Context. Criticisms of wildlife restocking operations typically focus on concerns that translocations can lead to the introduction of pathogens, and risk the integrity of locally adapted genetic diversity. Restocking programs aiming to stabilise population declines of European brown hares using captive-bred individuals have been carried out in several European countries, including Greece.

Aims. To assess the potential for imported hares to introduce novel strains of European brown hare syndrome virus (EBHSV) during restocking operations, by (1) inferring the origin of wild Greek hares on the basis of their mitochondrial DNA (mtDNA) haplotype, (2) screening the hares to detect and characterise EBHSV, and (3) determining whether certain hare origin–EBHSV combinations occur in the wild.

Methods. RNA extraction, polymerase chain reaction (PCR) amplification, and sequence and phylogenetic analyses of EBHSV were performed on 53 hares. Diagnostic RFLP markers of the mtDNA were used to infer the origins of sampled hares.

Key results. Thirty-three hares had ‘typical’ native Greek haplotypes and 20 had mtDNA haplotypes matching those found in imported and released hares. Twelve of the latter and none of the former were positive for EBHSV. Phylogenetic analysis showed that nine virus isolates formed a single genetic lineage distinct from northern–central European ones. Three virus sequences from three imported reared-and-released hares, from Chalkidiki, were closely related to the northern–central European EBHS viruses.

Conclusions. Alien strains of EBHSV are co-introduced with released captive-bred animals, possibly resulting in negative impacts on populations of Greek hares that have not evolved resistance to these novel virus strains.

Implications. The identification of these allopatric EBHSV strains has led the authorities to ban captures and transportations of local brown hares for any restocking operation. We consider it imperative to reinforce microbiological and genetic controls before further releases of captive-bred game species in the wild in Greece.