Browsing by Author "Whitfield, Michael G."

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    Comparative Performance of Genomic Methods for the Detection of Pyrazinamide Resistance and Heteroresistance in Mycobacterium tuberculosis
    (American Society for Microbiology, 2021) Whitfield, Michael G.; Engelthaler, David M.; Allender, Christopher; Folkerts, Megan; Heupink, Tim H.; Limberis, Jason; Warren, Robin M.; Van Rie, Annelies; Metcalfe, John Z.
    Pyrazinamide is an important component of both drug-susceptible and drug-resistant tuberculosis treatment regimens. Although approximately 50% of rifampin- resistant isolates are also resistant to pyrazinamide, pyrazinamide susceptibility testing is not routinely performed due to the challenging nature of the assay. We investigated the diagnostic accuracy of genotypic and phenotypic methods and explored the occurrence of pyrazinamide heteroresistance. We assessed pyrazinamide susceptibility among 358 individuals enrolled in the South African EXIT-RIF cohort using Sanger and targeted deep sequencing (TDS) of the pncA gene, whole-genome sequencing (WGS), and phenotypic drug susceptibility testing. We calculated the diagnostic accuracy of the different methods and investigated the prevalence and clinical impact of pncA heteroresistance. True pyrazinamide susceptibility status was assigned to each isolate using the Köser classification and expert rules. We observed 100% agreement across genotypic methods for detection of pncA fixed mutations; only TDS confidently identified three isolates (0.8%) with minor variants. For the 355 (99.2%) isolates that could be assigned true pyrazinamide status with confidence, phenotypic DST had a sensitivity of 96.5% (95% confidence interval [CI], 93.8 to 99.3%) and specificity of 100% (95% CI, 100 to 100%), both Sanger sequencing and WGS had a sensitivity of 97.1% (95% CI, 94.6 to 99.6%) and specificity of 97.8% (95% CI, 95.7 to 99.9%), and TDS had sensitivity of 98.8% (95% CI, 97.2 to 100%) and specificity of 97.8% (95% CI, 95.7 to 99.9%). We demonstrate high sensitivity and specificity for pyrazinamide susceptibility testing among all assessed genotypic methods. The prevalence of pyrazinamide heteroresistance in Mycobacterium tuberculosis isolates was lower than that identified for other first-line drugs.
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    A global perspective on pyrazinamide resistance: systematic review and meta-analysis
    (Public Library of Science, 2015) Whitfield, Michael G.; Soeters, Heidi M.; Warren, Robin M.; York, Talita; Sampson, Samantha L.; Streicher, Elizabeth M.; Van Helden, Paul D.; Van Rie, Annelies
    Background: Pyrazinamide (PZA) is crucial for tuberculosis (TB) treatment, given its unique ability to eradicate persister bacilli. The worldwide burden of PZA resistance remains poorly described. Methods Systematic PubMed, Science Direct and Scopus searches for articles reporting phenotypic (liquid culture drug susceptibility testing or pyrazinamidase activity assays) and/or genotypic (polymerase chain reaction or DNA sequencing) PZA resistance. Global and regional summary estimates were obtained from random-effects meta-analysis, stratified by presence or risk of multidrug resistant TB (MDR-TB). Regional summary estimates were combined with regional WHO TB incidence estimates to determine the annual burden of PZA resistance. Information on single nucleotide polymorphisms (SNPs) in the pncA gene was aggregated to obtain a global summary. Results: Pooled PZA resistance prevalence estimate was 16.2% (95% CI 11.2-21.2) among all TB cases, 41.3% (29.0-53.7) among patients at high MDR-TB risk, and 60.5% (52.3-68.6) among MDR-TB cases. The estimated global burden is 1.4 million new PZA resistant TB cases annually, about 270,000 in MDR-TB patients. Among 1,815 phenotypically resistant isolates, 608 unique SNPs occurred at 397 distinct positions throughout the pncA gene. Interpretation: PZA resistance is ubiquitous, with an estimated one in six incident TB cases and more than half of all MDR-TB cases resistant to PZA globally. The diversity of SNPs across the pncA gene complicates the development of rapid molecular diagnostics. These findings caution against relying on PZA in current and future TB drug regimens, especially in MDR-TB patients.
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    Mycobacterium tuberculosis pncA polymorphisms that do not confer pyrazinamide resistance at a breakpoint concentration of 100 micrograms per milliliter in MGIT
    (American Society for Microbiology, 2015) Whitfield, Michael G.; Warren, Robin M.; Streicher, Elizabeth M.; Sampson, Samantha L.; Sirgel, Frik A.; Van Helden, Paul D.; Mercante, Alexandra; Willby, Melisa; Hughes, Kelsey; Birkness, Kris; Morlock, Glenm; Van Rie, Annelies; Posey, James E.
    Sequencing of the Mycobacterium tuberculosis pncA gene allows for pyrazinamide susceptibility testing. We summarize data on pncA polymorphisms that do not confer resistance at a susceptibility breakpoint of 100 μg/ml pyrazinamide in MGIT within a cohort of isolates from South Africa and the U.S. Centers for Disease Control and Prevention.