Genomic Processes in Hybridization: Implications for Species Taxonomy

GENERAL INTRODUCTION

The red wolf was first described in 1851 as a subspecies of the gray wolf. In 1937, Edward A. Goldman elevated it to species status (Canis rufus) based on analyses of cranial and dental traits. However, the species classification of the red wolf remains controversial as genetic studies have shown that modern red wolves contain some amount of coyote ancestry. Numerous animal studies have shown that many recognized species, including humans, acquired genetic variation from other species through hybridization without undermining the distinctness of the species involved.

Conservation policy – including the decision to list a given group of animals as an endangered species – relies heavily on species designations. If the modern red wolf is not a valid species, then efforts on extant captive and managed populations devoted to recover the red wolf in the United States might be seen as misplaced.

The presence of coyote ancestry in the genome of modern red wolves may have resulted from any of three different evolutionary processes (Figure 1), or some combination thereof:

1. Incomplete Lineage Sorting: In this scenario, the red wolf diverged from coyotes recently enough that they continue to share alleles, even though each species has its own forward evolutionary trajectory.
2. Hybridization with Gene Introgression: In this scenario, red wolves and coyotes mate and yield hybrid offspring with mixed ancestry. If these hybrids mate with members of either or both species, a process called backcrossing, some alleles that originated in coyotes could be found in red wolves, and vice versa. This could have happened historically, be happening at present, or both. The significance of this genetic exchange to the identity of the red wolf as a species may vary depending on when and to what extent this exchange occurred.
3. Hybrid Speciation: In this scenario, red wolves were initially formed through hybridization between coyotes and another canid (e.g., the gray wolf), but are now a distinct evolutionary lineage.

In March 2019, the National Academies of Sciences, Engineering, and Medicine (NASEM) issued a report, Evaluating the Taxonomic Status of the Mexican Gray Wolf and the Red Wolf, which addressed the taxonomic status and evolutionary history of the red wolf. In October 2020, the NASEM released a follow-up report, A Research Strategy to Examine the Taxonomy of the Red Wolf . The reports were products of two separate studies that were conducted upon the request of the U.S. Fish and Wildlife Service (fws.gov).

This lesson uses the red wolf as an example to explore the genomic processes involved during hybridization, and to examine how to interpret these outcomes and their implications for the taxonomic status of hybridizing species. Most of the text used here has been taken with some modifications from the two reports described above.

HISTORICAL AND PRESENT DISTRIBUTION OF RED WOLVES AND COYOTES IN THE UNITED STATES

The earliest fossil of an animal identified as a red wolf, retrieved from a cave in Florida, was dated to 10,000 years ago. Discovery of other red wolf specimens indicated that they lived in the southeastern United States and lower Mississippi Valley, areas distinct from those inhabited by coyotes and gray wolves. However, later analyses of all available cranial specimens of wild canids from North America extended the red wolf’s inferred distribution throughout the East Coast of the United States, keeping it south of the Great Lakes but as far north as the Gaspe Peninsula in Quebec. The extended range placed a significant part of the red wolf’s distribution atop the expected distribution of gray wolves.

Before European settlement, red wolves and coyotes only overlapped on the western and eastern boundaries of their ranges, respectively. Accounts of these animals and modern observations indicate that red wolves inhabited forests while coyotes were found in more open mixed woods and fields. Around 1920, as red wolves and other large predators were being heavily hunted as part of predator eradication programs, and forested habitat in the southeastern United States was disappearing, coyotes expanded their range eastward, overlapping more and more with the range of the red wolf.

By 1980, red wolves in the United States were considered extinct in the wild, but a captive population, established since the 1970s from a small number of founder individuals, remained. In 1987, some individuals from the captive breeding program (at the time composed of 245 red wolves housed at 43 different breeding facilities across the United States) were used by U.S. Fish and Wildlife Service to re-introduce red wolves into a protected area in North Carolina. The re-introduced individuals established a population in the Alligator River National Wildlife Refuge in North Carolina and is managed by the U.S. Fish and Wildlife Service.

Recently, researchers have identified alleles in two canid populations (one in southwestern Louisiana and the other one in Galveston, Texas) that have not been observed in either the coyotes or the captive or managed populations of red wolves. Researchers concluded that these alleles could have been characteristic of the red wolf population before they were extirpated from the wild, and then lost during the selection of the founders of the captive population. Although ongoing research may soon shed some light on the plausibility of this hypothesis, the fact that hybridization has been, and continues to be, part of the history of the red wolf population points to a complex role of hybridization in the origin and maintenance of species and its implications for conservation policy.

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HOW DOES HYBRIDIZATION WORK AND WHY IT IS IMPORTANT TO UNDERSTAND IT WHEN DEFINING SPECIES?

Hybridization refers to the mating of two individuals from different species to yield offspring that have ancestry from both parents. In all eukaryotic organisms, the DNA in the nucleus of each cell is divided and organized into a number of segments called chromosomes. In wolves and other mammals, each cell contains two copies of each chromosome, one inherited from the mother and one from the father. These chromosomes are called homologous because they contain the same genes in the same locations, although the copies of a given gene inherited from the mother and from the father may not have identical sequences—that is, they may have different alleles. Thus, a hybrid individual will have one set of chromosomes from one species and the other set of homologous chromosomes from another.

During the production of eggs and sperm, the pair of homologous chromosomes, one from each parent, exchange segments through genetic recombination, producing new chromosome copies composed of combinations of the DNA from the original chromosomes. If hybrid offspring are fertile, they pass on to their own offspring individual chromosomes that contain DNA segments from both parental species. With each successive generation, chromosomal segments coming for each parental species are broken into smaller and smaller pieces through recombination, producing increasingly complex mosaics of DNA from the two parental species. Therefore, hybridization will produce specific patterns in the genomes of individuals of the species that interbreed.

Mating between individuals of two different species does not always result in sustained hybridization. In some cases, individuals of different species are unable to produce offspring together.  The offspring may not be viable, or they may survive but be infertile. In other cases, a first generation (F1) hybrid can produce fertile offspring, but when it mates with a member of the parental species or with other hybrids its offspring are infertile. Reduced hybrid viability and fertility would both impose a strict barrier to gene flow between the two species. It can also drive the evolution of pre-mating isolation, wherein individuals have more offspring if they choose a mate from their own species.

If hybrid individuals are able to produce fertile offspring, however, this can lead to the development of an admixed population, one that contains genetic contributions from more than one species. Individuals in this population can have genomes with varying proportions of genetic contributions from each parental species. These interspecific genetic combinations in an admixed population can lead to a number of long-term outcomes:

1. Survival of hybrid individuals and mating with one of the parental species can result in introgression, the movement of specific regions of the genome of one parental species into the homologous chromosomes of some individuals of the other species. If these chromosome regions contain differences in their sequence between species (i.e., different alleles), the presence of alleles coming from the other species may or may not affect the fitness of the individuals who possess them. These alleles will remain in the population if they provide advantage or at least are not detrimental to the individuals who carry them.
2. Chromosome regions that carry alleles that are extremely detrimental when combined with the genome of the opposite species (e.g., if the hybrids that have those alleles die or cannot reproduce) will not introgress and be removed from the population by natural selection.
3. Chromosomal regions with alleles that only moderately reduce fitness can linger for a long time or even become fixed, especially in small populations, where natural selection is weak as compared to genetic drift.

Therefore, hybridization can lead to the introgression of some parts of the genome, even while other parts remain distinct between species.

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In the specific case of the red wolf, genomic regions that do not introgress might be associated to specific adaptations and could diagnose the red wolf as a distinct species from coyotes or other canids, despite the presence of some genetic contributions from coyotes.