Early Detection of Mycobacterium tuberculosis in BACTEC Cultures by Ligase Chain Reaction

The LCx Mycobacterium tuberculosis ligase chain reaction system (Abbott Diagnostic Division, Abbott Park, Ill.) was used to detect M. tuberculosis in 150 consecutive BACTEC vials on the day on which a positive growth index (GI) was recorded. By LCx, M. tuberculosis DNA was detected in BACTEC vials on average 2.6 days before the presence of acid-fast bacilli could be confirmed by microscopic examination. A total of 106 of 108 M. tuberculosis isolates were detected without centrifugation from bottles presenting very low GIs (average, 70; median, 33). No false-positive result was obtained from nontuberculous mycobacteria or from isolates with contaminants. Rapid diagnosis of tuberculosis is an important component of measures to control the disease (13), which is an objective made more urgent by the appearance of multidrug-resistant strains (7). Nucleic acid amplification techniques, which allow detection of Mycobacterium tuberculosis directly from samples in a matter of hours, apparently represent the ultimate answer to this issue. They lack sensitivity for smear-negative specimens, and their use is generally restricted to selected samples. Culture remains the only mycobacterial investigation performed, along with microscopy, for the majority of clinical specimens. The radiometric method (BACTEC; Becton Dickinson, Towson, Md.) is at present the fastest and most sensitive cultural tool for the diagnosis of mycobacterial infections (9). The sensitivity of the radiometric assay allows a culture to reach a growth index (GI) of $10, which is considered the positivity threshold, in a very short time, often within a week. However, a BACTEC vial flagged as positive does not necessarily harbor mycobacteria, since other organisms may overgrow despite the decontamination procedure. Confirmation of the presence of acid-fast bacilli is required. Although microscopic tests are very easy to perform, they are characterized by a very low sensitivity, and examination of smears prepared from BACTEC vials with early positive signals has proven to be so frustrating that a delay in examination until the GI becomes greater than 100 is recommended. Several days of further incubation are often required, thus delaying the detection of mycobacteria. Furthermore, a microscopically confirmed positive culture may be due either to M. tuberculosis or to a nontuberculous mycobacterium (MOTT), which poses important dilemmas both for therapy and for the measures to be adopted to prevent spread of the infection. There is indeed interest in a procedure that would enable early recognition of the M. tuberculosis complex as the organism responsible for the positive BACTEC signal; such a method would in fact be consistent with the recommendations of the Centers for Disease Control for a timely identification of isolates to the species level (13). For this purpose, we investigated the reliability of a commercial ligase chain reaction (LCx M. tuberculosis; Abbott Di