The human Toll-like receptor 2 (TLR2) response during pathogenicLeptospirainfection
Authors: Kappagoda CN , Senevirathne RMISK , Jayasundara D , Warnasekara YPJN , Srimantha LASM , De Silva LAPNF , Agampodi SB 
Published: November 2023

AbstractBackgroundHuman innate immune responses are triggered through the interaction of human pattern recognition receptors and pathogen-associated molecular patterns. The role of toll-like receptor2 (TLR2) in mice innate immune response to leptospirosis is well established, while human studies are limited. The present study aimed to determine the TLR2 response among confirmed cases of leptospirosis.Methodology/Principle findingsThe study has two components. Clinically suspected patients of leptospirosis were confirmed using a previously validated qPCR assay. Total RNA was extracted from patients’ RNA-stabilized whole blood samples. Human TLR2 gene expression (RT-qPCR) analysis was carried out using an exon-exon spanning primer pair, using CFX Maestro™ software. The first set of patient samples was used to calculate the Relative Normalized Expression (ΔΔCq value) of the TLR2 gene in comparison to a healthy control sample and normalized by the reference gene GAPDH (Glyceraldehyde-3-phosphate dehydrogenase). Secondly, recruited patient samples were subjected to TLR2 gene expression analysis and compared to healthy controls and normalized by the reference genes Beta-2-microglobulin(B2M), Hypoxanthine phosphoribosyltransferase 1 (HPRT 1).In the initial cohort of 64 confirmed leptospirosis cases, 18 were selected for human TLR2 gene expression analysis based on criteria of leptospiremia and RNA yield. Within this group, one individual exhibited a down-regulation of TLR2 gene (Expression/ΔΔCq=0.01352), whereas the remaining subjects presented no significant change in gene expression. In a subsequent cohort of 23 confirmed cases, 13 were chosen for similar analysis. Among these, three patients demonstrated down-regulation of TLR2 gene expression, with Expression/ΔΔCq values of 0.86574, 0.47200, and 0.28579, respectively. No TLR2 gene expression was noted in the other patients within this second group.ConclusionsOur investigation into the acute phase of leptospirosis using human clinical samples has revealed a downregulation of TLR2 gene expression. This observation contrasts to the upregulation commonly reported in the majority of in-vitro and in-vivo studies ofLeptospirainfection. These preliminary findings prompt a need for further research to explore the mechanisms underlying TLR2’s role in the pathogenesis of leptospirosis, which may differ in clinical settings compared to laboratory models.Author SummaryThe human immune system employs pattern recognition receptors like toll-like receptor 2 (TLR2) to detect and combat infections such as leptospirosis. While TLR2’s role is well-documented in mice, its function in the human response to leptospirosis remains unclear. Our study evaluated TLR2 activity in patients with confirmed leptospirosis. We conducted a genetic analysis of blood samples from these patients, comparing TLR2 gene activity against healthy individuals, with standard reference genes for accuracy. Contrary to expectations and existing laboratory data, we observed a decrease in TLR2 activity in some patients. This suggests that human TLR2 responses in actual infections may diverge from established laboratory models. These findings indicate a need for further study to understand the human immune response to leptospirosis, which may significantly differ from that observed in controlled experimental settings.