Sequencing of mitochondrial NADH dehydrogenase subunit 1 (nad1) gene among Fasciola spp.and phylogenetic reconstruction

In order to examine sequence variation in the mitochondrial(mt) NADH dehydrogenase subunit 1(nad1) gene among Fasciola spp.from China,Niger,France and USA and to reconstruct their phylogenetic relationship using nad1 sequences,the partial nad1(pnad1) was amplified from each Fasciola samples,sequenced and compared with relevant sequences available in GenBankTM.Phylogenetic relationship was reconstructed using the neighbour-joining,maximum parasimony and maximum likelihood methods.The lengths of all pnad1 sequences was 446 bp.Sequence comparison revealed that the variation in pnad1 sequences among F.hepatica isolates ranged from 0.2% to 2.0%,while the variation among F.gigantica isolates ranged from 1.8% to 3.4%.The pnad1 sequences of the intermediate Fasciola were similar to that of F.gigantica.Phylogenetic analysis revealed that the intermediate Fasciola was genetically closer to F.gigantica than to F.hepatica.Sequence differences in pnad1 sequences were significantly greater than nucleotide variation within species,indicating that nad1 sequence can be used as genetic marker for the speci-fic differentiation among Fasciola spp.and studying genetic variation within each Fasciola spp..This study also provided novel mtDNA evidence for the existence of the intermediate Fasciola in China.