Construction and Analysis of the Complete Genome Sequence of Leprosy Agent Mycobacterium lepromatosis

Abstract

Leprosy is caused by Mycobacterium leprae and Mycobacterium lepromatosis. We report construction and analyses of the complete genome sequence of M. lepromatosis FJ924. The genome contained 3,271,694 nucleotides to encode 1,789 functional genes and 1,564 pseudogenes. It shared 1,420 genes and 885 pseudogenes (71.4%) with M. leprae but differed in 1,281 genes and pseudogenes (28.6%). In phylogeny, the leprosy bacilli started from a most recent common ancestor (MRCA) that diverged ~30 million years ago (Mya) from environmental organism Mycobacterium haemophilum. The MRCA then underwent reductive evolution with pseudogenization, gene loss, and chromosomal rearrangements. Analysis of the shared pseudogenes estimated the pseudogenization event ~14 Mya, shortly before species bifurcation. Afterwards, genomic changes occurred to lesser extent in each species. Like M. leprae, four major types of highly repetitive sequences were detected in M. lepromatosis, contributing to chromosomal rearrangements within and after MRCA. Variations in genes and copy numbers were noted, such as three copies of the gene encoding bifunctional diguanylate cyclase/phosphodiesterase in M. lepromatosis, but single copy in M. leprae; 6 genes encoding the TetR family transcriptional regulators in M. lepromatosis, but 11 such genes in M. leprae; presence of hemW gene in M. lepromatosis, but absence in M. leprae; and others. These variations likely aid unique pathogenesis, such as diffuse lepromatous leprosy associated with M. lepromatosis, while the shared genomic features should explain the common pathogenesis of dermatitis and neuritis in leprosy. Together, these findings and the genomic data of M. lepromatosis may facilitate future research and care for leprosy. IMPORTANCE Leprosy is a dreaded infection that still affects millions of people worldwide. Mycobacterium lepromatosis is a recently recognized cause in addition to the well-known Mycobacterium leprae. M. lepromatosis is likely specific for diffuse lepromatous leprosy, a severe form of the infection and endemic in Mexico. This study constructed and annotated the complete genome sequence of M. lepromatosis FJ924 and performed comparative genomic analyses with related mycobacteria. The results afford new and refined insights into the genome size, gene repertoire, pseudogenes, phylogenomic relationship, genome organization and plasticity, process and timing of reductive evolution, and genetic and proteomic basis for pathogenesis. The availability of the complete M. lepromatosis genome may prove to be useful for future research and care for the infection.

Keywords: Mycobacterium leprae; Mycobacterium lepromatosis; genomics; leprosy; reductive evolution.

FIG 1

Phylogenomic tree of selected mycobacterial species. ML tree was inferred using a concatenated conserved protein alignment (316,716 amino acid positions) under a JTT+F+R3 substitution model. Support values were obtained with 1000 ultrafast bootstraps (right node labels) and 1000 SH-aLRT (left node labels).

FIG 2

Synteny breaks and repeat sequences identified in M. lepromatosis FJ924. The outside circle displays the 24 synteny breaks (black) detected in the genome comparing with M. leprae. The inner circle displays the positions of repeat sequences: RLPM (purple), LPMREP (red), REPLPM (blue), LPMRPT (yellow) repeat families, and other unidentified repeats (orange). Arrowheads mark the coincidences between the positions of breaks and repeat sequences.

FIG 3

Synteny among mycobacterial genomes. (Left) Genome rearrangement phylogeny obtained with the Neighbor joining method and a distance matrix showing the minimal number of inversion events required to explain the differences between any pair of genomes obtained with GRIMM. Branch lengths are the numbers of inversion events estimated with the phylogenetic method. (Right) Graphic linear representation of the genome rearrangements observed comparing the four mycobacterial genomes. The graph was obtained with Mauve and displayed with genoPlotR.

FIG 4

Comparative analyses of mycobacterial proteomes. UpSet plot showing the computed coding gene clusters (intersections) derived from the proteomes encoded in the complete genomes of M. lepromatosis FJ924, M. leprae Br4923, M. haemophilum DSM44634, and M. tuberculosis H37Rv. Shared coding gene clusters (core) and M. lepromatosis FJ924 specific coding gene clusters are highlighted in yellow and blue, respectively.

FIG 5

Relative ages of pseudogenes. Histogram with the relative ages estimated for four types of pseudogenes. A value of 1 corresponds to the time of divergence between M. haemophilum and the leprosy bacilli. A value of 0 is present time. From left to right: M. leprae shared pseudogenes (with an orthologous pseudogene in M. lepromatosis, green color), M. lepromatosis shared pseudogenes (with an orthologous pseudogene in M. leprae, orange color), M. leprae unique pseudogenes (with a CDS annotated in M. lepromatosis, blue color) and M. lepromatosis unique pseudogenes (with a CDS annotated in M. leprae, pink color).

FIG 6

Divergence times of mycobacterial species. A timetree analysis using the RelTime method with a concatenated alignment (316,716 sites amino acid sites) was used. Evolutionary model was JTT+G+I+F. The calibration node is marked (red). Divergence time in millions of years ago.

FIG 7

Phylogenetic reconstruction of diguanylate cyclase/phosphodiesterase genes in selected mycobacterial species. Maximum likelihood phylogeny obtained with the amino acid alignment (601 sites), using the evolutionary model JTT+G. Numbers at the nodes indicate bootstrap values obtained with 500 replicates.