Mycobacterium terramassiliense, Mycobacterium rhizamassiliense and Mycobacterium numidiamassiliense sp. nov., three new Mycobacterium simiae complex species cultured from plant roots

Abstract

Three slowly growing mycobacteria named strain AB308, strain AB215 and strain AB57 were isolated from the tomato plant roots. The 16S rRNA and rpoB gene sequence analyses suggested that each strain was representative of one hitherto unidentified slowly-growing Mycobacterium species of the Mycobacterium simiae complex. Genome sequencing indicated that each strain contained one chromosome of 6.015-6.029 Mbp. A total of 1,197, 1,239 and 1,175 proteins were found to be associated with virulence and 107, 76 and 82 proteins were associated with toxin/antitoxin systems for strains AB308, AB215 and AB57, respectively. The three genomes encode for secondary metabolites, with 38, 33 and 46 genes found to be associated with polyketide synthases/non-ribosomal peptide synthases and nine, seven and ten genes encoding for bacteriocins, respectively. The genome of strain AB308 encodes for one questionable prophage and three incomplete prophages, while only incomplete prophages were predicted in AB215 and AB57 genomes. Genetic and genomic data indicate that strains AB308, AB215 and AB57 are each representative of a new Mycobacterium species that we respectively named Mycobacterium terramassiliense, Mycobacterium numidiamassiliense and Mycobacterium rhizamassiliense.

Figure 1

ESI-MS spectra of mycolic acids [M − H]⁻ ions. (A) M. rhizamassiliense AB57^T, (B) M. numidiamassiliense AB215^T, (C) M. terramassiliense AB308^T, (D) M. tuberculosis H37Rv.

Figure 2

Phylogenetic tree based on the 16S rRNA gene sequence showing the phylogenetic position of AB308, AB57 and AB 215 strains within the M. simiae complex of mycobacteria including other mycobacteria species and Mycobacterium tuberculosis H37Rv as an out group. Sequences were aligned using CLUSTLE W implemented on MEGA7 33. The analysis includes 35 nucleotide sequences. Positions containing gaps and missing data were eliminated. There were a total of 1237 positions in the final dataset. Phylogenetic inferences obtained using the maximum likelihood method based on the Tamura and Nei model (bootstrapped 1000 times). Bootstrap values > 50% are given at nodes. Bar, 0.005 substitutions per nucleotide position.

Figure 3

Transmission electron microscopy of M. terramassiliense strain AB308^T (A), M. rhizamassiliense strain AB57^T (B) and M. numidiamassiliense strain AB215^T (C). The scale bar represents 200 nm.

Figure 4

Graphical circular maps of the M. terramassiliense AB308^T, M. rhizamassiliense AB57^T and M. numidiamassiliense AB215^T genomes. From outside to the center: Contigs (red/grey), COGs category of genes on forward strand (three circles), genes on forward strand (blue circle), genes on reverse strand (red circle), COGs category on reverse strand (three circles), GC content. COGs, Clusters of Orthologous Groups database.

Figure 5

Genomic organization of M. terramassiliense AB308^T, M. numidiamassiliense AB215^T and M. rhizamassiliense AB57^T prophages.

Figure 6

Heatmap fenerated with OrthoANI values of M. terramassiliense AB308^T, M. numidiamassiliense AB215^T and M. rhizamassiliense AB57^T strains and other closest species of M. simiae complex calculated from the OAT software.