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In mainland Europe, mouflon were first introduced in the 18th and 19th centuries, mainly in Germany, Austria, the Czech Republic, Slovakia, and Hungary. In the early 20th century, mouflon were introduced in Croatia, while all introductions in Slovenia occurred in the 1950s and 1960s. Since the introduction, populations in both countries have remained largely stable, with occasional declines and increases in some areas. Due to several up-following introduction events, also using individuals with unknown origin, the genetic pool of the species might be very admixed; however, no genetic study has been made to date. Therefore, our aim was to: i) determine the origin of introduced mouflon in Slovenia and Croatia, ii) compare the neutral and adaptive genetic make-up of introduced populations. In genetic analyses, which were performed at neutral loci (partial fragment of mitochondrial control region, mtDNA CR) and adaptive major histocompatibility complex (MHC; DRB exon 2) using the next generation sequencing approach, we also included individuals from Germany, France (Corsica), Italy (Sardinia), and the Czech Republic. The haplotype network based on mtDNA CR showed that most analysed mouflon from Slovenia shared the same haplotypes or were closely related to mouflon from Germany and Sardinia, and a few shared the same haplotypes as most mouflon from Croatia. Croatian mouflon from all studied populations shared the same or close haplotypes with individuals from the first Croatian population established in Brijuni Island National Park (northern Adriatic Sea) in the early 20th century. Similar results for populations in both countries were revealed by MHC genes. According to the genetic data, the population origins are generally in agreement with the written historical records, although there are indications of the introduction or release of extra individuals into certain colonies where historical data is lacking. Furthermore, our findings validate that both mitochondrial and MHC genetic diversity are useful tools for investigating the paths of translocation.

The Balkan chamois (Rupicapra rupicapra balcanica) is widespread on the Balkan Peninsula, along mountain massifs from Croatia in the north to Greece in the south and Bulgaria in the east. Knowledge on the genetic structure of Balkan chamois populations is limited and restricted to local studies. Therefore, the main objective of this study was to use nuclear (16 microsatellites) and mitochondrial (partial 376 base pairs control region) markers to investigate the genetic structure of this chamois subspecies throughout its distribution range and to obtain information on the degree of connectivity of the different (sub)populations. We extracted DNA from bone, dried skin and muscle tissue and successfully genotyped 92 individuals of Balkan chamois and sequenced the partial control region in 44 individuals. The Bayesian analysis suggested 3 genetic clusters and assigned individuals from Serbia and Bulgaria to two separate clusters, while individuals from the other countries belonged to the same cluster. Thirty new haplotypes were obtained from partial mitochondrial DNA sequences, with private haplotypes in all analyzed populations and only two haplotypes shared among populations, indicating the possibility of past translocations. The subspecies genetic composition presented here provides the necessary starting point to assess the conservation status of the Balkan chamois and allows the development of conservation strategies necessary for its sustainable management and conservation.

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European wildlife has been subjected to intensifying levels of anthropogenic impact throughout the Holocene, yet the main genetic partitioning of many species is thought to still reflect the late-Pleistocene glacial refugia. We analyzed 26,342 nuclear SNPs of 464 wild boar (Sus scrofa) across the European continent to infer demographic history and reassess the genetic consequences of natural and anthropogenic forces. We found that population fragmentation, inbreeding and recent hybridization with domestic pigs have caused the spatial genetic structure to be heterogeneous at the local scale. Underlying local anthropogenic signatures, we found a deep genetic structure in the form of an arch-shaped cline extending from the Dinaric Alps, via Southeastern Europe and the Baltic states, to Western Europe and, finally, to the genetically diverged Iberian peninsula. These findings indicate that, despite considerable anthropogenic influence, the deeper, natural continental structure is still intact. Regarding the glacial refugia, our findings show a weaker signal than generally assumed, but are nevertheless suggestive of two main recolonization routes, with important roles for Southern France and the Balkans. Our results highlight the importance of applying genomic resources and framing genetic results within a species’ demographic history and geographic distribution for a better understanding of the complex mixture of underlying processes. Nasl. z nasl. zaslona. Opis vira z dne 3. 2. 02023. Skupno št. avtorjev: 21. Bibliografija: str. 142-143.

Recent technological advances in the field of genomics offer conservation managers and practitioners new tools to explore for conservation applications. Many of these tools are well developed and used by other life science fields, while others are still in development. Considering these technological possibilities, choosing the right tool(s) from the toolbox is crucial and can pose a challenging task. With this in mind, we strive to inspire, inform and illuminate managers and practitioners on how conservation efforts can benefit from the current genomic and biotechnological revolution. With inspirational case studies we show how new technologies can help resolve some of the main conservation challenges, while also informing how implementable the different technologies are. We here focus specifically on small population management, highlight the potential for genetic rescue, and discuss the opportunities in the field of gene editing to help with adaptation to changing environments. In addition, we delineate potential applications of gene drives for controlling invasive species. We illuminate that the genomic toolbox offers added benefit to conservation efforts, but also comes with limitations for the use of these novel emerging techniques.

The translocation of wild animal species became a common practice worldwide to re-establish local populations threatened with extinction. Archaeological data confirm that chamois once lived in the Biokovo Mountain but, prior to their reintroduction in the 1960s, there was no written evidence of their recent existence in the area. The population was reintroduced in the period 1964–1969 when 48 individuals of Balkan chamois from the neighbouring mountains in Bosnia and Herzegovina were released. The main objective of this study was to determine the accuracy of the existing historical data on the origin of the Balkan chamois population from the Biokovo Mountain and to assess the genetic diversity and population structure of the source and translocated populations 56 years after the first reintroduction. We used 16 microsatellite loci to analyse the genetic structure of three source chamois populations from Prenj, Čvrsnica and Čabulja Mountains and from Mt. Biokovo. Both STRUCTURE and GENELAND analyses showed a clear separation of the reintroduced population on Biokovo from Prenj’s chamois and considerable genetic similarity between the Biokovo population and the Čvrsnica–Čabulja population. This suggests that the current genetic composition of the Biokovo populations does not derive exclusively from Prenj, as suggested by the available literature and personal interviews, but also from Čvrsnica and Čabulja. GENELAND analysis recognized the Balkan chamois from Prenj as a separate cluster, distinct from the populations of Čvrsnica and Čabulja. This suggests that the Neretva River and the state M17 road are geographic barriers for the species dispersal, as they form a genetic boundary.

With an accelerating negative impact of anthropogenic actions on natural ecosystems, non-invasive biodiversity assessments are becoming increasingly crucial. As a consequence, the interest in the application of environmental DNA (eDNA) survey techniques has increased. The use of eDNA extracted from faeces from generalist predators, have recently been described as "biodiversity capsules" and suggested as a complementary tool for improving current biodiversity assessments. In this study, using faecal samples from two generalist omnivore species, the Eurasian badger and the red fox, we evaluated the applicability of eDNA metabarcoding in determining dietary composition, compared to macroscopic diet identification techniques. Subsequently, we used the dietary information obtained to assess its contribution to biodiversity assessments. Compared to classic macroscopic techniques, we found that eDNA metabarcoding detected more taxa, at higher taxonomic resolution, and proved to be an important technique to verify the species identification of the predator from field collected faeces. Furthermore, we showed how dietary analyses complemented field observations in describing biodiversity by identifying consumed flora and fauna that went unnoticed during field observations. While diet analysis approaches could not substitute field observations entirely, we suggest that their integration with other methods might overcome intrinsic limitations of single techniques in future biodiversity surveys.

The chamois, genus Rupicapra, is the most abundant mountain-dwelling ungulate in Europe and the Near East and is currently recognized to be divided into two species: R. rupicapra (Northern chamois) and R. pyrenaica (Southern chamois) further divided into seven and three subspecies, respectively. While neither of the species is of conservation concern, some subspecies are protected at the national level in (part of) their distribution ranges. However, for most subspecies and populations the information necessary to assess the conservation status are lacking and several populations were subject to translocations and reintroductions, further complicating their general taxonomic picture. Here we combine nine newly sequenced mitochondrial genomes (mitogenomes) of four subspecies with previously published mitogenomes of Rupicapra (5) and related (sub)species in the Caprini, Ovibovini and Pantilopini families (39). The Bayesian and maximum likelihood phylogenetic analyses concordantly confirmed the monophyly of the genus Rupicapra and its positioning within the Caprini tribe. Both analyses identified the subdivision of the subspecies into three lineages in accordance with their geographical distributions (Eastern, Central and Western). The Central and Western lineages were more closely related, confirming previously reported results based on partial mitochondrial DNA regions. Within the Eastern lineage, R. r. balcanica was the most differentiated and included all individuals coming from the northern part of the Dinaric Karst mountain range, Mt. Velebit (Croatia), a known human-caused contact zone between R. r. rupicapra and R. r. balcanica, whereas R. r. tatrica and R. r. rupicapra showed a sister group relationship. Overall, our results, while providing an increased resolution, confirmed the phylogeny previously reconstructed using both whole and partial mitochondrial DNA sequences. The discrepancy among nuclear, mitochondrial and Y-chromosome markers in phylogenetic reconstruction of the genus thus remains. Further studies combining multiple markers and possibly including archeological remains are therefore warranted to solve the controversy.

The Eurasian otter (Lutra lutra) is a semi-aquatic predator that lives in a wide range of aquatic habitats. Knowledge of the diet of predators is essential for the management and conservation of the species and their habitats. We assessed the diet of otters in fifty-three faecal samples (spraints) by morphological analysis of undigested prey remains and by DNA metabarcoding approaches with two different blocking primers. The blocking primers were shown to be similarly effective in repressing host DNA, improving amplification of non-host taxa. The analyses were conducted to compare the accuracy of the two methods. The prey items were categorized to the highest possible taxonomic resolution, and relative frequency of occurrence was applied in order to compare the morphological and DNA results. The bulk score for difference prey items in the spraints was based on volume estimates in macroscopic observations. The two methods had an overlap of identified prey species ranging from 69.9 to 72.9% (depending on the kind of primers) meaning that a large portion of prey items found by one method was replaced by other prey items in the other method and that most prey items found through DNA-metabarcoding are considered new when compared with the morphological analysis. Therefore, applying both morphological and DNA analysis is crucial in order to determine the diet of otters and, possibly, other predatory species.

Simple Summary The major histocompatibility complex is a family of genes of central importance in vertebrate adaptive immunity. Genes of the major histocompatibility complex are among the most polymorphic genes ever described in vertebrates, with their number differing greatly among species. Due to their functional significance and exceptional diversity, they represent excellent markers in evolutionary ecology and conservation. Traditional methods were commonly used for their genotyping, but the introduction of next-generation sequencing facilitated more accurate and reproducible genotyping of such polymorphic gene families. Nevertheless, due to the high polymorphism of the major histocompatibility complex genes, genotyping is inherently difficult. Here, we compared the performance of traditional and next-generation sequencing on genotyping of the major histocompatibility complex genes in chamois. Although the major histocompatibility complex system in chamois is quite simple, we found genotyping discrepancies between the two methods in 25% of individuals. Our results show that next-generation sequencing has a higher detection capacity and thus allows for more accurate genotyping of highly polymorphic genes. Abstract Genes of the major histocompatibility complex (MHC) code for cell surface proteins essential for adaptive immunity. They show the most outstanding genetic diversity in vertebrates, which has been connected with various fitness traits and thus with the long-term persistence of populations. In this study, polymorphism of the MHC class II DRB locus was investigated in chamois with Single-Strand Conformation Polymorphism (SSCP)/Sanger genotyping and Ion Torrent S5 next-generation sequencing (NGS). From eight identified DRB variants in 28 individuals, five had already been described, and three were new, undescribed alleles. With conventional SSCP/Sanger sequencing, we were able to detect seven alleles, all of which were also detected with NGS. We found inconsistencies in the individual genotypes between the two methods, which were mainly caused by allelic dropout in the SSCP/Sanger method. Six out of 28 individuals were falsely classified as homozygous with SSCP/Sanger analysis. Overall, 25% of the individuals were identified as genotyping discrepancies between the two methods. Our results show that NGS technologies are better performing in sequencing highly variable regions such as the MHC, and they also have a higher detection capacity, thus allowing a more accurate description of the genetic composition, which is crucial for evolutionary and population genetic studies.

There is an increased interest in the possibility to use large animals in the restoration of degraded ecosystems and to increase the capacity of natural areas to sustain richer biodiversity. We quantify the dietary differences and similarities between five moose and 22 red deer introduced in a 2100 ha fenced area under restoration (Lille Vildmose, Denmark) and their potential in the restoration process. Moose and red deer were selected based on their biogeographic affinity and existing knowledge of their diet and feeding behaviour, and are expected to counteract encroachment on open habitats. Using eDNA metabarcoding of dung collected in the field and species identification through saliva recovered from browsed twigs, we investigated the species’ diets and diet overlap after the moose were released into the area, and assessed how their diets match the expected ecosystem function. Despite the partial overlap of some dietary items, such as Betulaceae and Salicaceae, we report a differential use in height of tree species and distinctive use of other food resources between the two species, with red deer eating more herbaceous plants and the moose diet also containing aquatic plants. This study exemplifies a monitoring approach to restoration projects where large herbivores are expected to provide a key ecosystem function through their browsing and grazing.

In 2009, a frozen mummy of the steppe bison (SB) (Bison priscus) was discovered between the lower Kolyma River and the Alazeya River in northeast Siberia, Russia. The specimen was dated with 14C and estimated to have lived more than 48,000 14C years before present (BP). The relationship between SB and the European Bison (EB) (Bison bonasus), also known as wisent or European wood bison, is unresolved and it is unclear whether the SB and EB overlapped in space and time. The aim of our study was to compare genetic variability between the SB specimen and modern EB. We expected higher SB variability due to substantial bottlenecks in the EB approximately one century ago when it became extinct in the wild. The EB (n = 167) and the SB specimen were genotyped with the Illumina BovineHD BeadChip with 777,962 single-nucleotide polymorphism (SNP) markers. Steppe bison DNA was extracted and genotyped six times to account for genotyping errors due to low-quality DNA. We obtained a final set of 7786 SNPs. The mean number of private alleles in EB was 0.027 (± 0.0002) and in SB, it was 0.288 (± 0.0006). This could be explained by factors including differences between species, spatiotemporal divergence, and bottleneck effects. Investigation of historic EB samples could help resolve phylogenetic relationships, the role of the recent bottleneck, and provide information for conservation management to reduce the incidence of disease in the population and maintain its evolutionary potential.

Hybridization between domestic and wild species is known to widely occur and it is reported to be one of the major causes of the current biodiversity crisis. Despite this, poor attention has been deserved to the behavioural ecology of hybrids, in particular in relation to their social behaviour. We carried out a camera trap study to assess whether phenotypically anomalous colouration in wild boar, i.e. potentially introgressed with domestic pigs, affected the hierarchical structure of wild boar social groups. Chromatically anomalous wild boars (CAWs) were detected in 32 out of 531 wild boar videos. In most videos (75%) CAWs were the latest of the group, independently from their age class and group composition. Aggressions by their wild type fellows were recorded in 31.25% videos; by contrast, no aggression among wild type individuals was observed. The lack of camouflage may expose CAWs, and thus their group, to a higher predation risk, compared to wild type groups. This individual loss of local adaptation may increase predation risk by the wolf or detection by hunters, being maladaptive for the whole social group.

The Carpathian Mountains provide critical wildlife habitat in central Europe, and previous genome-wide studies have found western Carpathian Mountain wolves (Canis lupus) to be a separate population. Whereas differentiation to the north may be explained by a lowland-mountain transition and habitat fragmentation, the eastern Carpathian Mountains extending through Romania appear to offer continuous wildlife habitat southward. Our objective was to assess gene flow patterns and population connectivity among wolves in Romania, western Ukraine, and the Republic of Moldova. We sought to determine if the Carpathian Mountain region is best described by a north–south gradient in genetic profiles, or whether Romanian wolves show population structure with northern individuals clustering with western Ukraine. We genotyped 48 individuals with 170 000 single nucleotide polymorphism markers, and successful profiles from Romania (n = 27) and Moldova (n = 2) were merged with existing data from western Ukraine (n = 10). Expected heterozygosity was 0.234 (SE 0.001) for Romania and 0.229 (SE 0.001) for western Ukraine, whereas observed heterozygosity values were 0.230 (SE 0.001) versus 0.231 (SE 0.001). Population structure analyses with a maximum likelihood method supported K = 1 population, followed by K = 2 where Romania formed one cluster, and western Ukraine and Moldova formed another. Principal component analysis results were broadly consistent with K = 2. Pairwise FST between western Ukraine and Romania was 0.042 (p = 0.001). Our findings indicated weak population differentiation, and future research may clarify whether the spatial distribution of genetic diversity in the region is associated with environmental and ecological factors such as terrain ruggedness and the distribution of prey species.

During the early 1900s, Northern chamois (Rupicapra rupicapra) populations in the northern Dinaric Mountains were extirpated. During the 1960s and 1970s there were several reintroductions of individuals from two Northern chamois subspecies (Alpine chamois, R. r. rupicapra and Balkan chamois, R. r. balcanica) from neighbouring areas in the attempt to re-establish the population. Accurate taxonomic classification, at subspecies level, of the autochthonous extirpated population was not known. To clarify which subspecies was present before reintroduction, we genotyped four male chamois skulls originating from Velebit Mountain, collected around 25 years before the population local extinction. DNA was successfully extracted from middle layer and outer sheath of horns. Assignment based on microsatellite loci, using both Bayesian clustering in STRUCTURE (with q values between 0.55 and 0.73) and DAPC (with individual membership probabilities of 0.99 and 1.00) indicated higher assessed likelihood for the Alpine subspecies.

Necrotic inflammation of the prepuce (NIP, posthitis) is probably the most serious threat to the males of European bison (Bison bonasus) population living in the Białowieża National Park. Approximately 6% of the males have been affected every year. The aim of this study was to explore genetic regions potentially affecting this disease. The population was divided into two groups: healthy (25) and affected by posthitis (112). All individuals were genotyped with the Illumina BovineHD BeadChip. Tests performed on a single-locus mixed model (EMMAX) resulted in one SNP on the 25th chromosome (rs134172402) showing statistically important association with the posthitis incidence. The closest candidate gene located near this SNP was the periplakin (PPL) coding protein associated with skin development and some autoimmunity skin diseases such as pemphigus foliaceus or atopic dermatitis. Therefore, we conclude that PPL might be the candidate gene for posthitis, but further studies are necessary to confirm its expression and role in affected tissues.

The original version of this article unfortunately contained a mistake. The Electronic Supplementary Material was missing in the online version. The ESM is available in the online version of this correction article.

Hybridisation and gene introgression are important sources of diversification, the relevance of which in the evolutionary processes is well recognised. Their fitness consequences in animal populations, however, are not sufficiently well understood, despite hybridisation rates becoming increasingly important worldwide following human-related activities such as domestication, game management and habitat alteration. In Europe, the density and distribution of native ungulates have largely been influenced by humans since pre-historic times. This, alongside the introduction of non-native and domesticated species, may bear major consequences at the genetic and population levels. We provide an updated overview of recent hybridisation events in wild European ungulates; we describe their ecological drivers, extent, current distribution, potential consequences and proposed management strategies. We reviewed the scientific literature published between 2000 and 2018 and found that confirmed hybridisation was described in 75 of the 89 references we included, involving nearly all the species that we investigated. Most researchers relied on genetic information for hybrid identification, which often involved a domestic counterpart. However, introductions and translocations also led to crossbreeding between wild ungulate (sub)species. Only 43 papers provided management recommendations, mostly focused on preventing hybridisation and removing hybrids. Hybridisation proved to be relatively common in several ungulate taxa in Europe. Despite reported changes in phenotype and fitness-related traits in some species, the consequences of hybridisation for adaptation, life history, and evolutionary potential remain largely unknown. The current conservation paradigm aims to prevent the spread of domestic or non-native genes in native populations; accordingly, conservation plans should: 1) determine the genetic origin of possible source populations; 2) protect native populations from the risk of crossbreeding with non-native ones, and 3) establish permanent monitoring.

After a strong demographic decline before World War II, wild boar populations are expanding and the species is now the second-most abundant ungulate in Europe. This increase raises concerns due to wild boar impact on crops and natural ecosystems and as potential vector of diseases. Additionally, wild boar can hybridize with domestic pigs, which could increase health risks and alter wild boar adaptive potential. We analysed 47,148 Single Nucleotide Polymorphisms in wild boar from Europe (292) and the Near East (16), and commercial (44) and local (255) pig breeds, to discern patterns of hybridization across Europe. We identified 33 wild boars with more than 10% domestic ancestry in their genome, mostly concentrated in Austria, Bosnia and Herzegovina, Bulgaria and Serbia. This difference is probably due to contrasting practices, with free-ranging vs. industrial farming but more samples would be needed to investigate larger geographic patterns. Our results suggest hybridization has occurred over a long period and is still ongoing, as we observed recent hybrids. Although wild and domestic populations have maintained their genetic distinctiveness, potential health threats raise concerns and require implementation of management actions and farming practices aimed at reducing contact between wild and domestic pigs.

[Background]: Pigs were domesticated independently in Eastern and Western Eurasia early during the agricultural revolution, and have since been transported and traded across the globe. Here, we present a worldwide survey on 60K genome-wide single nucleotide polymorphism (SNP) data for 2093 pigs, including 1839 domestic pigs representing 122 local and commercial breeds, 215 wild boars, and 39 out-group suids, from Asia, Europe, America, Oceania and Africa. The aim of this study was to infer global patterns in pig domestication and diversity related to demography, migration, and selection., [Results]: A deep phylogeographic division reflects the dichotomy between early domestication centers. In the core Eastern and Western domestication regions, Chinese pigs show differentiation between breeds due to geographic isolation, whereas this is less pronounced in European pigs. The inferred European origin of pigs in the Americas, Africa, and Australia reflects European expansion during the sixteenth to nineteenth centuries. Human-mediated introgression, which is due, in particular, to importing Chinese pigs into the UK during the eighteenth and nineteenth centuries, played an important role in the formation of modern pig breeds. Inbreeding levels vary markedly between populations, from almost no runs of homozygosity (ROH) in a number of Asian wild boar populations, to up to 20% of the genome covered by ROH in a number of Southern European breeds. Commercial populations show moderate ROH statistics. For domesticated pigs and wild boars in Asia and Europe, we identified highly differentiated loci that include candidate genes related to muscle and body development, central nervous system, reproduction, and energy balance, which are putatively under artificial selection., [Conclusions]: Key events related to domestication, dispersal, and mixing of pigs from different regions are reflected in the 60K SNP data, including the globalization that has recently become full circle since Chinese pig breeders in the past decades started selecting Western breeds to improve local Chinese pigs. Furthermore, signatures of ongoing and past selection, acting at different times and on different genetic backgrounds, enhance our insight in the mechanism of domestication and selection. The global diversity statistics presented here highlight concerns for maintaining agrodiversity, but also provide a necessary framework for directing genetic conservation., This study is supported by National Production Technology System for the Pig Industry in China (nycytx-008) and Outstanding Talents and Innovation Team of Agricultural Science (2011-81) to LH, AGL2010-14822 and AGL2013-41834-R (Ministry of Economy and Science, Spain) to MPE. LI received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No. 656697. HJM received funding from the IMAGE project (Horizon 2020, No. 677353).

Hybridization between wild boar (Sus scrofa) and domestic pig occurred in the past and still occurs today, having great evolutionary and management implications. In fact, genetic introgression from the domestic form may alter traits like behavior, reproduction rate, and immunology in wild populations, with likely demographic impacts. Thus, it is crucial to understand under what conditions hybridization occurs in S. scrofa. Captive crosses with domestic pigs (released or escaped) have been suggested to constitute the major source of the spread of domestic genes into wild boar populations. However, to date, few studies have assessed the degree of admixture in farmed animals in comparison to the surrounding wild populations. With this purpose, we analyzed microsatellite loci in wild boar sampled in breeding stations and in the local wild population in two Italian regions (Sardinia and Piedmont). Both captive populations had lower allelic richness than the corresponding wild population, but a similar expected heterozygosity. In Piedmont, introgression from the domestic form into the wild population seems to be extremely low, while there are significant signs of admixture in the sampled breeding stations. In Sardinia, instead, the captive sample did not differ significantly from the wild population, which showed moderate signs of introgression. We conclude that hybridization in nature seems to play the key role in Sardinia, while intentional hybridization in captivity is the major source of introgression in Piedmont. Our findings emphasize the need for a routine genetic monitoring of wild boar captive populations, coupled with reference data on the neighboring wild populations.

After a strong demographic decline before World War II, wild boar populations are expanding and the species is now the second-most abundant ungulate in Europe. This increase raises concerns due to wild boar impact on crops and natural ecosystems and as potential vector of diseases. Additionally, wild boar can hybridize with domestic pigs, which could increase health risks and alter wild boar adaptive potential. We analysed 47,148 Single Nucleotide Polymorphisms in wild boar from Europe (292) and the Near East (16), and commercial (44) and local (255) pig breeds, to discern patterns of hybridization across Europe. We identified 33 wild boars with more than 10% domestic ancestry in their genome, mostly concentrated in Austria, Bosnia and Herzegovina, Bulgaria and Serbia. This difference is probably due to contrasting practices, with free-ranging vs. industrial farming but more samples would be needed to investigate larger geographic patterns. Our results suggest hybridization has occurred over a long period and is still ongoing, as we observed recent hybrids. Although wild and domestic populations have maintained their genetic distinctiveness, potential health threats raise concerns and require implementation of management actions and farming practices aimed at reducing contact between wild and domestic pigs.

[Aim]: Climate changes in the past had a deep impact on the evolutionary history of many species and left genetic signatures that are often still detectable today. We investigated the geographical pattern of mitochondrial DNA diversity in the European wild boar (Sus scrofa). Our final aims were to clarify the influence of present and past climatic conditions, infer the geographical position of glacial refugia, and suggest post-glacial spatial dynamics. [Location]: Europe. [Methods]: D-loop sequences were obtained for 763 individuals from Portugal to western Russia. Phylogenetic, multivariate and interpolation methods were used to describe the genetic and geographical patterns. Climatic suitability during the Last Glacial Maximum (LGM) was predicted using MaxEnt. The effect of present and past suitability on the observed patterns of diversity was evaluated by multiple linear regression. [Results]: We confirmed the existence of a ubiquitous mitochondrial clade in Europe (E1), an endemic clade in Italy (E2) and a few East Asian haplotypes (A), presumably introgressed from domestic pigs. No Near Eastern haplotypes were detected. Genetic divergence was not simply related to geographical distance. A clear south-north decreasing gradient of diversity was observed, with maximum levels in putative glacial refugia. Latitudinal variation in climatic conditions during the LGM was shown to be a good predictor of current genetic diversity. Moreover, an unexpected similarity between Iberia and eastern Europe was observed, while central European populations showed a higher affinity to the Italian gene pool. [Main conclusions]: The current distribution of mitochondrial genetic diversity was highly influenced by past climatic events, especially those related to the LGM, and is consistent with a major contribution of the Italian peninsula and the Balkans to the post-glacial recolonization of northern areas. More recent processes, such as restocking and extensive hunting, probably acted at rather local scales, without great impact on the global pattern of mitochondrial diversity., Sequencing activities were funded by the Sardinian Regional Government (L.R. 7/2007 Promozione della Ricerca Scientifica e dell’Innovazione Tecnologica in Sardegna). D.B. had a PhD grant financed by the European Social Fund, Operational Programme 2007/2013 – Axis IV – Human Capital Investment. S.T.V. was supported by a PhD Fellowship for International students of the University of Ferrara. B.J. andT.B. were supported by grants N N304 253935 from the Polish Ministry of Science and Higher Education and Research Potential in Conservation and Sustainable Management of Biodiversity – BIOCONSUS from the EC’s 7th Framework Programme (contract no. 245737). Mobility between labs in Sassari and Kiel was financed by the Italy–Germany Vigoni cooperation program 2010–2011.

In many areas, the management of overexploited populations of brown hare (Lepus europaeus) is based on annual restocking. While in some cases exotic hares are introduced, in some others hares are captured locally within protected areas and subsequently released into hunting grounds. We evaluated the genetic effects of this management regime in an Italian province where the brown hare population has recovered in the last few decades, by sequencing the hypervariable domain 1 of the mitochondrial control region and by genotyping eight autosomal microsatellites in hares sampled in both hunting and non-hunting areas. Both nuclear (H e = 0.68 and H o = 0.65) and mitochondrial variability (h = 0.853 and π = 0.012) were in line with other European populations. When comparing our data with mitochondrial sequences retrieved from GenBank, out of the 21 detected haplotypes, 14 were private to our study area. While 4.6 % of the individuals were found to carry haplotypes attributable to past introductions, 41.5 % grouped within a well-supported lineage, previously identified with a presumed native Italian taxon, L. e. meridiei. Despite the detectable geographic partitioning of mitochondrial haplotypes across the province, no genetic structure resulted from microsatellites analysis, indicating that no reproductive barriers exist among hares carrying different mitochondrial lineages. In conclusion, the local management seems to have contributed to the recovery of the species and to a full admixture of nuclear genes in the province. However, neither the extensive translocations nor the possible introductions of exotic heads seem to have completely undermined the local mitochondrial lineages.

Introgression of domestic genes into wild populations is considered to be a threat to biodiversity leading to loss of local adaptation and spread of human selected gene variants. Additionally, the contact between wild and domestic stocks can result in the transmission of infectious diseases. Thus, a better understanding of hybridization patterns in Europe might have important implications for conservation and management of both wild populations and local breeds, as well as for the contingency of infectious diseases.Here we present the results for the analysis of 235 wild boars (WB; from 22 areas) and 149 domestic pigs (DP; 49 from 5 commercial lines and 100 from 9 local breeds) with the Porcine SNP60 Beadchip.Principal component analysis shows a clear separation between domestic and wild populations. The network analysis concordantly identifies two main clusters (corresponding to WB and DP) but, additionally, highlights the presence of several individuals, of both ancestries, with intermediate positions. This result was confirmed by Admixture analysis that detected the presence of hybrid individuals in both WB and local domestic pig breeds.The introgression level varies considerably among populations, from non-detectable to very high. This result might be due to breeding practices, population history and WB management, but it arises questions on domestication and selective processes as well. Additional analyses to identify the presence of human or natural selected regions will be needed to disentangle the observed results.

The evolution of island populations in natural systems is driven by local adaptation and genetic drift. However, evolutionary pathways may be altered by humans in several ways. The wild boar (WB) (Sus scrofa) is an iconic game species occurring in several islands, where it has been strongly managed since prehistoric times. We examined genomic diversity at 49 803 single-nucleotide polymorphisms in 99 Sardinian WBs and compared them with 196 wild specimens from mainland Europe and 105 domestic pigs (DP; 11 breeds). High levels of genetic variation were observed in Sardinia (80.9% of the total number of polymorphisms), which can be only in part associated to recent genetic introgression. Both Principal Component Analysis and Bayesian clustering approach revealed that the Sardinian WB population is highly differentiated from the other European populations (F ST =0.126-0.138), and from DP (F ST =0.169). Such evidences were mostly unaffected by an uneven sample size, although clustering results in reference populations changed when the number of individuals was standardized. Runs of homozygosity (ROHs) pattern and distribution in Sardinian WB are consistent with a past expansion following a bottleneck (small ROHs) and recent population substructuring (highly homozygous individuals). The observed effect of a non-random selection of Sardinian individuals on diversity, F ST and ROH estimates, stressed the importance of sampling design in the study of structured or introgressed populations. Our results support the heterogeneity and distinctiveness of the Sardinian population and prompt further investigations on its origins and conservation status.

We investigated local gene flow in a high-density wolf (Canis lupus) population of the Italian Apennines, where no effective barrier to wolf dispersal was present. From 1998 to 2004 we examined wolf carcasses and non-invasively collected samples, focusing on three mountain districts, separated by two valleys, where wolf packs showed high spatial stability. Using nine autosomal microsatellites we successfully genotyped 177 samples, achieving the identification of 74 wolves. Genetic relatedness steeply decreased with increasing distance between sampling areas, thus suggesting that short-distance interpack migration is infrequent in this population. In addition, no individual from a central pack under intensive monitoring was sampled in the range of the surrounding packs over a 4-year period. The limited short-distance gene flow resulted in a cryptic genetic structure, which was revealed by Bayesian analysis. A different genetic cluster was found in each of the three mountain areas, and a small proportion of first-generation immigrants was detected. Overall, the present study suggests that local genetic differentiation in Italian wolves might arise from high spatial stability of packs and can be favoured by a combination of long-range dispersal, the attitude to mate between unrelated individuals and a high young mortality rate.

1. Despite the vast literature on genetic variation in the domestic pig Sus scrofa , little is known about genetic differentiation in wild boar populations. 2. Here we present an up-to-date review of published data on the past and recent history of the European wild boar, its genetic diversity and the spatial distribution of genetic variation throughout the continent. 3. The phylogeography of the species seems to be shaped mostly by past large-scale events (like postglacial recolonization) rather than by more recent human manipulation. Genetic differentiation is observed both on a continental and a regional scale, and non-intuitive barriers to gene flow occur. 4. From an indirect estimate, hybridization between wild boar and domestic pigs is seemingly a minor source of genetic variation for wild boar populations, yet risks are still linked to the release of captive hybrids in some areas. 5. Finally, we present future perspectives concerning the development of powerful molecular tools and their possible application to the study and management of this species.

We investigated the social organization of wild boars (Sus scrofa) using genetic and spatial data from a study population in Tuscany, Italy. In total, 120 wild boars of different sexes and age classes were captured and monitored from 2002 to 2006. All of them were genetically analyzed by using 10 polymorphic microsatellites (HE = 0.693, k = 6.6) and a matrix of pairwise relatedness was calculated. In addition, a reference sample of fully related individuals was created by genotyping 11 adult females and their fetuses (n = 56). Spatial data were gathered for 65 animals that had been fitted with either radiocollars or ear transmitters. Sixteen social units were identified by capture data and confirmed by observations and telemetry. A correlation between interindividual spatial distance and relatedness was observed only in summer-early autumn and seemed to be associated to the presence of piglets. The prediction of matrilinearity in wild boar social units was not confirmed, because a low degree of relatedness among boars was observed within groups. Aggregations of unrelated adult females (with their litters) were detected in the study population. The high turnover in the population due to human-caused mortality seems to be the main factor responsible for this altered social structure. Accordingly, we suggest that the observed social organization would result from grouping of unrelated survivors that is promoted by the presence of wolves in the area.