pncA gene expression and prediction factors on pyrazinamide resistance in Mycobacterium tuberculosis

Abstract

Summary Background Mutations in the pyrazinamidase (PZAse) coding gene, pncA, have been considered as the main cause of pyrazinamide (PZA) resistance in Mycobacterium tuberculosis. However, recent studies suggest there is no single mechanism of resistance to PZA. The pyrazinoic acid (POA) efflux rate is the basis of the PZA susceptibility Wayne test, and its quantitative measurement has been found to be a highly sensitive and specific predictor of PZA resistance. Based on biological considerations, the POA efflux rate is directly determined by the PZAse activity, the level of pncA expression, and the efficiency of the POA efflux pump system. Objective This study analyzes the individual and the adjusted contribution of PZAse activity, pncA expression and POA efflux rate on PZA resistance. Methods Thirty M. tuberculosis strains with known microbiological PZA susceptibility or resistance were analyzed. For each strain, PZAse was recombinantly produced and its enzymatic activity measured. The level of pncA mRNA was estimated by quantitative RT-PCR, and the POA efflux rate was determined. Mutations in the pncA promoter were detected by DNA sequencing. All factors were evaluated by multiple regression analysis to determine their adjusted effects on the level of PZA resistance. Findings Low level of pncA expression associated to mutations in the pncA promoter region was observed in pncA wild type resistant strains. POA efflux rate was the best predictor after adjusting for the other factors, followed by PZAse activity. These results suggest that tests which rely on pncA mutations or PZAse activity are likely to be less predictive of real PZA resistance than tests which measure the rate of POA efflux. This should be further analyzed in light of the development of alternate assays to determine PZA resistance.

Multicenter Study of the Emergence and Genetic Characteristics of Pyrazinamide-Resistant Tuberculosis in China

“…Most interestingly, the majority of those cases were associated with mutations in three specific regions of the pncA gene (codon 51 to 76, codon 130 to 142, and codon 163 to 180). As previously reported (25), these three pncA regions related to high-level PZA resistance might be associated with the activity of PZase. As structural models of the PZase have revealed, these three specific regions are located in loop 4, loop 7, and loop 10, which have been proven to be the part of PZase active-site region (loop 7) and the metal coordination site (loop 4) (26).…”

The Rate of Drug-Resistant Tuberculosis in Korean Children and Adolescents Since 2007

“…In a recent study, the prevalence of PZA resistance was 0%–9% in cases of drug-sensitive TB, and 31%–89% among MDR-TB isolates. The study found a significant increase in the prevalence of PZA-resistant TB with increased resistance to other drugs (3031). Further studies studying the prevalence, cause, and relation between MDR-TB and PZA-resistant TB are warranted.…”

The Efflux Pump Inhibitor Timcodar Improves the Potency of Antimycobacterial Agents

“…tuberculosis drug-resistant strains have been described to increase the expression of efflux pumps to enable their survival and persistence in the presence of antibiotics (41)(42)(43)(44). Although we did not explore drug-resistant strains in this study, additional research to try and reverse drug resistance using TIM and clinical isolates of M. tuberculosis and evaluation of these in broth and infected macrophage cultures, as well as in vivo studies, are warranted.…”