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

Robert Geffers,Boyke Bunk,Sahlawati Mustakim,Sabin Bhuju,Michael Jarek,Marga Goris,Siti Roszilawati Ramli,Cathrin Spröer,Frank Pessler

doi:10.3390/pathogens10091198

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 chromosomal core genome of 3318 coding sequences and strain-specific accessory genomes of 49–179 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.

Highlights

Pathogenic Leptospira spp. colonize the proximal renal tubules of reservoir animal hosts and are excreted through urine into the external environment
The results underscore the typical transmission of L. interrogans in moist environments and the broad spectrum of clinical manifestations and severity and identify 1530 as the strain associated with the mildest disease severity
The six L. interrogans strains were assigned to the traditional serogroups by microagglutination test (MAT), using a panel of 43 polyclonal rabbit anti-Leptospira reference sera, representing 24 pathogenic and two saprophytic serogroups (Table S2)

Summary

Introduction

Pathogenic Leptospira spp. colonize the proximal renal tubules of reservoir animal hosts and are excreted through urine into the external environment. The persistence of Leptospirae in moist environments and animal hosts combined with their ability to cause clinically highly variable disease in humans has made leptospirosis one of the most common zoonotic diseases worldwide [2]. Leptospirae span a broad spectrum of virulence and hosts, ranging from low pathogenic strains in the environment to those that cause severe disease in farm animals and humans [5]. L. interrogans is probably the most important species causing human leptospirosis worldwide. It can infect or colonize all mammals and was reported to be the most common Leptospira species among rodents in South East Asia [6]. Of the multitude of serovars and serogroups of this species, Icterohaemorrhagiae is the one most commonly associated with severe leptospirosis in humans [7]

Methods

Results

Conclusion