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Background: Pretomanid is a key component of new regimens for the treatment of drug-resistant tuberculosis (TB) which are being rolled out globally. However, there is limited information on the prevalence of pre-existing resistance to the drug. Methods: To investigate pretomanid resistance rates in China and its underlying genetic basis, as well as to generate additional minimum inhibitory concentration (MIC) data for epidemiological cutoff (ECOFF)/breakpoint setting, we performed MIC determinations in the Mycobacterial Growth Indicator Tube™ (MGIT) system, followed by WGS analysis, on 475 Mycobacterium tuberculosis (MTB) isolated from Chinese TB patients between 2013 and 2020. Results: We observed a pretomanid MIC distribution with a 99% ECOFF equal to 0.5 mg/L. Of the 15 isolates with MIC values > 0.5 mg/L, one (MIC = 1 mg/L) was identified as MTB lineage 1 (L1), a genotype previously reported to be intrinsically less susceptible to pretomanid, two were borderline resistant (MIC = 2–4 mg/L) and the remaining 12 isolates were highly resistant (MIC ≥ 16 mg/L) to the drug. Five resistant isolates did not harbor mutations in the known pretomanid resistant genes. Conclusions: Our results further support a breakpoint of 0.5 mg/L for a non-L1 MTB population, which is characteristic of China. Further, our data point to an unexpected high (14/475, 3%) pre-existing pretomanid resistance rate in the country, as well as to the existence of yet-to-be-discovered pretomanid resistance genes. [ABSTRACT FROM AUTHOR]

Mycobacterium bovis is the cause of bovine tuberculosis, and it can also cause disease in humans, with symptoms similar to those caused by M. tuberculosis. However, our understanding of its genomic diversity, biogeography, and drug resistance remains incomplete. We performed a comparative and phylogenetic analysis of 3228 M. bovis genomes from 24 countries. Following drug susceptibility testing, we applied a bacterial genome‐wide association study to capture associations between genomic variation and drug resistance in 74 newly isolated strains from China. The data show that the cattle‐adapted M. bovis were divided into six lineages with a strong phylogeographical population structure. Lineages 1 and 6 are the most widespread globally, while others show a strong geographical restriction. Note that 17.39% of M. bovis isolates were resistant to at least one drug in China. Furthermore, we identify genomic variations associated with an increased risk of resistance acquisition. This study furthers our knowledge of M. bovis diversity, biogeography, and drug resistance and will facilitate more deeply informed genomic tracking and surveillance to minimize its threat to human health, as a cause of zoonotic tuberculosis. [ABSTRACT FROM AUTHOR]

Background: Glycerol kinase (glpK) is essential for the first step of glycerol catabolism in Mycobacterium tuberculosis. However, Mycobacterium bovis has been known to grow poorly in glycerol media because of a base insertion in the glpK gene. Methods: We analyzed the glpK gene sequences of 60 clinical M. bovis isolates, and determined the minimum inhibitory concentration of 14 drugs by microdilution method to evaluate the effect of frameshift mutations on drug sensitivity. The effect of M. bovis growth rate on its drug sensitivity was investigated using bacteria grown on glycerol or pyruvate. Results: A total of 44 (73.33%) clinical M. bovis isolates have frameshift mutations in a homopolymeric tract of 7 cytosines in the glpK gene. 15.00% M. bovis isolates showed phenotypic drug resistance. Glycerol metabolism-deficient M. bovis showed reduced susceptibility to 9 out of 14 tested drugs. Mutations in the glpK gene can lead to impaired growth in glycerol-based media, while the minimal inhibitory concentration values of slow-growing M. bovis were higher. Conclusion: Mutations in the glpK gene can lead to slowed growth and reduced susceptibility to drugs in M. bovis, which may contribute to the emergence of drug-resistant M. bovis and pose a threat to human health owing to the zoonotic capacity of M. bovis. [ABSTRACT FROM AUTHOR]

Objectives: For the past decade, the epidemic of multidrug resistance tuberculosis (MDR-TB) stays high in China. We investigated the possible driving forces behind the epidemics from phylogenetic and historical perspectives.Methods: 420 representative strains were selected from the first national drug resistance survey based on their genotypes, drug susceptibility patterns and geographic information. We reconstructed the phylogeny by whole genome sequencing and compared it to the global phylogeny including MDR outbreaks reported in other settings. We estimated the historical trajectory of population dynamics by Bayesian Skygrid plot for all strains and MDR-TB alone. Integrating geographic information and mutations in drug resistance related genes, we investigated the spatial scale of transmission, recent selection of drug resistant mutant, and mechanism for fitness restoration.Results: Three new subgroups within Beijing clade are described for the first time, but none of the MDR-TB outbreak strains reported in other high MDR-TB burden settings is identified. The overall epidemics experienced two successive phases of expansion at different rates between 1660s and 1950s, followed by a sharp decline till today. Four fifths of the clustered MDR-TB strains suggest transmission of DR strains and nearly half suggest recent selection of (additional) mutations in rpoB. Among all identified transmission events, about one fifth occurred between far distant locations. Possible intergenic and intragenic compensatory mutations both presented in our dataset at comparable frequencies.Conclusions: MDR-TB epidemic in China is not yet driven by the spread of a few highly successful clonal expansions but by repeated emergence of smaller and currently less successful clusters. However, internal migration and undertreatment could escalate MDR-TB epidemic. To prevent generating of drug resistance and restoration of fitness as well as to stop transmission of MDR-TB at early stage, national TB control program needs to strengthen management of floating populations and promote universal drug susceptibility testing in China. [ABSTRACT FROM AUTHOR]

Phenotypic drug susceptibility testing for prediction of tuberculosis (TB) drug resistance is slow and unreliable, limiting individualized therapy and monitoring of national TB data. Our study evaluated whole-genome sequencing (WGS) for its predictive accuracy, use in TB drug-resistance surveillance and ability to quantify the effects of resistance-associated mutations on MICs of anti-TB drugs. We used WGS to measure the susceptibility of 4880 isolates to ten anti-TB drugs; for pyrazinamide, we used BACTEC MGIT 960. We determined the accuracy of WGS by comparing the prevalence of drug resistance, measured by WGS, with the true prevalence, determined by phenotypic susceptibility testing. We used the Student–Newman–Keuls test to confirm MIC differences of mutations. Resistance to isoniazid, rifampin and ethambutol was highly accurately predicted with at least 92.92% (95% confidence interval [CI], 88.19–97.65) sensitivity, resistance to pyrazinamide with 50.52% (95% CI, 40.57–60.47) sensitivity, and resistance to six second-line drugs with 85.05% (95% CI, 80.27–89.83) to 96.01% (95% CI, 93.89–98.13) sensitivity. The rpoB S450L, katG S315T and gyrA D94G mutations always confer high-level resistance, while rpoB L430P, rpoB L452P, fabG1 C-15T and embB G406S often confer low-level resistance or sub-epidemiological cutoff (ECOFF) MIC elevation. WGS can predict phenotypic susceptibility with high accuracy and could be a valuable tool for drug-resistance surveillance and allow the detection of drug-resistance level; It can be an important approach in TB drug-resistance surveillance and for determining therapeutic schemes. [ABSTRACT FROM AUTHOR]