The Griffon vulture was once a widespread species across the region of Southeast Europe, but it is now endangered and in some parts is completely extinct. In the Balkan Peninsula the largest Griffon vulture inland population inhabits the territory of Serbia. We present, for the first time, the genetic data of this valuable population that could be a source for future reintroduction programs planned in South-eastern Europe. To characterize the genetic structure of this population we used microsatellite markers from ten loci. Blood samples were collected from 57 chicks directly in the nests during the ongoing monitoring program. We performed a comparative analysis of the obtained data with the existing data from three native populations from French Pyrenees, Croatia, and Israel. We have assessed the genetic differentiation between different native populations and determined the existence of two genetic clusters that differentiate the populations from the Balkan and Iberian Peninsulas. Furthermore, we analysed whether the recent bottleneck events influenced the genetic structure of the populations studied, and we found that all native populations experienced a recent bottleneck event, and that the population of Israel was the least affected. Nevertheless, the parameters of genetic diversity suggest that all analysed populations have retained a similar level of genetic diversity and that the Griffon vulture population from Serbia exhibits the highest value for private alleles. The results of this study suggest that the Griffon vulture populations of the Balkan Peninsula are genetically differentiated from the populations of the Iberian Peninsula, which is an important information for future reintroduction strategies.

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Summary The Eurasian Griffon Vulture Gyps fulvus is a large Palearctic, Indohimalayan and Afrotropical Old-World vulture. The species’ range is vast, encompassing territories from the Pyrenees to the Himalayas. We reviewed and analysed a long-term data set for Griffon Vulture in the Balkans to estimate the change in its population size and range between 1980 and 2019. After a large historical decline, the Griffon Vulture population slightly increased in the last 39 years (λ = 1.02) and reached 445–565 pairs in 2019. We recorded a gradual increase of Griffon Vulture subpopulations in Serbia (λ = 1.08 ± 0.003), Bulgaria (λ = 1.08 ± 0.003) and Croatia (λ = 1.05 ± 0.005) and steep to a moderate decline of the species subpopulations in Greece (λ = 0.88 ± 0.005) and North Macedonia (λ = 0.94 ± 0.01). However, species range contracted to half of its former range in the same period. It occurred in 42 UTM squares in the 1980–1990 period and only 20 UTM squares between 2011 and 2019 and concentrated into three source subpopulations in Bulgaria, Serbia, and Croatia. Following reintroductions of the Griffon Vulture in Bulgaria, new colonies were formed at three novel localities after 2010. Regular movements of individuals between the different subpopulations exist nowadays. Therefore, preservation of both current and former core areas used for breeding and roosting is essential for species conservation in the region. However, the Griffon Vulture still faces severe threats and risk of local extinction. Various hazards such as poisoning, collision with energy infrastructure, disturbance and habitat alteration are depleting the status of the Balkan population and its full recovery. Further studies should analyse age-specific survival and mortality, recruitment, genetic relatedness, spatial use to inform the viability of this population in the future.

The knowledge in the behaviour and movement of endangered species is of key importance for the precise targeting and assessing the efficiency of nature conservation actions, especially considering vultures, which explore vast areas to locate ephemeral and unpredictable food resources. Therefore, a total of 51 Griffon Vultures (Gyps fulvus) from both the re-introduced population and the autochthonous Balkan Peninsula (Balkans) colonies have been tagged with GPS/GSM transmitters in recent years, in order to study their seasonal and spatial distribution. The current study presents the analysis of the high-resolution GPS location data, acquired between January 2016 and March 2021. A total of 1,138,383 locations (an average number of 23,716 ± 18,886 positions per bird, ranged between 2,515 and 76,431 of total fixes per bird; n=48) were used to estimate the home range size and identify the traditional foraging areas and roosting sites of the birds during the wintering, migration/roaming and summering periods. Our results reveal that Griffon Vultures movement activity and home range size varied considerably throughout the annual cycle, especially between their wintering and summering grounds, while exhibiting significant overlapping amongst the tracked individuals. Specifically, immature Griffon Vultures travel long distances across all Balkan Peninsula countries, but always gather with conspecifics, showing strong fidelity to active breeding/roosting sites. The total home range 95% area of the Griffon Vulture population on the Balkans was estimated at 39,986.4 km² and the 50% core area at 1,545.42 km² (n = 48). All tracked birds were found to either visit or frequently use (> 95% of the time) the same seven vulture key zones on the Balkan Peninsula – one in Serbia, one shared between North Macedonia and Bulgaria, one shared between Bulgaria and Greece, two entirely lying in Bulgaria, one in western Greece and one shared between Kvarner Archipelago islands in Croatia and the Julian Alps - Italy, Austria and Slovenia. Several smaller sub-zones were also defined within these general ones. The seven key zones form a coherent network and are used as stepping stones for Griffon Vultures during their migration movements and roaming, but also wintering and summering. The observed concentration tendency of Griffon Vultures on the Balkans and the predictability of their temporal and spatial presence should be used to precisely target, address and substantially increase the efficiency of the conservation measures in this marginal and, thus, still vulnerable meta-population.

Once a widespread and common species across the region of southeast Europe, the Griffon vulture is now confined to small and isolated populations across the Balkan Peninsula. The population from Serbia with 290 couples represents its biggest and most viable population that can serve as an important reservoir of genetic diversity from which the birds can be used for the region’s reintroduction or recolonization programs. To estimate the level of genetic diversity, the mitochondrial Cytb gene from 58 unrelated birds sampled during the marking in the nests was sequenced and compared to the homologous Griffon vulture sequences available in publicly accessible online databases. Phylogeographic analysis based on Cytb sequences showed that the most frequent haplotype is found in all Griffon vulture populations and that each population possesses private haplotypes. Our data suggest that the Griffon vulture population from Serbia should be used as a source population for restocking and reintroduction programs in the region. The observed genetic differentiation between the populations from the Iberian and Balkan Peninsulas suggest that the introduction of foreign birds from remote populations should be avoided and that birds from indigenous or neighboring populations, if available, should be used instead.