Isolation of 18 polymorphic microsatellite loci from the North American red squirrel,Tamiasciurus hudsonicus (Sciuridae,Rodentia), and their cross‐utility in other species

We isolated 18 polymorphic microsatellite loci to be used for pedigree analysis in a wild population of North American red squirrels, Tamiasciurus hudsonicus. Allelic diversity and observed heterozygosity ranged from six to 13 and 0.39 to 0.89, respectively, in a sample of 93 individuals. Up to 13 sets of primers also amplify in other rodent species.     

Subspecific identity and a comparison of genetic diversity between wild and ex situ wildebeest

The original North American ex situ wildebeest population was believed to originate from the white-bearded wildebeest (Connochaetes taurinus albojubatus), which is both morphologically distinct and geographically separated from the brindled wildebeest (C. t. taurinus). However, after an import of wildebeest into North America in 2001, managers have suspected that white-bearded and brindled wildebeest were mixed in herds at multiple institutions. We sequenced the mitochondrial control region (d-loop) from a portion of the managed North American population and compared our sequences with previously published sequences from wild individuals to determine the subspecific identity and genetic diversity of our ex situ population. We were able to confidently identify C. t. albojubatus as the subspecies identity of the sampled portion of our population. Within our population, haplotype and nucleotide diversity were low (0.169 and 0.001, respectively) with a single common haplotype (H1) containing 41 of the 45 individuals sequenced, while two rare haplotypes (H2 and H3) were derived from three individuals and a single individual, respectively. Nucleotide and haplotype diversity were greater overall in the wild populations compared with our managed population. However, C. t. albojubatus was found to exhibit lower nucleotide diversity in both wild and ex situ populations when compared to other wild subspecies. Though the overall goal of the North American wildebeest population is for public education and not reintroduction, maintaining genetic diversity is vital for the long-term viability of this managed population, which may benefit from periodic supplementation of wild animals.     

The cryptic genetic structure of the North American captive gorilla population

Western lowland gorillas (Gorilla gorilla gorilla) were imported from across their geographical range to North American zoos from the late 1800s through 1974. The majority of these gorillas were imported with little or no information regarding their original provenance and no information on their genetic relatedness. Here, we analyze 32 microsatellite loci in 144 individuals using a Bayesian clustering method to delineate clusters of individuals among a sample of founders of the captive North American zoo gorilla collection. We infer that the majority of North American zoo founders sampled are distributed into two distinct clusters, and that some individuals are of admixed ancestry. This new information regarding the existence of ancestral genetic population structure in the North American zoo population lays the groundwork for enhanced efforts to conserve the evolutionary units of the western lowland gorilla gene pool. Our data also show that the genetic diversity estimates in the founder population were comparable to those in wild gorilla populations (Mondika and Cross River), and that pairwise relatedness among the founders is no different from that expected for a random mating population. However, the relatively high level of relatedness (R = 0.54) we discovered in a pair of known breeding pairs reveals the need for incorporating genetic relatedness estimates in the captive management of western lowland gorillas.     

Genetic diversity of North American captive‐born gorillas (Gorilla gorilla gorilla)

Western lowland gorillas (Gorilla gorilla gorilla) are designated as critically endangered and wild populations are dramatically declining as a result of habitat destruction, fragmentation, diseases (e.g., Ebola) and the illegal bushmeat trade. As wild populations continue to decline, the genetic management of the North American captive western lowland gorilla population will be an important component of the long‐term conservation of the species. We genotyped 26 individuals from the North American captive gorilla collection at 11 autosomal microsatellite loci in order to compare levels of genetic diversity to wild populations, investigate genetic signatures of a population bottleneck and identify the genetic structure of the captive‐born population. Captive gorillas had significantly higher levels of allelic diversity (t₇ = 4.49, P = 0.002) and heterozygosity (t₇ = 4.15, P = 0.004) than comparative wild populations, yet the population has lost significant allelic diversity while in captivity when compared to founders (t₇ = 2.44, P = 0.04). Analyses suggested no genetic evidence for a population bottleneck of the captive population. Genetic structure results supported the management of North American captive gorillas as a single population. Our results highlight the utility of genetic management approaches for endangered nonhuman primate species.     

Assessing changes in genomic divergence following a century of human‐mediated secondary contact among wild and captive‐bred ducks

Along with manipulating habitat, the direct release of domesticated individuals into the wild is a practice used worldwide to augment wildlife populations. We test between possible outcomes of human‐mediated secondary contact using genomic techniques at both historical and contemporary timescales for two iconic duck species. First, we sequence several thousand ddRAD‐seq loci for contemporary mallards (Anas platyrhynchos) throughout North America and two domestic mallard types (i.e., known game‐farm mallards and feral Khaki Campbell's). We show that North American mallards may well be becoming a hybrid swarm due to interbreeding with domesticated game‐farm mallards released for hunting. Next, to attain a historical perspective, we applied a bait‐capture array targeting thousands of loci in century‐old (1842–1915) and contemporary (2009–2010) mallard and American black duck (Anas rubripes) specimens. We conclude that American black ducks and mallards have always been closely related, with a divergence time of ~600,000 years before present, and likely evolved through prolonged isolation followed by limited bouts of gene flow (i.e., secondary contact). They continue to maintain genetic separation, a finding that overturns decades of prior research and speculation suggesting the genetic extinction of the American black duck due to contemporary interbreeding with mallards. Thus, despite having high rates of hybridization, actual gene flow is limited between mallards and American black ducks. Conversely, our historical and contemporary data confirm that the intensive stocking of game‐farm mallards during the last ~100 years has fundamentally changed the genetic integrity of North America's wild mallard population, especially in the east. It thus becomes of great interest to ask whether the iconic North American mallard is declining in the wild due to introgression of maladaptive traits from domesticated forms. Moreover, we hypothesize that differential gene flow from domestic game‐farm mallards into the wild mallard population may explain the overall temporal increase in differentiation between wild black ducks and mallards, as well as the uncoupling of genetic diversity and effective population size estimates across time in our results. Finally, our findings highlight how genomic methods can recover complex population histories by capturing DNA preserved in traditional museum specimens.     

Breeding program for the lesser kudu (Tragelaphus imberbis) in North America

The lesser kudu (Tragelaphus imberbis) has been kept in North American zoological parks since 1930 but has never been a common species in collections. In 1987 this population totaled 28 animals: 15 males and 13 females. A pedigree evaluation in 1987 of the existing population indicated that eight effective founders and one potential founder were represented in the North American herd. Three new potential founders from European captive populations were added to the population in 1987 to increase the number of existing founder lines to 12 animals. As this species is not endangered or threatened in its native habitat, it is not a high priority to qualify for designation as an SSP species. Because of this, the institutions holding lesser kudu in North America decided to join informally and draft a breeding program to better manage this small captive population. This program was designed to minimize inbreeding and equalize genetic representation of founder animals to maximize genetic diversity. It requires a shift in management philosophy to establish stable groups of breeding females at participating institutions while rotating appropriate breeder males through these herds in a controlled manner to ensure minimization of inbreeding and maximization of genetic diversity. It is hoped that this program can serve as a model for the management of other small captive populations of non-SSP species.     

Rapid expansion of the distributional range and the population genetic structure of the freshwater amphipod <Emphasis Type="Italic">Crangonyx floridanus</Emphasis> in Japan

The freshwater amphipod Crangonyx floridanus (Amphipoda: Crangonyctidae) is considered to have been recently introduced from North America to Japan, and the recorded sites at which it has been collected now cover nearly all of Japan except for the northern part. In this study, we surveyed further areas outside of its known distribution range, and examined the population genetic structure and the phylogenetic relationships between Japanese and North American populations of this species based on nuclear (18S rRNA) and mitochondrial (COI) DNA sequences. We found that this amphipod has already reached Hokkaido, northernmost Japan, which suggests that it has undergone rapid expansion in a pattern of concentric circles from the central part of Japan. Genetic analysis showed that the Japanese population is genetically homogeneous, in contrast to the genetic diversification of this species seen in North American Crangonyx populations. The process of introducing, establishing, and expanding this amphipod in Japan may be explained as follows. A limited number of individuals from a North American native population were probably inadvertently introduced and established somewhere within the Kanto region. The local population size then increased and its distribution range expanded rapidly across Japan.     

Evidence for multiple introductions of Centaurea stoebe micranthos (spotted knapweed,Asteraceae) to North America

Invasive species’ success may depend strongly on the genetic resources they maintain through the invasion process. We ask how many introductions have occurred in the North American weed Centaurea stoebe micranthos (Asteraceae), and explore whether genetic diversity and population structure have changed as a result of introduction. We surveyed individuals from 15 European native range sites and 11 North American introduced range sites at six polymorphic microsatellite loci. No significant difference existed in the total number of alleles or in the number of private alleles found in each range. Shannon–Weaver diversity of phenotype frequencies was also not significantly different between the ranges, while expected heterozygosity was significantly higher in the invasive range. Population structure was similar between the native range and the invasive range, and isolation by distance was not significant in either range. Traditional assignment methods did not allocate any North American individuals to the sampled European populations, while Bayesian assignment methods grouped individuals into nine genetic clusters, with three of them shared between North America and Europe. Invasive individuals tended to have genetically admixed profiles, while natives tended to assign more strongly to a single cluster. Many North American individuals share assignment with Romania and Bulgaria, suggesting two separate invasions that have undergone gene flow in North America. Samples from three other invasive range sites were genetically distinct, possibly representing three other unique introductions. Multiple introductions and the maintenance of high genetic diversity through the introduction process may be partially responsible for the invasive success of C. stoebe micranthos.     

Evaluation of the genetic management of the endangered black‐footed ferret (Mustela nigripes)

Empirical support for the genetic management strategies employed by captive breeding and reintroduction programs is scarce. We evaluated the genetic management plan for the highly endangered black‐footed ferret (Mustela nigripes) developed by the American Zoo and Aquarium Associations (AZA) as a part of the species survival plan (SSP). We contrasted data collected from five microsatellite loci to predictions from a pedigree‐based kinship matrix analysis of the captive black‐footed ferret population. We compared genetic diversity among captive populations managed for continued captive breeding or reintroduction, and among wild‐born individuals from two reintroduced populations. Microsatellite data gave an accurate but only moderately precise estimate of heterozygosity. Genetic diversity was similar in captive populations maintained for breeding and release, and it appears that the recovery program will achieve its goal of maintaining 80% of the genetic diversity of the founder population over 25 years. Wild‐born individuals from reintroduced populations maintained genetic diversity and avoided close inbreeding. We detected small but measurable genetic differentiation between the reintroduced populations. The model of random mating predicted only slightly lower levels of heterozygosity retention compared to the SSP strategy. The random mating strategy may be a viable alternative for managing large, stable, captive populations such as that of the black‐footed ferret. Zoo Biol 22:287–298, 2003. © 2003 Wiley‐Liss, Inc.     

Genetic diversity in the US ex situ populations of the endangered Puerto Rican Boa,Chilabothrus inornatus

The Puerto Rican Boa (Chilabothrus inornatus) was placed on the US Endangered Species List in 1970. Progress has been made since to clarify the recovery status of this species, though the design of a new recovery plan must include information regarding genetic variation within and among populations of this species. While measures of genetic diversity in wild populations of this species are finally becoming available, relative genetic diversity represented in ex situ populations is unknown, which hampers efforts to develop an ex situ species management plan. Here, we provide an analysis of genetic diversity in US public and private collections (zoos and breeders) using mitochondrial sequence data and five highly polymorphic nuclear microsatellite loci. We analyzed 50 boas from the US ex situ population and determined overall genetic diversity and relatedness among these individuals. We then compared these data to mitochondrial and microsatellite data obtained from 176 individuals from wild populations across the native range of the species. We found little inbreeding and a large amount of retained genetic diversity in the US ex situ population of C. inornatus relative to wild populations. Genetic diversity in the ex situ population is similar to that found in wild populations. Ours is only the second explicit attempt to characterize genetic diversity at the molecular level in ex situ populations of boid snakes. We anticipate that these results will inform current breeding strategies as well as offer additional information that will facilitate the continuation of ex situ conservation breeding or management in boas.     

Clonal diversity and genetic differentiation revealed by SSR markers in wild Vaccinium macrocarpon and Vaccinium oxycoccos

American cranberry (Vaccinium macrocarpon) is a perennial, woody plant species, native to North American bogs and wetlands. Cranberries represent one of the few agriculturally important native plants in which wild gene pools are still readily available within the undeveloped wetlands of the northern US and Canada. Earlier studies have reported low genetic variation in V. macrocarpon at the species and population level. However, in this study, we characterised 229 individuals of wild V. macrocarpon and V. oxycoccos (small cranberry) from Wisconsin and 22 accessions using microsatellite markers and observed substantial genetic variation and differentiation within and among populations and species. While V. macrocarpon was analysed using 108 alleles from 11 microsatellite loci revealing 42 unique genotypes, V. oxycoccos was analysed using 156 alleles from eight loci revealing 28 unique genotypes. There were a total of 182 alleles found in both species combined with 156 of those alleles present in V. oxycoccos and 84 alleles found in V. macrocarpon. All eight loci possessed species‐specific alleles with V. oxycoccos possessing 98 private alleles versus 26 private alleles found V. macrocarpon, and 58 alleles were found in common between both species. Our data will be valuable not only for future wild cranberry diversity and population genetics research, but for other cranberry breeding and genetics studies.     

Invasion history of North American Canada thistle,Cirsium arvense

Aim Canada thistle (Cirsium arvense– Cardueae, Asteraceae) is one of the worst invasive plants world‐wide. Native to Eurasia, its unintentional introduction into North America now threatens the native flora and is responsible for enormous agricultural losses. The goals of this study are to: (1) reconstruct the evolutionary history of C. arvense and estimate how often it may have colonized North America, (2) compare the genetic diversity between European and North American populations to detect signs of demographic bottlenecks and/or patterns of population admixture, and (3) conduct bioclimatic comparisons to infer eventual niche shifts following this species’ introduction into North America. Location Europe and North America. Methods A total of 1522 individuals from 58 populations were investigated with six microsatellite markers. Estimates of heterozygosity (H_E) and allelic richness (R_S) were quantified for each population, and population structure was inferred via analyses of molecular variance (AMOVAs), principal components analyses (PCAs), Mantel tests and Bayesian clustering analyses. Climatic niche spaces were based on 19 bioclimatic variables extracted from approximately 32,000 locations covering the entire range, and compared using PCA and hierarchical cluster analysis. Results Although there is evidence of multiple introductions from divergent European lineages, North American populations of C. arvense exhibited significantly lower levels of genetic diversity than their putative ancestors. Bioclimatic comparisons pointed to a high degree of niche conservatism during invasion, but indicated that genotypes from the former USSR and Central European mountain chains were probably best adapted to invade North America upon entry into the continent. Main conclusions Genetic and historical data suggest that C. arvense first entered North America from Western Europe with the first European settlers, and was later introduced from Eastern Europe into the prairie states during the agricultural boom. The species went through a significant bottleneck following its introduction into the New World, but the level of genetic diversity remained high owing to admixture between genetically differentiated lineages and to a highly efficient outcrossing breeding system.     

One-step reconstruction of multi-generation pedigree networks in apple (Malus × domestica Borkh.) and the parentage of Golden Delicious

The increasing availability of genomic tools improves our ability to investigate the patterns of genetic diversity and relatedness among individuals. The pedigrees of many apple cultivars are completely unknown, often reducing the efficiency of breeding programs. Using a multilocus simple sequence repeat dataset, we applied a novel multi-generation pedigree-network reconstruction procedure based on the software FRANz in a Malus × domestica collection (101 cultivated and 22 wild apples) with partially known pedigree relationships. The procedure produced 78 parent–offspring relationships organized into three networks and showed high power for detecting real pedigree links (98.5 %) and a low false-positive rate (9.0 %). The largest reconstructed pedigree network spanned four generations and involved 65 cultivars. The availability of detailed pedigree connections confirmed that recent genealogical relationships affect population genetic structure in apple. Finally, our analysis enabled us to confirm or discard several pedigrees known only anecdotically, among which the cultivar Grimes Golden was validated as a parent of the widely grown cultivar Golden Delicious. The pedigree reconstruction protocol here described will be of broad applicability to other collections and crop species.     

Genetic assessment of a conservation breeding program of the houbara bustard (Chlamydotis undulata undulata) in Morocco,based on pedigree and molecular analyses

Protection and restoration of species in the wild may require conservation breeding programs under genetic management to minimize deleterious effects of genetic changes that occur in captivity, while preserving populations' genetic diversity and evolutionary resilience. Here, through interannual pedigree analyses, we first assessed the efficiency of a 21-year genetic management, including minimization of mean kinship, inbreeding avoidance, and regular addition of founders, of a conservation breeding program targeting on Houbara bustard (Chlamydotis undulata undulata) in Morocco. Secondly, we compared pedigree analyses, the classical way of assessing and managing genetic diversity in captivity, to molecular analyses based on seven microsatellites. Pedigree-based results indicated an efficient maintenance of the genetic diversity (99% of the initial genetic diversity retained) while molecular-based results indicated an increase in allelic richness and an increase in unbiased expected heterozygosity across time. The pedigree-based average inbreeding coefficient F remained low (between 0.0004 and 0.003 in 2017) while the proportion of highly inbred individuals (F > .1) decreased over time and reached 0.2% in 2017. Furthermore, pedigree-based F and molecular-based individual multilocus heterozygosity were weakly negatively correlated, (Pearson's r = −.061 when considering all genotyped individuals), suggesting that they cannot be considered as alternatives, but rather as complementary sources of information. These findings suggest that a strict genetic monitoring and management, based on both pedigree and molecular tools can help mitigate genetic changes and allow to preserve genetic diversity and evolutionary resilience in conservation breeding programs.     

Weedy adaptation in Setaria spp. II. Genetic diversity and population genetic structure in S. glauca,S. geniculata,and S. faberii (Poaceae)

Setaria glauca (yellow foxtail), S. geniculata (knotroot foxtail), and S. faberii (giant foxtail) are important cosmopolitan weeds of temperate and tropical regions. Isozyme markers were used to investigate genetic diversity and population genetic structure in 94 accessions of yellow foxtail, 24 accessions of knotroot foxtail, and 51 accessions of giant foxtail, collected mainly from North America and Eurasia. Giant foxtail populations were nearly identical genetically, with only one population exhibiting isozyme polymorphism. Yellow and knotroot foxtail populations had low genetic diversity but marked population differentiation. Although the latter species are similar morphologically, they are readily distinguished electrophoretically, with Nei's genetic identity being 0.83. In both species, genetic divergence between accessions from Eurasia and North America was minimal. Populations from the native ranges had slightly greater genetic diversity than those from the respective introduced ranges. Yellow foxtail populations genetically clustered into Asian, European, and North American groups. Within North America, yellow foxtail populations from Iowa were genetically diverse whereas populations collected from other North American locations were nearly monomorphic for the same multilocus genotype. Knotroot foxtail populations in North America were genetically differentiated into northern and southern groups on either side of a line at ≈37° N latitude. No genetic patterning was evident in knotroot foxtail populations from Eurasia. In both yellow and knotroot foxtail, patterns of population genetic structure have been influenced by several factors, including genetic bottlenecks associated with founder events, genetic drift, and natural selection.     

Genetic structure,admixture and invasion success in a Holarctic defoliator,the gypsy moth (Lymantria dispar,Lepidoptera: Erebidae)

Characterizing the current population structure of potentially invasive species provides a critical context for identifying source populations and for understanding why invasions are successful. Non‐native populations inevitably lose genetic diversity during initial colonization events, but subsequent admixture among independently introduced lineages may increase both genetic variation and adaptive potential. Here we characterize the population structure of the gypsy moth (Lymantria dispar Linnaeus), one of the world's most destructive forest pests. Native to Eurasia and recently introduced to North America, the current distribution of gypsy moth includes forests throughout the temperate region of the northern hemisphere. Analyses of microsatellite loci and mitochondrial DNA sequences for 1738 individuals identified four genetic clusters within L. dispar. Three of these clusters correspond to the three named subspecies; North American populations represent a distinct fourth cluster, presumably a consequence of the population bottleneck and allele frequency change that accompanied introduction. We find no evidence that admixture has been an important catalyst of the successful invasion and range expansion in North America. However, we do find evidence of ongoing hybridization between subspecies and increased genetic variation in gypsy moth populations from Eastern Asia, populations that now pose a threat of further human‐mediated introductions. Finally, we show that current patterns of variation can be explained in terms of climate and habitat changes during the Pleistocene, a time when temperate forests expanded and contracted. Deeply diverged matrilines in Europe imply that gypsy moths have been there for a long time and are not recent arrivals from Asia.     

Examining the genetic diversity of the orange blossom wheat midge,Sitodiplosis mosellana (Géhin), across North America

The orange blossom wheat midge, Sitodiplosis mosellana (Géhin) (Diptera: Cecidomyiidae), is a significant pest of wheat (Triticum spp.) grown in the Northern Hemisphere. It was accidently introduced to North America over 200 years ago and has subsequently spread throughout the northern Great Plains. Since 2010, several Canadian spring wheat varieties containing the resistance gene Sm1 have been released. Due to the potential of wheat midge populations to evolve virulent biotypes to Sm1, cultivars containing Sm1 are grown with a susceptible cultivar in an interspersed refuge. An understanding of the genetic diversity of wheat midge populations could provide important information on the potential development of resistance to Sm1. In the current study, we used two mitochondrial genes (CO1 and ND4) from wheat midge collected across the northern Great Plains and Québec in North America to assess population structure and genetic diversity. We found limited genetic diversity and population structure across the sampled North American populations. We also assessed North American haplotype similarity to wheat midge collected from Europe and China and found high similarity between North American and European populations, although sampling in Europe was limited. This supports the hypothesis that North American populations originated from Europe.     

Genetic diversity and population structure in cultivated sunflower and a comparison to its wild progenitor, <Emphasis Type="Italic">Helianthus annuus</Emphasis> L

Crop germplasm collections are valuable resources for ongoing plant breeding efforts. To fully utilize such collections, however, researchers need detailed information about the amount and distribution of genetic diversity present within collections. Here, we report the results of a population genetic analysis of the primary gene pool of sunflower (Helianthus annuus L.) based on a broad sampling of 433 cultivated accessions from North America and Europe, as well as a range-wide collection of 24 wild sunflower populations. Gene diversity across the cultivars was 0.47, as compared with 0.70 in the wilds, indicating that cultivated sunflower harbors roughly two-thirds of the total genetic diversity present in wild sunflower. Population structure analyses revealed that wild sunflower can be subdivided into four genetically distinct population clusters throughout its North American range, whereas the cultivated sunflower gene pool could be split into two main clusters separating restorer lines from the balance of the gene pool. Use of a maximum likelihood method to estimate the contribution of the wild gene pool to the cultivated sunflower germplasm revealed that the bulk of the cultivar diversity is derived from two wild sunflower population genetic clusters that are primarily composed of individuals from the east-central United States, the same general region in which sunflower domestication is believed to have occurred. We also identified a nested subset of accessions that capture as much of the allelic diversity present within the sampled cultivated sunflower germplasm collection as possible. At the high end, a core set of 288 captured nearly 90% of the alleles present in the full set of 433, whereas a core set of just 12 accessions was sufficient to capture nearly 50% of the total allelic diversity present within this sample of cultivated sunflower.     

Evaluating the ability of Bayesian clustering methods to detect hybridization and introgression using an empirical red wolf data set

Bayesian clustering methods have emerged as a popular tool for assessing hybridization using genetic markers. Simulation studies have shown these methods perform well under certain conditions; however, these methods have not been evaluated using empirical data sets with individuals of known ancestry. We evaluated the performance of two clustering programs, baps and structure , with genetic data from a reintroduced red wolf (Canis rufus) population in North Carolina, USA. Red wolves hybridize with coyotes (C. latrans), and a single hybridization event resulted in introgression of coyote genes into the red wolf population. A detailed pedigree has been reconstructed for the wild red wolf population that includes individuals of 50–100% red wolf ancestry, providing an ideal case study for evaluating the ability of these methods to estimate admixture. Using 17 microsatellite loci, we tested the programs using different training set compositions and varying numbers of loci. structure was more likely than baps to detect an admixed genotype and correctly estimate an individual's true ancestry composition. However, structure was more likely to misclassify a pure individual as a hybrid. Both programs were outperformed by a maximum‐likelihood‐based test designed specifically for this system, which never misclassified a hybrid (50–75% red wolf) as a red wolf or vice versa. Training set composition and the number of loci both had an impact on accuracy but their relative importance varied depending on the program. Our findings demonstrate the importance of evaluating methods used for detecting admixture in the context of endangered species management.     

Mallard–Black Duck Hybridization and Population Genetic Structure in North Carolina

North Carolina, USA, represents the southern extent of the American black duck's (Anas rubripes) breeding range. Mallards (A. platyrhynchos) are present on the breeding grounds of the American black duck and hybridization is observed between these species; therefore, we assessed the genetic integrity, hybridization rates, and population structure of this local breeding population. We extracted genomic and mitochondrial DNA from chorioallantoic membranes and contour feathers from monitored black duck nests. We then prepared the extracted DNA for analysis using high-throughput DNA sequencing methods (ddRAD-seq). First, we assessed nuclear and mitochondrial population structure, genetic diversity, and differentiation across samples from North Carolina, and compared them against 199 genetically vetted mallards, black ducks, and mallard × black duck hybrids that served as genetic references. Next, we tested for parentage and sibling relationship and overall relatedness of black ducks in North Carolina. We recovered strong population structure and high co-ancestry across genetic markers due to interrelatedness among sampled nests in North Carolina and concluded that black ducks have been locally breeding in this area for a prolonged period of time. Despite a high level of interrelatedness among our samples, nucleotide diversity was similar to the reference continental black duck population, suggesting little effect of genetic drift, including inbreeding. Additionally, we conclude that molecular diversity of black ducks in North Carolina is maintained at reference population levels through the influx of genetic material from unrelated, migrating male black ducks. Finally, we report a hybridization level of 47.5%, covering 3 filial generations. Of identified hybrids, 54.7% and 53% were the direct result of interbreeding between black ducks and captive-reared or wild mallards, respectively. We conclude that because of high rates of interspecific hybridization and successive backcrossing events, introgression from wild and feral mallards is occurring into this population of breeding black ducks and requires careful consideration in future management efforts. © 2021 The Wildlife Society.