Comparative genomic analysis of Mycobacterium tuberculosis drug resistant strains from Russia

Abstract

Tuberculosis caused by multidrug-resistant (MDR) and extensively drug-resistant (XDR) Mycobacterium tuberculosis (MTB) strains is a growing problem in many countries. The availability of the complete nucleotide sequences of several MTB genomes allows to use the comparative genomics as a tool to study the relationships of strains and differences in their evolutionary history including acquisition of drug-resistance. In our work, we sequenced three genomes of Russian MTB strains of different phenotypes--drug susceptible, MDR and XDR. Of them, MDR and XDR strains were collected in Tomsk (Siberia, Russia) during the local TB outbreak in 1998-1999 and belonged to rare KQ and KY families in accordance with IS6110 typing, which are considered endemic for Russia. Based on phylogenetic analysis, our isolates belonged to different genetic families, Beijing, Ural and LAM, which made the direct comparison of their genomes impossible. For this reason we performed their comparison in the broader context of all M. tuberculosis genomes available in GenBank. The list of unique individual non-synonymous SNPs for each sequenced isolate was formed by comparison with all SNPs detected within the same phylogenetic group. For further functional analysis, all proteins with unique SNPs were ascribed to 20 different functional classes based on Clusters of Orthologous Groups (COG). We have confirmed drug resistant status of our isolates that harbored almost all known drug-resistance associated mutations. Unique SNPs of an XDR isolate CTRI-4(XDR), belonging to a Beijing family were compared in more detail with SNPs of additional 14 Russian XDR strains of the same family. Only type specific mutations in genes of repair, replication and recombination system (COG category L) were found common within this group. Probably the other unique SNPs discovered in CTRI-4(XDR) may have an important role in adaptation of this microorganism to its surrounding and in escape from antituberculosis drugs treatment.

Figure 1. Venn diagram of single nucleotide polymorphisms in three studied isolates.

Figure 2. Comparative phylogenetic analysis of strains under study and 22 whole genomes from the NCBI database.

Tree based on all SNPs of genomes. Phylogenetic tree was constructed using the Neighbor-Joining algorithm. Evolutionary distances were calculated using p-distance method. The bootstrap test was for 1000 replicates. Branches corresponding to partitions reproduced in less than 50% bootstrap replicates are collapsed.

Figure 3. The relative distribution of individual SAPs found in proteins belonged to certain COGs.

C - energy production and conversion; D - cell cycle control, cell division, chromosome partitioning; E - amino acid transport and metabolism; F - nucleotide transport and metabolism; G - carbohydrate transport and metabolism; H - coenzyme transport and metabolism, I - lipid transport and metabolism, J - translation, ribosomal structure and biogenesis; K – transcription; L - replication, recombination and repair; M - cell wall/membrane/envelope biogenesis, N - cell motility, O - posttranslational modification, protein turnover, chaperones; P - inorganic ion transport and metabolism; Q - secondary metabolites biosynthesis, transport, and catabolism; R - general function prediction only; S - function unknown; T - signal transduction mechanisms; U - intracellular trafficking, secretion, and vesicular transport; V - defense mechanisms.