Sampling and analytical method development for qualitative assessment of airborne mycobacterial species of the Mycobacterium tuberculosis complex.

Abstract

This article presents a novel, qualitative approach for detecting airborne M. tuberculosis. Culturing or sample purification is not required. A DNA diagnostic method involving the polymerase chain reaction (PCR) coupled to an enzymatically generated color reaction was used for direct detection of M. bovis BCG (Bacillus of Calmette-Guérin), a surrogate for pathogenic M. tuberculosis. Fewer than 10 mycobacteria were detected with no culturing using this bioanalytical method. Analysis was completed in 1 to 1.5 days, in contrast to traditional culturing methods requiring a minimum of 2-3 weeks. To evaluate an air sampling method coupled to a PCR bioanalytical method, liquid cultures of the surrogate were aerosolized and collected for PCR analyses using 37-mm filter cassettes containing polytetrafluoroethylene filters. An Andersen six-stage (viable) particle sizing sampler was employed as a reference sampler. Aerosolized BCG impacted onto Andersen agar plates required incubation periods of 6-8 weeks before small colony forming units could be detected and enumerated. Although the BCG mean length of the rod-shaped particles was 8.3 microns, the airborne BCG particles were collected predominantly on the Andersen 4-6 stages, representing aerodynamic diameters 0.7 to 3.3 microns. Approximately 25 mycobacteria were detected without culturing using the PCR-filter cassette method. This approach could be used to detect airborne mycobacterial species of the M. tuberculosis complex and could permit the early detection of contaminated indoor air. Also, the efficacy of environmental controls could be evaluated and monitored. This approach could also be used to study the expulsion of infectious particles from patients and may permit risk assessment in regard to personal respiratory protection.