Genomic Comparison Among Global Isolates of L. interrogans Serovars Copenhageni and Icterohaemorrhagiae Identified Natural Genetic Variation Caused by an Indel

Abstract

Leptospirosis is a worldwide zoonosis, responsible for more than 1 million cases and 60,000 deaths every year. Among the 13 pathogenic species of the genus Leptospira, serovars belonging to L. interrogans serogroup Icterohaemorrhagiae are considered to be the most virulent strains, and responsible for majority of the reported severe cases. Serovars Copenhageni and Icterohaemorrhagiae are major representatives of this serogroup and despite their public health relevance, little is known regarding the genetic differences between these two serovars. In this study, we analyzed the genome sequences of 67 isolates belonging to L. interrogans serovars Copenhageni and Icterohaemorrhagiae to investigate the influence of spatial and temporal variations on DNA sequence diversity. Out of the 1072 SNPs identified, 276 were in non-coding regions and 796 in coding regions. Indel analyses identified 258 indels, out of which 191 were found in coding regions and 67 in non-coding regions. Our phylogenetic analyses based on SNP dataset revealed that both serovars are closely related but showed distinct spatial clustering. However, likelihood ratio test of the indel data statistically confirmed the presence of a frameshift mutation within a homopolymeric tract of lic12008 gene (related to LPS biosynthesis) in all the L. interrogans serovar Icterohaemorrhagiae strains but not in the Copenhageni strains. Therefore, this internal indel identified can genetically distinguish L. interrogans serovar Copenhageni from serovar Icterohaemorrhagiae with high discriminatory power. To our knowledge, this is the first study to identify global sequence variations (SNPs and Indels) in L. interrogans serovars Copenhageni and Icterohaemorrhagiae.

Detection of New Leptospira Genotypes Infecting Symptomatic Dogs: Is a New Vaccine Formulation Needed?

“…Interestingly, the IZSLER database revealed that Leptospira ST17 occurs in different hosts (mouse, cat, hedgehog, horse, goat and cow), which indicates its ability to cause infection in a wide range of animal species. The sequencing of the lic12008 gene, which was recently reported by Santos et al [ 54 ] as useful in discriminating Icterohaemorrhagiae and Copenhageni serovars, allowed us to determine that the major portion (94%) of samples typed as ST17 represented serovar Icterohaemorrhagiae, and only a small portion (6%) represented serovar Copenhageni. The reliability of this molecular test for serovar discrimination is still unclear, and further studies are necessary, given that the serovar determinations of two isolates, as assessed by mAbs and by lic12008 sequencing, were not in agreement ( Table 5 ).…”

“…A tract of the lic12008 gene of samples genotyped as ST17 (characterized as L. interrogans serogroup Icterohaemorrhagiae) was sequenced. The tract was recently found to be involved in the genetic distinction between serovars Icterohaemorrhagiae and Copenhageni [ 54 ], which are indistinguishable by MLST and MLVA. The analysis determines the presence of a single base insertion of a thymine nucleotide within a poly-thymine tract (9-bp long) in the lic12008 of all L. interrogans serovar Icterohaemorrhagiae isolates but not in Copenhageni strains.…”

A Validated Multiplex Real-Time PCR Assay for the Diagnosis of Infectious Leptospira spp.: A Novel Assay for the Detection and Differentiation of Strains From Both Pathogenic Groups I and II

“…The fact that the 16S RNA gene has been vastly supported as a good phylogenetic marker for Leptospira spp. classification (Fouts et al, 2016;Santos et al, 2018;Vincent et al, 2019) guarantees that this gene contains regions enabling the differentiation between the major groups of the Letospira spp. Moreover, using all sequences available at the GenBank database ensured that the design of the primers and probes in the current study had a broad and selective spectrum for the detection of all infectious Leptospira spp., with the exclusion of all saprophytic group and the novel clade of environmental Leptospira spp.…”

Insight into motility-dependent pathogenicity of the zoonotic spirocheteLeptospira

“…Pathogenic serovars of L. interrogans serovar Icterohaemorrhagiae (strain WFA135), an isolate from Rattus norvegicus in Tokyo, Japan, serovar Manilae (strain UP-MMC-NIID) 31,32 and a saprophytic L. biflexa serovar Patoc (strain Patoc I) were used in this study. The serovar of WFA135 was determined by multiple loci variable number of tandem repeats analysis (MLVA) and DNA sequencing of the lic12008 gene 32,33 . Bacteria were cultured in enriched Ellinghausen-McCullough-Johnson-Harris (EMJH) liquid medium (BD Difco, NJ, USA) containing 25 μg/mL spectinomycin at 30 °C for 2 ( L. biflexa ) or 4 ( L. interrogans ) days until the stationary phase.…”