title: Mutation of tlyA confers capreomycin resistance in Mycobacterium tuberculosis.
abstract: Capreomycin, an important drug for the treatment of multidrug-resistant tuberculosis, is a macrocyclic peptide antibiotic produced by Saccharothrix mutabolis subspecies capreolus. The basis of resistance to this drug was investigated by isolating and characterizing capreomycin-resistant strains of Mycobacterium smegmatis and Mycobacterium tuberculosis. Colonies resistant to capreomycin were recovered from a library of transposon-mutagenized M. smegmatis. The transposon insertion site of one mutant was mapped to an open reading frame in the unfinished M. smegmatis genome corresponding to the tlyA gene (Rv1694) in the M. tuberculosis H37Rv genome. In M. smegmatis spontaneous capreomycin-resistant mutants, the tlyA gene was disrupted by one of three different naturally occurring insertion elements. Genomic DNAs from pools of transposon mutants of M. tuberculosis H37Rv were screened by PCR by using primers to the tlyA gene and the transposon to detect mutants with an insertion in the tlyA gene. One capreomycin-resistant mutant was recovered that contained the transposon inserted at base 644 of the tlyA gene. Complementation with the wild-type tlyA gene restored susceptibility to capreomycin in the M. smegmatis and M. tuberculosis tlyA transposon mutants. Mutations were found in the tlyA genes of 28 spontaneous capreomycin-resistant mutants generated from three different M. tuberculosis strains and in the tlyA genes of capreomycin-resistant clinical isolates. In in vitro transcription-translation assays, ribosomes from tlyA mutant but not tlyA(+) strains resist capreomycin inhibition of transcription-translation. Therefore, TlyA appears to affect the ribosome, and mutation of tlyA confers capreomycin resistance.
authors.affiliation: Program in Microbiology and Molecular Genetics, Emory University, Atlanta, Georgia, USA.
authors.forename: Courtney E
authors.initials: CE
authors.lastname: Maus
authors.forename: Bonnie B
authors.initials: BB
authors.lastname: Plikaytis
authors.forename: Thomas M
authors.initials: TM
authors.lastname: Shinnick
mesh_terms: D000904:Antibiotics, Antitubercular / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics*; D015139:Blotting, Southern; D002207:Capreomycin / Q000494:pharmacology*; D003001:Cloning, Molecular; D004251:DNA Transposable Elements / Q000235:genetics; D004269:DNA, Bacterial / Q000096:biosynthesis / Q000235:genetics; D024881:Drug Resistance, Bacterial; D015723:Gene Library; D006460:Hemolysin Proteins / Q000235:genetics*; D006801:Humans; D008826:Microbial Sensitivity Tests; D016296:Mutagenesis; D009154:Mutation / Q000235:genetics*; D020102:Mycobacterium smegmatis / Q000187:drug effects / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics*; D010957:Plasmids; D014176:Protein Biosynthesis; D012329:RNA, Bacterial / Q000235:genetics; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic; D014376:Tuberculosis / Q000382:microbiology; D014756:Viomycin / Q000494:pharmacology
chemical_list: D000904:Antibiotics, Antitubercular; D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D006460:Hemolysin Proteins; D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S; C112817:TlyA protein, Mycobacterium tuberculosis; D002207:Capreomycin; D014756:Viomycin