Using biochemical and molecular approaches, we have identified a 9.8 kDa protein in the saliva of Ixodes scapularis that inhibits the intrinsic pathway of coagulation. The 9.8 kDa anticoagulant ...protein was purified by reverse‐phase HPLC and its N‐terminal amino acid sequence determined. The N‐terminal sequence showed homology with Salp14, an immuno‐dominant antigen present in the saliva of engorging I. scapularis nymphs. Recombinant Salp14 expressed in Escherichia coli prolonged the activated partial thromboplastin time (APTT) of human plasma in a dose‐dependent manner and was a specific inhibitor of factor Xa. A cDNA encoding a 9.3 kDa protein, Salp9Pac, was subsequently isolated from an I. scapularis salivary gland cDNA library. Salp9Pac showed 93% identity to the N‐terminal sequence of the anticoagulant purified by HPLC. These data indicate that the anticoagulant protein purified by HPLC, Salp9Pac and Salp14 are members of a family of novel coagulation protease inhibitors present in tick saliva. While recombinant Salp9Pac did not show biological activity in the assays tested currently, it is likely to be mechanistically different from its paralogues. This raises the possibility that ticks may enhance their adaptive ability to cope with a wide spectrum of proteases, by transcribing such structurally related anticoagulant proteins with different functions.
Hookworms are bloodfeeding nematodes that reside in the intestinal mucosa. These parasites secrete proteins that induce robust systemic immune responses in humans and experimental animals. By ...contrast, mucosal immune responses in and around the site of attachment are not described as well. This paper presents data from studies aimed at examining hookworm-specific mucosal antibody responses in a hamster model of Ancylostoma ceylanicum infection. Intestinal flush prepared from infected hamsters was analysed by ELISA and shown to be enriched in IgA-specific for A. ceylanicum excretory-secretory (ES) products. Evaluation of mucosal IgA responses by immunoblot demonstrated that infected hamsters recognized a broad range of ES proteins. Hamsters repeatedly exposed to drug-terminated infections were shown to have enhanced serum IgG and mucosal IgA responses, as well as a high level of protection from challenge infection. Parasite-specific IgA was also detected in the faeces of hamsters undergoing a primary infection, and increasing faecal IgA responses were coincident with significant reductions in intestinal worm burdens and faecal ES output over time. Together these results suggest that secretory IgA may act in concert with other components of the mucosal and systemic immune response to promote protective immunity against hookworm infection and/or disease.
Hookworm infection is associated with anaemia and malnutrition in many resource-limited countries. Ancylostoma hookworms have previously been shown to modulate host cellular immune responses through ...multiple mechanisms, including reduced mitogen-mediated lymphocyte proliferation, impaired antigen presentation/processing, and relative reductions in CD4⁺ T cells in the spleen and mesenteric lymph nodes. Syrian hamsters were depleted of CD4⁺ for up to 9 days following intraperitoneal injection (200 μg) of a murine anti-mouse CD4 monoclonal IgG (clone GK1·5). CD4⁺ T-cell-depleted hamsters infected with the hookworm Ancylostoma ceylanicum exhibited a threefold higher mean intestinal worm burden and more severe anaemia than animals that received isotype control IgG. In addition, depletion of CD4⁺ T cells was associated with impaired cellular and humoral (serum and mucosal) immune responses to hookworm antigens. These data demonstrate an effector role for CD4⁺ T cells in hookworm immunity and disease pathogenesis. Ultimately, these studies may yield important insights into the relationship between intestinal nematode infections and diseases that are associated with CD4⁺ T-cell depletion, including HIV.
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