A novel enhancer of Agouti contributes to parallel evolution of cryptically colored beach mice

Identifying the genetic basis of repeatedly evolved traits provides a way to reconstruct their evolutionary history and ultimately investigate the predictability of evolution. Here, we focus on the oldfield mouse (Peromyscus polionotus), which occurs in the southeastern United States, where it exhibits considerable coat-color variation. Dorsal coats range from dark brown in mice inhabiting mainland habitat to near white on the white-sand beaches of the southeastern US, where light pelage has evolved independently on Florida's Gulf and Atlantic coasts as an adaptation to visually hunting predators. To facilitate genomic analyses in this species, we first generated a high-quality, chromosome-level genome assembly of P. polionotus subgriseus. Next, in a uniquely variable mainland population that occurs near beach habitat (P. p. albifrons), we scored 23 pigment traits and performed targeted resequencing in 168 mice. We find that variation in pigmentation is strongly associated with a ~2 kb region approximately 5 kb upstream of the Agouti-signaling protein (ASIP) coding region. Using a reporter-gene assay, we demonstrate that this regulatory region contains an enhancer that drives expression in the dermis of mouse embryos during the establishment of pigment prepatterns. Moreover, extended tracts of homozygosity in this region of Agouti indicate that the light allele has experienced recent and strong positive selection. Notably, this same light allele appears fixed in both Gulf and Atlantic coast beach mice, despite these populations being separated by >1,000km. Given the evolutionary history of this species, our results suggest that this newly identified Agouti enhancer allele has been maintained in mainland populations as standing genetic variation and from there has spread to, and been selected in, two independent beach mouse lineages, thereby facilitating their rapid and parallel evolution.

Automated tracking reveals the social network of beach mice and their burrows

Evolutionary biologists have long sought to understand the selective pressures driving phenotypic evolution. While most experimental data come from the study of morphological evolution, we know much less about the ultimate drivers of behavioral variation. Among the most striking examples of behavioral evolution are the long, complex burrows constructed by oldfield mice ( Peromyscus polionotus ssp.). Yet how these mice use burrows in the wild, and whether burrow length may affect fitness, remains unknown. A major barrier to studying behavior in the wild has been the lack of technologies to continuously monitor — in this case, nocturnal and underground — behavior. Here, we designed and implemented a novel radio frequency identification (RFID) system to track patterns of burrow use in a natural population of beach mice. We combine RFID monitoring with burrow measurements, genetic data, and social network analysis to uncover how these monogamous mice use burrows under fully natural ecological and social conditions. We first found that long burrows provide a more stable thermal environment and have higher juvenile activity than short burrows, underscoring the likely importance of long burrows for rearing young. We also find that adult mice consistently use multiple burrows throughout their home range and tend to use the same burrows at the same time as their genetic relatives, suggesting that inclusive fitness benefits may accrue for individuals that construct and maintain multiple burrows. Our study highlights how new automated tracking approaches can provide novel insights into animal behavior in the wild.

Differentiating Footprints of Sympatric Rodents in Coastal Dune Communities: Implications for Imperiled Beach Mice

Identifying techniques for conducting frequent, effective, and inexpensive monitoring of small mammals can be challenging. Traditional approaches such as livetrapping can be laborious, expensive, detrimental to animal health, and ineffective. Passive approaches such as tracking (e.g., from tracks on the ground or footprints collected at a tracking station) have been shown to lessen those burdens, but a problem with tracking, particularly for rodents, is the uncertainty in identifying species from footprints. To address the need for a more accurate method of identifying small mammal tracks, we measured footprints from live-captured rodents and developed a classification tree for distinguishing between subspecies and species using footprint widths treated as having known or unknown identification. We captured rodents within or near the coastal dunes of Florida and Alabama with a focus on areas occupied by threatened and endangered beach mice Peromyscus polionotus subspp., whose populations warrant regular monitoring but whose tracks are not easily distinguished from those of some sympatric species. We measured 6,996 front and hind footprints from 540 individuals across eight species. The overall accuracy of our classification tree was 82.6% and we achieved this using only the front footprint width. Footprint width cutoffs for species identification were < 5.5 mm for house mice Mus musculus, 5.5–6.7 mm for beach mice, and 6.7–8.3 mm for cotton mice Peromyscus gossypinus. We were most successful in confirming the identity of beach mice: we correctly classified approximately 94% of beach mice, while we misclassified fewer than 6% as house mice and fewer than 1% as cotton mice. When we input a beach mouse individual into the classification tree as of an unknown species, we correctly identified 78.1% of individuals as beach mice from their tracks, and most incorrect identifications were of house mouse tracks. Our study demonstrates that researchers can identify sympatric rodent species in coastal dune communities from tracks using quantitative classification based on footprint width. Accurate identification of beach mice or other imperiled species from tracks has important management implications. Not only can wildlife managers determine the presence of a species accurately, but they can monitor populations with considerably less effort than livetrapping requires. Although our study was specific to coastal dune communities, our methods could be adapted for the creation of a classification tree for identifying tracks from suites of species in other areas.

Increasing Detection by Reducing Disturbance and Excluding Nontarget Small Mammal Species: An Occupancy Study Approach

Track tubes are a noninvasive, efficient method to monitor populations of small mammals that can be implemented on a large landscape scale and are a cost-effective approach for certain sampling situations. As with all field sampling tools, modifications are made depending on research objectives, habitat being sampled, and target species. We conducted two experiments with the objective to increase efficiency and decrease labor while retaining high detection probabilities as part of an Annual Multi-Agency Regional Southeastern Beach Mouse Peromyscus polionotus niveiventris Habitat Occupancy Survey. We conducted studies along a contiguous 72-km coastline of the Cape Canaveral Barrier Island Complex in east-central Florida, USA, that includes the Kennedy Space Center, Cape Canaveral Air Force Station, and Canaveral National Seashore. We conducted the experiments to address several issues that had regularly compromised our survey data. One experiment assessed the ideal length of deployment times for track tubes in various habitat types with multiple species where southeastern beach mice were previously detected. Another experiment, determined how to restrict certain meso-mammal species (raccoons Procyon lotor, and eastern spotted skunks Spilogale putorius) from knocking over the tubes or reaching into the tubes, thus reducing disturbance that results in missing data; or from entering the tubes (cotton rats Sigmodon hispidus), thereby obscuring any potential or actual footprints of the southeastern beach mouse or other targeted species. We observed an increase in the detection rate of southeastern beach mice in track tubes with increased nights of deployment (likelihood ratio test χ2 = 18.71, df = 3, P < 0.001) with the greatest increase between 3 nights and 6 nights and apparent leveling off between nights 9 and 12 and a large decrease in the detection rate of southeastern beach mice in track tubes with 0.5-in. (1.3-cm) excluders compared with the other-size excluders (likelihood ratio test χ2 = 167.89, df = 5, P < 0.001). We also found that 1-in. (2.5-cm) excluders prevented access by meso-mammals or entry by cotton rats and did not adversely affect the detection of either beach mice or cotton mice Peromyscus gossypinus, but did reduce disturbance and resulted in fewer missing data. These statistically significant process improvements have application for others involved with small mammal monitoring and species management.

Evolution and genetics of precocious burrowing behavior in Peromyscus mice

A central challenge in biology is to understand how innate behaviors evolve between closely related species. One way to elucidate how differences arise is to compare the development of behavior in species with distinct adult traits. Here, we report that Peromyscus polionotus is strikingly precocious with regard to burrowing behavior, but not other behaviors, compared to its sister species P. maniculatus . In P. polionotus , burrows were excavated as early as 17 days of age, while P. maniculatus did not build burrows until 10 days later. Moreover, the well-known differences in burrow architecture between adults of these species -- P. polionotus adults excavate long burrows with an escape tunnel, while P. maniculatus dig short, single-tunnel burrows -- were intact in juvenile burrowers. To test whether this juvenile behavior is influenced by early-life environment, pups of both species were reciprocally cross-fostered. Fostering did not alter the characteristic burrowing behavior of either species, suggesting these differences are genetic. In backcross F2 hybrids, we show that precocious burrowing and adult tunnel length are genetically correlated, and that a single P. polionotus allele in a genomic region linked to adult tunnel length is predictive of precocious burrow construction. The co-inheritance of developmental and adult traits indicates the same genetic region -- either a single gene with pleiotropic effects, or closely linked genes -- acts on distinct aspects of the same behavior across life stages. Such genetic variants likely affect behavioral drive (i.e. motivation) to burrow, and thereby affect both the development and adult expression of burrowing behavior.

Effects of a maternal high-fat diet on offspring behavioral and metabolic parameters in a rodent model

Maternal diet-induced obesity can cause detrimental developmental origins of health and disease in offspring. Perinatal exposure to a high-fat diet (HFD) can lead to later behavioral and metabolic disturbances, but it is not clear which behaviors and metabolic parameters are most vulnerable. To address this critical gap, biparental and monogamous oldfield mice (Peromyscus polionotus), which may better replicate most human societies, were used in the current study. About 2 weeks before breeding, adult females were placed on a control or HFD and maintained on the diets throughout gestation and lactation. F1 offspring were placed at weaning (30 days of age) on the control diet and spatial learning and memory, anxiety, exploratory, voluntary physical activity, and metabolic parameters were tested when they reached adulthood (90 days of age). Surprisingly, maternal HFD caused decreased latency in initial and reverse Barnes maze trials in male, but not female, offspring. Both male and female HFD-fed offspring showed increased anxiogenic behaviors, but decreased exploratory and voluntary physical activity. Moreover, HFD offspring demonstrated lower resting energy expenditure (EE) compared with controls. Accordingly, HFD offspring weighed more at adulthood than those from control fed dams, likely the result of reduced physical activity and EE. Current findings indicate a maternal HFD may increase obesity susceptibility in offspring due to prenatal programming resulting in reduced physical activity and EE later in life. Further work is needed to determine the underpinning neural and metabolic mechanisms by which a maternal HFD adversely affects neurobehavioral and metabolic pathways in offspring.