Molecular approaches to understand vector-host and vector-pathogen interactions

National Institute Of Allergy And Infectious Diseases

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Abstract

The accomplishment of the section are: 1. In this study we investigated the potential of a tick secreted salivary serine protease inhibitor, IxsS7, as a novel antigenic biomarker of I. scapularis exposure in humans. We demonstrate that the IxsS7 protein-coding sequence is highly conserved (>90 % identity) among other important Ixodes species (e.g., Ixodes ricinus, Ixodes persulcatus, and Ixodes pacificus) and poorly conserved (<50 % identity) with homologs from other tick genera, such as Amblyomma spp., Dermacentor spp., Rhipicephalus spp., and Haemaphysalis spp. Antibodies in sera from rabbits immunized with recombinant IxsS7 (rIxsS7) strongly recognize native IxsS7 when challenged with salivary gland homogenate (SGH) from blood-fed I. scapularis females, while showing minimal cross-reactivity with SGH from other hard tick (Ixodidae) genera. Western blot and ELISA analyses revealed that human subjects who reported recent prior exposure to ticks possessed IgG antibodies that recognized rIxsS7, highlighting its potential as a biomarker of exposure specifically against I. scapularis. Further development of serological tools that can measure human antibody responses to Ixodes-specific salivary antigens is essential to better quantify individual- and population-level risk of important tick-borne diseases such as Lyme disease. 2.In this study we used the Delftia tsuruhatensis TC1 strain, that inhibits the development of Plasmodium in mosquitoes, but in the context of Leishmania-infected sand flies. We show that D. tsuruhatensis TC1 colonizes the midgut of Phlebotomus duboscqi sand flies and impacts the development of L. major parasites, independently of the colonization timing. This phenotype is likely an indirect consequence of D. tsuruhatensis colonization, related with the induction of sand fly gut dysbiosis. Importantly, Leishmania-infected, D. tsuruhatensis-fed sand flies are less able to transmit L. major parasites and cause disease in mice. Modelling supports the disruption of disease endemicity in the field, highlighting D. tsuruhatensis as a promising agent for the control of leishmaniasis. 3. Antibodies to sand fly salivary antigens are biomarkers for vector-host exposure in leishmaniasis-endemic regions. Ph. argentipes transmits Leishmania donovani in Sri Lanka, primarily causing cutaneous leishmaniasis (CL). Our study compared the performance of salivary gland homogenate (SGH) from a lab-reared local strain of Ph. argentipes females to a composite recombinant salivary biomarker (rPagSP02 + rPagSP06) in a CL-endemic population. Sera from 546 healthy individuals, 30 CL patients, and 15 non-endemic individuals were collected. Western blot analysis of Ph. argentipes SGH identified immunogenic bands between 15 kDa and 67 kDa, with bands of predicted molecular weight ∼of 15 kDa (SP02) and ∼28-30 kDa (SP06) as the major antibody targets. Indirect ELISAs using SGH or rPagSP02 + rPagSP06 antigens showed high sensitivity (96.7%) and specificity (100%), detecting comparable seropositivity in endemic populations. rPagSP02 + rPagSP06 exhibited enhanced discriminatory ability, supported by a strong positive correlation (r = 0.869) with SGH. Our findings indicate that the composite rPagSP02 + rPagSP06 salivary biomarker effectively identifies Ph. argentipes exposure in individuals living in Sri Lanka, showing promising potential for use in surveillance. These findings should be further validated to confirm the epidemiological applications in leishmaniasis-endemic regions. 4.The brown dog tick (Rhipicephalus sanguineus) is the vector of Rickettsia rickettsii, the causative agent of Rocky Mountain spotted fever (RMSF) in Northern Mexico and Southwestern United States. The immune response to a tick protein in the sera of humans or animals may reveal the zones with a high propensity to acquire RMSF, and vector control strategies may be focused on these zones. Arginine kinase (AK) is a highly antigenic invertebrate protein that may serve as a marker for tick exposure. We used R. sanguineus recombinant AK in an indirect ELISA assay with RMSF-positive patient sera. The response to AK was significantly higher against the sera of RMSF patients than the control sera from healthy participants without contact with dogs. To validate the antigenicity of tick AK, we mutated one predicted conformational epitope to alanine residues, which reduced the recognition by RMSF patients' immunoglobulins. This preliminary result opens a perspective towards the development of a complimentary technique based on RsAK as an antigen biomarker for vector serological surveillance for Rickettsia RMSF prevention. 5. In this study we collaborated with a research group in Sri Lanka to test if our Ph. argentipes composite recombinant salivary biomarker (rPagSP02 + rPagSP06) developed at NIAID can be used to determine if Long-lasting insecticide-treated bed nets (LLINs) are an efficacious vector control strategy to prevent sand fly bites in humans. This study assessed the efficacy of LLINs compared to untreated nets in reducing vector-human contact and density of Phlebotomus argentipes, the vector of CL in Sri Lanka. Post-intervention, antibodies against the salivary biomarker significantly decreased by 28.8% at 6 months (p=0.00002) and 44.9% at 12 months (p=0.00001) in intervention group compared to controls. LLINs effectively reduced vector exposure is a promising intervention for control of CL in Sri Lanka.