Complete Genome Sequencing of Leptospira interrogans Isolates from Malaysia Reveals Massive Genome Rearrangement but High Conservation of Virulence-Associated Genes

Abstract

The ability of Leptospirae to persist in environments and animal hosts but to cause clinically highly variable disease in humans has made leptospirosis the most common zoonotic disease. Considering the paucity of data on variation in complete genomes of human pathogenic Leptospirae, we have used a combination of Single Molecule Real-Time (SMRT) and Illumina sequencing to obtain complete genome sequences of six human clinical L. interrogans isolates from Malaysia. All six contained the larger (4.28–4.56 Mb) and smaller (0.34–0.395 Mb) chromosome typical of human pathogenic Leptospirae and 0–7 plasmids. Only 24% of the plasmid sequences could be matched to databases. We identified a core genome of 3271 coding sequences and strain-specific accessory genomes of 50–352 coding sequences. These sequences enabled detailed genomic strain typing (Genome BLAST Distance Phylogeny, DNA–DNA hybridization, and multi locus sequence typing) and phylogenetic classification (whole-genome SNP genotyping). Even though there was some shared synteny and collinearity across the six genomes, there was evidence of major genome rearrangement, likely driven by horizontal gene transfer and homologous recombination. Mobile genetic elements were identified in all strains in highly varying numbers, including in the rfb locus, which defines serogroups and contributes to immune escape and pathogenesis. On the other hand, there was high conservation of virulence-associated genes including those relating to sialic acid, alginate, and lipid A biosynthesis. These findings suggest (i) that the antigenic variation, adaption to various host environments, and broad spectrum of virulence of L. interrogans are in part due to a high degree of genomic plasticity and (ii) that human pathogenic strains maintain a core set of genes required for virulence.

Whole genome sequencing data of Leptospira weilii and Leptospira kirschneri isolated from human subjects of Sri Lanka

“…2.0. g-tubes TM from Covaris (Woburn, MA, USA) was used to shear 1 g of genomic DNA, and AMPure PB beads from Pacific Bioscience were used to concentrate the DNA. The DNA was finally repaired following an overnight ligation to an affixed barcoded 8A adaptor (Pacific Bioscience) [ 4 , 5 ]. The size selection instructions provided by the manufacturer, Sage Science, Beverly, Massachusetts, USA, for Blue Pippin TM 4 kb or more were adhered to.…”

Cleavage of cell junction proteins as a host invasion strategy in leptospirosis

“…During the last decade, research work targeting the pathogenesis mechanism of leptospirosis has seen an upsurge in the identification of components involved in the adhesion, colonization, immune evasion, and establishment of pathogens in the host system. Even though the complete genome is available for many pathogenic and non-pathogenic strains (Ramli et al 2021 ; Vincent et al 2019 ; Thibeaux et al 2018 ) and many studies comparing the genomes are published, a clear picture of the pathogenesis mechanism is not available. In Leptospira , as per the current data, nearly 10 proteins were found to act on the cell junction proteins of the host system and play a role in the invasion/colonization process (Evangelista et al 2014a , b ; Eshghi et al 2019 ; Pinne et al 2010 and many more).…”

Sequencing Approaches for Understanding the Pathogenic Features in Leptospira Genome

“…LipL32 is also highly conserved in pathogenic, and it is expressed during infection. Ramli et al [53] concluded that L. interrogans is a dynamic pathogen and there are still many virulence features to be discovered.…”