Dramatic reduction of culture time of Mycobacterium tuberculosis

Abstract

Mycobacterium tuberculosis culture, a critical technique for routine diagnosis of tuberculosis, takes more than two weeks. Here, step-by-step improvements in the protocol including a new medium, microaerophlic atmosphere or ascorbic-acid supplement and autofluorescence detection dramatically shortened this delay. In the best case, primary culture and rifampicin susceptibility testing were achieved in 72 hours when specimens were inoculated directly on the medium supplemented by antibiotic at the beginning of the culture.

Figure 1. Comparison of different protocols for the culture of M. tuberculosis.

A total of 20 sputum specimens were cultured using different protocols, and the time of detection of M. tuberculosis in primary cultures was measured. Step-by-step modifications included medium composition, atmospheric control and ultimate detection of autofluorescent microcolonies. *P < 0.05, Student's t test.

Figure 2. Comparison of the three methods to detect M. tuberculosis colonies.

(a), Size and mycobacterial load when detecting microcolonies by autofluorescence, by binocular microscope and by naked eye. (b), Confocal microscopic image of a 90-μm autofluorescent microcolony of M. tuberculosis grown on MOD4 medium.