Fingerprinting mycobacteria quicker

MANTRA, a rapid genotyping method for Mycobacterium tuberculosis by multiplex PCR and microfluidic labchip. Merritt AJ, Keehner T, O’Reilly LC, McInnes RL, Inglis TJ.  J Clin Microbiol. 2010 Aug 11. [Epub ahead of print]

In this short report we described a method for rapid field genotyping of Mycobacterium tuberculosis. The name MANTRA was chosen to indicate that this is only a nominal tandem repeat analysis, and not the definitive analysis of M. tuberculosis tandem repeats on which MIRU-15 and MIRU-24 are based. Our intent is to use this easily run, single tube method for preliminary field work in resource-limited settings that lack Mycobacterial reference laboratory support close by.

The drive to come up with a field genotyping method was prompted by experience in places where tuberculosis is much commoner than in Australia, and where clinical laboratory services are limited to microscopy and culture – i.e. most places where tuberculosis is common. Publication of the method implies an intent to use the MANTRA method in future overseas development work such as the capacity-building projects we’ve been involved in before. It also leaves us with a need to investigate how best to run this field genotyping method alongside molecular detection/identification assays. Direct application to smear-positive sputa is a priority so that both detection and genotyping assays can be run much closer to the patient in endemic locations.

Definitive M. tuberculosis genotyping methods other than VNTR/MIRU are in current use in Mycobacteriology reference laboratories. These include IS6110 ribotyping and spoligotyping. It is common practice in the developed world to combine at least two different genotyping methods when delineating isolate clusters.

Leaving aside the potential application of whole bacterial genome sequencing to molecular epidemiology, the immediate need in M. tuberculosis genotyping is to combine shorter time-to-genotype with genotyping of a higher proportion of clinical isolates. VNTR/MIRU typing has helped on both counts but is currently hampered by a discussion on whether to use MIRU-15 or MIRU-24. While MIRU-24 may be a little more discriminatory, recent work shows that MIRU-24 and spoligotyping both miss some strain lineages.

Until the optimal method has been identified there will be a continuing discussion over genotype nomenclature.  That discussion will inevitably slow down the roll-out of a standard initial M. tuberculosis genotyping method.  While it was not our intention to replace either reference method of VNTR/MIRU typing, the MANTRA approach may have some merit due to its pragmatic avoidance of genotype labels. It is a de facto plea to molecular epidemiologists to concentrate their collective efforts on populations that suffer the bulk of the global tuberculosis burden.

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