Fascioliasis is a serious disease of sheep and cattle worldwide. Fasciola species parasitize a variety of mammals, caused by Fasciola hepatica, and Fasciola gigantica. Traditional morphological techniques to differentiate the two
species can be inaccurate, especially when hybrid forms are present. The definitive identification of Fasciola species,
including their hybrids, has been made possible by advanced molecular techniques. Our study aimed to estimate the
prevalence of Fascioliasis and identify the phenotypic features of Fasciola that infecting sheep, cattle, buffaloes, goats,
and camels in Qualyobia, Egypt. The genetic identity of Fasciola species was examined by the analysis of forward and
reverse sequences of the ITS2 of the rDNA gene that amplified in 300bp. Out of 286 slaughtered animals {88 sheep,
26 goats, 68 cattle, 25 buffaloes and 79 camels} in the regions of Qualyobia, 51 (17.8%) had Fasciola spp. morphologically {27/51(52.9%) were identical to F. hepatica while 18/51(35.3%) were identical to F. gigantica, 3/51(3.9%) were
larval stages and 3/51(3.9%) mixed infections}. DNA from 10 flukes extracted, amplified, and analyzed to identify
species using the ITS2 locus. Ten flukes were identified as F. gigantica and F. hepatica. PCR products from 2 flukes
were sequenced for phylogenetic analysis. The sequence of ITS2 gene isolates obtained from the present investigation
were compared with GenBank reference sequences of F. hepatica, F. gigantica. The phylogeny based on ITS2 revealed
two distinct clades separating F. hepatica from F. gigantica with snaps and indel in one isolate. So concluded that safety
measures should be done because the disease has zoonotic effect on public health. The prevalence of Fascioliasis was
high in Qualyobia, Egypt, a lot of liver was condemned, which resulted in financial losses for affected farmers. In order
to control the disease and raise farmer awareness, it is vital to implement the appropriate preventive measures.