Genetic characterization of paramphistomes of bufalo by HAT-RAPD analysis

Abstract

Paramphistomosis is widely prevalent in domestic farm animals, incurring heavy economic losses in several countriesacross the world. As the identifcation of these parasites based on morphological features is very difcult, molecular characterizationis necessary to discriminate among diferent species. We performed polymorphic DNA fngerprint analysis of 3 diferent species ofamphistomes collected from the rumen and bile ducts of bufaloes slaughtered at a local abattoir in Anand, Gujarat, India. The ruminalamphistomes were identifed as Paramphistomum cervi and Gastrothylax indicus, while the hepatic amphistomes were identifed asGigantocotyle bathycotyle . High annealing temperature-random amplifed polymorphic DNA (HAT-RAPD) analysis with 4 primers,OPA2, OPA4, OPA8, and OPA9, identifed species-specifc banding patterns for each species. The unique bands specifc for specieswere sequenced, resulting in identifcation of 15 sequences, which included 6 sequences for G. bathycotyle, 3 sequences for G. indicus,and 6 sequences for P. cervi. The dendrogram analysis of RAPD bands revealed genetic distance of 0.5798 between G. bathycotyleand G. indicus, compared to a distance of 1.0986 and 0.9163 between G. bathycotyle and P. cervi and between G. indicus and P. cervi,respectively . The primers derived from sequenced RAPD amplicons generated one species-specifc sequence characterized amplifedregion marker for the identifcation of G. indicus.