title: Accurate mapping of mutations of pyrazinamide-resistant Mycobacterium tuberculosis strains with a scanning-frame oligonucleotide microarray.
abstract: The increasing emergence of drug-resistant Mycobacterium tuberculosis poses significant threat to the treatment of tuberculosis. Conventional susceptibility testing for the front-line tuberculosis drug pyrazinamide (PZA) is difficult, because of the requirement for acid pH for the drug to show activity. Resistance to PZA in M. tuberculosis is caused by mutations in the pncA gene, and detection of pncA mutations can be an indicator of PZA resistance. In this study, we examined the feasibility of a microarray-based approach exploiting short overlapping oligonucleotides (sliding-frame array) to rapidly detect pncA mutations (substitutions, deletions, and insertions) in multiple strains of PZA-resistant M. tuberculosis. The genetic mapping of these mutations is necessary to link the gene sequence to the protein function defined by mutant phenotype. Microarray analysis was performed in a blind manner using 57 isolates of M. tuberculosis for which the sequence of the pncA gene was previously determined. Our results showed that all mutations could be unambiguously detected, suggesting that microarray can be a routine and valuable tool for rapid identification of drug-resistant M. tuberculosis isolates. We expect that mutation mapping with a sliding-frame microarray will accelerate the molecular analysis of drug-resistant M. tuberculosis bacteria and the microorganism populations.
authors.affiliation: Department of Molecular Microbiology and Immunology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, MD, USA.
authors.forename: Mary Margaret
authors.initials: MM
authors.lastname: Wade
authors.forename: Dmitriy
authors.initials: D
authors.lastname: Volokhov
authors.forename: Mike
authors.initials: M
authors.lastname: Peredelchuk
authors.forename: Vladimir
authors.initials: V
authors.lastname: Chizhikov
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhang
mesh_terms: D004269:DNA, Bacterial / Q000032:analysis; D024881:Drug Resistance, Bacterial; D020869:Gene Expression Profiling; D006801:Humans; D008826:Microbial Sensitivity Tests; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000187:drug effects / Q000235:genetics* / Q000302:isolation & purification; D020411:Oligonucleotide Array Sequence Analysis; D010597:Pharmacogenetics; D011718:Pyrazinamide / Q000494:pharmacology* / Q000627:therapeutic use; D012494:Sampling Studies; D012680:Sensitivity and Specificity; D018088:Tuberculosis, Multidrug-Resistant / Q000175:diagnosis / Q000188:drug therapy
chemical_list: D004269:DNA, Bacterial; D011718:Pyrazinamide