Multi-omics analysis of human patient samples identifies key immune factors in Leptospira infection.

Leptospirosis is a globally neglected zoonotic disease caused by pathogenic Leptospira species and characterized by diverse clinical manifestations. However, the molecular immune responses occurring in circulating human immune cells during acute infection remain incompletely understood. Although several leptospiral virulence determinants have been investigated, a detailed characterization of host immune signaling during human infection is still limited. To characterize host molecular responses during acute leptospirosis, we performed integrated transcriptomic and proteomic profiling of peripheral blood mononuclear cells (PBMCs) from laboratory-confirmed leptospirosis patients and healthy controls. PBMCs were selected because they contain circulating immune cells involved in pathogen recognition and cytokine signaling, enabling focused analysis of host immune responses. Multi-omics network analysis was used to identify conserved immune pathways, and key host factors were further examined using ex vivo human whole-blood infection models and in vitro infection of human macrophages and monocytes. Transcriptomic profiling identified over 5,800 differentially expressed genes, predominantly upregulated, and associated with inflammatory signaling, including tumor necrosis factor signaling, interferon responses, and cytokine-mediated immune activation. Proteomic analyses detected 362 differentially expressed proteins in clinical PBMC samples and 818 differentially expressed proteins in the experimental infection model, with overlapping proteins defining a conserved host response. Comparative analysis identified 16 consistently upregulated host factors enriched in pathways related to neutrophil activation, hemostasis, and cytokine signaling. Quantitative RT-PCR confirmed increased expression of SERPINA1, BASP1, ORM1, NAMPT, and GCA, supporting their roles in inflammatory signaling and acute-phase immune responses. Together, these findings reveal conserved immune pathways activated during acute Leptospira infection and provide molecular insights into host-pathogen interactions in human leptospirosis.IMPORTANCELeptospirosis is a globally important zoonotic disease caused by bacteria of the genus Leptospira. It can lead to severe systemic complications such as kidney failure, lung injury, and multi-organ dysfunction in affected individuals. Accumulating evidence indicates that these severe clinical manifestations are closely associated with dysregulated host immune and inflammatory responses triggered during Leptospira infection. However, the molecular mechanisms underlying these immune responses, particularly the transcriptional and proteomic alterations occurring in human immune cells during infection, remain incompletely understood. In this study, we analyzed immune cells from patients with leptospirosis and identified key host molecules and immune pathways activated during infection. Experiments using human blood and immune cell models confirmed the involvement of several inflammatory host factors. These findings improve our understanding of how the human immune system responds to Leptospira infection and identify host-response molecules that may help guide future biomarker discovery and therapeutic strategies.