New Approach Developed to Diagnose Tuberculosis

The diagnosis of pulmonary tuberculosis (TB) usually includes laboratory analysis of sputum, a viscous material derived from deep in the airways of patients with active disease which can be difficult to collect and analyze by microbiological and molecular techniques.

An alternative, less invasive sample matrix could greatly simplify TB diagnosis, and as Mycobacterium tuberculosis cells or DNA accumulate on the oral epithelia of pulmonary TB patients, these can be collected and detected by using oral (buccal) swabs.

Scientists at the University of Washington (Seattle, WA, USA) working with colleagues in South Africa, collected three swabs each from 20 subjects with active pulmonary TB and from 20 healthy controls. The swabs (OmniSwab, Whatman; Maidstone, UK) were brushed along the inside of the subject's cheek for about 10 seconds (7 to 8 times) to collect cells and saliva. The head of each swab was ejected into a tube containing suitable solutions.

Samples were tested by using a polymerase chain reaction (qPCR) specific to the M. tuberculosis IS6110 insertion element and qPCR was performed on the StepOnePlus Real-Time PCR system (Applied Biosystems; Foster City, CA, USA) using the protocol of initial incubation at 95 °C for 10 minutes and 45 cycles of 95 °C for 15 seconds (denaturation) and 60 °C for one minute (annealing/extension).

Upon testing by oral swab PCR (OSP), 18 of the 20 case subjects (90%) yielded at least two swabs positive for M. tuberculosis DNA either in partial volume or full volume analyses. Of the 60 swab samples collected from the case group, 44 (73.3%) were positive. All control subjects were negative in all three swab samples per subject, in both the partial volume and full volume analyses (OSP). Sputum smear (acid fast) microscopy results were available for 17 of the 20 case subjects, of which 10 (59%) were smear positive. Specificity of OSP was 100% relative to the presumed disease-negative status of healthy controls.

The authors stressed that the study is merely a proof of principle, limited by its small size. Efforts are under way to expand this initial study into a large-scale controlled trial. Yet the preliminary success offers hope for greatly improved TB detection and control, especially in nations with limited public health resources, OSP could simplify the molecular diagnosis of TB and potentially be used for active TB case finding, made possible by the simple, noninvasive nature of the sampling method. The study was published on March 2, 2015, in the journal Scientific Reports.

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University of Washington
Whatman
Applied Biosystems