DETECTION OF FASCIOLA-HEPATICA IN INFECTED INTERMEDIATE HOSTS USING RT-PCR

Fasciola hepatica, the common bile duct fluke, is an economically impo rtant parasite of domestic livestock. Current research interest is dir ected toward an understanding of the parasite's biology at the interme diate host level. To permit study of seasonal transmission patterns an d parasite/intermediate host interactions, a fasciolid-specific assay has been developed to detect infected snail vectors. This assay uses t he reverse transcriptase-polymerase chain reaction (RT-PCR) to amplify specifically a region of F. hepatica small-subunit rRNA, followed by hybridization to an F. hepatica-specific probe. The assay does not cro ss-react with 2 trematodes outside of the Fasciolidae but does detect Fascioloides magna rRNA. Sequence alignment with additional small-subu nit rRNAs shows Fasciolopsis buski would also cross-react with the ass ay. The detection limit of the assay is 10 fg of fluke total RNA with 5 mu g of snail RNA added as background. Additionally, the assay detec ts individual infected snails immediately after miracidial exposure an d throughout the parasite's development period.