Prunus necrotic ringspot virus (PNRSV) was detected by PCR in Tunisian stone fruit germplasm collections. Peach had the highest infection rate reaching 50%, followed by apricot (46.8%), almond (35.5%) and plum (9.5%). The genetic variability of the CP gene of 10 PNRSV isolates was evaluated by SSCP analysis of PCR products and analysis of their sequence. In phylogenetic trees, Tunisian PNRSV isolates clustered in the three major groups, PV96, PV32 and PE5. However, an almond isolate (AlTun8) segregated as an out-group. This isolate is phylogenetically distant from other PNRSV isolates from different parts of the world and displays nucleic and amino acid identities of 83-86% and 65-74%, respectively, with PV96, PE5 and PV32. This is the first report on the molecular variability of PNRSV in different Prunus hosts in Tunisia.