Phlebotominae sand flies (Diptera:Phlebotominae) are vectors of Leishmania parasites. During blood feeding sand flies deposit into the host skin saliva containing proteins with antigenic properties which elicit specific antibody response in bitten hosts. These anti-saliva antibodies enable to estimate the host exposure to sand flies and, in leishmaniasis endemic areas, also the risk for Leishmania infections. However, the use of whole salivary gland homogenates as antigen has several limitations and therefore recombinant salivary proteins or immunodominant peptides are tested to replace saliva in antibody detection assays.
Main aims of this study is to develop a diagnostic test based on salivary antigenic proteins of Phlebotomus perniciosus for determination of host exposure to sand fly bites and estimation of the risk of Leishmania transmission.
Immunodominant sand fly salivary antigens, obtained in a recombinant form, and peptides which mimmic immunodominant sand fly salivary antigens will be tested with sera of animals exposed to sand fly bites using ELISA and other techniques.
Recombinant proteins and peptides with highest diagnostic potential will be used to develop a new diagnostic test which will be used for phase II study in L. infantum foci.
Background: TThe IgG antibody response to sand fly salivary glands was shown to correlate with the number of sand fly bites in laboratory conditions. In the field such test would be useful to screen domestic animals to estimate their exposure to Phlebotomus perniciosus, main vector of L. infantum. The test would be useful also to evaluate the effectivity of vector control campaigns or effectivity of protection of dogs by insecticides. However, ELISA tests used at present are based on salivary gland homogenates which requires time-consuming maintenance of sand fly colonies and laborious dissections of insects.
Methodology: Immunodominant salivary antigens of Phlebotomus perniciosus obtained using E. coli expression system will be tested with sera of dogs repeatedly exposed to sand fly bites by ELISA using sand fly salivary gland homogenate as a positive control. Similar technique will be used to identify phage-displayed random peptides which mimmic immunodominant sand fly salivary antigens. The selected recombinant proteins and peptides will be characterized and optimized, and their sensitivity and specificity will be studied with animal sera already available at CUNI cryobank. Proteins and peptides with highest diagnostic potential will be used to develop a new diagnostic test. This test will be used for phase II study in L. infantum foci in Spain (IS Global).