Whole genome sequencing revealed host adaptation-focused genomic plasticity of pathogenic Leptospira

Abstract

Leptospirosis, caused by pathogenic Leptospira spp., has recently been recognized as an emerging infectious disease worldwide. Despite its severity and global importance, knowledge about the molecular pathogenesis and virulence evolution of Leptospira spp. remains limited. Here we sequenced and analyzed 102 isolates representing global sources. A high genomic variability were observed among different Leptospira species, which was attributed to massive gene gain and loss events allowing for adaptation to specific niche conditions and changing host environments. Horizontal gene transfer and gene duplication allowed the stepwise acquisition of virulence factors in pathogenic Leptospira evolved from a recent common ancestor. More importantly, the abundant expansion of specific virulence-related protein families, such as metalloproteases-associated paralogs, were exclusively identified in pathogenic species, reflecting the importance of these protein families in the pathogenesis of leptospirosis. Our observations also indicated that positive selection played a crucial role on this bacteria adaptation to hosts. These novel findings may lead to greater understanding of the global diversity and virulence evolution of Leptospira spp.

Figure 1. Accumulation curves for the pan-genome

(A) and core-genome (B) of 18 Leptospira species. Circles represented number of ortholog clusters for the different strain combinations. The red curve of panel A was a least squares fit of the power law y = k x^γ to medians, with the exponent γ > 0 indicating an open pan-genome. The red curve of panel B was least squares fit of the exponential decay y = kc exp[−x/tc] + Ω to medians, with Ω representing extrapolated core genome size. The equation was illustrated in each panel.

Figure 2. Phylogenetic analysis based on the maximum likelihood of the concatenated core genes of the Leptospira genome with Leptonema illini as the outgroup.

(A) All spirochete species genomes were included. (B) The enlarged pathogenic group of from the phylogenetic tree. Numbers before and after slash showed the numbers of gains and losses, respectively (G: gain; L: loss). There were gene gain and loss events in the evolution from the root to the lineages. Scale bar indicated an evolutionary distance of 0.05 amino acid substitutions per position.

Figure 3. Role of horizontal gene transfer (HGT), gene loss and gene duplication in pathogenic Leptospira evolution.

Scatter plots of duplication genes, HGT genes and most recent common ancestor (MRCA) lost genes versus the total number of genes in nine pathogenic Leptospira species were present. The abscissa represented the gene number of each species, and the ordinate represented the gained or lost gene number.

Figure 4. Phylogenetic analysis based on the concatenated orthologous proteins of 18 Leptospira species and the outgroup Leptonema illini using maximum likelihood method.

The complete tree was shown in Supplementary Figure 3. To the right of the tree, average numbers of 14 specific protein families that were not universally shared across all pathogenic species were indicated by different colored circles. White colored: no copies; green colored: 1 copy; orange colored: 2–4 copies; red colored: ≥5 copies. Scale bar indicated an evolutionary distance of 0.2 amino acid substitutions per position.

Figure 5. Positively selected virulence genes among the nine pathogenic Leptospira species.

Moving toward the inside, the first and second circles separately represented the 42 positively selected genes in the plus and minus strands of chromosome I of L. interrogans serovar Lai str.56601, and the gene names or loci were labeled outside of the circle. The 3^rd and 4^th circles represented GC content and GC skew, respectively. Precise molecular details for all mutations are shown in Supplementary dataset S16, S17 and S19.