Drug-resistant strains of Mycobacterium tuberculosis, though not a novel phenomenon, are emerging worldwide. According to the latest figures of the World Health Organization and the International Union against Tuberculosis and Lung Diseases, drug resistance, in particular acquired resistance, has poured additional fuel on the fire of global tuberculosis (TB) (18). Several outbreaks of multidrug-resistant TB (7) were characterized by delayed diagnoses, inadequate treatment regimens, high rates of mortality, and significant rates of transmission and have taught us two lessons: first, the days are definitely gone where full susceptibility of TB bacilli to front-line drugs can be taken for granted. Second, rapid detection of drug resistance is paramount, not only for effective treatment of TB patients but also for initiating adequate public health measures.In the quest for new nonradiometric, culture-based strategies which allow both rapid detection of acid-fast bacilli and testing of susceptibility to antimicrobial agents, new liquid medium-based systems, such as the MB/BacT (Organon-Teknika, Durham, N.C.), ESP Culture System II (AccuMed International, Westlake, Ohio), MB Redox (Biotest, Dreieich, Germany), and the Mycobacteria Growth Indicator Tube 960 (MGIT 960, Becton Dickinson Microbiology Systems, Sparks, Md.), have become available. They all aim not only at recovering mycobacteria from clinical specimens but also at generating antimicrobial susceptibility testing (AST) data with a shorter turnaround time than that observed with the current "gold standard," the agar proportion method (11). The performance of a new system should be comparable with that of the BACTEC 460 TB system, with elimination of the two core problems associated with the old BACTEC 460 TB technology, i.e., the risk of needle punctures and disposal of radioactive waste. Preliminary studies utilizing those new systems report good overall agreement of AST results with those generated with established methods (1, 3-5, 8, 10, 12-13, 15).Recent automation of the MGIT 960 technology was another step forward, as it allows continuous monitoring of positive fluorescence, which is based on bacterial growth. It is noninvasive and eliminates potential reading difficulties during visual judging of the tubes, apart from saving labor. The threshold algorithms help in determining the susceptibility automatically.In this multicenter study we have evaluated the reproducibility and reliability of the BACTEC MGIT 960 instrument for testing of M. tuberculosis susceptibility to isoniazid (INH), rifampin (RIF) ethambutol (EMB), and streptomycin (STR) and have compared the results to those obtained by the radiometric procedure. Discordant results were resolved by testing the strains with the agar proportion method using Löwenstein-Jensen (LJ) medium (6). This was done by an additional site which thus acted as an independent arbiter. Last, in order to address safety, we performed drug susceptibility testing in plastic MGITs, in addition to the glass tubes.

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Since its introduction in 1952, pyrazinamide (PZA) has remained one of the most important components in an effective treatment regimen for tuberculosis, mainly affecting semidormant, intracellular Mycobacterium tuberculosis (7). The drug is, however, active at a lower pH value only, making drug susceptibility testing in the clinical mycobacteriology laboratory more demanding. Based on a modified 7H12 medium at pH 6.0, the radiometric BACTEC 460TB technique is, to date, the only culture-based method for PZA susceptibility testing (see the BACTEC 460TB system product and procedure manual [publication MA-0029 of Becton Dickinson and Company, Sparks, Md.] by S. H. Siddiqi) which is recommended by the National Committee for Clinical Laboratory Standards (6).To eliminate the hazards associated with the use of 14 Clabeled substrates and the use of needles by laboratory personnel, nonradiometric culture systems have been established in the past few years which offer, in parallel, rapid, growthbased testing of susceptibility of M. tuberculosis to front-line drugs. Among those novel methods is the MGIT (Mycobacteria Growth Indicator Tube) technology, for which results on susceptibility testing are available for isoniazid (INH), rifampin (RMP), ethambutol (EMB), and streptomycin (STR) (1, 8), but not for PZA, due to the well-known drawbacks of measuring the drug's activity at a more acidic pH.We report here the results of susceptibility testing of M. tuberculosis to PZA performed in the fully automated BACTEC MGIT 960 system (Becton Dickinson), utilizing the novel BACTEC MGIT PZA medium (Becton Dickinson) which contains a modified 7H9 broth with a pH value adjusted to 5.9. Our study consisted of five phases during which results were compared on a one-to-one basis with those generated by the standard PZA susceptibility testing by the BACTEC 460TB system. MATERIALS AND METHODSStudy sites. PZA testing was done by the Swiss National Center for Mycobacteria (center 1) and the German Reference Center for Mycobacteria (center 2). Two additional laboratories (California Department of Health Services, Berkeley, Calif., and VA Medical Center, West Haven, Conn.), acted as independent arbiter sites to which strains were sent for retesting if BACTEC MGIT 960 results were discrepant from BACTEC 460TB results.Reagents. The BACTEC MGIT 960 PZA medium has recently been developed by Becton Dickinson. The tube contains 7 ml of a modified 7H9 broth adjusted to pH 5.9 and 110 l of a fluorescent indicator (Tris 4,7-diphenyl-1,10 phenanthroline ruthenium chloride pentahydrate) in a silicone rubber base. In the PZA kit there are two vials of lyophilized PZA and six vials of BACTEC MGIT PZA supplement containing bovine serum albumin, dextrose, catalase, polyoxyethylene stearate, and oleic acid.Drug concentration. The PZA concentration was 100 g/ml for both BACTEC MGIT 960 and BACTEC 460TB tests.Identification of strains. All M. tuberculosis strains included in this study had been identified by classical biochemical criteria (5).Preparation of inoc...

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