A particular strain of M. abscessus ssp. massiliense was associated with a prolonged epidemic of postsurgical infections in seven Brazilian states, suggesting that this strain may be distributed in Brazilian territory and better adapted to cause surgical-site infections.

Mutations related to streptomycin resistance in the rpsL and rrs genes are well known and can explain about 70% of this phenotypic resistance. Recently, the gidB gene was found to be associated with low-level streptomycin resistance in Mycobacterium tuberculosis. Mutations in gidB have been reported with high frequency, and this gene appears to be very polymorphic, with frameshift and point mutations occurring in streptomycinsusceptible and streptomycin-resistant strains. In this study, mutations in gidB appeared in 27% of streptomycin-resistant strains that contained no mutations in the rpsL or rrs genes, and they were associated with low-level streptomycin resistance. However, the association of certain mutations in gidB with streptomycin resistance needs to be further investigated, as we also found mutations in gidB in streptomycin-susceptible strains. This occurred only when the strain was resistant to rifampin and isoniazid. Two specific mutations appeared very frequently in this and other studies of streptomycin-susceptible and -resistant strains; these mutations were not considered related to streptomycin resistance, but as a polymorphism. We stratified the strains according to the different phylogenetic lineages and showed that the gidB 16 polymorphism (16G allele) was exclusively present in the Latin American-Mediterranean (LAM) genotype, while the gidB 92 polymorphism (92C allele) was associated with the Beijing lineage in another population. In the sample studied, the two characterized single-nucleotide polymorphisms could distinguish LAM and Beijing lineages from the other lineages.

The MIC for streptomycin in the presence of efflux pump (EP) inhibitors and the sequencing of rpsL, rrs, and gidB genes provided evidence for the possible participation of EP in low-level streptomycin (STR) resistance of some isolates without mutations. Mutation in the gidB gene and an EP could act synergistically to confer low STR resistance.Streptomycin (STR), the first antibiotic used to control tuberculosis, acts by binding to the 30S ribosomal subunit, inhibiting polypeptide synthesis (5, 24). Mutations in genes rpsL and rrs, encoding the ribosomal protein S12 and the 16S rRNA, respectively, are responsible for most of the high-level STR resistance in Mycobacterium tuberculosis (7,9,24). However, a low level of resistance is found in approximately one-third of the clinical isolates resistant to STR without mutations in these genes (7,11,17). Recently, a new STR resistance locus (gidB) that encodes a conserved 7-methylguanosine methyltransferase specific for the 16S rRNA was found to confer a low level of STR resistance (12).Also possibly involved with a low level of STR resistance is the efflux system. This system can grant to bacteria a mechanism which favors their survival in a hostile environment (e.g., the presence of antibiotics), and those bacteria that overexpress efflux pumps (EPs) can be selected (23). It has been shown that the M. tuberculosis genome encodes multiple putative EPs (2, 6), and reports have suggested that EPs may also be involved in transporting fluoroquinolones, aminoglycosides, tetracycline, and possibly isoniazid and ethambutol (4,14,18,22).In this study we have evaluated the possible role of the efflux mechanism as a molecular basis of STR resistance in clinical isolates of M. tuberculosis, using carbonyl cyanide m-chlorophenylhydrazone (CCCP) and verapamil as efflux pump inhibitors (EPI) (1, 10).A total of 79 clinical isolates from the state of Rio Grande do Sul, Brazil, were studied. Resistance to drugs other than STR was found in 62 isolates (see Table S1 in the supplemental material). The resazurin microtiter assay (13) was used for MIC determination. Microplates were divided into three equal parts: one with 7H9-oleic acid-albumin-dextrose-catalase medium, one with medium and verapamil (100 M), and one with medium and CCCP (11 M). Serial 1:2 dilutions of STR (250 g/ml to 0.25 g/ml) were performed in each column. The breakpoint to determine STR resistance was a MIC of Ն8 g/ml.DNA was isolated from mycobacterial cultures by the lysozyme/proteinase K cetyltrimethylammonium bromide procedure (21). A 306-bp fragment of the M. tuberculosis rpsL gene (GenBank accession number L08011) and the 530 loop (238 bp) and 912 region (238 bp) of the rrs gene (GenBank accession number X52917) were amplified as described by Tracevska et al. (20). A 675-bp fragment of the gene gidB (GenBank accession number AAK48404) was amplified using primers gidB1 (5ЈGTCCCTCCACTCGCCATC3Ј) and gidB2 (5ЈGCGGAGTGCGTAATGTCTC3Ј). PCR amplification was performed by the following steps: initial denaturation at 92°C ...

The presence of mutations in specific regions of the katG, inhA, and ahpC genes was analyzed with 69 Mycobacterium tuberculosis isoniazid-resistant isolates from three Brazilian states. Point mutations in codon 315 of the katG gene were observed in 87.1, 60.9, and 60% of the isolates from Rio Grande do Sul, Rio de Janeiro, and São Paulo, respectively. Mutations in the inhA gene were identified only in one isolate from RJ State, and the ahpC promoter region revealed mutations in distinct positions in 12.9, 21.7, and 6.7% of the isolates from RS, RJ and SP, respectively.

BackgroundMutations associated with resistance to rifampin or streptomycin have been reported for W/Beijing and Latin American Mediterranean (LAM) strain families of Mycobacterium tuberculosis. A few studies with limited sample sizes have separately evaluated mutations in katG, ahpC and inhA genes that are associated with isoniazid (INH) resistance. Increasing prevalence of INH resistance, especially in high tuberculosis (TB) prevalent countries is worsening the burden of TB control programs, since similar transmission rates are noted for INH susceptible and resistant M. tuberculosis strains.ResultsWe, therefore, conducted a comprehensive evaluation of INH resistant M. tuberculosis strains (n = 224) from three South American countries with high burden of drug resistant TB to characterize mutations in katG, ahpC and inhA gene loci and correlate with minimal inhibitory concentrations (MIC) levels and spoligotype strain family. Mutations in katG were observed in 181 (80.8%) of the isolates of which 178 (98.3%) was contributed by the katG S315T mutation. Additional mutations seen included oxyR-ahpC; inhA regulatory region and inhA structural gene. The S315T katG mutation was significantly more likely to be associated with MIC for INH ≥2 μg/mL. The S315T katG mutation was also more frequent in Haarlem family strains than LAM (n = 81) and T strain families.ConclusionOur data suggests that genetic screening for the S315T katG mutation may provide rapid information for anti-TB regimen selection, epidemiological monitoring of INH resistance and, possibly, to track transmission of INH resistant strains.

The occurrence of tuberculosis (TB)

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