RESUMO
The plasmid-borne mph(A) gene that confers resistance to azithromycin and has recently emerged in Shigella sonnei is present in multidrug- and non-multidrug-resistant Escherichia coli isolates from 4 continents. Further spread of mph(A) to Shigella and Salmonella spp. may be expected.
Assuntos
Farmacorresistência Bacteriana Múltipla/genética , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Macrolídeos/farmacologia , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Regulação Bacteriana da Expressão Gênica , Testes de Sensibilidade MicrobianaRESUMO
OBJECTIVES: To determine the prevalence of macrolide and streptogramin resistance in Streptococcus mitis isolates from neutropenic patients and to identify mechanisms of macrolide and streptogramin resistance in resistant isolates. METHODS: MICs of erythromycin, spiramycin, lincomycin and pristinamycin were determined for S. mitis isolates. Macrolide-resistance genes were characterized by PCR and ribosomal mutations by sequencing. RESULTS: A total of 169 S. mitis isolates were recovered from 66 patients at the Tunisian Bone Marrow Transplant Centre. Of these, 120 (70%) were non-susceptible to erythromycin and one was resistant to pristinamycin; 48.5% of isolates had an MLSB phenotype with cross-resistance between erythromycin, spiramycin and lincomycin, 4% had a dissociated MLSB phenotype with resistance to erythromycin and spiramycin but apparent susceptibility to lincomycin and 47.5% displayed the M phenotype. Resistance determinants were characterized in 33 isolates. Ten of 14 isolates with the cross MLSB resistance contained an ermB-like gene and four a combination of ermB- and mefA-like genes. Four of the five isolates with a dissociated MLSB phenotype contained ermB-like and one a combination of ermB- and mefA-like genes. All the 14 isolates with an M phenotype contained mefA-like genes. The pristinamycin-resistant strain had G105 and A108 substitutions in the conserved C terminus of the L22 ribosomal protein. CONCLUSIONS: The prevalence of macrolide resistance is high in S. mitis from neutropenic patients and is due to the spread of ermB- or mefA-like genes alone or combined. Resistance to streptogramins is rare and in this case associated with ribosomal mutation.
Assuntos
Antibacterianos/farmacologia , Macrolídeos/farmacologia , Neutropenia/microbiologia , Infecções Estreptocócicas/microbiologia , Streptococcus mitis/efeitos dos fármacos , Streptococcus mitis/genética , Estreptograminas/farmacologia , Sequência de Aminoácidos , Transplante de Medula Óssea , Primers do DNA , Farmacorresistência Bacteriana , Eritromicina/farmacologia , Genótipo , Humanos , Hospedeiro Imunocomprometido , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Fenótipo , Pristinamicina/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Mutations in genes coding for L4 (rplD) or L22 (rplV) ribosomal proteins or in 23S rRNA (rrl gene) are reported as a cause of macrolide resistance in streptococci and staphylococci. This study was aimed at evaluating a denaturing high-performance liquid chromatography (DHPLC) technique as a rapid mutation screening method. Portions of these genes were amplified by PCR from total DNA of 48 strains of Streptococcus pneumoniae (n = 22), Staphylococcus aureus (n = 16), Streptococcus pyogenes (n = 6), Streptococcus oralis (n = 2), and group G streptococcus (n = 2). Thirty-seven of these strains were resistant to macrolides and harbored one or several mutations in one or two of the target genes, and 11 were susceptible. PCR products were analyzed by DHPLC. All mutations were detected, except a point mutation in a pneumococcal rplD gene. The method detected one mutated rrl copy out of six in S. aureus. This automated method is promising for screening of mutations involved in macrolide resistance in gram-positive cocci.
Assuntos
Cocos Gram-Positivos/genética , Macrolídeos/farmacologia , Mutação/genética , Desnaturação Proteica/genética , Ribossomos/genética , Cromatografia Líquida de Alta Pressão , Farmacorresistência Bacteriana/genética , Deleção de Genes , Fosfolipase D/genética , RNA Bacteriano/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Staphylococcus aureus/genética , Streptococcus/genéticaRESUMO
The mechanism of resistance to the streptogramin antibiotics quinupristin and dalfopristin was studied in a Staphylococcus aureus clinical isolate selected under quinupristin-dalfopristin therapy, in four derivatives of S. aureus RN4220 selected in vitro, and in a mutant selected in a model of rabbit aortic endocarditis. For all strains the MICs of erythromycin, quinupristin, and quinupristin-dalfopristin were higher than those for the parental strains but the MICs of dalfopristin and lincomycin were similar. Portions of genes for domains II and V of 23S rRNA and the genes for ribosomal proteins L4 and L22 were amplified and sequenced. All mutants contained insertions or deletions in a protruding beta hairpin that is part of the conserved C terminus of the L22 protein and that interacts with 23S rRNA. Susceptible S. aureus RN4220 was transformed with plasmid DNA encoding the L22 alteration, resulting in transformants that were erythromycin and quinupristin resistant. Synergistic ribosomal binding of streptogramins A and B, studied by analyzing the fluorescence kinetics of pristinamycin I(A)-ribosome complexes, was abolished in the mutant strain, providing an explanation for quinupristin-dalfopristin resistance.
Assuntos
Proteínas de Bactérias/genética , Resistência Microbiana a Medicamentos/genética , Quimioterapia Combinada/farmacologia , Mutação , Proteínas de Ligação a RNA/genética , Proteínas Ribossômicas , Staphylococcus aureus/efeitos dos fármacos , Virginiamicina/farmacologia , Sequência de Aminoácidos , Humanos , Dados de Sequência Molecular , RNA Ribossômico 23S/genética , Proteínas de Ligação a RNA/química , Ribossomos/metabolismo , Staphylococcus aureus/genéticaRESUMO
Mechanisms of resistance were studied in 22 macrolide-resistant mutants selected in vitro from 5 parental strains of macrolide-susceptible Streptococcus pneumoniae by serial passage in various macrolides (T. A. Davies, B. E. Dewasse, M. R. Jacobs, and P. C. Appelbaum, Antimicrob. Agents Chemother., 44:414-417, 2000). Portions of genes encoding ribosomal proteins L22 and L4 and 23S rRNA (domains II and V) were amplified by PCR and analyzed by single-strand conformational polymorphism analysis to screen for mutations. The DNA sequences of amplicons from mutants that differed from those of parental strains by their electrophoretic migration profiles were determined. In six mutants, point mutations were detected in the L22 gene (G95D, P99Q, A93E, P91S, and G83E). The only mutant selected by telithromycin (for which the MIC increased from 0.008 to 0.25 microg/ml) contained a combination of three mutations in the L22 gene (A93E, P91S, and G83E). L22 mutations were combined with an L4 mutation (G71R) in one strain and with a 23S rRNA mutation (C2611A) in another strain. Nine other strains selected by various macrolides had A2058G (n = 1), A2058U (n = 2), A2059G (n = 1), C2610U (n = 1), and C2611U (n = 4) mutations (Escherichia coli numbering) in domain V of 23S rRNA. One mutant selected by clarithromycin and resistant to all macrolides tested (MIC, >32 microg/ml) and telithromycin (MIC, 4 microg/ml) had a single base deletion (A752) in domain II. In six remaining mutants, no mutations in L22, L4, or 23S rRNA could be detected.
