Functional Transcriptomics of Wild-Caught Lutzomyia intermedia Salivary Glands: Identification of a Protective Salivary Protein against Leishmania braziliensis Infection

Abstract

Background Leishmania parasites are transmitted in the presence of sand fly saliva. Together with the parasite, the sand fly injects salivary components that change the environment at the feeding site. Mice immunized with Phlebotomus papatasi salivary gland (SG) homogenate are protected against Leishmania major infection, while immunity to Lutzomyia intermedia SG homogenate exacerbated experimental Leishmania braziliensis infection. In humans, antibodies to Lu. intermedia saliva are associated with risk of acquiring L. braziliensis infection. Despite these important findings, there is no information regarding the repertoire of Lu. intermedia salivary proteins.Methods and FindingsA cDNA library from the Salivary Glands (SGs) of wild-caught Lu. intermedia was constructed, sequenced, and complemented by a proteomic approach based on 1D SDS PAGE and mass/mass spectrometry to validate the transcripts present in this cDNA library. We identified the most abundant transcripts and proteins reported in other sand fly species as well as novel proteins such as neurotoxin-like proteins, peptides with ML domain, and three small peptides found so far only in this sand fly species. DNA plasmids coding for ten selected transcripts were constructed and used to immunize BALB/c mice to study their immunogenicity. Plasmid Linb-11—coding for a 4.5-kDa protein—induced a cellular immune response and conferred protection against L. braziliensis infection. This protection correlated with a decreased parasite load and an increased frequency of IFN-γ-producing cells.ConclusionsWe identified the most abundant and novel proteins present in the SGs of Lu. intermedia, a vector of cutaneous leishmaniasis in the Americas. We also show for the first time that immunity to a single salivary protein from Lu. intermedia can protect against cutaneous leishmaniasis caused by L. braziliensis.

Immunity to Lutzomyia whitmani Saliva Protects against Experimental Leishmania braziliensis Infection

“…These components could play important roles in exposure to sand flies and highlight the importance of testing vaccine candidates against the challenge of infected sand fly bites [38]. Together, our data reinforce the possibility of employing sand fly salivary molecules as components for a leishmaniasis vaccine [9,10,40,45]. In parallel, we also envisage the development of Lu .…”

“…Regarding differences in the antigenicity of salivary proteins, the sialome of Lu . intermedia has been described [40] but characterization of the salivary proteins from Lu . whitmani is still lacking so we cannot, presently, probe for possible differences that could explain the distinct outcomes.…”

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

“…Members of the Lufaxin family were detected in saliva of various sand flies [26]–[28], [31], [50], [52], [54]–[56], but not in other bloodsucking insects. Sequences coding for a Lufaxin-like protein, were detected in the cDNA library of MW P. orientalis colony (PorMSP78/KC170976) (Table 2).…”

Ayadualin, a novel RGD peptide with dual antihemostatic activities from the sand fly Lutzomyia ayacuchensis, a vector of Andean-type cutaneous leishmaniasis

“…This unique form of the RGD peptide is found only in the saliva of Lutzomyia species [18][19][20][21], and ayadualin is the first salivary protein from sand flies to be found to have a dual inhibitory effect on hemostasis. In addition, this peptide inhibits platelet aggregation by a different mechanism from apyrase, the only known antiplatelet component from sand fly saliva, and is the first inhibitor of the intrinsic coagulation pathway from sand flies.…”

“…ayacuchensis, and Lu. intermedia [18][19][20][21]. In the salivary transcriptome analysis of [22][23][24] and L. (Viannia) peruviana in Peruvian Andes [24,25], a peptide containing an RGD (Arg-Gly-Asp) sequence flanked by cysteine residues in the C-terminal end was identified as the most abundant transcript.…”