Detecting introgressive hybridization between free-ranging domestic dogs and wild wolves (Canis lupus) by admixture linkage disequilibrium analysis

Occasional crossbreeding between free-ranging domestic dogs and wild wolves (Canis lupus) has been detected in some European countries by mitochondrial DNA sequencing and genotyping unlinked microsatellite loci. Maternal and unlinked genomic markers, however, might underestimate the extent of introgressive hybridization, and their impacts on the preservation of wild wolf gene pools. In this study, we genotyped 220 presumed Italian wolves, 85 dogs and 7 known hybrids at 16 microsatellites belonging to four different linkage groups (plus four unlinked microsatellites). Population clustering and individual assignments were performed using a Bayesian procedure implemented in structure 2.1, which models the gametic disequilibrium arising between linked loci during admixtures, aiming to trace hybridization events further back in time and infer the population of origin of chromosomal blocks. Results indicate that (i) linkage disequilibrium was higher in wolves than in dogs; (ii) 11 out of 220 wolves (5.0%) were likely admixed, a proportion that is significantly higher than one admixed genotype in 107 wolves found previously in a study using unlinked markers; (iii) posterior maximum-likelihood estimates of the recombination parameter r revealed that introgression in Italian wolves is not recent, but could have continued for the last 70 (+/- 20) generations, corresponding to approximately 140-210 years. Bayesian clustering showed that, despite some admixture, wolf and dog gene pools remain sharply distinct (the average proportions of membership to wolf and dog clusters were Q(w) = 0.95 and Q(d) = 0.98, respectively), suggesting that hybridization was not frequent, and that introgression in nature is counteracted by behavioural or selective constraints.     

Genome‐wide sequence information reveals recurrent hybridization among diploid wheat wild relatives

Many conflicting hypotheses regarding the relationships among crops and wild species closely related to wheat (the genera Aegilops, Amblyopyrum, and Triticum) have been postulated. The contribution of hybridization to the evolution of these taxa is intensely discussed. To determine possible causes for this, and provide a phylogeny of the diploid taxa based on genome‐wide sequence information, independent data were obtained from genotyping‐by‐sequencing and a target‐enrichment experiment that returned 244 low‐copy nuclear loci. The data were analyzed using Bayesian, likelihood and coalescent‐based methods. D statistics were used to test if incomplete lineage sorting alone or together with hybridization is the source for incongruent gene trees. Here we present the phylogeny of all diploid species of the wheat wild relatives. We hypothesize that most of the wheat‐group species were shaped by a primordial homoploid hybrid speciation event involving the ancestral Triticum and Am. muticum lineages to form all other species except Ae. speltoides. This hybridization event was followed by multiple introgressions affecting all taxa except Triticum. Mostly progenitors of the extant species were involved in these processes, while recent interspecific gene flow seems insignificant. The composite nature of many genomes of wheat‐group taxa results in complicated patterns of diploid contributions when these lineages are involved in polyploid formation, which is, for example, the case for tetraploid and hexaploid wheats. Our analysis provides phylogenetic relationships and a testable hypothesis for the genome compositions in the basic evolutionary units within the wheat group of Triticeae.     

Biodiversity friend or foe: land use by a top predator,the dingo in contested landscapes of the Australian Wet Tropics

Dingoes (Canis dingo) in the coastal lowlands of Australia's Wet Tropics are perceived as a major threat to biodiversity and subjected to broad‐spectrum lethal control. However, evidence of their impacts is equivocal, and control programmes generally ignore the ecological benefits that dingoes might provide. Previous diet analysis has shown that dingoes in the Wet Tropics primarily prey on common, terrestrial mammals. However, little is known of dingo habitat use or prey acquisition in the region despite these activities having major implications for biodiversity conservation. We investigated land use by dingoes in the lowland Wet Tropics to enable predictions of potential prey types, relative prey use and modes of prey acquisition. Nine dingoes were tracked for 3–6 months. Home ranges and resting areas were estimated using multiple estimators, and habitat use was analysed using compositional analysis of habitat use and generalized additive models. Dingo ranging behaviour suggested that anthropogenic food subsidies were infrequently used. Each territory comprised several sclerophyll forest rest areas with adjacent sugarcane‐grassland high activity areas. Individuals used each rest‐activity area for extended durations before moving on to another. Sclerophyll and rainforests, which contain the fauna species of primary conservation concern, were generally used for rest/sleep, or movement between rest‐activity areas. Activity patterns were consistent with dingoes hunting in open sugarcane‐grassland habitats during daylight hours. Dingo activity was low in areas where fauna species of conservation concern occur, which suggests that dingoes do not pose a threat to their survival. Consequently, current broad‐spectrum lethal control may have minimal benefits or even incur costs for biodiversity. Maximizing the ecosystem services provided by dingoes while simultaneously minimizing their negative impacts requires a more targeted location‐specific management approach, one that assesses and mitigates impacts specifically where background circumstances suggest particular packs may be either a conservation or economic threat.     

A genome‐wide survey reveals abundant rice blast R genes in resistant cultivars

Plant resistance genes (R genes) harbor tremendous allelic diversity, constituting a robust immune system effective against microbial pathogens. Nevertheless, few functional R genes have been identified for even the best‐studied pathosystems. Does this limited repertoire reflect specificity, with most R genes having been defeated by former pests, or do plants harbor a rich diversity of functional R genes, the composite behavior of which is yet to be characterized? Here, we survey 332 NBS‐LRR genes cloned from five resistant Oryza sativa (rice) cultivars for their ability to confer recognition of 12 rice blast isolates when transformed into susceptible cultivars. Our survey reveals that 48.5% of the 132 NBS‐LRR loci tested contain functional rice blast R genes, with most R genes deriving from multi‐copy clades containing especially diversified loci. Each R gene recognized, on average, 2.42 of the 12 isolates screened. The abundant R genes identified in resistant genomes provide extraordinary redundancy in the ability of host genotypes to recognize particular isolates. If the same is true for other pathogens, many extant NBS‐LRR genes retain functionality. Our success at identifying rice blast R genes also validates a highly efficient cloning and screening strategy.     

Low neutral genetic variability in a specialist puffin hunter: the Norwegian Lundehund

The genetic variability of 125 Norwegian Lundehund and 27 Nova Scotia Duck Tolling Retriever was analysed using a set of 26 microsatellite markers. In Lundehund, the average number of alleles per locus was 1.73, and average observed (H_O) and expected (H_E) heterozygosity were 0.07. In Toller, all measures of genetic diversity were much higher than in Lundehund and similar to studies on other dog breeds. The cluster analysis correctly assigned individuals to their respective breed. The low genetic variability in Lundehund was not surprising, given the two strong bottlenecks in the 1940s and the 1960s. The relatedness of Lundehund to other Nordic small spitzes should be investigated in the view of possible outcrossing.     

Conservation and ecological implications of the use of space by chilla foxes and free‐ranging dogs in a human‐dominated landscape in southern Chile

We investigated whether domestic dogs (Canis lupus familiaris) influenced the use of space by chilla foxes (Lycalopex griseus) in southern Chile and tested the hypothesis that dogs interfere with chillas. We used scent stations and occurrence of scats within systematically placed plots to assess habitat use by both species and to test whether dogs were associated to the use of space by chillas. Activity data were obtained from captures for both species, and telemetry for foxes. Diet of both species was analysed in order to tease out the potential existence of exploitation competition. We found that, when active, chillas used prairies more and native forest less than expected according to availability, whereas inactive chillas (during the day) preferred native forest, the only habitat type that dogs did not use. The odds of dog occurrence increased at shorter distance to human houses, whereas the inverse pattern was observed for chillas. Poisson models showed that the number of chilla visits to scent stations was negatively correlated with the number of dog visits. We observed dogs persecuting and/or killing chillas which was also corroborated by local people that used dogs to prevent chilla‐related poultry losses, supporting the idea that dogs harass foxes actively. The analysis of scats showed that dogs fed mainly on house food and domestic ruminant carcasses whereas foxes fed on hare, hens, mice and wild birds, thus suggesting that exploitation competition is not a strong alternative hypothesis as a proximate cause for the patterns observed. We conclude that dogs seem to constrain the use of space by wild carnivores via interference.     

Size‐assortative choice and mate availability influences hybridization between red wolves (Canis rufus) and coyotes (Canis latrans)

Anthropogenic hybridization of historically isolated taxa has become a primary conservation challenge for many imperiled species. Indeed, hybridization between red wolves (Canis rufus) and coyotes (Canis latrans) poses a significant challenge to red wolf recovery. We considered seven hypotheses to assess factors influencing hybridization between red wolves and coyotes via pair‐bonding between the two species. Because long‐term monogamy and defense of all‐purpose territories are core characteristics of both species, mate choice has long‐term consequences. Therefore, red wolves may choose similar‐sized mates to acquire partners that behave similarly to themselves in the use of space and diet. We observed multiple factors influencing breeding pair formation by red wolves and found that most wolves paired with similar‐sized conspecifics and wolves that formed congeneric pairs with nonwolves (coyotes and hybrids) were mostly female wolves, the smaller of the two sexes. Additionally, we observed that lower red wolf abundance relative to nonwolves and the absence of helpers increased the probability that wolves consorted with nonwolves. However, successful pairings between red wolves and nonwolves were associated with wolves that maintained small home ranges. Behaviors associated with territoriality are energetically demanding and behaviors (e.g., aggressive interactions, foraging, and space use) involved in maintaining territories are influenced by body size. Consequently, we propose the hypothesis that size disparities between consorting red wolves and coyotes influence positive assortative mating and may represent a reproductive barrier between the two species. We offer that it may be possible to maintain wild populations of red wolves in the presence of coyotes if management strategies increase red wolf abundance on the landscape by mitigating key threats, such as human‐caused mortality and hybridization with coyotes. Increasing red wolf abundance would likely restore selection pressures that increase mean body and home‐range sizes of red wolves and decrease hybridization rates via reduced occurrence of congeneric pairs.     

Canid hybridization: contemporary evolution in human‐modified landscapes

Contemporary evolution through human‐induced hybridization occurs throughout the taxonomic range. Formerly allopatric species appear especially susceptible to hybridization. Consequently, hybridization is expected to be more common in regions with recent sympatry owing to human activity than in areas of historical range overlap. Coyotes ( Canis latrans) and gray wolves ( C. lupus) are historically sympatric in western North America. Following European settlement gray wolf range contracted, whereas coyote range expanded to include eastern North America. Furthermore, wolves with New World (NW) mitochondrial DNA (mtDNA) haplotypes now extend from Manitoba to Québec in Canada and hybridize with gray wolves and coyotes. Using mtDNA and 12 microsatellite markers, we evaluated levels of wolf‐coyote hybridization in regions where coyotes were present (the Canadian Prairies, n = 109 samples) and absent historically (Québec, n = 154). Wolves with NW mtDNA extended from central Saskatchewan (51°N, 69°W) to northeastern Québec (54°N, 108°W). On the Prairies, 6.3% of coyotes and 9.2% of wolves had genetic profiles suggesting wolf‐coyote hybridization. In contrast, 12.6% of coyotes and 37.4% of wolves in Québec had profiles indicating hybrid origin. Wolves with NW and Old World ( C. lupus) mtDNA appear to form integrated populations in both regions. Our results suggest that hybridization is more frequent in historically allopatric populations. Range shifts, now expected across taxa following climate change and other human influence on the environment, might therefore promote contemporary evolution by hybridization.     

Compound heterozygosity for TNXB genetic variants in a mixed‐breed dog with Ehlers‐Danlos syndrome

The Ehlers‐Danlos syndromes (EDSs) are a heterogeneous group of inherited connective tissue disorders characterized by skin hyperextensibility, joint hypermobility and tissue fragility. Inherited disorders similar to human EDS have been reported in different mammalian species. In the present study, we investigated a female mixed‐breed dog with clinical signs of EDS. Whole‐genome sequencing of the affected dog revealed two missense variants in the TNXB gene, encoding the extracellular matrix protein tenascin XB. In humans, TNXB genetic variants cause classical‐like EDS or the milder hypermobile EDS. The affected dog was heterozygous at both identified variants. Each variant allele was transmitted from one of the case's parents, consistent with compound heterozygosity. Although one of the variant alleles, XM_003431680.3:c.2012G>A, p.(Ser671Asn), was private to the family of the affected dog and absent from whole‐genome sequencing data of 599 control dogs, the second variant allele, XM_003431680.3:c.2900G>A, p.(Gly967Asp), is present at a low frequency in the Chihuahua and Poodle population. Given that TNXB is a functional candidate gene for EDS, we suggest that compound heterozygosity for the identified TNXB variants may have caused the EDS‐like phenotype in the affected dog. Chihuahuas and Poodles should be monitored for EDS cases, which might confirm the hypothesized pathogenic effect of the segregating TNXB variant.     

Low rates of hybridization between European wildcats and domestic cats in a human‐dominated landscape

Hybridization between wild species and their domestic congeners is considered a major threat for wildlife conservation. Genetic integrity of the European wildcat, for instance, is a concern as they are outnumbered by domestic cats by several orders of magnitude throughout its range. We genotyped 1,071 individual wildcat samples obtained from hair traps and roadkills collected across the highly fragmented forests of western Central Europe, in Germany and Luxembourg, to assess domestic cat introgression in wildcats in human‐dominated landscapes. Analyses using a panel of 75 autosomal SNPs suggested a low hybridization rate, with 3.5% of wildcat individuals being categorized as F1, F2, or backcrosses to either parental taxon. We report that results based on a set of SNPs were more consistent than on a set of 14 microsatellite markers, showed higher accuracy to detect hybrids and their class in simulation analyses, and were less affected by underlying population structure. Our results strongly suggest that very high hybridization rates previously reported for Central Europe may be partly due to inadequate choice of markers and/or sampling design. Our study documents that an adequately selected SNP panel for hybrid detection may be used as an alternative to commonly applied microsatellite markers, including studies relying on noninvasively collected samples. In addition, our finding of overall low hybridization rates in Central European wildcats provides an example of successful wildlife coexistence in human‐dominated, fragmented landscapes.     

Extensive hybridization and past introgression between divergent lineages in a quasi‐clonal hermaphroditic fish: Ramifications for species concepts and taxonomy

Extreme inbreeding is expected to reduce the incidence of hybridization, serving as a prezygotic barrier. Mangrove rivulus is a small killifish that reproduces predominantly by self‐fertilization, producing highly homozygous lines throughout its geographic range. The Bahamas and Caribbean are inhabited by two highly diverged phylogeographic lineages of mangrove rivulus, Kryptolebias marmoratus and a ‘Central clade’ closely related to K. hermaphroditus from Brazil. The two lineages are largely allopatric, but recently were found in syntopy on San Salvador, Bahamas, where a single hybrid was reported. To better characterize the degree of hybridization and the possibility of secondary introgression, here we conducted a detailed genetic analysis of the contact zone on San Salvador. Two mixed populations were identified, one of which contained sexually mature hybrids. The distribution of heterozygosity at diagnostic microsatellite loci in hybrids showed that one of these hybrids was an immediate offspring from the K. marmoratus x Central clade cross, whereas the remaining five hybrids were products of reproduction by self‐fertilization for 1–3 generations following the initial cross. Two hybrids had mitochondrial haplotypes of K. marmoratus and the remaining four hybrids had a haplotype of the Central clade, indicating that crosses go in both directions. In hybrids, alleles of parental lineages were represented in equal proportions suggesting lack of recent backcrossing to either of the parental lineages. However, sympatric populations of two lineages were less diverged than allopatric populations, consistent with introgression. Results are discussed in terms of applicability of the biological species concept for isogenic, effectively clonal, organisms.     

Influence of body mass and environmental conditions on winter mortality risk of a northern ungulate: Evidence for a late‐winter survival bottleneck

1. A relationship between winter weather and survival of northern ungulates has long been established, yet the possible roles of biological (e.g., nutritional status) and environmental (e.g., weather) conditions make it important to determine which potential limiting factors are most influential.
2. Our objective was to examine the potential effects of individual (body mass and age) and extrinsic (winter severity and snowmelt conditions) factors on the magnitude and timing of mortality for adult (>2.5 years old) female white‐tailed deer (Odocoileus virginianus [Zimmerman, 1780]) during February–May in the Upper Peninsula of Michigan, USA.
3. One hundred and fifty deer were captured and monitored during 2009–2015 in two areas with varying snowfall. February–May survival ranged from 0.24 to 0.89 (mean = 0.69) across years. Mortality risk increased 1.9% with each unit increase in cumulative winter severity index, decreased 8.2% with each cumulative snow‐free day, and decreased 4.3% with each kg increase in body mass. Age and weekly snow depth did not influence weekly deer survival. Predation, primarily from coyote (Canis latrans [Say, 1823]) and wolves (Canis lupus [L., 1758]), accounted for 78% of known‐cause mortalities.
4. Our results suggest that cumulative winter severity, and possibly to a lesser degree deer condition entering winter, impacted deer winter survival. However, the timing of spring snowmelt appeared to be the most influential factor determining late‐winter mortality of deer in our study. This supports the hypothesis that nutrition and energetic demands from weather conditions are both important to northern ungulate winter ecology. Under this model, a delay of several weeks in the timing of spring snowmelt could exert a large influence on deer survival, resulting in a survival bottleneck.

     

On the path to extinction: Inbreeding and admixture in a declining grey wolf population

Allee effects reduce the viability of small populations in many different ways, which act synergistically to lead populations towards extinction vortexes. The Sierra Morena wolf population, isolated in the south of the Iberian Peninsula and composed of just one or few packs for decades, represents a good example of how diverse threats act additively in very small populations. We sequenced the genome of one of the last wolves identified (and road‐killed) in Sierra Morena and that of another wolf in the Iberian Wolf Captive Breeding Program and compared them with other wolf and dog genomes from around the world (including two previously published genome sequences from northern Iberian wolves). The results showed relatively low overall genetic diversity in Iberian wolves, but diverse population histories including past introgression of dog genes. The Sierra Morena wolf had an extraordinarily high level of inbreeding and long runs of homozygosity, resulting from the long isolation. In addition, about one‐third of the genome was of dog origin. Despite the introgression of dog genes, heterozygosity remained low because of continued inbreeding after several hybridization events. The results thus illustrate the case of a small and isolated wolf population where the low population density may have favoured hybridization and introgression of dog alleles, but continued inbreeding may have resulted in large chromosomal fragments of wolf origin completely disappearing from the population, and being replaced by chromosomal fragments of dog origin. The latest population surveys suggest that this population may have gone extinct.     

Viral gut metagenomics of sympatric wild and domestic canids,and monitoring of viruses: Insights from an endangered wolf population

Animal host–microbe interactions are a relevant concern for wildlife conservation, particularly regarding generalist pathogens, where domestic host species can play a role in the transmission of infectious agents, such as viruses, to wild animals. Knowledge on viral circulation in wild host species is still scarce and can be improved by the recent advent of modern molecular approaches. We aimed to characterize the fecal virome and identify viruses of potential conservation relevance of diarrheic free‐ranging wolves and sympatric domestic dogs from Central Portugal, where a small and threatened wolf population persists in a highly anthropogenically modified landscape. Using viral metagenomics, we screened diarrheic stools collected from wolves (n = 8), feral dogs (n = 4), and pet dogs (n = 6), all collected within wolf range. We detected novel highly divergent viruses as well as known viral pathogens with established effects on population dynamics, including canine distemper virus, a novel bocavirus, and canine minute virus. Furthermore, we performed a 4‐year survey for the six wolf packs comprising this endangered wolf population, screening 93 fecal samples from 36 genetically identified wolves for canine distemper virus and the novel bocavirus, previously identified using our metagenomics approach. Our novel approach using metagenomics for viral screening in noninvasive samples of wolves and dogs has profound implications on the knowledge of both virology and wildlife diseases, establishing a complementary tool to traditional screening methods for the conservation of threatened species.     

Genome‐wide analysis of allele frequency change in sunflower crop–wild hybrid populations evolving under natural conditions

Crop‐wild hybridization occurs in numerous plant species and could alter the genetic structure and evolutionary dynamics of wild populations. Studying crop‐derived alleles in wild populations is also relevant to assessing/mitigating the risks associated with transgene escape. To date, crop‐wild hybridization has generally been examined via short‐term studies, typically within a single generation, focusing on few traits or genetic markers. Little is known about patterns of selection on crop‐derived alleles over multiple generations, particularly at a genome‐wide scale. Here, we documented patterns of natural selection in an experimental crop × wild sunflower population that was allowed to evolve under natural conditions for two generations at two locations. Allele frequencies at a genome‐wide collection of SNPs were tracked across generations, and a common garden experiment was conducted to compare trait means between generations. These data allowed us to identify instances of selection on crop‐derived alleles/traits and, in concert with QTL mapping results, test for congruence between our genotypic and phenotypic results. We found that natural selection overwhelmingly favours wild alleles and phenotypes. However, crop alleles in certain genomic regions can be favoured, and these changes often occurred in parallel across locations. We did not, however, consistently observe close agreement between our genotypic and phenotypic results. For example, when a trait evolved towards the wild phenotype, wild QTL alleles associated with that trait did not consistently increase in frequency. We discuss these results in the context of crop allele introgression into wild populations and implications for the management of GM crops.     

netview p: a network visualization tool to unravel complex population structure using genome‐wide SNPs

Network‐based approaches are emerging as valuable tools for the analysis of complex genetic structure in wild and captive populations. netview p combines data quality control with the construction of population networks through mutual k‐nearest neighbours thresholds applied to genome‐wide SNPs. The program is cross‐platform compatible, open‐source and efficiently operates on data ranging from hundreds to hundreds of thousands of SNPs. The pipeline was used for the analysis of pedigree data from simulated (n = 750, SNPs = 1279) and captive silver‐lipped pearl oysters (n = 415, SNPs = 1107), wild populations of the European hake from the Atlantic and Mediterranean (n = 834, SNPs = 380) and grey wolves from North America (n = 239, SNPs = 78 255). The population networks effectively visualize large‐ and fine‐scale genetic structure within and between populations, including family‐level structure and relationships. netview p comprises a network‐based addition to other population analysis tools and provides user‐friendly access to a complex network analysis pipeline through implementation in python .     

A genome‐wide search for local adaptation in a terrestrial‐breeding frog reveals vulnerability to climate change

Terrestrial‐breeding amphibians are likely to be vulnerable to warming and drying climates, as their embryos require consistent moisture for successful development. Adaptation to environmental change will depend on sufficient genetic variation existing within or between connected populations. Here, we use Single Nucleotide Polymorphism (SNP) data to investigate genome‐wide patterns in genetic diversity, gene flow and local adaptation in a terrestrial‐breeding frog (Pseudophryne guentheri) subject to a rapidly drying climate and recent habitat fragmentation. The species was sampled across 12 central and range‐edge populations (192 samples), and strong genetic structure was apparent, as were high inbreeding coefficients. Populations showed differences in genetic diversity, and one population lost significant genetic diversity in a decade. More than 500 SNP loci were putatively under directional selection, and 413 of these loci were correlated with environmental variables such as temperature, rainfall, evaporation and soil moisture. One locus showed homology to a gene involved in the activation of maturation in Xenopus oocytes, which may facilitate rapid development of embryos in drier climates. The low genetic diversity, strong population structuring and presence of local adaptation revealed in this study shows why management strategies such as targeted gene flow may be necessary to assist isolated populations to adapt to future climates.     

Real‐time assessment of hybridization between wolves and dogs: combining noninvasive samples with ancestry informative markers

Wolves and dogs provide a paradigmatic example of the ecological and conservation implications of hybridization events between wild and domesticated forms. However, our understanding of such implications has been traditionally hampered by both high genetic similarity and the difficulties in obtaining tissue samples (TS), which limit our ability to assess ongoing hybridization events. To assess the occurrence and extension of hybridization in a pack of wolf–dog hybrids in northwestern Iberia, we compared the power of 52 nuclear markers implemented on TS with a subset of 13 ancestry informative markers (AIMs) typed in noninvasive samples (NIS). We demonstrate that the 13 AIMs are as accurate as the 52 markers that were chosen without regard to the power to differentiate between wolves and dogs, also having the advantage of being rapidly screened on NIS. The efficiency of AIMs significantly outperformed ten random sets of similar size and an additional commercial set of 18 markers. Bayesian clustering analysis implemented on AIMs and NIS identified nine hybrids, two wolves and two dogs. Four hybrids were unambiguously assigned to F1xWolf backcrosses. Our approach (AIMs + NIS) overcomes previous difficulties related to sample availability and informative power of markers, allowing a quick identification of wolf–dog hybrids in the first phases of hybridization episodes. This provides managers with a reliable tool to evaluate hybridization and estimate the success of their actions. This approach may be easily adapted for other pairs of wild/domesticated species, thus improving our understanding of the introgression of domestication genes into natural populations.