Salivary Antigen SP32 Is the Immunodominant Target of the Antibody Response to Phlebotomus papatasi Bites in Humans

Abstract

BackgroundZoonotic cutaneous leishmaniasis (ZCL) due to Leishmania major is highly prevalent in Tunisia and is transmitted by a hematophagous vector Phlebotomus papatasi (P. papatasi). While probing for a blood meal, the sand fly injects saliva into the host's skin, which contains a variety of compounds that are highly immunogenic. We recently showed that the presence of anti-saliva antibodies was associated with an enhanced risk for leishmaniasis and identified the immunodominant salivary protein of Phlebotomus papatasi as a protein of approximately 30 kDa.Methodology/Principal FindingsWe cloned and expressed in mammalian cells two salivary proteins PpSP30 and PpSP32 with predicted molecular weights close to 30 kDa from the Tunisian strain of P. papatasi. The two recombinant salivary proteins were purified by two-step HPLC (High-Performance Liquid Chromatography) and tested if these proteins correspond to the immunodominant antigen of 30 kDa previously shown to be recognized by human sera from endemic areas for ZCL and exposed naturally to P. papatasi bites. While recombinant PpSP30 (rPpSP30) was poorly recognized by human sera from endemic areas for ZCL, rPpSP32 was strongly recognized by the tested sera. The binding of human IgG antibodies to native PpSP32 was inhibited by the addition of rPpSP32. Consistently, experiments in mice showed that PpSP32 induced the highest levels of antibodies compared to other P. papatasi salivary molecules while PpSP30 did not induce any detectable levels of antibodies.ConclusionsOur findings demonstrate that PpSP32 is the immunodominant target of the antibody response to P. papatasi saliva. They also indicate that the recombinant form of PpSP32 is similar to the native one and represents a good candidate for large scale testing of human exposure to P. papatasi bites and perhaps for assessing the risk of contracting the disease.

Comparative Analysis of Salivary Gland Transcriptomes of Phlebotomus orientalis Sand Flies from Endemic and Non-endemic Foci of Visceral Leishmaniasis

“…Recombinant P. ariasi D7-related protein (AAX55749) elicited the production of specific humoral response in immunized mice [27]. Anti- P. papatasi saliva antibodies reacted with the 30 kDa recombinant P. papatasi D7-related protein (AAL11049) [5], but the same protein was not recognized by the human sera from an endemic area of CL in Tunisia [100]. Moreover, recombinant 28 kDa D7-related protein from P. papatasi saliva (AAL11048) was not targeted by the specific antibodies of immunized mice [5].…”

A sand fly salivary protein vaccine shows efficacy against vector-transmitted cutaneous leishmaniasis in nonhuman primates

“…Substantiating evidence is demonstrated by the cross-protection of P. papatasi-exposed mice against P. duboscqi salivary gland extract plus L. major (21). Notably, 90% of 200 children from a CL endemic area in Tunisia, where L. major is transmitted by P. papatasi, also recognized a protein of 15 kD, likely PpSP15, by Western blot (22,23). Also important to vaccine development, PdSP15 is foreign to humans.…”

In Silico Identification of a Candidate Synthetic Peptide (Tsgf118–43) to Monitor Human Exposure to Tsetse Flies in West Africa

“…Resort to recombinant proteins has proved to be an efficient way to develop standardized immunoassays to a variety of disease vectors such as ticks, sanflies, triatomines or to Afrotropical malaria vectors [27], [28], [30], [31], [51]. Nevertheless, the production and the storage/shipment of recombinant proteins are often problematic and require good facilities which limit their use in many contexts such as in developing countries.…”