Potato cyst nematodes PCNs are the most economically damaging pest of potato crop worldwide. During 2013, a survey was carried out in Ain Defla region of Algeria. 81 soil samples collected from potato fields of 14 localities were submitted to nematological analysis which revealed the presence of these nematodes in 22.22% of the prospected fields. Sixteen PCN populations from five localities were characterized by combination of features the perineal regions of cysts and those of second stage juveniles. The morphological identification has been confirmed by the analysis of the ITS-RFLP profiles, sequencing and phylogenetic analysis of the ITS region. The results revealed that the two species Globodera rostochiensis and G. pallida are present in this region occurring separately or in mixed populations. However, we noted a dominance of G. pallida since only 12.25% of the populations have been identified like G. rostochiensis whereas 31.5% were G. pallida and 56, 25% of the populations were constituted of a mixture of the two species and among these mixed populations, 77.77% presented a dominance of G. pallida. The predominance of G. pallida has been noted in the sites of Ain Defla, El Amra, Mekhatria and Arib. G. rostochiensis was dominant in Rouina. Intraspecific variation was noted between populations of G. rostochiensis and G. pallida. Because of the high divergence among Algerian populations of G. pallida and G. rostochiensis it can be assumed that they were multi-introduced in Algeria. The most divergent population of G. pallida, that formed a well separated group with some populations from Chile and Peru, suggests a later or independent introduction of this population in Algeria.