Authors: Stronen, Astrid Vik; Mattucci, Federica ;Fabbri, Elena;Galaverni, Marco;Cocchiararo, Berardino ;Skrbinšek, Tomaž ;Vlasseva, Albena ;Caniglia, Romolo ;

Human-induced hybridization is a global threat to wildlife, and must be distinguished from natural intra- and interspecific gene flow. Genetic markers to differentiate among these processes are therefore essential for conservation. Genetically distinct populations of grey wolves (Canis lupus) are now reconnecting within Europe. However, the species is affected by hybridization with domestic dogs (C. l. familiaris), which are also known to hybridize with golden jackals (C. aureus) in south-eastern Europe. We used existing genomic resources to identify discriminant single nucleotide polymorphism (SNP) markers, with focus on the Iberian, Italian and Dinaric wolf populations, to (1) separate wolves, dogs, and golden jackals, and (2) identify their first two hybrid generations. We selected 192 SNPs that identified the five canid groups, all simulated first-generation (F1) hybrids (0.482 ≤ Qi ≤ 0.512 between their respective parental groups) and all first backcross (BC1) individuals (0.723≤ Qi ≤ 0.827 to parental groups). An assay design and test with invasive and non-invasive samples performed successfully for 178 SNPs, which correctly categorized the five groups and simulated hybrids. The identified SNPs permit differentiation between inter-specific hybridization and natural population admixture and promote data sharing among laboratories, facilitating evolutionary research, monitoring, and conservation of European canid populations.