Genetic sequencing for surveillance of drug resistance in tuberculosis in highly endemic countries: a multi-country population-based surveillance study.

BACKGROUND: In many countries, regular monitoring of the emergence of resistance to anti-tuberculosis drugs is hampered by the limitations of phenotypic testing for drug susceptibility. We therefore evaluated the use of genetic sequencing for surveillance of drug resistance in tuberculosis. METHODS: Population-level surveys were done in hospitals and clinics in seven countries (Azerbaijan, Bangladesh, Belarus, Pakistan, Philippines, South Africa, and Ukraine) to evaluate the use of genetic sequencing to estimate the resistance of Mycobacterium tuberculosis isolates to rifampicin, isoniazid, ofloxacin, moxifloxacin, pyrazinamide, kanamycin, amikacin, and capreomycin. For each drug, we assessed the accuracy of genetic sequencing by a comparison of the adjusted prevalence of resistance, measured by genetic sequencing, with the true prevalence of resistance, determined by phenotypic testing. FINDINGS: Isolates were taken from 7094 patients with tuberculosis who were enrolled in the study between November, 2009, and May, 2014. In all tuberculosis cases, the overall pooled sensitivity values for predicting resistance by genetic sequencing were 91% (95% CI 87-94) for rpoB (rifampicin resistance), 86% (74-93) for katG, inhA, and fabG promoter combined (isoniazid resistance), 54% (39-68) for pncA (pyrazinamide resistance), 85% (77-91) for gyrA and gyrB combined (ofloxacin resistance), and 88% (81-92) for gyrA and gyrB combined (moxifloxacin resistance). For nearly all drugs and in most settings, there was a large overlap in the estimated prevalence of drug resistance by genetic sequencing and the estimated prevalence by phenotypic testing. INTERPRETATION: Genetic sequencing can be a valuable tool for surveillance of drug resistance, providing new opportunities to monitor drug resistance in tuberculosis in resource-poor countries. Before its widespread adoption for surveillance purposes, there is a need to standardise DNA extraction methods, recording and reporting nomenclature, and data interpretation. FUNDING: Bill & Melinda Gates Foundation, United States Agency for International Development, Global Alliance for Tuberculosis Drug Development.

The TB Portals: an Open-Access, Web-Based Platform for Global Drug-Resistant-Tuberculosis Data Sharing and Analysis.

The TB Portals program is an international consortium of physicians, radiologists, and microbiologists from countries with a heavy burden of drug-resistant tuberculosis working with data scientists and information technology professionals. Together, we have built the TB Portals, a repository of socioeconomic/geographic, clinical, laboratory, radiological, and genomic data from patient cases of drug-resistant tuberculosis backed by shareable, physical samples. Currently, there are 1,299 total cases from five country sites (Azerbaijan, Belarus, Moldova, Georgia, and Romania), 976 (75.1%) of which are multidrug or extensively drug resistant and 38.2%, 51.9%, and 36.3% of which contain X-ray, computed tomography (CT) scan, and genomic data, respectively. The top Mycobacterium tuberculosis lineages represented among collected samples are Beijing, T1, and H3, and single nucleotide polymorphisms (SNPs) that confer resistance to isoniazid, rifampin, ofloxacin, and moxifloxacin occur the most frequently. These data and samples have promoted drug discovery efforts and research into genomics and quantitative image analysis to improve diagnostics while also serving as a valuable resource for researchers and clinical providers. The TB Portals database and associated projects are continually growing, and we invite new partners and collaborations to our initiative. The TB Portals data and their associated analytical and statistical tools are freely available at https://tbportals.niaid.nih.gov/.

Population-based resistance of Mycobacterium tuberculosis isolates to pyrazinamide and fluoroquinolones: results from a multicountry surveillance project.

BACKGROUND: Pyrazinamide and fluoroquinolones are essential antituberculosis drugs in new rifampicin-sparing regimens. However, little information about the extent of resistance to these drugs at the population level is available. METHODS: In a molecular epidemiology analysis, we used population-based surveys from Azerbaijan, Bangladesh, Belarus, Pakistan, and South Africa to investigate resistance to pyrazinamide and fluoroquinolones among patients with tuberculosis. Resistance to pyrazinamide was assessed by gene sequencing with the detection of resistance-conferring mutations in the pncA gene, and susceptibility testing to fluoroquinolones was conducted using the MGIT system. FINDINGS: Pyrazinamide resistance was assessed in 4972 patients. Levels of resistance varied substantially in the surveyed settings (3·0-42·1%). In all settings, pyrazinamide resistance was significantly associated with rifampicin resistance. Among 5015 patients who underwent susceptibility testing to fluoroquinolones, proportions of resistance ranged from 1·0-16·6% for ofloxacin, to 0·5-12·4% for levofloxacin, and 0·9-14·6% for moxifloxacin when tested at 0·5 µg/mL. High levels of ofloxacin resistance were detected in Pakistan. Resistance to moxifloxacin and gatifloxacin when tested at 2 µg/mL was low in all countries. INTERPRETATION: Although pyrazinamide resistance was significantly associated with rifampicin resistance, this drug may still be effective in 19-63% of patients with rifampicin-resistant tuberculosis. Even though the high level of resistance to ofloxacin found in Pakistan is worrisome because it might be the expression of extensive and unregulated use of fluoroquinolones in some parts of Asia, the negligible levels of resistance to fourth-generation fluoroquinolones documented in all survey sites is an encouraging finding. Rational use of this class of antibiotics should therefore be ensured to preserve its effectiveness. FUNDING: Bill & Melinda Gates Foundation, United States Agency for International Development, Global Alliance for Tuberculosis Drug Development.