In order to investigate the genetic heterogeneity of ovine pestivirus isolates in France, 32-clinical samples collected between 1985 and 2006, previously detected positive by indirect immuno-fluorescence for the presence of Border disease virus (BDV), were tested by reverse transcription-polymerase chain reaction (RT-PCR). Parts of the 5'-non-coding region (5'-NCR) and the Npro-coding region were successfully amplified from 23 samples. An internal control was designed and used to assess the validity of negative RT-PCR results, which could be explained by RNA degradation during the storage of samples at -20 degrees C for at least 16 years. A phylogenetic study was performed by using sequences obtained from the two loci. French isolates clustered into four distinct groups. Six isolates were assigned to the previously described BDV-3 group. However, some isolates could not be assigned to any existing phylogenetic BDV groups, and therefore, tentative new phylogenetic groups are suggested. The AV strain, isolated in 1984 from sheep showing a severe hemorrhagic syndrome in the rearing region of Aveyron in France and sequenced during this study, should be considered as the strain model of the BDV-5 group. Nine viral sequences clustered in a set distinct from all other groups were assigned to the BDV-6 group. Two viral sequences were distinct from the BDV phylum and composed the last set assigned to the group of unclassified pestivirus that had been previously isolated in Tunisia. The marked diversity of pestiviruses might reflect the sheep trade in France and with foreign countries.