Whole genome SNP analysis suggests unique virulence factor differences of the Beijing and Manila families of Mycobacterium tuberculosis found in Hawaii

Abstract

While tuberculosis (TB) remains a global disease, the WHO estimates that 62% of the incident TB cases in 2016 occurred in the WHO South-East Asia and Western Pacific regions. TB in the Pacific is composed predominantly of two genetic families of Mycobacterium tuberculosis (Mtb): Beijing and Manila. The Manila family is historically under-studied relative to the families that comprise the majority of TB in Europe and North America (e.g. lineage 4), and it remains unclear why this lineage has persisted in Filipino populations despite the predominance of more globally successful Mtb lineages in most of the world. The Beijing family is of particular interest as it is increasingly associated with drug resistance throughout the world. Both of these lineages are important to the State of Hawaii, where they comprise over two-thirds of TB cases. Here, we performed whole genome sequencing on 82 Beijing family, Manila family, and outgroup clinical Mtb isolates from Hawaii to identify lineage-specific SNPs (SNPs found in all isolates from their respective families, and exclusively in those families) in established virulence factor genes. Six non-silent lineage-specific virulence factor SNPs were found in the Beijing family, including mutations in alternative sigma factor sigG and polyketide synthases pks5 and pks7. The Manila family displayed more than eleven non-silent lineage-specific and characteristic virulence factor mutations, including in genes coding for MCE-family protein Mce1B, two mutations in fatty-acid-AMP ligase FadD26, and virulence-regulating transcriptional regulator VirS. This study further identified an ancient clade that shared some virulence factor mutations with the Manila family, and investigated the relationship of those and other “Manila-like” spoligotypes to the Manila family with this SNP dataset. This work identified a set of virulence genes that are worth pursuing to determine potential differences in transmission or virulence displayed by these two Mtb families.

Virulence of Mycobacterium tuberculosis Clinical Isolates Is Associated With Sputum Pre-treatment Bacterial Load, Lineage, Survival in Macrophages, and Cytokine Response

“…This was further supported by the observation that there was an increased bacterial number in sputum of patients infected with virulent Mtb defined by rapid macrophage lysis, as bacterial load in sputum is one of the risk factors for TB transmission. A recent study has suggested that different Mtb lineages may carry their unique lineage-specific genetic markers in virulence genes (Koster et al, 2018 ). Therefore, the association of Mtb genetics and the virulence defined by macrophage lysis will be investigated in the next stage of this work.…”

Connection between two historical tuberculosis outbreak sites in Japan, Honshu, by a new ancestral Mycobacterium tuberculosis L2 sublineage

“…In brief, regarding the set-up of the platform, the processes are the following: SRAs of interest are selected and kept only provided a certain number of conditions that together reinforce the reliability of the data: they must have read length>75 bp, clean reads file must be at least 100 Mo, and CRISPR could be reconstructed using CRISPRbuilder-TB [ 54 ]. For each SRA, in addition to reconstructing the CRISPR-Cas region using CRISPRbuilder-TB and apart from collecting NCBI information on the genomes, two scripts successively perform the following tasks: (1) search for SNPs according to reference catalogues (Supplementary Table S1) totalling more than 50 000 SNPs, including drug-resistance related SNPs, phylogenetic SNPs as per 26 studies contributing to SNP-based classification [ 26 , 28 – 30 , 33 , 55 – 75 ]; (2) search for additional SNPs in each isolate based on H37Rv reference sequence; (3) look for the presence/absence of H37Rv genes as annotated in mycobrowser ( ), (4) look for the presence/absence of deletion regions; (5) identify insertion sites of all known insertion sequences in MTBC. The CRISPR locus is rebuilt semi-automatically using a dedicated and previously published script (43/68/360 Spacers format) with an assignment of a Spoligo-International-Type (SIT) tag; the application produces an ordered list of spacers/repeat with variants and IS 6110 insertion sequences if present [ 54 , 76 , 77 ].…”

Genomic sequencing is required for identification of tuberculosis transmission in Hawaii

“…Making full use of the resulting WGS dataset, we further examined isolates from clusters that WGS identified to represent actual transmission events and investigated which genes or regions were developing mutations that differentiated individual isolates in a cluster. Our previous work has identified virulence factor mutations in the Beijing and Manila families that may be involved in virulence or latency, and this work seeks to help us further characterize these historically under-studied families [ 29 , 30 ].…”