Morphological and Molecular Identification of Paramphistomum epiclitum from Buffaloes in Pakistan

Little is known about the genetic and morphological characters of Paramphistomum epiclitum. For the first time in Pakistan, adult flukes were morphologically characterized and the sequence variation in the nuclear ribosomal DNA (rDNA) region, including the first internal transcribed spacers (ITS1) and the 5.8S gene of the Paramphistomum epiclitum were studied. Adult amphistomes were examined by light microscopy and sequences of ITS1 and 5.88S rDNA genes were obtained. Twenty adult flukes were measured, 13.17 ± 1.19 mm in length and 5.28 ± 1.34 mm in width. Seventeen adult flukes were sequenced and high sequence variability was observed in 5′ end of ITS1 region. The 5.8S and 3′ end of ITS1 sequences had 100% identity among the samples. A comparative analysis revealed that different types and numbers of repeats were found within each ITS1 region. The 3′ end of ITS1 region from P. epiclitum showed 98% homology with P. cervi from China and formed a subclade with genetic distance of 0.1663. The 5.8S gene showed 100% identity within Paramphistomidae family and formed a sub-clade with P. epiclitum, P. leydeni, P. cervi and Cotylophoron cotylophorum species isolated from China, India and Uruguay. This work provides new information on morphological identity and genetics of P. epiclitum from Pakistan.