Molecular genetics of Mycobacterium tuberculosis resistant to aminoglycosides and cyclic peptide capreomycin antibiotics in Korea
ABSTRACT Aminoglycosides are key drugs for the treatment of multidrug-resistant tuberculosis. A total of 97 extensively drug-resistant (XDR) and 29 pan-susceptible Mycobacterium tuberculosis isolates from Korean tuberculosis patients were analyzed to characterize mutations within the rrs, rpsL, gidB, eis and tlyA genes. Thirty (56.6 %) of the 53 streptomycin (STR)-resistant strains had a rpsL mutation and eight strains (15.1 %) had a rrs (514 or 908 site) mutation, whereas 11 (20.8 %) of the 53 STR-resistant strains had a gidB mutation without rpsL or either rrs mutation. Most of the gidB mutations conferred low-level STR resistance, and 22 of these mutations were novel. Mutation at position 1401 in rrs lead to resistance to kanamycin (80/95 = 84.2 %; KAN), amikacin (80/87 = 92.0 %; AMK), and capreomycin (74/86 = 86.0 %; CAP). In this study, 13.7 % (13/95) of KAN-resistant strains showed eis mutations, including 4 kinds of novel mutations. Isolates with eis structural gene mutations were cross-resistant to STR, KAN, CAP, and AMK. Here, 5.8 % (5/86) of the CAP-resistant strains harbored a tlyA mutation that included 3 different novel point mutations. Detection of the A1401G mutation appeared to be 100 % specific for the detection of resistance to KAN and AMK. These data establish the presence of phenotypic XDR strains using molecular profiling and are helpful to understanding of aminoglycoside resistance at the molecular level.
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ABSTRACT: Currently, mutations in three genes, namely rrs, rpsL, and gidB, encoding 16S rRNA, ribosomal protein S12, and 16S rRNA-specific methyltransferase, respectively, are considered to be involved in conferring resistance to streptomycin (STR) in Mycobacterium tuberculosis. The aim of this study was to investigate the spectrum and frequency of these mutations in M. tuberculosis clinical isolates, both resistant and susceptible to STR. Sixty-four M. tuberculosis isolates recovered from as many TB patients from Poland in 2004 were included in the study. Within the sample were 50 multidrug-resistant (32 STR-resistant and 18 STR-susceptible) and 14 pan-susceptible isolates. Preliminary testing for STR resistance was performed with the 1% proportion method. The MICs of STR were determined by the Etest method. Mutation profiling was carried out by amplifying and sequencing the entire rrs, rpsL, and gidB genes. Non-synonymous mutations in either rrs or rpsL gene were detected in 23 (71.9%) of the STR-resistant and none of the STR-susceptible isolates. Mutations in the gidB gene were distributed among 12 (37.5%) STR-resistant and 13 (40.6%) STR-susceptible isolates. Four (12.5%) STR-resistant isolates were wild-type at all three loci examined. None of the rrs, rpsL or gidB mutations could be linked to low, intermediate or high level of STR resistance. In accordance with previous findings, the gidB 47T→G (L16R) mutation was associated with the Latin American-Mediterranean genotype family, whereas 276A→C (E92D) and 615A→G (A205A) mutations of the gidB gene were associated with the Beijing lineage. The study underlines the usefulness of rrs and rpsL mutations as molecular markers for STR resistance yet not indicative of its level. The gidB polymorphisms can serve as phylogenetic markers.PLoS ONE 06/2014; 9(6):e100078. DOI:10.1371/journal.pone.0100078 · 3.53 Impact Factor
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ABSTRACT: As the prevalence of multidrug-resistant and extensively drug-resistant tuberculosis strains continues to rise, so does the need to develop accurate and rapid molecular tests to complement time consuming growth-based drug susceptibility testing. Performance of molecular methods relies on the association of specific mutations with phenotypic drug resistance and while considerable progress has been made for resistance detection of first-line antituberculosis drugs, rapid detection of resistance for second-line drugs lags behind. The rrs A1401G allele is considered a strong predictor of cross-resistance between the three second-line injectable drugs, capreomycin (CAP), kanamycin, and amikacin. However, discordance is often observed between the rrs A1401G mutation and CAP resistance, with up to 40% of rrs A1401G mutants being classified as CAP susceptible. We measured the minimal inhibitory concentrations (MICs) to CAP in 53 clinical isolates harboring the A1401G mutation and found that the CAP MICs ranged from 8 μg/ml to 40 μg/ml. These results were drastically different from engineered A1401G mutants generated in isogenic Mycobacterium tuberculosis, which exclusively exhibited high-level CAP MICs of 40 μg/ml. These data support prior studies suggesting the critical concentration of CAP (10 μg/ml) used to determine resistance by indirect agar proportion may be too high to detect all CAP resistant strains and suggests that a larger percentage of resistant isolates could be identified by lowering the critical concentration. These data also suggest that differences in resistance levels among clinical isolates are possibly due to second site or compensatory mutations located elsewhere in the genome.Antimicrobial Agents and Chemotherapy 11/2014; 59(1). DOI:10.1128/AAC.04438-14 · 4.57 Impact Factor
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ABSTRACT: The emergence of multidrug-resistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) makes the treatment and control of tuberculosis difficult. Rapid detection of drug-resistant strains is important for the successful treatment of drug-resistant tuberculosis; however, not all resistance mechanisms to the injectable second-line drugs such as amikacin (AK), kanamycin (KM), and capreomycin (CAP) are well understood. This study aims to validate the mechanisms associated with AK, KM, and CAP resistance in M. tuberculosis clinical strains isolated in Thailand.BMC Microbiology 06/2014; 14(1):165. DOI:10.1186/1471-2180-14-165 · 2.98 Impact Factor