title: Characterization of mutations in streptomycin-resistant Mycobacterium tuberculosis clinical isolates in the area of Barcelona.
abstract: OBJECTIVE
  To determine the proportion and type of mutations in Mycobacterium tuberculosis isolates resistant to streptomycin, and their relationship with the level of resistance and with the epidemiological molecular pattern of the isolates.
  
  
  METHODS
  Sixty-nine streptomycin-resistant isolates from a M. tuberculosis strain collection (1995-2005) from Barcelona were studied. The MIC of streptomycin for each isolate was determined using the proportions method with Middlebrook 7H11 medium. The entire rpsL gene and two specific fragments of the rrs gene (the 530 loop and the 912 region) were sequenced. IS6110-restriction fragment length polymorphism and spoligotyping were performed in each isolate.
  
  
  RESULTS
  Twenty-six (26/69, 37.7%) streptomycin-resistant isolates presented a mutation in either the rpsL gene and/or the rrs530 loop, with no mutation in the rrs912 region. Seventeen (24.6%) isolates showed rpsL mutations (codons 43 and 88) associated with high MIC levels. Nine (13.0%) isolates had alterations in the rrs gene (A513T, A513C and C516T). Nineteen isolates (19/64, 29.7%) were classified into seven clusters (containing 2-5 isolates per cluster). Nineteen different spoligotype patterns were found. All the LAM3 spoligotype isolates (10/67, 14.9%) were associated with a C491T change in the rrs gene, being also observed in all LAM3 streptomycin-susceptible isolates.
  
  
  CONCLUSIONS
  Mutations in the rpsL and rrs genes were detected in 37.7% of streptomycin-resistant M. tuberculosis isolates. High-level resistance was associated with mutations in the rpsL gene, whereas wild-type isolates showed low MIC levels. The presence of the C491T substitution in the rrs gene in streptomycin-susceptible and -resistant isolates demonstrates that this change is an epidemiological marker associated with LAM3 sublineage.
authors.affiliation: Servei de Microbiologia, CDB, Hospital Clínic de Barcelona-IDIBAPS, Universitat de Barcelona (UB), Barcelona, Spain.
authors.forename: Griselda
authors.initials: G
authors.lastname: Tudó
authors.forename: Emma
authors.initials: E
authors.lastname: Rey
authors.forename: Sònia
authors.initials: S
authors.lastname: Borrell
authors.forename: Fernando
authors.initials: F
authors.lastname: Alcaide
authors.forename: Gemma
authors.initials: G
authors.lastname: Codina
authors.forename: Pere
authors.initials: P
authors.lastname: Coll
authors.forename: Núria
authors.initials: N
authors.lastname: Martín-Casabona
authors.forename: Michel
authors.initials: M
authors.lastname: Montemayor
authors.forename: Raquel
authors.initials: R
authors.lastname: Moure
authors.forename: Angels
authors.initials: A
authors.lastname: Orcau
authors.forename: Margarita
authors.initials: M
authors.lastname: Salvadó
authors.forename: Eva
authors.initials: E
authors.lastname: Vicente
authors.forename: Julià
authors.initials: J
authors.lastname: González-Martín
mesh_terms: D000995:Antitubercular Agents / Q000494:pharmacology*; D015373:Bacterial Typing Techniques; D016172:DNA Fingerprinting; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics; D024881:Drug Resistance, Bacterial; D006801:Humans; D008826:Microbial Sensitivity Tests; D020125:Mutation, Missense; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics* / Q000302:isolation & purification; D012269:Ribosomal Proteins / Q000235:genetics; D013030:Spain; D013307:Streptomycin / Q000494:pharmacology*; D014376:Tuberculosis / Q000382:microbiology*
chemical_list: D000995:Antitubercular Agents; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D012269:Ribosomal Proteins; D013307:Streptomycin