Twenty-five isolates of Echinococcus granulosus protoscoleces from hydatid cysts were collected from 25 buffaloes in five different provinces of Iran. DNA was extracted from protoscoleces and used for phylogenetical analysis. For this aim, the DNA was amplified using specific primers derived from cox1 gene and, subsequently, the corresponding polymerase chain reaction products were sequenced. The sequence analysis showed five genotypes within the samples. Twenty-three isolates were identified as G1 and under grouped as separate genotypes: G1α, G1β, G1γ and G1δ, and two isolates were identified as G3 strain in Iran. G3 strain has not been already reported in buffaloes in Iran. The nucleotide and amino acid sequences of cox1 gene from corresponding isolates were documented in GenBank under accession numbers: HM130574, HM130575, HM130576, HM130577, HM130578, HM130579, HM130580, HM130581, HM130582, HM130583, HM130584, HM130585, HM130586, HM130587, HM130588, HM130589, HM130590, HM130591, HM130592, HM130593, HM130594, HM130595, HM130596, HM130597 and HM 130598.