[Evaluation of a fully automated mycobacteria culture system, MB/BacT using a newly developed digestion-decontamination procedure, semi-alkaline protease-N-acetyl-L-cysteine-NaOH (SAP-NALC-NaOH) method].

A fully automated non-radiometric mycobacteria culture system, MB/BacT (Organon Teknika, Durham, NC, U.S.A.), was recently introduced in Japan and evaluated for its ability to detect mycobacteria in clinical sputum specimens. A previous study yielded nearly a 40% contamination ratio from sputa treated with the standard N-acetyl-L-cysteine (NALC)-NaOH method. This study employed a mucolytic agent (semi-alkaline protease; SAP) in which the sputa were processed twice for digestion followed by decontamination at twice the standard volume of NALC-NaOH. The concentrated sediments were resuspended in phosphate buffer (0.067 M, pH 6.8), and inoculated into the MB/BacT Process Bottles supplemented with antibiotics. The bottles were incubated at 37 degrees C and monitored for up to fifty-six days. Recovery of mycobacteria was compared in three different egg-based Ogawa media in addition to a non-selective Middlebrook 7H10 agar. A total of 1, 124 clinical sputum specimens have been evaluated. Of these, 464 were positive for growth of mycobacteria, of which 447 (96.3%) were positive by the MB/BacT. False-positive alarms due to break through contamination, mainly by Pseudomonas aeruginosa and Candida spp., were observed in twenty-one specimens (1.9%). The three Ogawa media could detect only 283 (60.5%) to 353 (75.4%) positives, and Middlebrook 7H10 agar only 424 (90.6%) positives. The time to detect positive cultures of Mycobacterium tuberculosis complex by the MB/BacT ranged from 2.2 days to 52.3 days, and 50% of positive cultures were detected within 16.7 days of incubation. It can be concluded that the combination of SAP-NALC-NaOH digestion-decontamination procedure and the MB/BacT is particularly useful for the isolation of mycobacteria and has a faster time to detect than conventional methods. MB/BacT is a suitable alternative method for the detection of mycobacteria in Japan, where the radiometric Bactec System is not available.