Dataset Information

Genomic Analysis of the Evolution of Fluoroquinolone Resistance in Mycobacterium tuberculosis Prior to Tuberculosis Diagnosis.

ABSTRACT: Fluoroquinolones (FQs) are effective second-line drugs for treating antibiotic-resistant tuberculosis (TB) and are being considered for use as first-line agents. Because FQs are used to treat a range of infections, in a setting of undiagnosed TB, there is potential to select for drug-resistant Mycobacterium tuberculosis mutants during FQ-based treatment of other infections, including pneumonia. Here we present a detailed characterization of ofloxacin-resistant M. tuberculosis samples isolated directly from patients in Taiwan, which demonstrates that selection for FQ resistance can occur within patients who have not received FQs for the treatment of TB. Several of these samples showed no mutations in gyrA or gyrB based on PCR-based molecular assays, but genome-wide next-generation sequencing (NGS) revealed minority populations of gyrA and/or gyrB mutants. In other samples with PCR-detectable gyrA mutations, NGS revealed subpopulations containing alternative resistance-associated genotypes. Isolation of individual clones from these apparently heterogeneous samples confirmed the presence of the minority drug-resistant variants suggested by the NGS data. Further NGS of these purified clones established evolutionary links between FQ-sensitive and -resistant clones derived from the same patient, suggesting de novo emergence of FQ-resistant TB. Importantly, most of these samples were isolated from patients without a history of FQ treatment for TB. Thus, selective pressure applied by FQ monotherapy in the setting of undiagnosed TB infection appears to be able to drive the full or partial emergence of FQ-resistant M. tuberculosis, which has the potential to confound diagnostic tests for antibiotic susceptibility and limit the effectiveness of FQs in TB treatment.

SUBMITTER: Zhang D

PROVIDER: S-EPMC5075065 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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Genomic Analysis of the Evolution of Fluoroquinolone Resistance in Mycobacterium tuberculosis Prior to Tuberculosis Diagnosis.

Zhang Danfeng D Gomez James E JE Chien Jung-Yien JY Haseley Nathan N Desjardins Christopher A CA Earl Ashlee M AM Hsueh Po-Ren PR Hung Deborah T DT

Antimicrobial agents and chemotherapy 20161021 11

Fluoroquinolones (FQs) are effective second-line drugs for treating antibiotic-resistant tuberculosis (TB) and are being considered for use as first-line agents. Because FQs are used to treat a range of infections, in a setting of undiagnosed TB, there is potential to select for drug-resistant Mycobacterium tuberculosis mutants during FQ-based treatment of other infections, including pneumonia. Here we present a detailed characterization of ofloxacin-resistant M. tuberculosis samples isolated di ...[more]

PMID: 27572408

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Project description:RationaleFluoroquinolones are the most commonly prescribed antibiotic class in the United States. They have the potential to become first-line antituberculosis therapy, but the effect of fluoroquinolone use on fluoroquinolone resistance in Mycobacterium tuberculosis is not well characterized.ObjectivesTo determine the prevalence of and risk factors for fluoroquinolone-resistant tuberculosis in a large United States population.MethodsWe identified all people with culture-confirmed tuberculosis enrolled in TennCare (Medicaid) and reported to the Tennessee Department of Health from January 2002 to December 2006. People with fluoroquinolone-resistant M. tuberculosis isolates (cases) were compared with those with susceptible isolates (control subjects). Fluoroquinolone resistance was determined by agar proportion using ofloxacin 2 microg/ml. Outpatient fluoroquinolone exposure in the 12 months before tuberculosis diagnosis was ascertained from TennCare pharmacy data.Measurements and main resultsOf 640 study patients, 116 (18%) had fluoroquinolone exposure in the 12 months before diagnosis, and 16 (2.5%; 95% confidence interval [CI], 1.4-4.0%) M. tuberculosis isolates were fluoroquinolone resistant. Among the 54 patients with more than 10 days of fluoroquinolone exposure, 7 (13%) had fluoroquinolone resistance. In multivariable logistic regression analyses using propensity score to control for age, sex, race, HIV serostatus, and site of disease, more than 10 days of fluoroquinolone exposure before tuberculosis diagnosis was associated with fluoroquinolone resistance (odds ratio 7.0; 95% CI, 2.3-20.6; P = 0.001). Fluoroquinolone exposure for more than 10 days that occurred more than 60 days before tuberculosis diagnosis was associated with the highest risk of resistance (20.8%; odds ratio 17.0; 95% CI, 5.1-56.8; P < 0.001 compared with no exposure).ConclusionsOverall, fluoroquinolone resistance was relatively low. However, receipt of fluoroquinolones for more than 10 days, particularly more than 60 days before tuberculosis diagnosis, was associated with a high risk of fluoroquinolone-resistant tuberculosis.

Project description:Mycobacterium tuberculosis has acquired drug resistance to all drugs that have been used against it, including those only recently introduced into clinical practice. Compared to other bacteria, it has a well conserved genome due to its role as an obligate human pathogen that has adapted to a niche over five to ten thousand years. These features facilitate reconstruction and dating of M. tuberculosis phylogenies, giving key insights into how resistance has been acquired and spread globally. Resistance to each new drug has occurred within five to ten years of clinical use and has occurred even more rapidly with recently introduced drugs. In most cases, resistance-conferring mutations come with a fitness cost, but this can be overcome by compensatory mutations which restore fitness to that of wild-type bacteria. It is likely that M. tuberculosis acquires drug resistance while maintaining limited genomic variability due the generation of low frequency within-host variation, combined with ongoing purifying selection causing loss of variants without a clear fitness advantage. However, variants that do confer an advantage, such as drug resistance, can increase in prevalence amongst all bacteria within a host and become the dominant clone. These resistant strains can then be transmitted leading to primary drug resistant infection in a new host. As many countries move towards genomic methods for diagnosis of M. tuberculosis infection and drug resistance, it is important to be aware of the implications for the evolution of resistance. Currently, understanding of resistance-conferring mutations is incomplete, and some targeted genetic diagnostics create their own selective pressures. We discuss an example where a rifampicin resistance-conferring mutation which was not routinely covered by standard testing became dominant. Finally, resistance to new drugs such as bedaquiline and delamanid is caused by individually rare mutations occurring across a large mutational genomic target that have been detected over a short time, and do not provide statistical power for genotype-phenotype correlation - in contrast to longer-established drugs that form the backbone of drug-sensitive antituberculosis therapy. Therefore, we need a different approach to identify resistance-conferring mutations of new drugs before their resistance becomes widespread, abrogating their usefulness.

Project description:Fluoroquinolone antibiotics are among the most potent second-line drugs used for treatment of multidrug-resistant tuberculosis (MDR TB), and resistance to this class of antibiotics is one criterion for defining extensively drug resistant tuberculosis (XDR TB). Fluoroquinolone resistance in Mycobacterium tuberculosis has been associated with modification of the quinolone resistance determining region (QRDR) of gyrA. Recent studies suggest that amino acid substitutions in gyrB may also play a crucial role in resistance, but functional genetic studies of these mutations in M. tuberculosis are lacking. In this study, we examined twenty six mutations in gyrase genes gyrA (seven) and gyrB (nineteen) to determine the clinical relevance and role of these mutations in fluoroquinolone resistance. Transductants or clinical isolates harboring T80A, T80A+A90G, A90G, G247S and A384V gyrA mutations were susceptible to all fluoroquinolones tested. The A74S mutation conferred low-level resistance to moxifloxacin but susceptibility to ciprofloxacin, levofloxacin and ofloxacin, and the A74S+D94G double mutation conferred cross resistance to all the fluoroquinolones tested. Functional genetic analysis and structural modeling of gyrB suggest that M330I, V340L, R485C, D500A, D533A, A543T, A543V and T546M mutations are not sufficient to confer resistance as determined by agar proportion. Only three mutations, N538D, E540V and R485C+T539N, conferred resistance to all four fluoroquinolones in at least one genetic background. The D500H and D500N mutations conferred resistance only to levofloxacin and ofloxacin while N538K and E540D consistently conferred resistance to moxifloxacin only. Transductants and clinical isolates harboring T539N, T539P or N538T+T546M mutations exhibited low-level resistance to moxifloxacin only but not consistently. These findings indicate that certain mutations in gyrB confer fluoroquinolone resistance, but the level and pattern of resistance varies among the different mutations. The results from this study provide support for the inclusion of the QRDR of gyrB in molecular assays used to detect fluoroquinolone resistance in M. tuberculosis.

Dataset Information

Genomic Analysis of the Evolution of Fluoroquinolone Resistance in Mycobacterium tuberculosis Prior to Tuberculosis Diagnosis.

Publications

Genomic Analysis of the Evolution of Fluoroquinolone Resistance in Mycobacterium tuberculosis Prior to Tuberculosis Diagnosis.

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