Identification of vaccine candidates for experimental visceral leishmaniasis by immunization with sequential fractions of a cDNA expression library

Visceral leishmaniasis caused by the intracellular parasite Leishmania dono vani is a significant public health problem in many regions of the world. B ecause of its large genome and complex biology, developing a vaccine for th is pathogen has proved to be a challenging task and, to date, protective re combinant vaccine candidates have not been identified. To tackle this diffi cult problem, we adopted a reductionist approach with the intention of iden tifying cDNA sequences in an L. donovani amastigote cDNA library that colle ctively or singly conferred protection against parasite challenge in a muri ne model of visceral leishmaniasis. We immunized BALB/c mice with plasmid D NA isolated and pooled from 15 cDNA sublibraries (similar to 2,000 cDNAs/su blibrary). Following systemic challenge with L. donovani, mice immunized wi th 6 of these 15 sublibraries showed a significantly reduced (35- to 1,000- fold) hepatic parasite burden. Because of the complexity and magnitude of t he sequential fractionation-immunization-challenge approach, we restricted our attention to the two sublibraries that conferred the greatest in vivo p rotection. From one of these two sublibraries, we identified several groups of cDNAs that afforded protection, including a set of nine novel cDNAs and , surprisingly, a group of five cDNAs that encoded L. donovani histone prot eins. At each fractionation step, the cDNA sublibraries or the smaller DNA fractions that afforded in vivo protection against the parasite also induce d in vitro parasite-specific T helper 1 immune responses. Our studies demon strate that immunization with sequential fractions of a cDNA library is a p owerful strategy for identifying anti-infective vaccine candidates.