). Molecular confirmation of resistance to secondary antimicrobials may be available upon special request, please consult with the NRCM Program Manager/designate.

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 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, identification, and susceptibility results, if available. Requested testing for urgent submissions should be accompanied with justification.

Please notify the NRCM as to why the testing is required. 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.

If a documented mutation is present, confirmation of drug resistance can be provided. The lack of such a mutation is not sufficient to rule out resistance. Results will be determined by comparison of sequence data to 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. Campbell PJ, Morlock GP, Sikes RD, Dalton TL, Metchock B, Starks AM, Hooks DP, Cowan LS, Plikaytis BB, Posey JE. Molecular Detection of Mutations Associated with First and Second-Line Drug Resistance Compared with Conventional Drug Susceptibility Testing in M. tuberculosis. Antimicrob Agents Chemother.. 2011 Feb 7; doi:10.1128/AAC.01550-10

2. Feuerriegel, S., H. S. Cox, N. Zarkua, H. A. Karimovich, K. Braker, S. Rusch-Gerdes, and S. Niemann. 2009. Sequence analyses of just four genes to detect extensively drug-resistant Mycobacterium tuberculosis strains in multidrug-resistant tuberculosis patients undergoing treatment. Antimicrob Agents Chemother 53:3353-6.