Biochemical Characterization and Differential Expression of a 16.5-Kilodalton Tegument-Associated Antigen from the Liver Fluke Fasciola hepatica

Abstract

ABSTRACTA cDNA encoding a 16.5-kDa protein termed FhTP16.5 was identified by immunoscreening of a cDNA library fromFasciola hepaticaadult flukes using pooled sera from rabbits infected withF. hepaticafor 4 weeks. Quantitative reverse transcriptase PCR (qPCR) analysis revealed that FhTP16.5 is not expressed in unembryonated eggs. It is poorly expressed in miracidia and highly expressed at the juvenile and adult stages; however, significant differences were found between the expression levels of FhTP16.5 in juveniles versus adult flukes. Recombinant FhTP16.5 was expressed at high levels inEscherichia coli, purified by affinity chromatography, and used to raise anti-FhTP16.5 polyclonal antibodies in rabbits. Immunoblot analysis using the anti-FhTP16.5 IgG antibody identified FhTP16.5 in crude and tegumental extracts and in excretory-secretory products ofF. hepatica. The protein was not detected in crude extracts ofSchistosoma mansoniorSchistosomajaponicum. Antibodies to FhTP16.5 were detected in the sera of rabbits at 3 to 12 weeks ofF. hepaticainfection as well as in the sera of humans with chronic fascioliasis; these findings suggest that FhTP16.5 could be a good antigen for serodiagnosis of fascioliasis. Immunohistochemistry demonstrated that FhTP16.5 localizes to the surface of the tegument of various developmental stages and in parenchymal tissues of the adult fluke. Such specific localization makes FhTP16.5 an attractive target for immunoprophylaxis or chemotherapy.