National Reference Centre for Mycobacteriology

• Research
• Testing and Capabilities
• Detection and Identification Services
• Susceptibility Testing Services
• Genotyping Services

Program Manager: Joyce Wolfe

The National Reference Centre for Mycobacteriology (NRCM) program mandate is to promote basic research and training in the field of Mycobacteriology.  This involves the study of a genus of bacteria known as Mycobacterium, which includes Mycobacterium tuberculosis, and the provision of esoteric diagnostic, surveillance, and consultation services to clients and health care providers. These services are used in the investigation, control, and prevention of human disease within the population.

The NRCM operates a state-of-the art Containment Level 3 (CL3) laboratory that houses the national bio-repository for Mycobacterium strains, which includes a comprehensive collection of both clinical and reference strains of the M. tuberculosis complex and non-tuberculosis mycobacteria. This facility allows the program to provide reference services for health care laboratories and clients across Canada in the field of mycobacterial disease, particularly tuberculosis.  In addition, the program is currently involved in basic and applied research in areas of drug susceptibility testing, molecular targets for antimicrobial resistance, virulence factors in tuberculosis, and species characterization. These endeavors encourage and maintain a wide variety of partnerships, affiliations, and collaborations at the national and international level.

A large effort coordinated and administrated by the NRCM is the National Proficiency Program. This was implemented for Canadian provincial laboratories to ensure laboratory quality assurance for established and new technologies in the areas of mycobacterial species identification, drug susceptibility testing, and molecular genotyping. The NRCM also acts as an external reference for American state proficiency programs. At this time, the NRCM laboratory has completed accrediting their quality system to ISO 17025 and is now planning a second phase of accreditation.

Furthermore, the department provides leadership for the National Tuberculosis Surveillance Program, and responds to tuberculosis outbreaks with surveillance of drug resistance and strain genotyping.

In keeping with its mandate, the department also offers extensive teaching programs for medical infectious disease physicians, graduate and undergraduate students, and visiting scientists.

The NRCM includes;

• Research
• Testing and Capabilities
• Detection and Identification Services
• Susceptibility Testing Services
• Genotyping Services

Research

Detection and Identification Research

Research activities include the development and improvement of molecular techniques for rapid detection and identification of the Mycobacterium tuberculosis complex members and non-tuberculosis mycobacteria direct from clinical samples using real-time PCR technology, characterization and validation of clinically relevant novel Mycobacterium species, and the creation and maintenance of quality controlled sequence databases for rapid and accurate identification of clinical isolates.

Drug Susceptibility Research

Drug susceptibility research projects at the NRCM involve the characterization and validation of molecular targets conferring antimicrobial resistance of M. tuberculosis isolates to first line and second line drugs. Other applied research includes the validation of continuous monitoring systems for phenotypic susceptibility testing of primary and secondary antimicrobials for M. tuberculosis. The NRCM is also involved in the testing of newer antimicrobials against tuberculous and non-tuberculous mycobacteria.

Genotyping Research

Strain typing research activities at the NRCM include the development and validation of new molecular techniques to improve throughput and turnaround times. In conjunction, the development of a national database accessible to clients and users across Canada is in progress to join laboratory and epidemiology data with the aim to better provide public health personnel with accurate, real-time surveillance data. 

Fundamental Research

Fundamental research projects in progress at the NRCM include:

• Microarray technology: Identification of the key genes to differentiate between strains of M. tuberculosis complex.
• Host responses to M. tuberculosis (MTB): Testing strategies of vaccination against chronic diseases caused by mycobacterial or by Leishmania parasitic infections – submitted to CIHR as an operating grant.
• Proteomics of national MTB strains: Collaboration between NRCM, Health Sciences Centre, and Winnipeg Regional Health Authorities – integrated program
• Proteomics of international MTB strains: Collaboration between NRCM and Rogelio Hernandez-Pando, Mexico
• The effects of hyperimmune immune-enhancement on survival against lethal infection of mice with Mycobacterium tuberculosis: Collaboration between NRCM and Dr. Alexei Fedórovish Licea Navarro, Laboratorio de Inmunología Molecular y Biotoxinas, Ensenada, B.C. México

Testing and Capabilities

The NRCM provides a variety of reference services including identification, susceptibility testing and genotyping of tuberculous and non-tuberculous Mycobacterium species (NTMs).

Currently, the NRCM has achieved ISO accreditation for antimicrobial susceptibility testing of M. tuberculosis isolates against primary antimicrobials. The NRCM is actively pursuing ISO accreditation of other reference services including susceptibility testing of NTMsusing Trek® microtitre panels and sequence-based identifications.

Detection and Identification Services

The NRCM provides PCR and sequence-based identification for acid-fast organisms such as Mycobacterium sp., emerging bacterial pathogens, and bioterrorism agents. The NRCM employs the use of a variety of molecular targets for the speciation of both the M. tuberculosis complex members and NTMs. Identification to the species or subspecies level is facilitated by the maintenance of custom gene sequence databases.

Susceptibility Testing Services

The NRCM provides antimicrobial susceptibility testing services for all isolates of Mycobacterium speciesagainst primary and secondary antimicrobial drugs as recommended by the Clinical and Laboratory Standards Institute (formerly NCCLS). Phenotypic susceptibility testing of M. tuberculosis complex isolates is performed primarily with the use of the Bactec 460, as well as the MGIT 960 and MB Bact continuous monitoring systems. Susceptibility testing of non-tuberculous mycobacteria is performed using commercially prepared microtitre dilution plates.

Drug resistance and uncharacteristic drug susceptibility are confirmed through a secondary method such as molecular testing of drug resistance targets. With the characterization of newer molecular targets conferring drug resistance, the NRCM is looking to expand the range of molecular targets tested.

Genotyping Services

The NRCM/NML provides a variety of mycobacterial strain genotyping services, predominantly techniques for typing Mycobacterium tuberculosis complex (MTBC) members. MIRU-VNTR is now a recognized method for M. tuberculosis genotyping and the NRCM is actively pursuing automation of this method for high throughput and improved turnaround times. In addition to MIRU-VNTR, the NRCM utilizes IS6110 RFLP and spoligotyping as secondary genotyping methods for M. tuberculosis isolates as per CDC guidelines.

Through the use of national/international databases, strain typing acts as an effective supplement to standard tuberculosis epidemiology, such as contact tracing, for tuberculosis outbreak investigation, prevention, and control. Strain typing is also valuable for detecting cross-contamination in the laboratory and the prevention of misdiagnosis.