Field Evaluation of a Fluorogenic Probe-Based PCR Assay for Identification of a Visceral Leishmaniasis Gene Target
Abstract
Visceral leishmaniasis (VL) is a potentially fatal disease caused primarily by Leishmania donovani complex species in Old World endemic regions. Pathogenesis requires a virulent factor transcribed by a gene family designated A2. An A2 sequence specific fluorogenic probe hydrolysis (TaqMan) PCR assay was developed on a field-deployable assay platform for real-time screening of the sand fly vector for VL causative agents. Laboratory-based assay optimization and specificity testing were conducted with total nucleic acid extracted from L. donovani-infected Phlebotomus alexandri, the primary vector. Cross-reactivity did not occur when tested against total nucleic acid extract of L. tropica-infected P. sergenti, L. major-infected P. papatasi, clinically relevant organisms, diverse viral and bacterial species, vector and human genomic DNA. Field evaluations were conducted in south central Iraq with sand fly pools screened by a previously established Leishmania universal PCR assay. The Leishmania universal assay was field-formatted (lyophilized) prior to deploying and upon field-evaluation against cold chain maintained wet reagents results were 98% (55/56) concordant. Of 86 Leishmania-positive sand fly pools, six were identified as A2 positive with a field-formatted visceral genotype specific assay.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jun 01, 2004
- Accession Number
- ADA433725
Entities
People
- James A. Swaby
- James C. Mcavin
- Keith W. Blount
- Russell E. Coleman
Organizations
- United States Air Force School of Aerospace Medicine