Morphological and molecular identification of amphistomes occurring in cattle in Zimbabwe

This study characterized and identified amphistomes collected from cattle slaughtered at selected abattoirs in Zimbabwe, using both morphological and molecular techniques. Amphistomes were collected from 158 cattle originating from 14 localities, slaughtered in abattoirs in Harare and Bulawayo. One-hundred and twenty-one amphistome specimens were selected and sectioned into two portions, one piece for morphological characterization based on histological sectioning using haematoxylin and eosin staining, and the other piece for molecular identification based on the ITS-2 gene. Morphological characters classified the amphistomes into ten species as follows; Calicophoron (C.) calicophorum, C. microbothrium, C. clavula, C. raja, C. phillerouxi, C. sukari, Paramphistomum (P.) gracile, P. hibarniae, Carmyerius (Ca.) multivitellarius, Cotylophoron (Co.) cotylophorum, and unidentified Paramphistomum sp. Calicophoron was the most common genus, and C. microbothrium was the most common and widely distributed species recorded across the 14 localities. The ITS-2 gene was able to confirm the identification of C. microbothrium, C. clavula, C. raja, C. calicophorum and C. phillerouxi. There were no comparative ITS-2 sequences for C. sukari, P. gracile and P. hibarniae on the GenBank. Hence, specimens identified morphologically as C. sukari showed a 100% homology to multiple Calicophoron species, but grouped with C. raja. Furthermore, Ca. multivitellarius specimens confirmed the genus identity by forming sister clade with other Carmyerius spp. There were no species-level resolution for Paramphistomum sp., P. gracile and P. hibarniae, as these isolates formed a clade with Paramphistomum sp. (OQ459353.1). The study resulted in new geographical records for P. gracile, P. hibarniae and Ca. multivitellarius in Zimbabwe, hence adding new knowledge to the geographical distribution of these species within Africa.