Abstract
BACKGROUND: During leptospiral infection, the host innate immune response is initiated through recognition of pathogen-associated molecular patterns (PAMPs) by Toll-like receptor 2 (TLR2). Among these PAMPs, LipL32, Loa22, Lsa21, and lipopolysaccharide biosynthesis genes, are of particular interest. OBJECTIVE: This study aimed to investigate these molecules' genetic variability and evolutionary conservation in recently isolated clinical Leptospira strains from Sri Lanka. RESULTS: We analyzed the whole-genome sequences of 25 clinical Leptospira isolates obtained from patients across Sri Lanka, sequenced using long-read technology and annotated using a standardized pipeline. Genes encoding LipL32, Loa22, Lsa21, and enzymes within the lipopolysaccharide biosynthesis locus were extracted and analyzed for phylogenetic relationships and sequence variation. LipL32 and Loa22 were highly conserved across all isolates, with only a single amino acid substitution observed in each. In contrast, genes associated with lipopolysaccharide biosynthesis, specifically those encoding glycosyl transferase and a sodium-dependent anion transporter, exhibited notable genetic variation, including multiple single nucleotide polymorphisms leading to amino acid changes. The Lsa21 gene was present only in Leptospira interrogans strains and showed no protein-level variation. Leptospira borgpetersenii isolates demonstrated strong conservation across all gene targets at both nucleotide and protein levels. CONCLUSION: Our findings highlight the high conservation of LipL32 and Loa22, reinforcing their potential as stable targets for molecular diagnostics and serological assays. In contrast, the variability observed in lipopolysaccharide biosynthesis genes suggests a possible role in immune evasion or adaptation, warranting further functional investigation. The restricted presence of Lsa21 in specific species also raises questions about its contribution to pathogenicity.