Molecular analysis of the genera Hapalotrema Looss, 1899 and Learedius Price, 1934 (Digenea: Spirorchiidae) reveals potential cryptic species, with comments on the validity of the genus Learedius

Phoebe A. Chapman, Thomas H. Cribb, David Blair, Rebecca J. Traub, Myat T. Kyaw-Tanner, Mark Flint, Paul C. Mills

Adult blood flukes of the genera Hapalotrema Looss, 1899 and Learedius Price, 1934 were collected from turtles off Queensland and the Hawaiian Islands. Specimens were identified as Hapalotrema pambanensis Mehrotra, 1973, H. synorchis Luhman, 1935, H. postorchis Rao, 1976 and Learedius learedi Price, 1934 on the basis of morphology. No major morphological differences were found between specimens from this study and previously published descriptions. DNA was also extracted and sequences obtained using custom spirorchiid-specific primers for the ITS2 and 28S rDNA regions, in order to confirm species identification and investigate phylogenetic relationships. Intraspecific genetic variation was generally low. However the ITS2 region of H. postorchis and to a lesser extent that of L. learedi showed considerable variation between specimens from the Pacific and Atlantic oceans. Further studies will be required to determine whether this variation should be considered inter- or intra-specific. Maximum likelihood phylogenetic analyses were completed for both sequenced genes. Learedius learedi was unequivocally nested among species of Hapalotrema, suggesting that the status of the genus Learedius may need to be reassessed.