Molecular Evaluation of Fluoroquinolone Resistance in Serial Mycobacterium tuberculosis Isolates from Individuals Diagnosed with Multidrug-Resistant Tuberculosis

Abstract

Fluoroquinolones (FQ) are crucial components of multidrug-resistant tuberculosis (MDR TB) treatment. Differing levels of resistance are associated with specific mutations within the quinolone-resistance-determining region (QRDR) of gyrA We sequenced the QRDR from serial isolates of MDR TB patients in the Preserving Effective TB Treatment Study (PETTS) with baseline FQ resistance (FQ^R) or acquired FQ resistance (FQ^ACQR) using an Ion Torrent Personal Genome Machine (PGM) to a depth of 10,000× and reported single nucleotide polymorphisms in ≥1% of reads. FQ^R isolates harbored 15 distinct alleles with 1.3 (maximum = 6) on average per isolate. Eighteen alleles were identified in FQ^ACQR isolates with an average of 1.6 (maximum = 9) per isolate. Isolates from 78% of FQ^ACQR individuals had mutant alleles identified within 6 months of treatment initiation. Asp94Gly was the predominant allele in the initial FQ-resistant isolates followed by Ala90Val. Seventy-seven percent (36/47) of FQ^ACQR group patients had isolates with FQ resistance alleles prior to changes to the FQ component of their treatment. Unlike the individuals treated initially with other FQs, none of the 21 individuals treated initially with levofloxacin developed genotypic or phenotypic FQ resistance, although country of residence was likely a contributing factor since 69% of these individuals were from a single country. Initial detection of phenotypic resistance and genotypic resistance occurred simultaneously for most; however, phenotypic resistance occurred earlier in isolates harboring mixtures of alleles of very low abundance (<1% of reads), whereas genotypic resistance often occurred earlier for alleles associated with low-level resistance. Understanding factors influencing acquisition and evolution of FQ resistance could reveal strategies for improved treatment success.

Keywords: QRDR; acquired drug resistance; fluoroquinolone; levofloxacin; moxifloxacin; multidrug-resistant tuberculosis.