ObjectivesTo characterize 250 drug-resistant Mycobacterium tuberculosis (MTB) isolates in Hong Kong with respect to their drug susceptibility phenotypes to five common anti-tuberculosis drugs (ofloxacin, rifampicin, ethambutol, isoniazid and pyrazinamide) and the relationship between such phenotypes and the patterns of genetic mutations in the corresponding resistance genes (gyrA, rpoB, embB, katG, inhA, ahpC and pncA).MethodsThe MIC values of the aforementioned anti-tuberculosis drugs were determined for each of the 250 drug-resistant MTB clinical isolates by the absolute concentration method. Genetic mutations in the corresponding resistance genes in these MTB isolates were identified by PCR-single-stranded conformation polymorphism/multiplex PCR amplimer conformation analysis (SSCP/MPAC), followed by DNA sequencing of the purified PCR products.ResultsResistance to four or five drugs was commonly observed in these MTB isolates; such phenotypes accounted for over 34% of the 250 isolates. The most frequently observed phenotypes were those involving both rifampicin and isoniazid, with or without additional resistance to the other drugs. A total of 102 novel mutations, which accounted for 80% of all mutation types detected in the 7 resistance genes, were recovered. Correlation between phenotypic and mutational data showed that genetic changes in the gyrA, rpoB and katG genes were more consistently associated with a significant resistance phenotype. Despite this, however, a considerable proportion of resistant MTB isolates were found to harbour no detectable mutations in the corresponding gene loci.ConclusionsThese findings expand the spectrum of potential resistance-related mutations in MTB clinical isolates and help consolidate the framework for the development of molecular methods for delineating the drug susceptibility profiles of MTB isolates in clinical laboratories.

Pleural fluid IL-6, TNF-alpha and IFN-gamma assays are useful in the diagnosis of pleural TB. The initial IL-6 level correlates with the number of febrile days. The percentage change of cytokines after 2 weeks of treatment also helps to predict residual pleural scarring.

The postantibiotic effects (PAEs) of seven antimycobacterial agents, tested at their respective peak concentrations in serum alone and in different combinations, against Mycobacterium tuberculosis ATCC 27294 were studied with a radiometric culture system in parallel with the viable count method. Rifampin gave the longest PAE (67.8 h) among the drugs used alone, and combinations of first-line drugs generally gave PAEs longer than 120 h. The data obtained might help provide a better understanding of the scientific basis of intermittently administered antituberculosis chemotherapy.Postantibiotic effect (PAE) refers to the continued suppression of bacterial growth following limited exposure of organisms to an antimicrobial agent (1,15,21). A prolonged PAE may allow wider dosing intervals without the loss of therapeutic efficacy (3). For the treatment of tuberculosis, administration of drugs at wider intervals would reduce the costs and toxicities of drugs and facilitate directly observed antituberculosis chemotherapy, thus enhancing patient adherence (8) and leading to a favorable outcome. Earlier work on pulsed exposure to isoniazid, rifampin, and pyrazinamide for 6 to 96 h has provided hints about the suitability of intermittent administration of these drugs for the treatment of tuberculosis (4-7, 16). In an attempt to better understand the scientific basis of the efficacies of certain established intermittently administered antituberculosis regimens, as well as to gain pertinent knowledge about newer antimycobacterial agents, we embarked on a study that addresses the PAEs of various antituberculosis drugs in vitro.The standard strain of Mycobacterium tuberculosis chosen for the study, strain H37Rv (ATCC 27294), was susceptible to all drugs tested. The MICs of all single drugs except pyrazinamide were determined by the broth macrodilution method (25), while susceptibility testing with pyrazinamide was done by the absolute concentration method (12). Altogether, seven antituberculosis drugs were assessed by using concentrations equivalent to their respective peak concentrations in serum (C max ) in humans (17): amikacin (32 mg/liter), ethambutol (5 mg/ liter), isoniazid (4 mg/liter), ofloxacin (8 mg/liter), rifampin (16 mg/liter), streptomycin (40 mg/liter), and pyrazinamide (60 mg/liter). Stock solutions of the drugs (all drugs except ofloxacin were purchased from Sigma, St. Louis, Mo.; ofloxacin was a gift from Daiichi, Tokyo, Japan) were prepared in appropriate solvents, stored at Ϫ70°C in 1.0-ml aliquots, and used within 6 months. For each experiment, aliquots of the stock solutions were thawed and subsequently diluted in Middlebrook 7H9 broth supplemented with 2% glycerol and 10% oleic acid-dextrose-catalase (Difco Laboratories).To determine the PAE, a homogeneous suspension of cells whose turbidity matched that of a no. 1 McFarland standard was obtained from a 3-week-old culture and stored at Ϫ40°C in 1.0-ml aliquots. For each experiment, a single vial of cells was quickly thawed at 37°C and inoculated into 10...