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Genome sequencing plays a key role in understanding the genetic diversity of  Mycobacterium tuberculosis (M.tb) . The genotype-specific character of  M. tb  contributes to tuberculosis severity and emergence of drug resistance. Strains of  M. tb  complex can be classified into seven lineages. The Nonthaburi (NB) genotype, belonging to the Indo-Oceanic lineage (lineage 1), has a unique spoligotype and IS 6110 -RFLP pattern but has not previously undergone a detailed whole genome analysis. In addition, there is not much information available on the whole genome analysis of  M. tb  isolates from tuberculous meningitis (TBM) patients in public databases. Isolates CSF3053, 46-5069 and 43-13838 of NB genotype were obtained from the cerebrospinal fluids of TBM Thai patients in Siriraj Hospital, Bangkok. The whole genomes were subjected to high throughput sequencing. The sequence data of each isolate were assembled into draft genome. The sequences were also aligned to reference genome, to determine genomic variations. Single nucleotide polymorphisms (SNPs) were obtained and grouped according to the functions of the genes containing them. They were compared with SNPs from 1,601 genomes, representing the seven lineages of  M. tb  complex, to determine the uniqueness of NB genotype. Susceptibility to first-line, second-line and other antituberculosis drugs were determined and related to the SNPs previously reported in drug-resistant related genes. The assembled genomes have an average size of 4,364,461 bp, 4,154 genes, 48 RNAs and 64 pseudogenes. A 500 base pairs deletion, which includes  ppe50 , was found in all isolates. RD239, specific for members of Indo Oceanic lineage, and RD147c were identified. A total of 2,202 SNPs were common to the isolates and used to classify the NB strains as members of sublineage 1.2.1. Compared with 1,601 genomes from the seven lineages of  M. tb  complex, mutation G2342203C was found novel to the isolates in this study. Three mutations (T28910C, C1180580T and C152178T) were found only in Thai NB isolates, including isolates from previous study. Although drug susceptibility tests indicated pan-susceptibility, non-synonymous SNPs previously reported to be associated with resistance to anti-tuberculous drugs; isoniazid, ethambutol, and ethionamide were identified in all the isolates. Non-synonymous SNPs were found in virulence genes such as the genes playing roles in apoptosis inhibition and phagosome arrest. We also report polymorphisms in essential genes, efflux pumps associated genes and genes with known epitopes. The analysis of the TBM isolates and the availability of the variations obtained will provide additional resources for global comparison of isolates from pulmonary tuberculosis and TBM. It will also contribute to the richness of genomic databases towards the prediction of antibiotic resistance, level of virulence and of origin of infection.

peerj

Genetic signatures of<i>Mycobacterium tuberculosis</i>Nonthaburi genotype revealed by whole genome analysis of isolates from tuberculous meningitis patients in Thailand

Genetic signatures ofMycobacterium tuberculosisNonthaburi genotype revealed by whole genome analysis of isolates from tuberculous meningitis patients in Thailand