Mycobacterium tuberculosis complex lineage 5 exhibits high levels of within-lineage genomic diversity and differing gene content compared to the type strain H37Rv

Abstract

Pathogens of the Mycobacterium tuberculosis complex (MTBC) are considered to be monomorphic, with little gene content variation between strains. Nevertheless, several genotypic and phenotypic factors separate strains of the different MTBC lineages (L), especially L5 and L6 (traditionally termed Mycobacterium africanum ) strains, from each other. However, this genome variability and gene content, especially of L5 strains, has not been fully explored and may be important for pathobiology and current approaches for genomic analysis of MTBC strains, including transmission studies. By comparing the genomes of 355 L5 clinical strains (including 3 complete genomes and 352 Illumina whole-genome sequenced isolates) to each other and to H37Rv, we identified multiple genes that were differentially present or absent between H37Rv and L5 strains. Additionally, considerable gene content variability was found across L5 strains, including a split in the L5.3 sub-lineage into L5.3.1 and L5.3.2. These gene content differences had a small knock-on effect on transmission cluster estimation, with clustering rates influenced by the selected reference genome, and with potential overestimation of recent transmission when using H37Rv as the reference genome. We conclude that full capture of the gene diversity, especially high-resolution outbreak analysis, requires a variation of the single H37Rv-centric reference genome mapping approach currently used in most whole-genome sequencing data analysis pipelines. Moreover, the high within-lineage gene content variability suggests that the pan-genome of M. tuberculosis is at least several kilobases larger than previously thought, implying that a concatenated or reference-free genome assembly (de novo) approach may be needed for particular questions.

Understanding the Genetic Diversity of Mycobacterium africanum Using Phylogenetics and Population Genomics Approaches

“…L6 is known to be an opportunistic pathogen owing to mutations in genes essential for growth and contributes toward latent TB burden in West Africa ( de Jong et al, 2005 ; de Jong et al, 2010a ; Gehre et al, 2013 ; Ofori-Anyinam et al, 2017 ). Similar studies for L5 genomics are limited that has been highlighted earlier ( Yeboah-Manu et al, 2017 ; Ates et al, 2018 ; Coscolla et al, 2021 ; Sanoussi et al, 2021 ). A slower response to TB treatment for L6 was observed when compared to other sensu stricto lineages ( Diarra et al, 2018 ).…”

“…Lineage identification is of considerable significance in tuberculosis control, as it helps in quick diagnosis leading to effective treatment and prevention of potential future outbreaks ( Dou et al, 2017 ; Napier et al, 2020 ). Although extensive studies pertaining to the differentiation of L5 and L6 have been carried out in the past, recent reports revealed the existence of underlying sub-lineages ( Ates et al, 2018 ; Coscolla et al, 2021 ; Sanoussi et al, 2021 ). Methods such as phylogeny and PCA have been used to understand the lineage distribution of L5 and L6, with reports of incongruous clustering of L5 ( Coscolla et al, 2021 ).…”

“…Methods such as phylogeny and PCA have been used to understand the lineage distribution of L5 and L6, with reports of incongruous clustering of L5 ( Coscolla et al, 2021 ). Recent studies have also suggested clustering of L5 using “RD711” and other large sequence polymorphisms ( Comín et al, 2021 ; Coscolla et al, 2021 , Sanoussi et al, 2021 ). Taking these observations into cognizance, an attempt was made to understand the fine-level population stratification of Maf, especially L5 and its sub-lineages using SNPs with the aid of model- and non-model-based clustering approaches.…”

An updated evolutionary history and taxonomy ofMycobacterium tuberculosislineage 5, also calledM. africanum

“…Long-read sequencing on three geographically distant isolates of L5 suggested that using H37Rv as a reference genome is limiting and showed that the pangenome of L5 could be larger than expected previously (Sanoussi et al, 2021). Preliminary results using de novo alignment using Roary indeed suggests that some L5 may harbor specific genes that are not found in H37Rv (results not shown) (Page et al, 2015).…”

Tuberculosis caused by Mycobacterium africanum: Knowns and unknowns

“…Whole-genome analyses also revealed the existence of genetic diversity within M . africanum L5 and L6 strains [ 9 , 38 – 40 ] and allowed the construction of detailed MTBC phylogeny trees. It is interesting to note that strains of the M .…”