Estimating distribution and abundance of wide-ranging species with integrated spatial models: Opportunities revealed by the first wolf assessment in south-central Italy.

Estimating demographic parameters for wide-ranging and elusive species living at low density is challenging, especially at the scale of an entire country. To produce wolf distribution and abundance estimates for the whole south-central portion of the Italian wolf population, we developed an integrated spatial model, based on the data collected during a 7-month sampling campaign in 2020-2021. Data collection comprised an extensive survey of wolf presence signs, and an intensive survey in 13 sampling areas, aimed at collecting non-invasive genetic samples (NGS). The model comprised (i) a single-season, multiple data-source, multi-event occupancy model and (ii) a spatially explicit capture-recapture model. The information about species' absence was used to inform local density estimates. We also performed a simulation-based assessment, to estimate the best conditions for optimizing sub-sampling and population modelling in the future. The integrated spatial model estimated that 74.2% of the study area in south-central Italy (95% CIs = 70.5% to 77.9%) was occupied by wolves, for a total extent of the wolf distribution of 108,534 km2 (95% CIs = 103,200 to 114,000). The estimate of total population size for the Apennine wolf population was of 2557 individuals (SD = 171.5; 95% CIs = 2127 to 2844). Simulations suggested that the integrated spatial model was associated with an average tendency to slightly underestimate population size. Also, the main contribution of the integrated approach was to increase precision in the abundance estimates, whereas it did not affect accuracy significantly. In the future, the area subject to NGS should be increased to at least 30%, while at least a similar proportion should be sampled for presence-absence data, to further improve the accuracy of population size estimates and avoid the risk of underestimation. This approach could be applied to other wide-ranging species and in other geographical areas, but specific a priori evaluations of model requirements and expected performance should be made.

Estimating Wolf Population Size and Dynamics by Field Monitoring and Demographic Models: Implications for Management and Conservation.

We estimated the current size and dynamics of the wolf population in Tuscany and investigated the trends and demographic drivers of population changes. Estimates were obtained by two different approaches: (i) mixed-technique field monitoring (from 2014 to 2016) that found the minimum observed pack number and estimated population size, and (ii) an individual-based model (run by Vortex software v. 10.3.8.0) with demographic inputs derived from a local intensive study area and historic data on population size. Field monitoring showed a minimum population size of 558 wolves (SE = 12.005) in 2016, with a density of 2.74 individuals/100 km2. The population model described an increasing trend with an average annual rate of increase λ = 1.075 (SE = 0.014), an estimated population size of about 882 individuals (SE = 9.397) in 2016, and a density of 4.29 wolves/100 km2. Previously published estimates of wolf population were as low as 56.2% compared to our field monitoring estimation and 34.6% in comparison to our model estimation. We conducted sensitivity tests to analyze the key parameters driving population changes based on juvenile and adult mortality rates, female breeding success, and litter size. Mortality rates played a major role in determining intrinsic growth rate changes, with adult mortality accounting for 62.5% of the total variance explained by the four parameters. Juvenile mortality was responsible for 35.8% of the variance, while female breeding success and litter size had weak or negligible effects. We concluded that reliable estimates of population abundance and a deeper understanding of the role of different demographic parameters in determining population dynamics are crucial to define and carry out appropriate conservation and management strategies to address human-wildlife conflicts.

A multidisciplinary approach to estimating wolf population size for long-term conservation.

The wolf (Canis lupus) is among the most controversial of wildlife species. Abundance estimates are required to inform public debate and policy decisions, but obtaining them at biologically relevant scales is challenging. We developed a system for comprehensive population estimation across the Italian alpine region (100,000 km2 ), involving 1513 trained operators representing 160 institutions. This extensive network allowed for coordinated genetic sample collection and landscape-level spatial capture-recapture analyses that transcended administrative boundaries to produce the first estimates of key parameters for wolf population status assessment. Wolf abundance was estimated at 952 individuals (95% credible interval 816-1120) and 135 reproductive units (i.e., packs) (95% credible interval 112-165). We also estimated that mature individuals accounted for 33-45% of the entire population. The monitoring effort was spatially estimated thereby overcoming an important limitation of citizen science data. This is an important approach for promoting wolf-human coexistence based on wolf abundance monitoring and an endorsement of large-scale harmonized conservation practices.

Una estrategia multidisciplinaria para la estimación del tamaño poblacional de los lobos para la conservación a largo plazo Resumen El lobo (Canis lupus) está entre las especies de fauna más controversiales. Se requieren estimaciones de abundancia para informar al debate público y las decisiones políticas, pero es un reto obtenerlos en escalas con relevancia biológica. Desarrollamos un sistema para la estimación completa de la población en la región alpina de Italia (100,000 km2 ), con la participación de 1,513 operadores entrenados que representan a 160 instituciones. Esta red extensa permitió una colecta coordinada de muestras genéticas y análisis de captura-recaptura espacial que trascendieron las fronteras administrativas para así producir las primeras estimaciones de los parámetros clave para la evaluación del estado de la población de los lobos. Se estimó la abundancia en 952 individuos (95% intervalo de confianza 816-1120) y 135 unidades reproductivas (es decir, manadas) (95% intervalo de confianza 112-165). También estimamos que los individuos maduros representaban el 33-45% de toda la población. El esfuerzo de monitoreo se estimó espacialmente, por lo que sobrepasó una limitación importante de la ciencia ciudadana. Esta estrategia es importante para promover la coexistencia entre lobos y humanos con base en el monitoreo de la abundancia y el apoyo a las prácticas armonizadas de conservación a gran escala.

Possible origins and implications of atypical morphologies and domestication-like traits in wild golden jackals (Canis aureus).

Deciphering the origins of phenotypic variations in natural animal populations is a challenging topic for evolutionary and conservation biologists. Atypical morphologies in mammals are usually attributed to interspecific hybridisation or de-novo mutations. Here we report the case of four golden jackals (Canis aureus), that were observed during a camera-trapping wildlife survey in Northern Israel, displaying anomalous morphological traits, such as white patches, an upturned tail, and long thick fur which resemble features of domesticated mammals. Another individual was culled under permit and was genetically and morphologically examined. Paternal and nuclear genetic profiles, as well as geometric morphometric data, identified this individual as a golden jackal rather than a recent dog/wolf-jackal hybrid. Its maternal haplotype suggested past introgression of African wolf (Canis lupaster) mitochondrial DNA, as previously documented in other jackals from Israel. When viewed in the context of the jackal as an overabundant species in Israel, the rural nature of the surveyed area, the abundance of anthropogenic waste, and molecular and morphological findings, the possibility of an individual presenting incipient stages of domestication should also be considered.

How to Choose? Comparing Different Methods to Count Wolf Packs in a Protected Area of the Northern Apennines.

Despite a natural rewilding process that caused wolf populations in Europe to increase and expand in the last years, human-wolf conflicts still persist, threatening the long-term wolf presence in both anthropic and natural areas. Conservation management strategies should be carefully designed on updated population data and planned on a wide scale. Unfortunately, reliable ecological data are difficult and expensive to obtain and often hardly comparable through time or among different areas, especially because of different sampling designs. In order to assess the performance of different methods to estimate wolf (Canis lupus L.) abundance and distribution in southern Europe, we simultaneously applied three techniques: wolf howling, camera trapping and non-invasive genetic sampling in a protected area of the northern Apennines. We aimed at counting the minimum number of packs during a single wolf biological year and evaluating the pros and cons for each technique, comparing results obtained from different combinations of these three methods and testing how sampling effort may affect results. We found that packs' identifications could be hardly comparable if methods were separately used with a low sampling effort: wolf howling identified nine, camera trapping 12 and non-invasive genetic sampling eight packs. However, increased sampling efforts produced more consistent and comparable results across all used methods, although results from different sampling designs should be carefully compared. The integration of the three techniques yielded the highest number of detected packs, 13, although with the highest effort and cost. A common standardised sampling strategy should be a priority approach to studying elusive large carnivores, such as the wolf, allowing for the comparison of key population parameters and developing shared and effective conservation management plans.

Phylogenetic History and Phylogeographic Patterns of the European Wildcat (Felis silvestris) Populations.

Disentangling phylogenetic and phylogeographic patterns is fundamental to reconstruct the evolutionary histories of taxa and assess their actual conservation status. Therefore, in this study, for the first time, the most exhaustive biogeographic history of European wildcat (Felis silvestris) populations was reconstructed by typing 430 European wildcats, 213 domestic cats, and 72 putative admixed individuals, collected across the entire species' distribution range, at a highly diagnostic portion of the mitochondrial ND5 gene. Phylogenetic and phylogeographic analyses identified two main ND5 lineages (D and W) roughly associated with domestic and wild polymorphisms. Lineage D included all domestic cats, 83.3% of putative admixed individuals, and also 41.4% of wildcats; these latter mostly showed haplotypes belonging to sub-clade Ia, that diverged about 37,700 years ago, long pre-dating any evidence for cat domestication. Lineage W included all the remaining wildcats and putative admixed individuals, spatially clustered into four main geographic groups, which started to diverge about 64,200 years ago, corresponding to (i) the isolated Scottish population, (ii) the Iberian population, (iii) a South-Eastern European cluster, and (iv) a Central European cluster. Our results suggest that the last Pleistocene glacial isolation and subsequent re-expansion from Mediterranean and extra-Mediterranean glacial refugia were pivotal drivers in shaping the extant European wildcat phylogenetic and phylogeographic patterns, which were further modeled by both historical natural gene flow among wild lineages and more recent wild x domestic anthropogenic hybridization, as confirmed by the finding of F. catus/lybica shared haplotypes. The reconstructed evolutionary histories and the wild ancestry contents detected in this study could be used to identify adequate Conservation Units within European wildcat populations and help to design appropriate long-term management actions.

"Guess Who's Coming to Dinner": Molecular Tools to Reconstruct multilocus Genetic Profiles from Wild Canid Consumption Remains.

Non-invasive genetic sampling is a practical tool to monitor pivotal ecological parameters and population dynamic patterns of endangered species. It can be particularly suitable when applied to elusive carnivores such as the Apennine wolf (Canis lupus italicus) and the European wildcat (Felis silvestris silvestris), which can live in overlapping ecological contexts and sometimes share their habitats with their domestic free-ranging relatives, increasing the risk of anthropogenic hybridisation. In this case study, we exploited all the ecological and genetic information contained in a single biological canid faecal sample, collected in a forested area of central Italy, to detect any sign of trophic interactions between wolves and European wildcats or their domestic counterparts. Firstly, the faecal finding was morphologically examined, showing the presence of felid hair and claw fragment remains. Subsequently, total genomic DNA contained in the hair and claw samples was extracted and genotyped, through a multiple-tube approach, at canid and felid diagnostic panels of microsatellite loci. Finally, the obtained individual multilocus genotypes were analysed with reference wild and domestic canid and felid populations to assess their correct taxonomic status using Bayesian clustering procedures. Assignment analyses classified the genotype obtained from the endothelial cells present on the hair sample as a wolf with slight signals of dog ancestry, showing a qi = 0.954 (C.I. 0.780-1.000) to the wolf cluster, and the genotype obtained from the claw as a domestic cat, showing a qi = 0.996 (95% C.I. = 0.982-1.000) to the domestic cat cluster. Our results clearly show how a non-invasive multidisciplinary approach allows the cost-effective identification of both prey and predator genetic profiles and their taxonomic status, contributing to the improvement of our knowledge about feeding habits, predatory dynamics, and anthropogenic hybridisation risk in threatened species.

A reduced SNP panel to trace gene flow across southern European wolf populations and detect hybridization with other Canis taxa.

Intra- and inter-specific gene flow are natural evolutionary processes. However, human-induced hybridization is a global conservation concern across taxa, and the development of discriminant genetic markers to differentiate among gene flow processes is essential. Wolves (Canis lupus) are affected by hybridization, particularly in southern Europe, where ongoing recolonization of historic ranges is augmenting gene flow among divergent populations. Our aim was to provide diagnostic canid markers focused on the long-divergent Iberian, Italian and Dinaric wolf populations, based on existing genomic resources. We used 158 canid samples to select a panel of highly informative single nucleotide polymorphisms (SNPs) to (i) distinguish wolves in the three regions from domestic dogs (C. l. familiaris) and golden jackals (C. aureus), and (ii) identify their first two hybrid generations. The resulting 192 SNPs correctly 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 canid samples performed successfully for 178 SNPs. By separating natural population admixture from inter-specific hybridization, our reduced panel can help advance evolutionary research, monitoring, and timely conservation management.

Reliable wolf-dog hybrid detection in Europe using a reduced SNP panel developed for non-invasively collected samples.

BACKGROUND: Understanding the processes that lead to hybridization of wolves and dogs is of scientific and management importance, particularly over large geographical scales, as wolves can disperse great distances. However, a method to efficiently detect hybrids in routine wolf monitoring is lacking. Microsatellites offer only limited resolution due to the low number of markers showing distinctive allele frequencies between wolves and dogs. Moreover, calibration across laboratories is time-consuming and costly. In this study, we selected a panel of 96 ancestry informative markers for wolves and dogs, derived from the Illumina CanineHD Whole-Genome BeadChip (174 K). We designed very short amplicons for genotyping on a microfluidic array, thus making the method suitable also for non-invasively collected samples. RESULTS: Genotypes based on 93 SNPs from wolves sampled throughout Europe, purebred and non-pedigree dogs, and suspected hybrids showed that the new panel accurately identifies parental individuals, first-generation hybrids and first-generation backcrosses to wolves, while second- and third-generation backcrosses to wolves were identified as advanced hybrids in almost all cases. Our results support the hybrid identity of suspect individuals and the non-hybrid status of individuals regarded as wolves. We also show the adequacy of these markers to assess hybridization at a European-wide scale and the importance of including samples from reference populations. CONCLUSIONS: We showed that the proposed SNP panel is an efficient tool for detecting hybrids up to the third-generation backcrosses to wolves across Europe. Notably, the proposed genotyping method is suitable for a variety of samples, including non-invasive and museum samples, making this panel useful for wolf-dog hybrid assessments and wolf monitoring at both continental and different temporal scales.

Population Dynamics in Italian Canids between the Late Pleistocene and Bronze Age.

Dog domestication is still largely unresolved due to time-gaps in the sampling of regions. Ancient Italian canids are particularly understudied, currently represented by only a few specimens. In the present study, we sampled 27 canid remains from Northern Italy dated between the Late Pleistocene and Bronze Age to assess their genetic variability, and thus add context to dog domestication dynamics. They were targeted at four DNA fragments of the hypervariable region 1 of mitochondrial DNA. A total of 11 samples had good DNA preservation and were used for phylogenetic analyses. The dog samples were assigned to dog haplogroups A, C and D, and a Late Pleistocene wolf was set into wolf haplogroup 2. We present our data in the landscape of ancient and modern dog genetic variability, with a particular focus on the ancient Italian samples published thus far. Our results suggest there is high genetic variability within ancient Italian canids, where close relationships were evident between both a ~24,700 years old Italian canid, and Iberian and Bulgarian ancient dogs. These findings emphasize that disentangling dog domestication dynamics benefits from the analysis of specimens from Southern European regions.

Genomic evidence for the Old divergence of Southern European wolf populations.

The grey wolf (Canis lupus) is one of the most widely distributed mammals in which a variety of distinct populations have been described. However, given their currently fragmented distribution and recent history of human-induced population decline, little is known about the events that led to their differentiation. Based on the analysis of whole canid genomes, we examined the divergence times between Southern European wolf populations and their ancient demographic history. We found that all present-day Eurasian wolves share a common ancestor ca 36 000 years ago, supporting the hypothesis that all extant wolves derive from a single population that subsequently expanded after the Last Glacial Maximum. We also estimated that the currently isolated European populations of the Iberian Peninsula, Italy and the Dinarics-Balkans diverged very closely in time, ca 10 500 years ago, and maintained negligible gene flow ever since. This indicates that the current genetic and morphological distinctiveness of Iberian and Italian wolves can be attributed to their isolation dating back to the end of the Pleistocene, predating the recent human-induced extinction of wolves in Central Europe by several millennia.

A standardized approach to empirically define reliable assignment thresholds and appropriate management categories in deeply introgressed populations.

Anthropogenic hybridization is recognized as a major threat to the long-term survival of natural populations. While identifying F1 hybrids might be simple, the detection of older admixed individuals is far from trivial and it is still debated whether they should be targets of management. Examples of anthropogenic hybridization have been described between wolves and domestic dogs, with numerous cases detected in the Italian wolf population. After selecting appropriate wild and domestic reference populations, we used empirical and simulated 39-autosomal microsatellite genotypes, Bayesian assignment and performance analyses to develop a workflow to detect different levels of wolf x dog admixture. Membership proportions to the wild cluster (qiw) and performance indexes identified two q-thresholds which allowed to efficiently classify the analysed genotypes into three assignment classes: pure (with no or negligible domestic ancestry), older admixed (with a marginal domestic ancestry) and recent admixed (with a clearly detectable domestic ancestry) animals. Based on their potential to spread domestic variants, such classes were used to define three corresponding management categories: operational pure, introgressed and operational hybrid individuals. Our multiple-criteria approach can help wildlife managers and decision makers in more efficiently targeting the available resources for the long-term conservation of species threatened by anthropogenic hybridization.

Genomic approaches to identify hybrids and estimate admixture times in European wildcat populations.

The survival of indigenous European wildcat (Felis silvestris silvestris) populations can be locally threatened by introgressive hybridization with free-ranging domestic cats. Identifying pure wildcats and investigating the ancestry of admixed individuals becomes thus a conservation priority. We analyzed 63k cat Single Nucleotide Polymorphisms (SNPs) with multivariate, Bayesian and gene-search tools to better evaluate admixture levels between domestic and wild cats collected in Europe, timing and ancestry proportions of their hybrids and backcrosses, and track the origin (wild or domestic) of the genomic blocks carried by admixed cats, also looking for possible deviations from neutrality in their inheritance patterns. Small domestic ancestry blocks were detected in the genomes of most admixed cats, which likely originated from hybridization events occurring from 6 to 22 generations in the past. We identified about 1,900 outlier coding genes with excess of wild or domestic ancestry compared to random expectations in the admixed individuals. More than 600 outlier genes were significantly enriched for Gene Ontology (GO) categories mainly related to social behavior, functional and metabolic adaptive processes (wild-like genes), involved in cognition and neural crest development (domestic-like genes), or associated with immune system functions and lipid metabolism (parental-like genes). These kinds of genomic ancestry analyses could be reliably applied to unravel the admixture dynamics in European wildcats, as well as in other hybridizing populations, in order to design more efficient conservation plans.

The Sicilian Wolf: Genetic Identity of a Recently Extinct Insular Population.

Historically, many local grey wolf (Canis lupus) populations have undergone substantial reductions in size or become extinct. Among these, the wolf population once living in Sicily, the largest island in the Mediterranean Sea, was completely eradicated by human activity in the early decades of the 20th century. To gain a better understanding of the genetic identity of the Sicilian wolf, we used techniques for the study of ancient DNA to analyze the mitochondrial (mt) variability of six specimens stored in Italian museums. We were able to amplify a diagnostic mtDNA fragment of the control region (CR) in four of the samples. Two of the samples shared the same haplotype, differing by two substitutions from the currently most diffused Italian wolf haplotype (W14) and one substitution from the only other Italian haplotype (W16). The third sample showed a previously unreported wolf-like haplotype, and the fourth a haplotype commonly found in dogs. All of the wolf haplotypes analyzed in this study belonged to the mitochondrial haplogroup that includes haplotypes detected in all the known European Pleistocene wolves and in several modern southern European populations. Unfortunately, this endemic island population, which exhibited unique mtDNA variability, was definitively lost before it was possible to understand its taxonomic uniqueness and conservational value.

Old wild wolves: ancient DNA survey unveils population dynamics in Late Pleistocene and Holocene Italian remains.

BACKGROUND: The contemporary Italian wolf (Canis lupus italicus) represents a case of morphological and genetic uniqueness. Today, Italian wolves are also the only documented population to fall exclusively within the mitochondrial haplogroup 2, which was the most diffused across Eurasian and North American wolves during the Late Pleistocene. However, the dynamics leading to such distinctiveness are still debated. METHODS: In order to shed light on the ancient genetic variability of this wolf population and on the origin of its current diversity, we collected 19 Late Pleistocene-Holocene samples from northern Italy, which we analyzed at a short portion of the hypervariable region 1 of the mitochondrial DNA, highly informative for wolf and dog phylogenetic analyses. RESULTS: Four out of the six detected haplotypes matched the ones found in ancient wolves from northern Europe and Beringia, or in modern European and Chinese wolves, and appeared closely related to the two haplotypes currently found in Italian wolves. The haplotype of two Late Pleistocene samples matched with primitive and contemporary dog sequences from the canine mitochondrial clade A. All these haplotypes belonged to haplogroup 2. The only exception was a Holocene sample dated 3,250 years ago, affiliated to haplogroup 1. DISCUSSION: In this study we describe the genetic variability of the most ancient wolf specimens from Italy analyzed so far, providing a preliminary overview of the genetic make-up of the population that inhabited this area from the last glacial maximum to the Middle Age period. Our results endorsed that the genetic diversity carried by the Pleistocene wolves here analyzed showed a strong continuity with other northern Eurasian wolf specimens from the same chronological period. Contrarily, the Holocene samples showed a greater similarity only with modern sequences from Europe and Asia, and the occurrence of an haplogroup 1 haplotype allowed to date back previous finding about its presence in this area. Moreover, the unexpected discovery of a 24,700-year-old sample carrying a haplotype that, from the fragment here obtained, falls within the canine clade A, could represent the oldest evidence in Europe of such dog-rich clade. All these findings suggest complex population dynamics that deserve to be further investigated based on mitochondrial or whole genome sequencing.

Genetic Characterization of Canine Adenovirus Type 1 Detected by Real-Time Polymerase Chain Reaction in an Oral Sample of an Italian Wolf (Canis Lupus).

We report the detection of canine adenovirus type 1 DNA by real-time PCR technique in an oral sample of an Italian wolf (Canis lupus italicus). Genetic characterization of the virus revealed a strict relationship with viruses detected in dogs (Canis lupus familiaris), wolves, and red foxes (Vulpes vulpes), suggesting that transmission between wild animals and dogs had occurred.

Use of hidden Markov capture-recapture models to estimate abundance in the presence of uncertainty: Application to the estimation of prevalence of hybrids in animal populations.

Estimating the relative abundance (prevalence) of different population segments is a key step in addressing fundamental research questions in ecology, evolution, and conservation. The raw percentage of individuals in the sample (naive prevalence) is generally used for this purpose, but it is likely to be subject to two main sources of bias. First, the detectability of individuals is ignored; second, classification errors may occur due to some inherent limits of the diagnostic methods. We developed a hidden Markov (also known as multievent) capture-recapture model to estimate prevalence in free-ranging populations accounting for imperfect detectability and uncertainty in individual's classification. We carried out a simulation study to compare naive and model-based estimates of prevalence and assess the performance of our model under different sampling scenarios. We then illustrate our method with a real-world case study of estimating the prevalence of wolf (Canis lupus) and dog (Canis lupus familiaris) hybrids in a wolf population in northern Italy. We showed that the prevalence of hybrids could be estimated while accounting for both detectability and classification uncertainty. Model-based prevalence consistently had better performance than naive prevalence in the presence of differential detectability and assignment probability and was unbiased for sampling scenarios with high detectability. We also showed that ignoring detectability and uncertainty in the wolf case study would lead to underestimating the prevalence of hybrids. Our results underline the importance of a model-based approach to obtain unbiased estimates of prevalence of different population segments. Our model can be adapted to any taxa, and it can be used to estimate absolute abundance and prevalence in a variety of cases involving imperfect detection and uncertainty in classification of individuals (e.g., sex ratio, proportion of breeders, and prevalence of infected individuals).

Combining Bayesian genetic clustering and ecological niche modeling: Insights into wolf intraspecific genetic structure.

The distribution of intraspecific genetic variation and how it relates to environmental factors is of increasing interest to researchers in macroecology and biogeography. Recent studies investigated the relationships between the environment and patterns of intraspecific genetic variation across species ranges but only few rigorously tested the relation between genetic groups and their ecological niches. We quantified the relationship of genetic differentiation (F ST) and the overlap of ecological niches (as measured by n-dimensional hypervolumes) among genetic groups resulting from spatial Bayesian genetic clustering in the wolf (Canis lupus) in the Italian peninsula. Within the Italian wolf population, four genetic clusters were detected, and these clusters showed different ecological niches. Moreover, different wolf clusters were significantly related to differences in land cover and human disturbance features. Such differences in the ecological niches of genetic clusters should be interpreted in light of neutral processes that hinder movement, dispersal, and gene flow among the genetic clusters, in order to not prematurely assume any selective or adaptive processes. In the present study, we found that both the plasticity of wolves-a habitat generalist-to cope with different environmental conditions and the occurrence of barriers that limit gene flow lead to the formation of genetic intraspecific genetic clusters and their distinct ecological niches.

Wolf outside, dog inside? The genomic make-up of the Czechoslovakian Wolfdog.

BACKGROUND: Genomic methods can provide extraordinary tools to explore the genetic background of wild species and domestic breeds, optimize breeding practices, monitor and limit the spread of recessive diseases, and discourage illegal crossings. In this study we analysed a panel of 170k Single Nucleotide Polymorphisms with a combination of multivariate, Bayesian and outlier gene approaches to examine the genome-wide diversity and inbreeding levels in a recent wolf x dog cross-breed, the Czechoslovakian Wolfdog, which is becoming increasingly popular across Europe. RESULTS: Pairwise FST values, multivariate and assignment procedures indicated that the Czechoslovakian Wolfdog was significantly differentiated from all the other analysed breeds and also well-distinguished from both parental populations (Carpathian wolves and German Shepherds). Coherently with the low number of founders involved in the breed selection, the individual inbreeding levels calculated from homozygosity regions were relatively high and comparable with those derived from the pedigree data. In contrast, the coefficient of relatedness between individuals estimated from the pedigrees often underestimated the identity-by-descent scores determined using genetic profiles. The timing of the admixture and the effective population size trends estimated from the LD patterns reflected the documented history of the breed. Ancestry reconstruction methods identified more than 300 genes with excess of wolf ancestry compared to random expectations, mainly related to key morphological features, and more than 2000 genes with excess of dog ancestry, playing important roles in lipid metabolism, in the regulation of circadian rhythms, in learning and memory processes, and in sociability, such as the COMT gene, which has been described as a candidate gene for the latter trait in dogs. CONCLUSIONS: In this study we successfully applied genome-wide procedures to reconstruct the history of the Czechoslovakian Wolfdog, assess individual wolf ancestry proportions and, thanks to the availability of a well-annotated reference genome, identify possible candidate genes for wolf-like and dog-like phenotypic traits typical of this breed, including commonly inherited disorders. Moreover, through the identification of ancestry-informative markers, these genomic approaches could provide tools for forensic applications to unmask illegal crossings with wolves and uncontrolled trades of recent and undeclared wolfdog hybrids.

Studies of modern Italian dog populations reveal multiple patterns for domestic breed evolution.

Through thousands of years of breeding and strong human selection, the dog (Canis lupus familiaris) exists today within hundreds of closed populations throughout the world, each with defined phenotypes. A singular geographic region with broad diversity in dog breeds presents an interesting opportunity to observe potential mechanisms of breed formation. Italy claims 14 internationally recognized dog breeds, with numerous additional local varieties. To determine the relationship among Italian dog populations, we integrated genetic data from 263 dogs representing 23 closed dog populations from Italy, seven Apennine gray wolves, and an established dataset of 161 globally recognized dog breeds, applying multiple genetic methods to characterize the modes by which breeds are formed within a single geographic region. Our consideration of each of five genetic analyses reveals a series of development events that mirror historical modes of breed formation, but with variations unique to the codevelopment of early dog and human populations. Using 142,840 genome-wide SNPs and a dataset of 1,609 canines, representing 182 breeds and 16 wild canids, we identified breed development routes for the Italian breeds that included divergence from common populations for a specific purpose, admixture of regional stock with that from other regions, and isolated selection of local stock with specific attributes.