The goal of the proposed research is to develop reagents and procedures for detection of nucleic acids from Leishmania. Leishmania are grouped broadly into the New and Old World species. Throughout tropical and subtropical regions of the world, leishmaniasis is a serious public health concern. Effective treatments exist, yet diagnosis is difficult, owing to low abundance of parasites in the peripheral blood. Consensus probes and primers will be designed, based upon published mini circle kinetoplast sequences. Primers will incorporate inosines and/or mixed base positions to support amplification of Old and New World species. Non-radioactive microtiter plate assays have been developed for performing hybridization analysis of amplified DNA. These methods, together with simplified DNA extraction and amplification techniques will be adapted to a consensus DNA probe assay for Leishmania using laboratory strains. In Phase II, these procedures will be refined and extended to detection in clinical specimens.
