Gyrase Mutations Are Associated with Variable Levels of Fluoroquinolone Resistance in Mycobacterium tuberculosis

Abstract

Molecular diagnostics that rapidly and accurately predict resistance to fluoroquinolone drugs and especially later-generation agents promise to improve treatment outcomes for patients with multidrug-resistant tuberculosis and prevent the spread of disease. Mutations in the gyr genes are known to confer most fluoroquinolone resistance, but knowledge about the effects of gyr mutations on susceptibility to early- versus later-generation fluoroquinolones and about the role of mutation-mutation interactions is limited. Here, we sequenced the full gyrA and gyrB open reading frames in 240 multidrug-resistant and extensively drug-resistant tuberculosis strains and quantified their ofloxacin and moxifloxacin MIC by testing growth at six concentrations for each drug. We constructed a multivariate regression model to assess both the individual mutation effects and interactions on the drug MICs. We found that gyrB mutations contribute to fluoroquinolone resistance both individually and through interactions with gyrA mutations. These effects were statistically significant. In these clinical isolates, several gyrA and gyrB mutations conferred different levels of resistance to ofloxacin and moxifloxacin. Consideration of gyr mutation combinations during the interpretation of molecular test results may improve the accuracy of predicting the fluoroquinolone resistance phenotype. Further, the differential effects of gyr mutations on the activity of early- and later-generation fluoroquinolones requires further investigation and could inform the selection of a fluoroquinolone for treatment.

FIG 1

Estimated ofloxacin (OFX) and moxifloxacin (MXF) MIC distributions in isolates with the gyrA mutation A90V or D94A (there were no isolates with both mutations). MIC distributions are superimposed on a histogram of the observed MICs. The MXF MIC peaks lower than the lower limit of the peak MXF serum concentrations (dotted pink line; 3 mg/liter) (41); however, the OFX MIC peaks higher than the peak serum concentration for OFX (dotted blue line; 2 mg/liter) (42).

FIG 2

Histogram of gyr mutation frequency as a function of MIC by drug. MICs are plotted using a log₂ scale. The two extreme MIC bins were collapsed into the adjacent MIC bin. All nonsynonymous mutations were counted except for the lineage gyrA mutations E21Q, T80A, S95T, G247S, and G668D and the gyrB mutations I310M, V340L, and S700T, as the latter 3 were not found to be associated with drug resistance in the multivariate model.