Abstract
BackgroundNon-tuberculous mycobacteria recovered from respiratory tract specimens are emerging confounder organisms for the laboratory diagnosis of tuberculosis worldwide. There is an urgent need for new techniques to rapidly identify mycobacteria isolated in clinical practice. Matrix-assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF MS) has previously been proven to effectively identify mycobacteria grown in high-concentration inocula from collections. However, a thorough evaluation of its use in routine laboratory practice has not been performed.MethodologyWe set up an original protocol for the MALDI-TOF MS identification of heat-inactivated mycobacteria after dissociation in Tween-20, mechanical breaking of the cell wall and protein extraction with formic acid and acetonitrile. By applying this protocol to as few as 105 colony-forming units of reference isolates of Mycobacterium tuberculosis, Mycobacterium avium, and 20 other Mycobacterium species, we obtained species-specific mass spectra for the creation of a local database. Using this database, our protocol enabled the identification by MALDI-TOF MS of 87 M. tuberculosis, 25 M. avium and 12 non-tuberculosis clinical isolates with identification scores ≥2 within 2.5 hours.ConclusionsOur data indicate that MALDI-TOF MS can be used as a first-line method for the routine identification of heat-inactivated mycobacteria. MALDI-TOF MS is an attractive method for implementation in clinical microbiology laboratories in both developed and developing countries.
Highlights
Pulmonary tuberculosis is a deadly, contagious infection caused by mycobacteria belonging to the Mycobacterium tuberculosis complex (MTC) [1]
Our data indicate that MALDI-TOF MS can be used as a first-line method for the routine identification of heatinactivated mycobacteria
This task has been complicated by the emergence of non-tuberculous mycobacteria present in respiratory tract specimens from patients suspected of pulmonary tuberculosis, even in countries with a high incidence and burden of pulmonary tuberculosis [3]
Summary
Pulmonary tuberculosis is a deadly, contagious infection caused by mycobacteria belonging to the Mycobacterium tuberculosis complex (MTC) [1]. Tuberculosis remains in the top 10 health priorities in many developing countries, and with more than 1.3 million new cases yearly [2] and the emergence and rapid spread of multidrug-resistant and extensively drug-resistant M. tuberculosis strains in such countries, the rapid laboratory diagnosis of tuberculosis is essential This task has been complicated by the emergence of non-tuberculous mycobacteria present in respiratory tract specimens from patients suspected of pulmonary tuberculosis, even in countries with a high incidence and burden of pulmonary tuberculosis [3]. Confined real-time PCR has been shown to be effective for the rapid detection of tuberculosis, including rifampin-resistant tuberculosis, in respiratory tract specimens [13,14] These molecular techniques still require specific technical expertise and technically demanding, expensive equipment [15,16,17]. A thorough evaluation of its use in routine laboratory practice has not been performed
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