Southeastern Naturalist 283 B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 22001177 SOUTHEASTERN NATURALIST 1V6o(2l.) :1268,3 N–2o9. 62 Stable Isotope Analysis of Dietary Overlap between the Endangered Red Wolf and Sympatric Coyote in Northeastern North Carolina Brian S. Arbogast1,*, Anne-Marie C. Hodge1,2, and Joan Brenner-Coltrain3 Abstract - The only remaining wild Canis rufus (Red Wolf) are part of an experimental population inhabiting the Albemarle peninsula of northeastern North Carolina. This population was established in the late 1980s as part of the US Fish and Wildlife Service’s Red Wolf Recovery Program. Recently, controversy has arisen over whether to maintain, expand, or end the recovery program. This controversy is complex, but one source of concern about the program is the perception among some local stakeholders that, compared to the smaller, sympatric C. latrans (Coyote), Red Wolves put greater pressure on game species, such as Odocoileus viginianus (White-tailed Deer). However, previous research comparing fecal remains indicated a broad dietary overlap between sympatric populations of the 2 species. In this study, we investigated the question of dietary overlap between Red Wolves and Coyotes using stable isotope analysis. Our results are consistent with those based on fecal analyses in showing that sympatric populations of Red Wolves and Coyotes have similar diets. This finding has important conservation and management implications for Red Wolves because it suggests that: (1) this species does not prey upon game species, such as White-tailed Deer, to any greater degree than sympatric Coyotes; and (2) whereas the loss of the only wild population of Red Wolves would result in a reduction of phylogenetic diversity in northeastern North Carolina, it may not result in a loss of functional diversity if Coyotes or Coyote–Red Wolf hybrids are able to play a similar ecological role to that of Red Wolves. Introduction Canis rufus Audubon & Bachman (Red Wolf; see Paradiso and Nowak 1972) is one of the most critically endangered canids in the world, with an estimated wild population of ~45–60 individuals as of January 2017, down from an estimated 100 individuals in 2014 (US Fish and Wildlife [USFWS] Red Wolf Recovery Program 2014, 2017). Once widespread across the eastern and south-central US, the species was nearly exterminated during the 20th century, which prompted the USFWS to develop a captive-breeding program in the 1970s with a goal of eventually reestablishing Red Wolf populations in the wild (see Chambers et al 2012 for a review of the history of the Red Wolf Recovery Program). The first reintroduction of Red Wolves into northeastern North Carolina occurred in 1987. It consisted of 4 male– female pairs released into the Alligator River National Wildlife Refuge (ARNWR). 1Department of Biology and Marine Biology, University of North Carolina-Wilmington, Wilmington, NC 28403. 2Current address - Department of Zoology and Physiology, University of Wyoming, 1000 East University Avenue, Laramie, WY. 82070. 3Anthropology Department, University of Utah, Salt Lake City, UT. 84112. *Corresponding author - arbogastb@uncw.edu. Manuscript Editor: Michael V. Cove Southeastern Naturalist B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 284 Active reintrodutions and management of what was deemed an “experimental Red Wolf population” continued into 2014, and today, all known wild individuals of this species occur within a 5-county area that encompasses ~690,000 ha of the Albemarle Peninsula of northeastern North Carolina (hereafter, referred to as the Red Wolf Experimental Population Area). Although the taxonomic status of the Red Wolf has been controversial (Brzeski et al. 2014, Chambers et al 2012, Rutledge et al. 2010, von Holdt et al. 2011), the USFWS recognizes the Red Wolf as a distinct species (USFWS Red Wolf Recovery Program 2017). Under contract by the USFWS, the Wildlife Management Institute, Inc. (WMI; 2014), produced a review of the Red Wolf Recovery Program in late 2014. In June of the following year, USFWS (2015) announced it was suspending reintroductions of the Red Wolf into the wild while it gathered information and evaluated research into the feasibility of recovery for the species under the Endangered Species Act. Several concerns were cited as reasons for taking this action, including hybridization of the Red Wolf with C. latrans Say (Coyote; see Beckoff 1977). Historically, the geographic ranges of Red Wolves and Coyotes did not overlap in North Carolina, but over the last several decades the Coyote has expanded its range rapidly eastward and into the Red Wolf Experimental Population Area. This new sympatry between the 2 species in eastern North Carolina has led to documented hybridization between Red Wolves and Coyotes in the area (USFWS Red Wolf Recovery Program 2017), a clear threat to the persistence of the former in the wild. In an effort to curtail such hybridization, the Red Wolf Recovery Program had previously instituted a “place-holder” management approach, which consisted of capturing, sterilizing, and re-releasing Coyotes to serve as place-holders that would reduce the influx of new, fertile Coyotes into the area. In their review, WMI (2014) endorsed the validity of this approach but suggested that the effectiveness of the strategy had not been rigorously evaluated. The review identified issues regarding the practicality, expense, and unclear time-frame of the place-holder strategy as an effective long-term approach for maintaining a wild population of Red Wolves in northeastern North Carolina. WMI (2014) staff also administered an online survey to measure public perceptions about the reintroduction of the Red Wolf into northeastern North Carolina. Among the highest-ranking concerns of respondents that lived within the recovery zone was a decrease in the deer population. WMI (2014) also noted that “at the public meetings, we heard numerous statements of concern about the impact of Red Wolves and Coyotes on the deer population in the restoration area.” However, no rigorous scientific data are available to assess whether there has been a decrease in Odocoileus virginianus Zimmermann (White-tailed Deer; see Smith 1992) populations in the Red Wolf Experimental Population Area since reintroductions of Red Wolves began in 1987. What is clear is that the cooccurrence of Red Wolves and Coyotes in this area is a relatively recent phenomenon, and that it is worth studying how these 2 species of canids might be affecting populations of game species, such as White-tailed Deer. To date, the only detailed dietary analysis of sympatric Red Wolves and Coyotes in the Red Wolf Experimental Population Area is that of McVey et al. (2013). Those Southeastern Naturalist 285 B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 authors used fecal DNA analysis to identify whether Red Wolves or Coyotes deposited a given scat sample, and then identified prey species by comparing hair, bone, tooth, claw, and hoof fragments found in a scat to those in reference collections and identification manuals. McVey et al. (2013) concluded that there was no significant difference between the diets of sympatric Red Wolves and Coyotes. White-tailed Deer, rabbits, and small rodents were the most common prey items consumed by both species, and although not statistically significant, McVey et al. (2013) found White-tailed Deer remains in a slightly higher proportion of scats from Coyotes than in scats from Red Wolves. Overall, the authors concluded that the 2 species appeared to be affecting prey populations similarly within the Red Wolf Experimental Population Area. To better understand the ecology of sympatric Red Wolves and Coyotes and to independently evaluate McVey et al.’s (2013) findings based on scat analysis, we used a different method—stable-isotope mass spectrometry—to compare the diets of Red Wolves and Coyotes. This method can identify differences in the stableisotope chemistry of analyzed taxa, which in turn reflects the degree to which their diets differ in composition (Post et al. 2002). Stable-isotope ecology is a rapidly growing field that has opened many doors to increasing understanding of trophic structure, migration patterns, nutrient flow, dietary shifts, and even climate change within an ecosystem (Bershaw et al. 2010, Crawford et al. 2008, Fry 2006, Kelly 2000). Furthermore, stable-isotope analysis has been shown to be an extremely efficient method in terms of sampling. For example, Fox-Dobbs et al. (2007) showed that for Canis lupus L. (Gray Wolf), stable-isotope analysis of just 4–6 individuals provided mean stable-isotope values that were very close to that of the mean of the entire population. In this respect, if the goal is to compare overall dietary similarity and relative trophic level among species, stable isotope analysis can provide useful results with much lesss sampling effort than many other methods of dietary analysis. The most commonly used isotopic signatures used in ecological studies come from nitrogen (N) and carbon (C). Isotopic analysis of nitrogen enrichment provides insights into an organism’s trophic level, because enriched δ15N values (15N /14N) correlate positively with the proportion of tissues consumed from animals occupying higher trophic levels (e.g., primary and secondary consumers; Robbins et al. 2005). Trophic-level enrichment is typically 3–5‰ relative to prey taxa (Peterson and Fry 1987). Therefore, comparing the isotopic signatures of sympatric predator species can aid in elucidating the structure of trophic hierarchies (McCutchan et al. 2003, Roemer et al. 2002). In addition to determining relative trophic rank, stable isotope analysis makes it possible to determine whether specific prey are selected or utilized by a given species to a higher degree than other consumers in the same community, although this is predicated upon obtaining nitrogen-enrichment values for as many prey species as possible (Bluthgen et al. 2003, Hilderbrand et al. 1996, Post 2002, Stewart et al. 2003). In contrast to δ15N isotope values, δ13C carbon isotope values (13C/12C) are enriched ~1% as consumers increase in ranking within a trophic hierarchy. More importantly, the latter can be used to distinguish between tree- and shrub-based Southeastern Naturalist B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 286 (C3) and grass-based (C4) terrestrial food webs and to evaluate consumer reliance on marine versus terrestrial resources (Michener and Lajtha 2007). Thus, differences in this metric are often indicative of differences in foraging grounds and other geographic movement patterns (Hobson 1999). Comparisons of carbon enrichment between species reveal differences in the composition of plants forming the foundation of a given food web (Kelly 2000), taking advantage of different photosynthetic pathways between C3 and C4 plants (Ben-David and Flaherty 2012, Cerling et al. 1999). For example, this technique has been used to identify differences in the composition of herbivore species in predators’ diets in systems where grazers are primarily reliant on C4 savannah grasses and browsers forage on C3 forbs and shrubs (Codron et al. 2007). A variety of animal tissues—bone, blood, hair, claws/nails, or muscle—may be used to obtain isotopic signatures that reflect dietary composition. Consumed proteins fuel the synthesis of amino acids, and thus, connective tissues such as collagen are an excellent source for quantifying isotopic signatures. Despite the relatively short-term record of diet in dentine (tooth collagen), dentine isotope signals can be compared to those of scats, fur, or blood, which record even shorter-term dietary data (e.g., McVey et al. 2013). However, unlike bone collagen, tooth dentine does not turn over after formation (Hillson 1996); thus, dentine records dietary intake over the juvenile period during which it forms. Canids often provision their pups through regurgitation with prey caught by adults (Mech et al. 1999), pups’ diets after weaning are likely to reflect prey being consumed by adults in their social group. It has been shown that the permanent canines of Coyotes erupt at 4–5 months of age (Linhart and Knowlton 1967), well after the pups have been weaned (which occurs at 5–7 weeks of age; Bekoff 1977). Given that the rate of pup development between wild Canis species in North America is similar (Bekoff and Jameson 1975), it is appropriate to assume that Red Wolf and Coyote individuals do not differ significantly in the ages at which they reach similar stages of dental development. Thus, if samples are collected from individuals in the same geographic area, stable isotope analysis of tooth dentine in these species should produce values that are directly comparable. Comparison at this stage of development is appropriate because the purpose of our study was to compare the degree of dietary overlap between sympatric Red Wolf and Coyote populations and not to determine the specific dietary composition of each. Future studies that seek to use stable isotopes to detail dietary composition will require analysis of both tooth and bone collagen from Red Wolves and Coyotes as well as stable isotope analyses on a wide variety of potential prey species; those tasks were beyond the scope of the present study. In their recent study, McVey et al. (2013) found that, based on physical remains in feces, sympatric Coyotes and Red Wolves do not appear to have significantly different dietary compositions (at least over the time scale considered in their study). However, this question has not yet been approached using stable isotope analysis. In many ways, the stable isotope approach we employ is complementary to the fecal analysis approach used by McVey et al. (2013). In the present study, we thus sought Southeastern Naturalist 287 B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 to answer 2 main questions relevant to the effects of Red Wolf reintroduction in northeastern North Carolina: (1) Does the stable isotope approach indicate that sympatric Coyotes and Red Wolves use similar, or significantly different, dietary resources?; and (2) Are there significant differences in diet between males and females across the 2 species? Methods Red Wolf Recovery Program provided tooth samples (1 adult canine tooth) from 31 deceased individuals of Canis spp. (originally collected from the Red Wolf Experimental Population Area) (Table 1). The sampled individuals included 15 Red Wolves (6 female, 9 male) and 16 Coyotes (8 female, 8 male). Based on information in the Red Wolf Recovery Program pedigree database, all of the Red Wolf individuals were known to be adults at their time of death (variation in ages Table 1. US Fish and Wildlife identification numbers (USFW ID), species, sex, C:N ratios and 15N and 13C values for specimens of C. latrans (Coyote) and C. rufus (Red Wolf) examined in this study. USFW ID Species Sex δ15N ‰ δ13C‰ 20712 C. latrans M 10.4 -20.1 20724 C. latrans M 11.0 -16.3 20741 C. latrans M 11.5 -18.2 20746 C. latrans M 13.6 -19.3 20777 C. latrans M 11.6 -19.4 20673 C. latrans M 11.4 -18.5 20723 C. latrans M 11.4 -15.5 20779 C. latrans M 9.1 -18.7 20713 C. latrans F 11.5 -20.0 20719 C. latrans F 15.1 -16.6 20730 C. latrans F 11.7 -15.3 20731 C. latrans F 9.9 -20.3 20744 C. latrans F 10.4 -21.1 20757 C. latrans F 10.6 -19.5 20608 C. latrans F 11.6 -16.9 20636 C. latrans F 9.2 -20.6 11629 C. rufus M 11.2 -19.9 SB875 C. rufus M 13.5 -16.4 SB159 C. rufus M 9.0 -13.9 SB677 C. rufus M 7.7 -14.8 SB373 C. rufus M 11.5 -16.5 SB501 C. rufus M 11.1 -17.8 635M C. rufus M 10.7 -17.4 10933M C. rufus M 11.9 -15.6 883M C. rufus M 11.4 -18.2 11846 C. rufus F 10.7 -17.8 10878F C. rufus F 9.1 -17.9 10985F C. rufus F 10.6 -16.2 SB315 C. rufus F 8.9 -14.2 10797F C. rufus F 9.2 -19.1 10774F C. rufus F 10.8 -19.0 Southeastern Naturalist B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 288 = 1.7–12 y, average age = 4.3 y). We assume that Coyote individuals represented a similar demographic because in the study area, estimated survival rates for pups, juveniles, and adults are similar between the 2 species, and Coyotes are estimated to have only a slightly higher reproductive rate (Roth et al. 2008). These sample sizes are ~3 times greater than those needed to reliably estimate mean levels of nitrogen and carbon enrichment of the target Red Wolf and Coyote populations (i.e., 4–6 individuals), and high enough to do so for each sex of each species (Fox-Dobbs et al. 2007). Dentine (a form of collagen) was acid- and base-extracted, gelatinized, and filtered from each tooth by the Archaeological Center Research Facility within the University of Utah’s Department of Anthropology, Salt Lake City, UT. Mass spectrometry was performed on the resulting extractions at the University of Wyoming’s Stable Isotope Core Facility. We used the mass spectrometry results to determine the quality of preservation of the extracted samples (using atomic C:N ratio; Ambrose 1990) and to compare isotopic ratios (carbon enrichment and nitrogen enrichment) between species, as well as between females and males both across and within the 2 species (Tables 1, 2, 3). We created a simple linear model to compare the isotopic ratios to the mass of the canine teeth, which was used as an index of body size. We performed these comparisons between groups with Student’s t-tests in the stats package in Program R (R Core Team 2016). We considered relationships to be statistically significant at alpha < 0.05. Parental provisioning of pups could influence isotopic values obtained from dentine; thus, we used the Red Wolf Recovery Program pedigree database to insure that the Red Wolves we examined were neither siblings from the same litter nor siblings or half-siblings from different litters. We did not have similar pedigree data for the Coyotes. However, given that our Coyote samples came from a larger population pool than our Red Wolf samples, we suspect that the former also contained few, if any, siblings from the same litter. Results Atomic C:N ratios of all of our samples were 3.2, a value typical for high quality, modern bone collagen (Ambrose 1990). Results of the isotopic analysis are summarized in Tables 2 and 3. There was no significant difference in nitrogen isotope Table 3. Summary statistics for isotopic enrichment of male versus female Red Wolves and Coyotes. Sex Average δ15N‰ SD δ15N‰ Average δ13C‰ SD δ13C‰ Male 11.06 1.46 -17.44 1.84 Female 10.66 1.59 -18.18 2.11 Table 2. Summary statistics for isotopic enrichment of sympatric C. rufus (Red Wolf) and C. latrans (Coyote). Species Average δ15N‰ SD δ15N‰ Average δ13C‰ SD δ13C‰ C. rufus 10.5 1.4 -17.0 1.8 C. latrans 11.2 1.5 -18.4 1.9 Southeastern Naturalist 289 B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 chemistry (δ15N‰) between Red Wolves and Coyotes (t[29] = -1.43, P = 0.16; Fig. 1A) or between the males and females across both species (t[29] = -0.72, P = 0.48; Fig. 1B). There was a significant difference in carbon enrichment (δ13C0/00) between the two species (t[29) = 2.33, P = 0.03; Fig. 1C), although not between sexes across species (t[29] = -1.04; P = 0.31; Fig. 1D). Mean Coyote canine mass and the stable isotope values of males and females were not significantly different (t[14] = 0.97, Figure 1. (A) Comparison of nitrogen enrichment between 2 sympatric Canis species (difference not significant, t(29) = -1.43, P = 0.16);( B) Comparison of nitrogen enrichment between males and females across both species (difference not significant; t(29) = -0.72, P = 0.48); (C) Comparison of carbon enrichment between 2 sympatric Canis species (significant difference; t(29) = 2.33, P = 0.03); (D) Comparison of carbon enrichment between males and females across both species (difference not significant; t(29)= -1.04; P = 0.31). Southeastern Naturalist B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 290 P = 0.35), indicating that Coyote sub-adults of both sexes were similar in size over the period during which permanent canines formed and were, not surprisingly, consuming a similar range of prey taxa (presumably both from direct foraging and from being provisioned by adults) during that developmental stage. In contrast, the canine mass of Red Wolf males versus females was significantly different (t[13] = 2.160, P = 0.022). Red Wolves with larger canines exhibited enriched δ15N values (R2 = 0.4464, P = 0.006; Fig. 2A). The same is not true of Coyotes, in which canine weight was less varied and unrelated to δ15N (R2 = 0.004, P = 0.81; Fig. 2B). When we examined this relationship in male Red Wolves only, the R2 value increased with a corresponding increase in P-value, presumably due to a reduction in sample size (R2 = 0.5331, P = 0.03; Fig. 2C). There was no relationship between canine weight and δ13C values in either taxon (Red Wolves: R2 = > 0.001, P = 0.94; Coyotes: R2 = 0.10; P = 0.22). Finally, 2 Red Wolves (SB159, SB677) with the most depleted δ15N values (9‰ and 7.7‰, respectively) also exhibited the most enriched δ13C values (-13.9‰, -14.8‰, respectively). They also exhibited the lowest male canine weights suggesting they may be the smallest male sub-adults in the study and had diets proportionally more dependent on low trophic-level resources such as rodents, rabbits, or possibly agricultural crops. Approximate trophic position is visualized by plotting δ13C against δ15N (Fig. 3). Discussion By investigating potential dietary overlap between 2 sympatric Canis species using stable isotope analysis—an approach that had not previously been applied to this system—our study is complementary to previous analyses of scat samples from sympatric Red Wolves and Coyotes from the same geographic area (McVey et al. 2013). The sample sizes we used were ~3 times greater than those needed to reliably estimate mean levels of 15N and 13C enrichment in sympatric populations of Red Wolves and Coyotes in northeastern North Carolina, and large enough to obtain reliable population-level estimates for each sex of each species (Fox-Dobbs et al. 2007). This information is important for future biodiversity conservation efforts in this region and others because the USFWS (2016) plans to determine whether additional sites are appropriate for establishment of wild Red Wolf populations. The USFWS will base its decision on environmental assessments and collaboration with the public and other stakeholders. Therefore, there is a critical need to gather and disseminate information about the ecology of Red Wolves and sympatric Coyotes. Our data show that Red Wolves and Coyotes do not have significantly different nitrogen enrichment levels, suggesting that they exist at comparable levels of Figure 2 (following page). (A) Relationship between Red Wolf canine-tooth weight and enriched δ15N values (R2 = 0.4464, P = 0.006); (B) Relationship between Coyote canine-tooth weight and enriched δ13C values (R2 = 0.004; P = 0.81); (C) Relationship between male Red Wolf canine-tooth weight and enriched δ15N values (R2 = 0.5331, P = 0.03). Southeastern Naturalist 291 B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 Figure 2. [Caption on previous page]. Southeastern Naturalist B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 292 the trophic hierarchy in this ecosystem and appear to have similar diets. However, given a significant positive relationship between canine mass and δ15N in male Red Wolves, it is reasonable to hypothesize that as male Red Wolves mature in body size, they may be accessing higher trophic-level resources than those being accessed by Coyotes at maturity. In the future, stable isotope analyses could be used to elucidate the composition of Canis diets in more detail. However, such analyses will require a comparison of Canis isotopic signatures to signatures from potential prey items. At present, 2 independent lines of evidence—presented here and by McVey et al. (2013)—strongly suggest that the 2 species rely on the same basic prey base where they co-occur in the Red Wolf Recovery Area in northeastern North Carolina. Consequently, the 2 species appear to have very similar diets and trophic positions in this ecosystem, and stakeholders should carefully consider this information when debating the ecological implications of Red Wolf reintroductions and the potential loss of this species from the present ecosystem. Plotting δ13C against δ15N ratio (Fig. 3) can be used to assess an organism’s position in the Hutchinsonian “n-dimensional niche space” (Hutchinson 1957), following Hutchinson’s (1979) definitions of “bionomic” (resources consumed) and “scenopoetic” (the bioclimatic character of the environment, with a strong tie to spatial activity patterns) niche dimensions. Under this model, δ15N serves as a bionomic axis and δ13C serves as a scenopoetic axis. Figure 3 indicates significant overlap in the ecological niche of Red Wolves and Coyotes. A subset of Red Wolf samples, however, show enriched δ13C and depleted δ15N, in contrast to the scatter of other data points, while a subset of Coyotes showed relatively high δ15N and lower δ13C. Although these groupings may suggest some clustering of data points in Figure 3. Stable isotope signatures of sympatric Canis species. Average δ13C:δ15N ratio between species is similar (t[21] = 1.92, P = 0.07). Southeastern Naturalist 293 B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 each species (Fig. 3), the relatively wide variation in values we observed also could reflect inter-annual variation in climatic extremes because individuals reaching the same age may have experienced different seasonal conditions over the course of their lifetimes. Such seasonal differences in diet were observed by McVey et al. (2013) in their study of sympatric populations of Red Wolves and Coyotes. The difference in average carbon enrichment between the 2 species, although statistically significant, was relatively small (Table 2) and could be attributed to a number of factors, including the use of agricultural fields or the margins of human settlements as hunting or denning sites. If that is the case, these animals are likely subsisting on an omnivorous diet due to subsidization of crops and refuse from human activity (see Newsome et al. 2010). The Red Wolf Recovery Program is located near to both the North Carolina coast and inland agricultural areas, so this difference could indicate that the species are partitioning their foraging movements and home ranges within the region differently (for example, the species could be using coastal areas containing sea grasses to different degrees; see Peterson et al. 1980), or it could indicate that 1 of the species, possibly the larger-bodied Red Wolf (Nowak 1992, Thurber and Peterson 1991), occupies a wider variety of the available habitat types. Neither Red Wolves nor Coyotes showed significant intraspecific differences in diet between male and female individuals. This result is not unexpected for closely related species that tend to hunt in pairs and/or groups, and suggests that the sex ratios of reintroduced Red Wolf populations should not affect the level of predation pressure placed on the local prey base. Dentine formation is finalized before these 2 species begin to hunt for themselves; thus, an additional explanation for the lack of difference between sexes could be that adults provision pups similarly regardless of sex. Future studies comparing individuals of different age classes, through the comparison of bone collagen to tooth collagen (e.g., Bocherens et al. 1994), may identify ontogenetic shifts in diet selection and potentially reveal differences in diet between sexes during different periods of an individual’s life. Overall, our results correspond with those of McVey et al. (2013) in suggesting that sympatric Red Wolves and Coyotes have similar diets and are likely affecting prey populations similarly in the Red Wolf Experimental Recovery Area. This conclusion has important implications for conservation and management of Red Wolves in eastern North Carolina. Cessation of recovery efforts (i.e., allowing hybridization with Coyotes to occur freely), would very likely lead to the extinction of the only wild population of Red Wolves. Given that Red Wolves and Coyotes appear to have similar diets in eastern North Carolina, the loss of wild populations of the former may not lead to a decrease in functional diversity in the region. However, Red Wolf extirpation would lead to a decrease in taxonomic and phylogenetic diversity, and potentially to the loss of an important genetic component of biodiversity. Acknowledgments We thank David Rabon (formerly of the USFWS Red Wolf Recovery Program), Rebecca Harrison (USFWS), and W. David Webster (UNC Wilmington) for their help in providing Southeastern Naturalist B.S. Arbogast, A.M.C. Hodge, and J. Brenner-Coltrain 2017 Vol. 16, No. 2 294 access to specimens and associated pedigree data. 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