Leptospiral VM Protein Antigen Detection for Rapid Diagnosis of Leptospirosis

Summary Pathogenic members of the genus, Leptospira, in particular L. interrogans, cause leptospirosis, a zoonotic bacterial disease of global importance, which affects more than 1 million people annually with a conservatively estimated 5-20% case fatality rate. Early diagnosis of leptospirosis leading to specific antimicrobial treatment is thought to prevent progression to severe disease, which is characterized by fever, refractory shock, pulmonary (and other) hemorrhage, and acute oliguric renal failure. Because the initial manifestations of leptospirosis - acute undifferentiated fever — are non-specific, the diagnosis of this disease is often missed or delayed. The potential market for a diagnostic test that identifies early, acute leptospirosis is substantial and an unmet need, both in terms of clinical care as well as for quantifying the burden of leptospirosis disease that is of prime interest to public health authorities. In the developing tropical world, leptospirosis affects large swaths of endemic populations as well as those subject to major weather-related events such as flooding in large urban settings that occur with predictable regularity. Clinically-actionable tests to diagnose acute leptospirosis remain unavailable. This proposal aims to address this gap by developing tests capable of rapid, point-of-care, actionable diagnosis. The premise of this application is that tests based on leptospiral antigen in body fluids (whole blood, serum, urine) will rapidly and specifically diagnose leptospirosis, leading to improved clinical care and providing more precise public health data related to burden of disease policymaking. The hypothesis underlying this proposal is that detecting abundant leptospiral antigens including Virulence Modifying proteins (VMP) (based on new preliminary data), pathogenic Leptospira-specific LipL32, and Icterohaemorrhagiae serogroup lipopolysaccharide (LPS) in patient blood will lead to actionable therapeutic interventions and enable burden of disease studies to justify further leptospirosis product development such as vaccines and biologics. Specific Aims: 1) Optimize and validate solid phase VMP-based antigen-detection leptospirosis diagnostic tests. 2) Determine sensitivity and specificity of multiplex antigen-detection solid phase assays, adding LipL32 and LPS antigen-detection leptospirosis diagnostic tests. The performance of new ELISA and lateral flow assay prototypes will be determined using in vitro spiked specimens using normal human blood, serum and urine matrices, ex vivo specimens from acute and chronic animal models, and endemic country-obtained de-identified leptospirosis human samples. This Phase I STTR project will yield intellectual property and prototypes of solid phase leptospiral antigen-detecting assays for diagnosis of human leptospirosis. In a Phase II project, we will scale-up manufacturing for ELISA kits and LFA devices, and expand field-based clinical testing in humans in established leptospirosis-endemic regions and at US-based clinical reference laboratories.