The Mycobacterium tuberculosis Beijing family is often associated with multidrug resistance and large outbreaks. Conventional genotyping study of a community outbreak of multidrug-resistant tuberculosis (MDR-TB) that occurred in Kanchanaburi Province, Thailand was carried out. The study revealed that the outbreak was clonal and the strain was identified as a member of Beijing family. Although, the outbreak isolates showed identical spoligotyping and mycobacterial interspersed repetitive units-variable number tandem repeats patterns, a discrepancy regarding ethambutol resistance was observed. In-depth characterization of the isolates through whole genome sequencing of the first and the last three isolates from our culture collection showed them to belong to principal genetic group 1, single nucleotide polymorphism (SNP) cluster group 2, sequence type 10. Compared with the M. tuberculosis H37Rv reference genome, 1242 SNPs were commonly found in all isolates. The genomes of these isolates were shown to be clonal and highly stable over a 5-year period and two or three unique SNPs were identified in each of the last three isolates. Genes known to be associated with drug resistance and their promoter regions, where applicable, were analyzed. The presence of low or no fitness cost mutations for drug resistance and an additional L731P SNP in the rpoB gene was observed in all isolates. These findings might account for the successful transmission of this MDR-TB strain.

Tuberculosis disease (TB), caused by Mycobacterium tuberculosis , is a major public health issue in Thailand. The high prevalence of modern Beijing (Lineage 2.2.1) strains has been associated with multi- and extensively drug-resistant infections (MDR-, XDR-TB), complicating disease control. The impact of rarer proto-Beijing (L2.1) strains is less clear. In our study of thirty-seven L2.1 clinical isolates spanning thirteen years, we found a high prevalence of XDR-TB cases (32.4%). With ≤ 12 pairwise SNP distances, 43.2% of L2.1 patients belong to MDR-TB or XDR-TB transmission clusters suggesting a high level of clonal expansion across four Thai provinces. All XDR-TB (100%) were likely due to transmission rather than inadequate treatment. We found a 47 mutation signature and a partial deletion of the fadD14 gene in the circulating XDR-TB cluster, which can be used for surveillance of this rare and resilient M. tuberculosis strain-type that is causing increasing health burden. We also detected three novel deletion positions, a deletion of 1285 bp within desA3 (Rv3230c) , large deletions in the plcB, plcA, and ppe38 gene which may play a role in the virulence, pathogenesis or evolution of the L2.1 strain-type.

Introduction. Multidrug-resistant tuberculosis (MDR-TB) is a major public health problem globally, including in Indonesia. Whole-genome sequencing (WGS) analysis has rarely been used for the study of TB and MDR-TB in Indonesia.Aim. We evaluated the use of WGS for drug-susceptibility testing (DST) and to investigate the population structure of drugresistant Mycobacterium tuberculosis in Java, Indonesia.Methodology. Thirty suspected MDR-TB isolates were subjected to MGIT 960 system (MGIT)-based DST and to WGS. Phylogenetic analysis was done using the WGS data. Results obtained using MGIT-based DST and WGS-based DST were compared.Results. Agreement between WGS and MGIT was 93.33 % for rifampicin, 83.33 % for isoniazid and 76.67 % for streptomycin but only 63.33 % for ethambutol. Moderate WGS-MGIT agreement was found for second-line drugs including amikacin, kanamycin and fluoroquinolone (73.33-76.67 %). MDR-TB was more common in isolates of the East Asian Lineage (63.3%). No evidence of clonal transmission of DR-TB was found among members of the tested population.Conclusion. Our study demonstrated the applicability of WGS for DST and molecular epidemiology of DR-TB in Java, Indonesia. We found no transmission of DR-TB in Indonesia.

Mutations in genes involved in drug metabolism have been well-associated with drug resistance. Sequence analysis of known antimycobacterial drug-resistant genes is often used to predict resistance to antibiotics. However, some polymorphisms in such genes may serve a phylogenetic purpose rather than resistance to drugs. The Beijing family of Mycobacterium tuberculosis (MTB) is prevalent worldwide and has been associated with the emergence of multidrug resistance. Sequence type (ST) 10 of the Beijing family is the most predominant in countries like Peru, Taiwan and Thailand. A sequence analysis was performed of 81 previously reported drug-resistant associated genes in multidrug-resistant and pan-susceptible strains of the Beijing family sequence type 10 of MTB. This analysis revealed 10 synonymous and 12 nonsynonymous single nucleotide polymorphisms (SNPs) that are shared by all strains under study. One frameshift mutation was also observed to be common to all. These data might be useful in excluding some observed SNPs in drug-resistant-associated genes of MTB Beijing ST 10 when performing genotypic drug susceptibility assay.