Our inability to predict which mutations could result in antibiotic resistance has made it difficult to rapidly identify the emergence of resistance, identify pre-existing resistant populations, and manage our use of antibiotics to effectively treat patients and prevent or slow the spread of resistance. Here we investigated the potential for resistance against the new antitubercular nitroimidazole prodrugs pretomanid and delamanid to emerge in Mycobacterium tuberculosis, the causative agent of tuberculosis (TB). Deazaflavin-dependent nitroreductase (Ddn) is the only identified enzyme within M. tuberculosis that activates these prodrugs, via an F₄₂₀H₂-dependent reaction. We show that the native menaquinone-reductase activity of Ddn is essential for emergence from hypoxia, which suggests that for resistance to spread and pose a threat to human health, the native activity of Ddn must be at least partially retained. We tested 75 unique mutations, including all known sequence polymorphisms identified among ~15,000 sequenced M. tuberculosis genomes. Several mutations abolished pretomanid and delamanid activation in vitro, without causing complete loss of the native activity. We confirmed that a transmissible M. tuberculosis isolate from the hypervirulent Beijing family already possesses one such mutation and is resistant to pretomanid, before being exposed to the drug. Notably, delamanid was still effective against this strain, which is consistent with structural analysis that indicates delamanid and pretomanid bind to Ddn differently. We suggest that the mutations identified in this work be monitored for informed use of delamanid and pretomanid treatment and to slow the emergence of resistance. Bacterial pathogens often evolve resistance to antibiotics via mutations in the coding sequences of genes–frequently the target, an enzyme that metabolizes or transports the active drug, or an enzyme that activates a prodrug. In the case of tuberculosis, antibiotic resistance is a growing problem, with the rapid emergence of multi-drug resistant strains. New nitroimidazole-based antibiotic prodrugs, such as pretomanid and delamanid have the potential to help infected individuals, but we must guard against the evolution of resistance to these new compounds in Mycobacterium tuberculosis. In this report we use protein engineering to identify mutations that could potentially result in antibiotic resistance by knocking out the prodrug activating activity of the deazaflavin dependent nitroreductase (DDN), without completely abolishing its native menaquinone reductase activity. The retention of its native activity is important as DDN appears to be required for M. tuberculosis to emerge from hypoxia. Strikingly, when we analysed >15,000 M. tuberculosis genomes from clinical strains, we identified several that harboured mutations that we identified as abolishing prodrug activation in vitro. A hypervirulent Beijing strain N0008 from Vietnam (which had not been exposed to pretomanid in the clinic) was predicted, and confirmed, to be resistant to pretomanid, revealing that resistance to this drug has arisen through genetic drift and not selective pressure in this instance. These data show that by testing potential resistant mutations in the laboratory before large-scale use of antibiotics, we should be able to use them more judiciously in order to slow the spread of resistance.

A screening of more than 1,500 drug-resistant strains of Mycobacterium tuberculosis revealed evolutionary patterns characteristic of positive selection for three alanine racemase (Alr) mutations. We investigated these mutations using molecular modeling, in vitro MIC testing, as well as direct measurements of enzymatic activity, which demonstrated that these mutations likely confer resistance to d -cycloserine.

Drug-resistant tuberculosis (TB) is a major health threat in Myanmar. An initial study was conducted to explore the potential utility of whole-genome sequencing (WGS) for the diagnosis and management of drug-resistant TB in Myanmar. Fourteen multidrug-resistant Mycobacterium tuberculosis isolates were sequenced. Known resistance genes for a total of nine antibiotics commonly used in the treatment of drug-susceptible and multidrug-resistant TB (MDR-TB) in Myanmar were interrogated through WGS. All 14 isolates were MDR-TB, consistent with the results of phenotypic drug susceptibility testing (DST), and the Beijing lineage predominated. Based on the results of WGS, 9 of the 14 isolates were potentially resistant to at least one of the drugs used in the standard MDR-TB regimen but for which phenotypic DST is not conducted in Myanmar. This study highlights a need for the introduction of second-line DST as part of routine TB diagnosis in Myanmar as well as new classes of TB drugs to construct effective regimens.

Tuberculosis (TB) remains a significant cause of morbidity and mortality in Myanmar. The fourth National TB Prevalence Survey was conducted in 2017–2018 to determine the actual burden of TB not only at the national level but also for three subnational strata (the states, regions other than Yangon, and the Yangon region) and develop a more efficacious country strategy on TB care and control. One hundred and thirty eight clusters were selected by population proportionate sampling. Adult (≥15 years of age) residents having lived for 2 weeks or more in the households of the selected clusters were invited to participate in the survey. The survey participants were screened for TB by a questionnaire and digital chest X-ray (CXR) after providing written informed consent. Individuals with a positive symptom screen and/or chest X-ray suggestive of TB were asked to provide sputum samples to test for Mycobacterium tuberculosis (Mtb) by Ziehl-Neelsen direct light microscopy, Xpert MTB/RIF Ultra (Xpert), and culture (Ogawa media). Bacteriologically confirmed TB cases were defined by an expert panel. Of 75 676 eligible residents, 66 480 (88%) participated, and 10 082 (15%) screened positive for TB. Among these, 322 participants were defined as bacteriologically confirmed TB cases. Cough lasting for two weeks or longer, one of the criteria used for screening for symptoms, could detect only 14% (45/322) of the study cases. The estimated prevalence of bacteriologically confirmed adult pulmonary TB was 468 (95% CI: 391–546) per 100,000. The prevalence was much higher among males, the older age group, urban Yangon and remote villages. In-depth interview with the participants on TB treatment showed that none of them was diagnosed in a TB health centre (primary care facilities). The prevalence of TB in Myanmar is still high due to challenges such as uncontrolled urbanization, an ageing population, migration, and poor access to health facilities in remote areas. New screening and diagnostic tools might help to detect more TB patients. There is a need to lay greater emphasis on multisectoral approaches, decentralization and the integration of basic TB services into primary care facilities.

Background: This is the first survey to use the World Health Organization (WHO) methodology to document the magnitude and main drivers of tuberculosis (TB) patient costs in order to guide policies on cost mitigation and to produce a baseline measure for the percentage of TB-affected households experiencing catastrophic costs in Myanmar. Methods: A nationally representative cross-sectional survey was administered to 1000 TB patients in health facilities from December 2015 to February 2016, focusing on costs of TB treatment (direct and indirect), household income, and coping strategies. A total cost was estimated for each household by extrapolating reported costs and comparing them to household income. If the proportion of total costs exceeded 20% of the annual household income, a TB-affected household was deemed to have faced catastrophic costs. Results: 60% of TB-affected households faced catastrophic costs in Myanmar. On average, total costs were USD 759, and the largest proportion of this total was accounted for by patient time (USD 365), followed by food costs (USD 200), and medical expenses (USD 130). Low household wealth quintile and undergoing MDR-TB treatment were both significant predictors for households facing catastrophic costs. Conclusions: The high proportion of TB-affected households experiencing catastrophic costs suggests the need for TB-specific social protection programs in patient-centered healthcare. The survey findings have led the government and donors to increase support for MDR-TB patients. The significant proportion of total spending attributable to lost income and food or nutritional supplements suggests that income replacement programs and/or food packages may ameliorate the burdensome costs.

Worldwide, studies investigating the relationship between the lineage of Mycobacterium tuberculosis (MTB) across geographic areas has empowered the “End TB” program and understand transmission across national boundaries. Genomic diversity of MTB varies with geographical locations and ethnicity. Genomic diversity can also affect the emergence of drug resistance. In Myanmar, we still have limited genetic information about geographical, ethnicity, and drug resistance linkage to MTB genetic information. This study aimed to describe the geno-spatial distribution of MTB and drug resistance profiles in Myanmar–Thailand border areas. A cross-sectional study was conducted with a total of 109 sequenced isolates. The lineages of MTB and the potential associated socio-demographic, geographic and clinical factors were analyzed using Fisher’s exact tests. p value of statistically significance was set at < 0.05. We found that 67% of the isolates were lineage 1 (L1)/East-African-Indian (EAI) (n = 73), followed by lineage 2 (L2)/Beijing (n = 26), lineage 4 (L4)/European American (n = 6) and lineage 3 (L3)/Delhi/Central Asian (n = 4). “Gender”, “type of TB patient”, “sputum smear grading” and “streptomycin resistance” were significantly different with the lineages of MTB. Sublineages of L1, which had never been reported elsewhere in Myanmar, were detected in this study area. Moreover, both ethnicity and lineage of MTB significantly differed in distribution by patient location. Diversity of the lineage of MTB and detection of new sublineages suggested that this small area had been resided by a heterogeneous population group who actively transmitted the disease. This information on distribution of lineage of MTB can be linked in the future with those on the other side of the border to evaluate cross-border transmission.