title: Characterisation of rpsL, rrs and embB mutations associated with streptomycin and ethambutol resistance in Mycobacterium tuberculosis.
abstract: In order to characterise molecular mechanisms of first-line drug resistance in Mycobacterium tuberculosis and to evaluate the use of molecular markers of resistance (gene point mutations), we analysed 66 multi-drug-resistant (MDR) isolates from Latvian tuberculosis patients. They were all resistant to rifampin (RIF), isoniazid (INH) and streptomycin (SM), and 33 were resistant to ethambutol (EMB). Enzymatic digestion by MboII and nucleotide sequencing of the rpsL gene fragment detected a single nucleotide substitution K43R in 40 (61%) of the 66 SM-resistant M. tuberculosis isolates. Of the other 26 SM-resistant isolates, 16 (24%) had mutations at positions 513A-->C and 516C-->T of the rrs gene and 10 (15%) had the wild-type sequence. The single-stranded DNA conformation polymorphism (SSCP) method was used to detect mutations in the embB gene associated with EMB resistance. Substitutions in the embB gene were found by SSCP analysis in 15 (45%) and by sequencing in 17 (52%) of the 33 EMB-resistant isolates. Surprisingly, SSCP revealed a nucleotide mutation at codon M306 in five (15%) of 33 in vitro EMB-susceptible MDR isolates.
authors.affiliation: Laboratory of Molecular Microbiology, Biomedical Research and Study Centre, University of Latvia, Ratsupites 1, Riga LV 1067, Latvia. tatjanap@biomed.lu.lv
authors.forename: Tatjana
authors.initials: T
authors.lastname: Tracevska
authors.forename: Inta
authors.initials: I
authors.lastname: Jansone
authors.forename: Anda
authors.initials: A
authors.lastname: Nodieva
authors.forename: Olgerts
authors.initials: O
authors.lastname: Marga
authors.forename: Girts
authors.initials: G
authors.lastname: Skenders
authors.forename: Viesturs
authors.initials: V
authors.lastname: Baumanis
mesh_terms: D000890:Anti-Infective Agents / Q000494:pharmacology*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D004977:Ethambutol / Q000494:pharmacology*; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics*; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D012336:RNA, Ribosomal, 16S / Q000737:chemistry / Q000235:genetics; D012269:Ribosomal Proteins / Q000737:chemistry / Q000235:genetics; D017422:Sequence Analysis, DNA; D013307:Streptomycin / Q000494:pharmacology*; D014376:Tuberculosis / Q000188:drug therapy / Q000382:microbiology*
chemical_list: D000890:Anti-Infective Agents; D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S; D012269:Ribosomal Proteins; C034212:ribosomal protein S12; D004977:Ethambutol; C024413:endodeoxyribonuclease MboII; D015252:Deoxyribonucleases, Type II Site-Specific; D013307:Streptomycin