Spoligotyping and Drug Resistance Analysis of Mycobacterium tuberculosis Strains from National Survey in China

Section: Discussionsupporting

confidence: 87%

Prevalence and Molecular Characteristics of Drug-Resistant Mycobacterium tuberculosis in Hunan, China

Zhao

Chen

et al. 2014

“…A total of 3,634 smear-positive patients were included in the study (15); 145 isolates were identified as OFX resistant by conventional DST. Out of the 145 OFX-resistant strains, 138 could be subcultured successfully for subsequent MIC analysis and DNA sequencing of the entire gyrA and gyrB genes.…”

Section: Resultsmentioning

confidence: 99%

“…OFX-resistant M. tuberculosis strains, identified by conventional susceptibility testing, were all obtained from a national tuberculosis drug resistance survey of China conducted in 2007 (10,15). The M. tuberculosis strains isolated from clusters were all transferred to the National Tuberculosis Reference Laboratory (NTRL), and drug susceptibility was determined by the proportion method in the NTRL with an OFX concentration of 0.2 g/ml.…”

Section: Methodsmentioning

confidence: 99%

Prevalence and Molecular Characterization of Fluoroquinolone-Resistant Mycobacterium tuberculosis Isolates in China

Zhang

Wang

et al. 2014

Self Cite

China is one of the countries with the highest burdens of multidrug-resistant (MDR) and fluoroquinolone (FQ)-resistant tuberculosis (TB) globally. Nevertheless, knowledge about the prevalence and molecular characterization of FQ-resistant Mycobacterium tuberculosis isolates from this region remains scant. In this study, 138 M. tuberculosis isolates determined by the agar proportion susceptibility method to be resistant to ofloxacin (OFX) were enrolled from a national drug resistance survey of China. All these strains were tested for susceptibility to ofloxacin, levofloxacin, moxifloxacin, gatifloxacin, and sparfloxacin using liquid Middlebrook 7H9 medium. The entire gyrA and gyrB genes conferring FQ resistance were sequenced, and spoligotyping was performed to distinguish different genotypes. Overall, the prevalence of resistance in China was highest for ofloxacin (3.76%), intermediate for levofloxacin (3.18%) and moxifloxacin (3.12%), and lowest for sparfloxacin (1.91%) and gatifloxacin (1.33%). Mutations in the gyrA gene were observed in 89 (64.5%) out of the 138 OFX-resistant M. tuberculosis strains. Positions 94 and 90 were the most frequent sites of mutation conferring FQ resistance on these strains, accounting for high-level FQ resistance. Furthermore, the Beijing genotype showed no association with high-level FQ resistance or distribution in hot spots in the quinolone resistance-determining region (QRDR) of gyrA. Our findings provide essential implications for the feasibility of genotypic tests relying on detection of mutations in the QRDR of gyrA and the shorter first-line treatment regimens based on FQs in China.

“…The spoligotyping method was performed using primers DRa and DRb as previously reported (23). ECL enhanced chemiluminescence detection liquid was used to detect hybridizing DNA, followed by exposure to X-ray film.…”

Section: Methodsmentioning

confidence: 99%

Study of the Rifampin Monoresistance Mechanism in Mycobacterium tuberculosis

Pang

Wang

et al. 2013

Self Cite

102

b Rifampin (RIF) susceptibility is a key factor in determining the treatment effectiveness of the standardized treatment regimens. In Mycobacterium tuberculosis, both target gene mutation and the efflux pump play major roles in the resistance to antituberculosis drugs. By eliminating RIF-resistant strains with rpoB mutation, the choice of RIF-monoresistant strains may allow us to identify the RIF-specific efflux pump genes. This study explored the RIF monoresistance mechanism in M. tuberculosis. Data from DNA sequencing and MIC measurements revealed that specific mutations, including Ser531Leu and His526Asp in RpoB, show high-level drug resistance. Three-dimensional structure modeling provided further evidence that the affinity between RIF and RpoB mutants was in accordance with the drug resistance level of the corresponding isolates. Furthermore, transcriptionlevel analysis among the nonmutated isolates indicated that three efflux pumps (Rv0783, Rv2936, and Rv0933) might be involved in exporting RIF from the cell. Compared to 8 g/ml for wild-type Escherichia coli, the MICs for the transgenic E. coli strains with either Rv0783 or Rv2936 were 32 and 16 g/ml, respectively. In conclusion, our study indicated that several RpoB mutant types, including Ser531Leu and His526Asp, show high-level RIF resistance attributed to low affinity between RpoB mutant proteins and RIF. In addition, this work demonstrates that Rv2936 and Rv0783 may be responsible for low-level resistance to RIF by exporting RIF from cells. The predicted structure of RpoB and the newly identified efflux pumps in this study will provide a novel approach to design new drugs and develop novel diagnosis technologies.