Genomic analysis of smooth tubercle bacilli provides insights into ancestry and pathoadaptation of Mycobacterium tuberculosis

Abstract

Global spread and genetic monomorphism are hallmarks of Mycobacterium tuberculosis, the agent of human tuberculosis. In contrast, Mycobacterium canettii, and related tubercle bacilli that also cause human tuberculosis and exhibit unusual smooth colony morphology, are restricted to East-Africa. Here, we sequenced and analyzed the genomes of five representative strains of smooth tubercle bacilli (STB) using Sanger (4-5x coverage), 454/Roche (13-18x coverage) and/or Illumina DNA sequencing (45-105x coverage). We show that STB are highly recombinogenic and evolutionary early-branching, with larger genome sizes, 25-fold more SNPs, fewer molecular scars and distinct CRISPR-Cas systems relative to M. tuberculosis. Despite the differences, all tuberculosis-causing mycobacteria share a highly conserved core genome. Mouse-infection experiments revealed that STB are less persistent and virulent than M. tuberculosis. We conclude that M. tuberculosis emerged from an ancestral, STB-like pool of mycobacteria by gain of persistence and virulence mechanisms and we provide genome-wide insights into the molecular events involved.

Key experimental evidence of chromosomal DNA transfer among selected tuberculosis-causing mycobacteria

“…Thus, phage-mediated DNA transfer can be excluded as a reason for the observed genome mosaicism. In addition, no prophage were detected in the genomes of M. canettii A and L, in contrast to M. canettii I, where a 55-kb prophage region was found (10). However, no recombinants were obtained when M. canettii I was used as a donor (SI Appendix, Table S1).…”

“…To assess various M. canettii and M. tuberculosis strains for their ability to exchange chromosomal DNA, green-fluorescent putative donor strains were constructed by transformation with pYUB412 or F10 integrating cosmids (20) as established before for M. smegmatis strains. Redfluorescent potential M. canettii recipient strains were obtained by electroporation with plasmid pMRF1-DsRed, which was successful for all strains except M. canettii J (CIPT 140070017), which exclusively harbors an eptABCD gene cluster (10,21), recently described as restricting plasmid maintenance in M. smegmatis (22).…”

“…First hints came from putative recombination tracts identified by in silico analyses in genomes of smooth tubercle bacilli of the M. canettii clade (10,11). M. canettii strains are remarkable, rare mycobacteria, which are almost exclusively isolated from patients in the region of the Horn of Africa (35, 36).…”

“…Consequently, this type of HGT was termed distributive conjugal transfer (DCT) (16). Detection of mosaic sequence arrangements in M. canettii genomes (10,11,17,18) raised the issue whether similar molecular mechanisms might be active in tuberculosis-causing mycobacteria and/or in mycobacteria, in general.…”

“…Recent whole genome sequence (WGS) analyses of early branching, smooth tubercle bacilli (STB) of the Mycobacterium canettii clade revealed genome-wide traces of putative interstrain recombination events (10,11), raising questions on the existence and functionality of particular HGT mechanisms in tubercle bacilli. An interesting observation in this respect was made for the nonvirulent, fast-growing model organism Mycobacterium smegmatis, which shows an unconventional conjugal DNA transfer (12,13), distinct from classical plasmid-mediated DNA transfer described for other bacteria (14,15).…”