Predicting the odds of chronic wasting disease with Habitat Risk software.

Chronic wasting disease (CWD) is a transmissible spongiform encephalopathy that was first detected in captive cervids in Colorado, United States (US) in 1967, but has since spread into free-ranging white-tailed deer (Odocoileus virginianus) across the US and Canada as well as to Scandinavia and South Korea. In some areas, the disease is considered endemic in wild deer populations, and governmental wildlife agencies have employed epidemiological models to understand long-term environmental risk. However, continued rapid spread of CWD into new regions of the continent has underscored the need for extension of these models into broader tools applicable for wide use by wildlife agencies. Additionally, efforts to semi-automate models will facilitate access of technical scientific methods to broader users. We introduce software (Habitat Risk) designed to link a previously published epidemiological model with spatially referenced environmental and disease testing data to enable agency personnel to make up-to-date, localized, data-driven predictions regarding the odds of CWD detection in surrounding areas after an outbreak is discovered. Habitat Risk requires pre-processing publicly available environmental datasets and standardization of disease testing (surveillance) data, after which an autonomous computational workflow terminates in a user interface that displays an interactive map of disease risk. We demonstrated the use of the Habitat Risk software with surveillance data of white-tailed deer from Tennessee, USA.

Large-scale assessment of genetic structure to assess risk of populations of a large herbivore to disease.

Chronic wasting disease (CWD) can spread among cervids by direct and indirect transmission, the former being more likely in emerging areas. Identifying subpopulations allows the delineation of focal areas to target for intervention. We aimed to assess the population structure of white-tailed deer (Odocoileus virginianus) in the northeastern United States at a regional scale to inform managers regarding gene flow throughout the region. We genotyped 10 microsatellites in 5701 wild deer samples from Maryland, New York, Ohio, Pennsylvania, and Virginia. We evaluated the distribution of genetic variability through spatial principal component analysis and inferred genetic structure using non-spatial and spatial Bayesian clustering algorithms (BCAs). We simulated populations representing each inferred wild cluster, wild deer in each state and each physiographic province, total wild population, and a captive population. We conducted genetic assignment tests using these potential sources, calculating the probability of samples being correctly assigned to their origin. Non-spatial BCA identified two clusters across the region, while spatial BCA suggested a maximum of nine clusters. Assignment tests correctly placed deer into captive or wild origin in most cases (94%), as previously reported, but performance varied when assigning wild deer to more specific origins. Assignments to clusters inferred via non-spatial BCA performed well, but efficiency was greatly reduced when assigning samples to clusters inferred via spatial BCA. Differences between spatial BCA clusters are not strong enough to make assignment tests a reliable method for inferring the geographic origin of deer using 10 microsatellites. However, the genetic distinction between clusters may indicate natural and anthropogenic barriers of interest for management.

Evaluation of DNA yield from various tissue and sampling sources for use in single nucleotide polymorphism panels.

Genetics studies are used by wildlife managers and researchers to gain inference into a population of a species of interest. To gain these insights, microsatellites have been the primary method; however, there currently is a shift from microsatellites to single nucleotide polymorphisms (SNPs). With the different DNA requirements between microsatellites and SNPs, an investigation into which samples can provide adequate DNA yield is warranted. Using samples that were collected from previous genetic projects from regions in the USA from 2014 to 2021, we investigated the DNA yield of eight sample categories to gain insights into which provided adequate DNA to be used in ddRADseq or already developed high- or medium-density SNP panels. We found seven sample categories that met the DNA requirements for use in all three panels, and one sample category that did not meet any of the three panels requirements; however, DNA integrity was highly variable and not all sample categories that met panel DNA requirements could be considered high quality DNA. Additionally, we used linear random-effects models to determine which covariates would have the greatest influence on DNA yield. We determined that all covariates (tissue type, storage method, preservative, DNA quality, time until DNA extraction and time after DNA extraction) could influence DNA yield.

Prion forensics: a multidisciplinary approach to investigate CWD at an illegal deer carcass disposal site.

Infectious prions are resistant to degradation and remain infectious in the environment for several years. Chronic wasting disease (CWD) has been detected in cervids inhabiting North America, the Nordic countries, and South Korea. CWD-prion spread is partially attributed to carcass transport and disposal. We employed a forensic approach to investigate an illegal carcass dump site connected with a CWD-positive herd. We integrated anatomic, genetic, and prion amplification methods to discover CWD-positive remains from six white-tailed deer (Odocoileus virginianus) and, using microsatellite markers, confirmed a portion originated from the CWD-infected herd. This approach provides a foundation for future studies of carcass prion transmission risk.

Variability in prion protein genotypes by spatial unit to inform susceptibility to chronic wasting disease.

Chronic wasting disease (CWD) is a fatal encephalopathy affecting North American cervids. Certain alleles in a host's prion protein gene are responsible for reduced susceptibility to CWD. We assessed for the first time variability in the prion protein gene of elk (Cervus canadensis) present in Pennsylvania, United States of America, a reintroduced population for which CWD cases have never been reported. We sequenced the prion protein gene (PRNP) of 565 elk samples collected over 7 years (2014-2020) and found two polymorphic sites (codon 21 and codon 132). The allele associated with reduced susceptibility to CWD is present in the population, and there was no evidence of deviations from Hardy-Weinberg equilibrium in any of our sampling years (p-values between 0.14 and 1), consistent with the lack of selective pressure on the PRNP. The less susceptible genotypes were found in a frequency similar to the ones reported for elk populations in the states of Wyoming and South Dakota before CWD was detected. We calculated the proportion of less susceptible genotypes in each hunt zone in Pennsylvania as a proxy for their vulnerability to the establishment of CWD, and interpolated these results to obtain a surface representing expected proportion of the less susceptible genotypes across the area. Based on this analysis, hunt zones located in the southern part of our study area have a low proportion of less susceptible genotypes, which is discouraging for elk persistence in Pennsylvania given that these hunt zones are adjacent to the deer Disease Management Area 3, where CWD has been present since 2014.

Fatal infection with Versteria sp. in a muskrat, with implications for human health.

The Taeniidae tapeworms are a family of helminths that have a similar life cycle, with intermediate hosts developing characteristic cysts in visceral organs. We describe here a case in Pennsylvania, USA, of fatal Versteria infection in a muskrat (Ondatra zibethicus), which, to our knowledge, has not been reported to develop disease associated with infection. Postmortem examination revealed widespread tissue loss and replacement by solid-bodied cestode larvae with minimal adjacent inflammation in many visceral organs, most severe in the lungs, liver, and brain. Key morphologic features via histology included cephalic structures and short rostellar hooklets, which are characteristic for the genus. Genetic characterization confirmed the cestode as being an undescribed lineage of Versteria that has been implicated as the cause of severe morbidity and mortality in humans and nonhuman primates in North America. Considering the zoonotic significance of this pathogen, our report expands on the limited literature regarding disease caused by Versteria and emphasizes the need to identify the causative tapeworm more accurately, especially in rodent intermediate hosts given that previous reports do not have molecular confirmation of species.

Surveillance for diseases, pathogens, and toxicants of muskrat (Ondatra zibethicus) in Pennsylvania and surrounding regions.

Using diagnostic data and contemporary sampling efforts, we conducted surveillance for a diversity of pathogens, toxicants, and diseases of muskrats (Ondatra zibethicus). Between 1977 and 2019, 26 diagnostic cases were examined from Kansas and throughout the Southeast and Mid-Atlantic, USA. We identified multiple causes of mortality in muskrats, but trauma (8/26), Tyzzer's disease (5/6), and cysticercosis (5/26) were the most common. We also conducted necropsies, during November 2018-January 2019 Pennsylvania muskrat trapping season, on 380 trapper-harvested muskrat carcasses after the pelt was removed. Tissue samples and exudate were tested for presence of or exposure to a suite of pathogens and contaminants. Gastrointestinal tracts were examined for helminths. Intestinal helminths were present in 39.2% of necropsied muskrats, with Hymenolepis spp. (62%) and echinostome spp. (44%) being the most common Molecular testing identified a low prevalence of infection with Clostridium piliforme in the feces and Sarcocystis spp. in the heart. We detected a low seroprevalence to Toxoplasma gondii (1/380). No muskrats were positive for Francisella tularensis or Babesia spp. Cysticercosis was detected in 20% (5/26) of diagnostic cases and 15% (57/380) of our trapper-harvested muskrats. Toxic concentrations of arsenic, cadmium, lead, or mercury were not detected in tested liver samples. Copper, molybdenum, and zinc concentrations were detected at acceptable levels comparative to previous studies. Parasite intensity and abundance were typical of historic reports; however, younger muskrats had higher intensity of infection than older muskrats which is contradictory to what has been previously reported. A diversity of pathogens and contaminants have been reported from muskrats, but the associated disease impacts are poorly understood. Our data are consistent with historic reports and highlight the wide range of parasites, pathogens and contaminants harbored by muskrats in Pennsylvania. The data collected are a critical component in assessing overall muskrat health and serve as a basis for understanding the impacts of disease on recent muskrat population declines.

Comparison of sample types from white-tailed deer (Odocoileus virginianus) for DNA extraction and analyses.

Collection of biological samples for DNA is necessary in a variety of disciplines including disease epidemiology, landscape genetics, and forensics. Quantity and quality of DNA varies depending on the method of collection or media available for collection (e.g., blood, tissue, fecal). Blood is the most common sample collected in vials or on Whatman Flinders Technology Associates (FTA) cards with short- and long-term storage providing adequate DNA for study objectives. The focus of this study was to determine if biological samples stored on Whatman FTA Elute cards were a reasonable alternative to traditional DNA sample collection, storage, and extraction. Tissue, nasal swabs, and ocular fluid were collected from white-tailed deer (Odocoileus virginianus). Tissue samples and nasal swabs acted as a control to compare extraction and DNA suitability for microsatellite analysis for nasal swabs and ocular fluid extracted from FTA Elute cards. We determined that FTA Elute cards improved the extraction time and storage of samples and that nasal swabs and ocular fluid containing pigmented fluid were reasonable alternatives to traditional tissue DNA extractions.

A Review of Pathogens, Diseases, and Contaminants of Muskrats (Ondatra zibethicus) in North America.

Over the last 50 years, significant muskrat (Ondatra zibethicus) harvest declines have been observed throughout North America. Several theories for the decline have been proposed, including increased parasite infections and disease within muskrat populations. No existing wholistic review of muskrat exposure to pathogens, contaminants, and diseases exists. To address this knowledge gap, we conducted a thorough review of existing literature on muskrat pathogens, contaminants, and diseases across their natural range. This review is comprised of 131 articles from 1915 to 2019 and from 27 U.S. states and 9 Canadian provinces. A wide diversity of contaminants, toxins, and pathogens were reported in muskrats, with the most common diseases being cysticercosis, tularemia, Tyzzer's disease, and biotoxin poisoning from cyanobacteria. This review provides a summary of muskrat pathogens, contaminants, and diseases over a century that has observed significant population declines throughout the species' range in North America. Such data provide a baseline for understanding the potential role of disease in these declines. In addition, these data highlight critical knowledge gaps that warrant future research efforts.

Assessment of spatial genetic structure to identify populations at risk for infection of an emerging epizootic disease.

Understanding the geographic extent and connectivity of wildlife populations can provide important insights into the management of disease outbreaks but defining patterns of population structure is difficult for widely distributed species. Landscape genetic analyses are powerful methods for identifying cryptic structure and movement patterns that may be associated with spatial epizootic patterns in such cases.We characterized patterns of population substructure and connectivity using microsatellite genotypes from 2,222 white-tailed deer (Odocoileus virginianus) in the Mid-Atlantic region of the United States, a region where chronic wasting disease was first detected in 2009. The goal of this study was to evaluate the juxtaposition between population structure, landscape features that influence gene flow, and current disease management units.Clustering analyses identified four to five subpopulations in this region, the edges of which corresponded to ecophysiographic provinces. Subpopulations were further partitioned into 11 clusters with subtle (F ST ≤ 0.041), but significant genetic differentiation. Genetic differentiation was lower and migration rates were higher among neighboring genetic clusters, indicating an underlying genetic cline. Genetic discontinuities were associated with topographic barriers, however.Resistance surface modeling indicated that gene flow was diffuse in homogenous landscapes, but the direction and extent of gene flow were influenced by forest cover, traffic volume, and elevational relief in subregions heterogeneous for these landscape features. Chronic wasting disease primarily occurred among genetic clusters within a single subpopulation and along corridors of high landscape connectivity.These results may suggest a possible correlation between population substructure, landscape connectivity, and the occurrence of diseases for widespread species. Considering these factors may be useful in delineating effective management units, although only the largest features produced appreciable differences in subpopulation structure. Disease mitigation strategies implemented at the scale of ecophysiographic provinces are likely to be more effective than those implemented at finer scales.

Can genetic assignment tests provide insight on the influence of captive egression on the epizootiology of chronic wasting disease?

Identifying the sources of ongoing and novel disease outbreaks is critical for understanding the diffusion of epizootic diseases. Identifying infection sources is difficult when few physical differences separate individuals with different origins. Genetic assignment procedures show great promise for assessing transmission dynamics in such situations. Here, we use genetic assignment tests to determine the source of chronic wasting disease infections in free-ranging white-tailed deer (Odocoileus virginianus) populations. Natural dispersal is thought to facilitate the geographic diffusion of chronic wasting disease, but egression from captive cervid populations represents an alternative source of infection that is difficult to detect due to physical similarities with wild deer. Simulated reference populations were created based on allele frequencies from 1,912 empirical microsatellite genotypes collected in four sampling subregions and five captive facilities. These reference populations were used to assess the likelihood of ancestry and assignment of 1,861 free-ranging deer (1,834 noninfected and 27 infected) and 51 captive individuals to captive or wild populations. The ancestry (Q) and assignment scores (A) for free-ranging deer to wild populations were high (average Q wild = 0.913 and average A wild = 0.951, respectively), but varied among subregions (Q wild = 0.800-0.947, A wild = 0.857-0.976). These findings suggest that captive egression and admixture are rare, but risk may not be spatially uniform. Ancestry and assignment scores for two free-ranging deer with chronic wasting disease sampled in an area where chronic wasting disease was previously unobserved in free-ranging herds indicated a higher likelihood of assignment and proportion of ancestry attributable to captive populations. While we cannot directly assign these individuals to infected facilities, these findings suggest that rare egression events may influence the epizootiology of chronic wasting disease in free-ranging populations. Continued disease surveillance and genetic analyses may further elucidate the relative disease risk attributable to captive and wild sources.

Identification and evaluation of a core microsatellite panel for use in white-tailed deer (Odocoileus virginianus).

BACKGROUND: Microsatellite loci have been used extensively over the past two decades to study the genetic characteristics of non-model species. The ease of microsatellite development and ability to adapt markers from related species has led to the proliferation of available markers for many commonly studied species. Because it is often infeasible to genotype individuals across all available loci, researchers generally rely on subsets of markers. Marker choice can bias inferences made using disparate suites of loci. This has been a primary motivation for efforts to identify uniform marker panels. Here, we use the geographic distribution of previous studies to identify microsatellite loci for white-tailed deer (Odocoileus virginianus) with the potential for widespread use, and we evaluate the effectiveness of this panel in a portion of the range where few previous studies have been conducted. The purpose was to consolidate the numerous genetic resources for this species into a manageable panel and to provide a uniform methodology that improves comparisons between past and future studies. RESULTS: We reviewed microsatellite panels from 58 previous or ongoing projects and identified 106 candidate loci. We developed a multiplex protocol and evaluated the efficacy of 17 of the most commonly used loci using 720 DNA samples collected from the Mid-Atlantic region of the United States of America. Amplification errors were detected in six of these loci. The 11 remaining loci were highly polymorphic, exhibited low frequencies of null alleles, and were easy to interpret with the aid of allele binning software. CONCLUSIONS: The development of broadly-applicable, core microsatellite panels has the potential to improve repeatability and comparative ability for commonly studied species. The properties of the consolidated 11 microsatellite panel suggest that they are applicable for many common research objectives for white-tailed deer. The geographic distribution of previous studies using these markers provides a greater degree of confidence regarding the robustness to common sources of error related to amplification anomalies, such as null alleles, relative to loci with more limited use. While this does not replace further evaluation of genotyping errors, it does provide a common platform that benefits future research studies.

Spatial heterogeneity of prion gene polymorphisms in an area recently infected by chronic wasting disease.

Genetic variability in the prion protein (Prnp) gene influences host susceptibility to many pathogenic prion diseases. Understanding the distribution of susceptible Prnp variants and determining factors influencing spatial genetic patterns are important components of many chronic wasting disease mitigation strategies. Here, we describe Prnp variability in white-tailed deer (Odocoileus virginianus) from the Mid-Atlantic region of the United States of America, an area with a recent history of infection and low disease incidence. This population is characterized by lower rates of polymorphism and significantly higher frequencies of the more susceptible 96GG genotype compared to previously surveyed populations. The prevalence of the most susceptible genotypes at disease-associated loci did vary among subregions, indicating that populations have innate differences in genotype-dictated susceptibility.

Seasonal home ranges and habitat selection of three elk (Cervus elaphus) herds in North Dakota.

Changes in land use have resulted in range shifts of many wildlife species, including those entering novel environments, resulting in the critical need to understand their spatial ecology to inform ecosystem effects and management decisions. Dispersing elk (Cervus elaphus) were colonizing areas of suitable habitat in the Northern Great Plains, USA, resulting in crop depredation complaints in these areas. Although state resource managers had little information on these elk herds, limited evidence suggested temporal movements into Canada. We collected and analyzed essential information on home range and habitat selection for 3 elk herds residing in North Dakota. We captured 5 adult female elk in each study area, affixed global positioning system collars, and monitored them for 1 year (2016-2017). We estimated diel period, seasonal, and hunting season home ranges using Brownian Bridge Movement Models for each individual. We analyzed habitat selection using multinomial logit models to test for differences in use of land classes, and for departures from proportionate use based on random sampling; our predictor variables included individual elk, diel period, and season. Home ranges differed between the 3 herds, seasons, and diel period; gun and winter season home ranges were both larger than in summer, as was night when compared with day. Female elk generally restricted themselves to cover during the day and entered open areas at night and during winter months. Our results also suggest that elk in our study areas tended to seek more cover, and in the case of our Turtle Mountain study area, some cross into Canada during gun season. Our study provides a better understanding of the spatial ecology of elk in the Northern Great Plains while highlighting the need for enhanced international cooperative management efforts.

Heterogeneity of a landscape influences size of home range in a North American cervid.

In the northeastern United States, chronic wasting disease has recently been detected in white-tailed deer (Odocoileus virginianus) populations, and understanding the relationship between landscape configuration and home range may improve disease surveillance and containment efforts. The objectives of our study were to compare size of home range for deer occupying a continuum of forested landscapes and to investigate relationships between size of home range and measures of landscape configuration. We used a movement-based kernel density estimator to estimate home range at five spatial scales among deer across study areas. We developed 7 linear regression models that used measures of the configuration of the forested landscape to explain size of home range. We observed differences in size of home range between sexes among areas that differed based on landscape configuration. We documented size of home range changed with various metrics that identifying connectivity of forested patches. Generally, size of home range increased with an increasing proportion of homogenous forest. Our results suggest that deer in our region occupy a landscape at hierarchically-nested scales that is controlled by the connectivity of the forested landscape across local or broad geographical regions.

Influence of Precipitation and Crop Germination on Resource Selection by Mule Deer (Odocoileus hemionus) in Southwest Colorado.

Mule deer (Odocoileus hemionus) populations in the western United States provide many benefits to local economies but can also cause considerable damage to agriculture, particularly damage to lucrative crops. Limited information exists to understand resource selection of mule deer in response to annual variation in crop rotation and climatic conditions. We tested the hypothesis that mule deer select certain crops, and in particular sunflower, based on annual climatic variability. Our objective was to use movements, estimates of home range, and resource selection analysis to identify resources selected by mule deer. We used annually-derived crop-specific datasets along with Global Positioning System collars to monitor 14 mule deer in an agricultural area near public lands in southwestern Colorado, USA. We estimated home ranges for two winter seasons that ranged between 7.68 and 9.88 km2, and for two summer seasons that ranged between 5.51 and 6.24 km2. Mule deer selected areas closer to forest and alfalfa for most periods during 2012, but selected areas closer to sunflower in a majority of periods during 2013. Considerable annual variation in climate patterns and precipitation levels appeared to influence selection by mule deer because of variability in crop rotation and success of germination of specific crops.

CWDPRNP: a tool for cervid prion sequence analysis in program R.

SUMMARY: Chronic wasting disease is a fatal, neurological disease caused by an infectious prion protein, which affects economically and ecologically important members of the family Cervidae. Single nucleotide polymorphisms within the prion protein gene have been linked to differential susceptibility to the disease in many species. Wildlife managers are seeking to determine the frequencies of disease-associated alleles and genotypes and delineate spatial genetic patterns. The CWDPRNP package, implemented in program R, provides a unified framework for analyzing prion protein gene variability and spatial structure. AVAILABILITY AND IMPLEMENTATION: The CWDPRNP package, manual and example data files are available at http://ecosystems.psu.edu/research/labs/walter-lab/additional-labs/population-genetics-lab. This package is available for all commonly used platforms. CONTACT: wlm159psu@gmail.com.

Seeded Amplification of Chronic Wasting Disease Prions in Nasal Brushings and Recto-anal Mucosa-Associated Lymphoid Tissues from Elk by Real-Time Quaking-Induced Conversion.

Chronic wasting disease (CWD), a transmissible spongiform encephalopathy of cervids, was first documented nearly 50 years ago in Colorado and Wyoming and has since been detected across North America and the Republic of Korea. The expansion of this disease makes the development of sensitive diagnostic assays and antemortem sampling techniques crucial for the mitigation of its spread; this is especially true in cases of relocation/reintroduction or prevalence studies of large or protected herds, where depopulation may be contraindicated. This study evaluated the sensitivity of the real-time quaking-induced conversion (RT-QuIC) assay of recto-anal mucosa-associated lymphoid tissue (RAMALT) biopsy specimens and nasal brushings collected antemortem. These findings were compared to results of immunohistochemistry (IHC) analysis of ante- and postmortem samples. RAMALT samples were collected from populations of farmed and free-ranging Rocky Mountain elk (Cervus elaphus nelsoni;n= 323), and nasal brush samples were collected from a subpopulation of these animals (n= 205). We hypothesized that the sensitivity of RT-QuIC would be comparable to that of IHC analysis of RAMALT and would correspond to that of IHC analysis of postmortem tissues. We found RAMALT sensitivity (77.3%) to be highly correlative between RT-QuIC and IHC analysis. Sensitivity was lower when testing nasal brushings (34%), though both RAMALT and nasal brush test sensitivities were dependent on both thePRNPgenotype and disease progression determined by the obex score. These data suggest that RT-QuIC, like IHC analysis, is a relatively sensitive assay for detection of CWD prions in RAMALT biopsy specimens and, with further investigation, has potential for large-scale and rapid automated testing of antemortem samples for CWD.

Antemortem Detection of Chronic Wasting Disease Prions in Nasal Brush Collections and Rectal Biopsy Specimens from White-Tailed Deer by Real-Time Quaking-Induced Conversion.

Chronic wasting disease (CWD), a transmissible spongiform encephalopathy of cervids, was first documented nearly 50 years ago in Colorado and Wyoming and has since spread to cervids in 23 states, two Canadian provinces, and the Republic of Korea. The expansion of this disease makes the development of sensitive diagnostic assays and antemortem sampling techniques crucial for the mitigation of its spread; this is especially true in cases of relocation/reintroduction of farmed or free-ranging deer and elk or surveillance studies of private or protected herds, where depopulation is contraindicated. This study sought to evaluate the sensitivity of the real-time quaking-induced conversion (RT-QuIC) assay by using recto-anal mucosa-associated lymphoid tissue (RAMALT) biopsy specimens and nasal brush samples collected antemortem from farmed white-tailed deer (n= 409). Antemortem findings were then compared to results from ante- and postmortem samples (RAMALT, brainstem, and medial retropharyngeal lymph nodes) evaluated by using the current gold standardin vitroassay, immunohistochemistry (IHC) analysis. We hypothesized that the sensitivity of RT-QuIC would be comparable to IHC analysis in antemortem tissues and would correlate with both the genotype and the stage of clinical disease. Our results showed that RAMALT testing by RT-QuIC assay had the highest sensitivity (69.8%) compared to that of postmortem testing, with a specificity of >93.9%. These data suggest that RT-QuIC, like IHC analysis, is an effective assay for detection of PrP(CWD)in rectal biopsy specimens and other antemortem samples and, with further research to identify more sensitive tissues, bodily fluids, or experimental conditions, has potential for large-scale and rapid automated testing for CWD diagnosis.

Is there a single best estimator? Selection of home range estimators using area-under-the-curve.

BACKGROUND: Global positioning system (GPS) technology for monitoring home range and movements of wildlife has resulted in prohibitively large sample sizes of locations for traditional estimators of home range. We used area-under-the-curve to explore the fit of 8 estimators of home range to data collected with both GPS and concurrent very high frequency (VHF) technology on a terrestrial mammal, the Florida panther Puma concolor coryi, to evaluate recently developed and traditional estimators. RESULTS: Area-under-the-curve was the highest for Florida panthers equipped with Global Positioning System (GPS) technology compared to VHF technology. For our study animal, estimators of home range that incorporated a temporal component to estimation performed better than traditional first- and second-generation estimators. CONCLUSIONS: Comparisons of fit of home range contours with locations collected would suggest that use of VHF technology is not as accurate as GPS technology to estimate size of home range for large mammals. Estimators of home range collected with GPS technology performed better than those estimated with VHF technology regardless of estimator used. Furthermore, estimators that incorporate a temporal component (third-generation estimators) appeared to be the most reliable regardless of whether kernel-based or Brownian bridge-based algorithms were used and in comparison to first- and second-generation estimators. We defined third-generation estimators of home range as any estimator that incorporates time, space, animal-specific parameters, and habitat. Such estimators would include movement-based kernel density, Brownian bridge movement models, and dynamic Brownian bridge movement models among others that have yet to be evaluated.