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Friday, July 12, 2013

ABSTRACTS OF PRESENTATIONS FROM THE 2013 Box Turtle Conservation Workshop


March 22-23, 2013- North Carolina Zoological Park, Asheboro, NC
Reprinted Courtesy of HerpDigest.org, with the permission of the North American Box Turtle Conservation Committer www.boxturtleconservation.org
Part 1
Abstract of Presentations

1) Epidemiology and Treatment of Ranaviral Disease in North American Chelonians
2) Development of a Health Screening Protocol and Incorporating it into an Existing Field Project
3) Survivorship and Home Range of Head-started Juvenile Eastern Box Turtles
4) Rural Box Turtles in Missouri Have Bigger Home Ranges and Lower Corticosterone Levels Than Urban Ones
5) Box Turtles: Connecting People to Nature
6) Identifying Layers of Opportunity: How Box Turtles Can Help Promote Partnerships Which Include Foresters, Researchers, Developers, Educators, Conservationists, and the General Public.
7) Current Status of the Box Turtle Terrapene coahuila in the Wetlands of Cuatrocienegas Coahuila
8) Impact of Railroad Tracks on Box Turtles (and Other Species of Turtles)
9) Home Range, Habitat Use, and Mortality of Hatchling and Juvenile Ornate Box Turtles (Terrapene ornata ornata) in Iowa
10) A Population Study of the Eastern Box Turtle (Terrapene carolina) in the Piedmont of North Carolina
11) Experimental Challenge Study of Ranavirus Infection in Previously Infected Eastern Box Turtles (Terrapene carolina carolina) to Assess Immunity
12) Landscape Use and Movement of Three-toed Box Turtles (Terrapene carolina triunguis) During Drought Conditions
13) Rangewide Population Genetics of the Eastern Box Turtle Terrapene c. carolina
14) Ornate Box Turtles in the Sand Hills of Colorado: Population Structure and Behavioral Ecology
15) The Georgia Sea Turtle Center: Connecting Eastern Box Turtle Rehabilitation, Population Health and Disease Monitoring, Education and Research
16) Establishment of a Reference Interval for Fibrinogen in Ornate Box Turtles (Terrepene ornata ornata)
17) Under Fire: Responses of Eastern Box Turtles to Prescribed Burns
18)Response of the Eastern Box Turtle (Terrapene carolina carolina) to Silviculture Treatments in the Valley and Ridge Province of East Tennessee
19) Population Characteristics and Habitat Use of Ornate Box Turtles (Terrapene ornata) within Restored and Remnant Tallgrass Prairies
20) A Dollar a Turtle or How Your State Can Gather Long-term, Widespread Data on a Budget
21)Reintroduction Program Reveals Homing Behavior and a Previously Unknown Nesting Strategy in Terrapene ornata
22) The Impacts of Intraspecific Variation on Phylogenetic Resolution in Terrapene
23) Husbandry Techniques Used during a Ranavirus Outbreak in Eastern Box Turtles (Terrapene carolina carolina) at the Maryland Zoo in Baltimore
____________________________________________________
This Workshop is dedicated to the memory of 
Dr. Susan Hagood
(December 27, 1956 - November 8, 2011)
Dr. Susan Hagood, herpetologist and conservationist, was a founding member of the North American Box Turtle Conservation Committee and she played a key role in organizing the box turtle conservation workshops. This fourth workshop, held in March 2013 in Asheboro, North Carolina, is dedicated to her memory.

Susan was a tireless worker and spokesperson on behalf of wildlife welfare and conservation. For more than twenty years, she served as a Wildlife Specialist with the Humane Society of the United States. Her work at HSUS focused on wildlife protection, particularly turtles and tortoises. She became a leader in efforts to develop innovative ways for wildlife to safely cross or travel under roadways.

While working at HSUS, Susan earned her Ph.D. in Ecology from  the University of Maryland, College Park. Competed in 2009, her dissertation, was titled: “Genetic Differentiation of Selected Eastern Box Turtle Populations in Fragmented Habitats, and a Comparison of Road-based Mortality Rates to Population Size.”

Susan discovered that Maryland box turtle populations that were separated by as much as 50 or 60 miles, were very similar in their genetic makeup. She concluded that roads and highways, which are potential barriers to population mixing, had not been in place long enough for populations to differentiate genetically. Susan also found that the rate of road mortality was about 5% annually for local populations; a seemingly low rate, but nonetheless high enough to lead to steady, long-term population declines. Susan was often accompanied in the field by Drew, her chocolate Labrador retriever, who she had trained to find box turtles. Together they logged hundreds if not thousands of hours in fields and woods during her studies.

Susan was a joy and an inspiration to many. Susan was consistently optimistic and positive towards the conservation projects she headed up and even towards her adversaries. She was a driving force behind the successful effort to find and translocate hundreds of box turtles from the path of an 18-mile long highway that was under construction in Montgomery County, Maryland. This project demonstrated that it is feasible to catch and release chelonians that face imminent destruction resulting from a development project. Susan earned the respect and support of the State Highway Department and Department of Natural Resources in this effort and the project generated much positive media coverage.

Susan leaves behind a legacy and a body of scientific work that will be used by all of us who work to sustain and protect turtle populations.


____________________________________________________
 1) Epidemiology and Treatment of Ranaviral Disease in North American Chelonians
Matthew Allender,1,3 Mark A. Mitchell,1 and Sherry Cox2
1Department of Comparative Biosciences, University of Illinois, Urbana, IL
2Department of Comparative Medicine, University of Tennessee, Knoxville, TN
Ranaviruses have been proposed as a major threat to amphibian biodiversity, and the impact of these pathogens on reptiles is less well understood. A quantitative PCR was developed using TaqMan that was 100% efficient in detecting the major capsid protein of frog virus 3 (FV3) in turtle samples. The overall prevalence of Ranaviruses in a study of 606 Eastern box turtles (Terrapene carolina carolina) across the SE US was 1.3%, with a non-significantly higher prevalence in juveniles than adults. The low prevalence found in this population supports the theory that this virus is associated with acute disease/death. Subsequently, red-eared sliders (Trachemys scripta elegans) were challenged with FV3 and demonstrated higher mortality rates when maintained at 22°C than at 28°C, supporting the theory that this virus is less virulent at higher temperatures. FV3 DNA was detected in whole blood, oral swabs, and cloacal swabs. The sensitivity and specificity of detection in both whole blood samples and cloacal swabs when compared to necropsy was 100%, while the sensitivity and specificity in oral swabs was found to be 83% and 100%, respectively. Clinical signs observed in experimentally infected red-eared sliders included lethargy, conjunctivitis, oral plaques, oral ulcers, and injection site swelling, while those in box turtles were fractures and diarrhea. Treatment of ranavirus with anti-viral therapy has been reported to have variably poor success, but was based on anecdotal dosing recommendations. Pharmacokinetic analysis in box turtles of a single oral dose of valcyclovir demonstrated measureable levels, and may prove useful against this virus.
2) Development of a Health Screening Protocol and Incorporating it into an Existing Field Project
Matthew Allender
Department of Comparative Biosciences, University of Illinois, Urbana, IL
Outbreaks and potential population declines due to disease or health related factors have been increasingly reported. The cause and progression of many of these diseases are unknown and relate to the lack of baseline knowledge of populations. Therefore, collecting baseline information on routine health parameters might be critical to identifying emerging threats and response to disease outbreaks. Several diagnostic modalities are available and might be quite confusing. A brief overview and interpretation of hematology, plasma biochemistries, disease epidemiology, serology, protein electrophoresis, and toxicology will be presented.
3) Survivorship and Home Range of Head-started Juvenile Eastern Box Turtles
Kimberly M. Andrews1,2,3, Joseph E. Colbert1, and Terry M. Norton1
1Jekyll Island Authority Georgia Sea Turtle Center, Jekyll Island, GA
2University of Georgia Savannah River Ecology Laboratory, Aiken, SC
Urbanization is increasing rapidly resulting in growing rates of human-wildlife interactions that are detrimental to native animals. Eastern box turtles, Terrapene carolina, are a priority example of species experiencing declines throughout their range. Effects from habitat loss, road mortality, injury by domestic pets, collection for the pet trade, and reduced health in wild populations have all been documented. Displaced, injured, and deceased animals are periodically admitted to the Jekyll Island State Park Authority’s Georgia Sea Turtle Center (GSTC) on Jekyll Island, GA. Eggs are recovered from injured and freshly-dead females and subsequently incubated in the hospital. Additionally, individuals that cannot be rehabilitated to the point of wild release are placed in our outdoor educational display where they reproduce with other captive adults. Eggs are allowed to incubate in situ in the display. Hatchlings are head-started at the GSTC until their second year and at least 130 grams in order to reduce the risk of predation upon release into the wild. Using health assessment and radio telemetry methodologies, we evaluate survivorship, growth, habitat use, and movement patterns in the wild. This assessment can be used to guide future rehabilitation and management priorities in determining whether head-started individuals can contribute to the viability of local populations and therefore, whether head-starting is an appropriate and responsible technology in these situations. To date, all nine juveniles have survived and continue to exhibit positive growth rates. Home range sizes are 2.49 ha on average but substantial intraspecific variation has been observed. We will additionally present future direction in research on Jekyll’s resident box turtle population and the continued monitoring of head-started juveniles.
4) Rural Box Turtles in Missouri Have Bigger Home Ranges and Lower Corticosterone Levels Than Urban Ones
Stephen Blake1,2,7, Corinne Kozlowski3, Jenny Fung4, Joanna Wang5, Sharon L. Deem6
1 Max Planck Institute for Ornithology in Radolfzell, Radolfzell, Germany
2Current Address: Whitney R. Harris World Ecology Center, Washington University in St. Louis, St. Louis, MO
3Research Dept., St. Louis Zoo, St. Louis, MO
4Biology Program, Washington University in St. Louis, St. Louis, MO
5Environmental Biology Program, Washington University in St. Louis, St. Louis, MO
6Institute for Conservation Medicine, St. Louis Zoo, St. Louis, MO
Box turtles were once ubiquitous through much of the eastern and central United States, but are now in steep decline due to a combination of habitat loss and fragmentation, road kill, disease and the live turtle trade for food and pets. Despite Archie Carr’s assertion that “everyone likes box turtles”, lack of understanding of these threats means that the public remains largely unaware of this conservation crisis. Since 1) populations of box turtles can occur close to urban centers, and 2) there is an urgent need for better data on trends in ranging, health, and demography of box turtles in relation to human activities, an opportunity exists to combine research, management and hands-on outreach in a unified conservation framework.
In 2012 we initiated a pilot project in St. Louis, Missouri, to quantify home range size and health status of box turtle populations in urban and rural conditions, generate a meaningful local outreach program, and assess the options for developing a long term conservation initiative. The project took place in Forest Park, in St. Louis city, and the Tyson Research Center, a rural site some 20km from the city limits. Forest Park is a 556ha mosaic of woodland fragments embedded in a traditional city parkscape of golf courses, playing fields, lakes, and other recreation areas traversed by roads and bike trails. Tyson Research Center is an 809ha fenced, protected oak-hickory forest block bisected by one tarmacked road and a network of tracks. We fitted VHF tags to ten adult turtles at each site and regularly recorded their locations using GPS through the summer until brumation. We completed biophysical exams and took blood samples from tagged individuals and a further 63 turtles at the two sites.
The mean home range (MCP) of urban turtles ( was two orders of magnitude smaller than that of rural turtles ( . The mean blood corticosterone level of urban turtles was over twice that of rural turtles .
We currently have no causal explanations for these results, though likely reasons include that the small forest fragments of Forest Park may limit turtle movements compared to the larger rural habitat which increases physiological stress. Moreover, the heavy human use of Forest Park may lead to high levels of disturbance for these urban turtles.
Outreach activities included introducing several groups of local school children to the turtles of Forest Park, and integrating undergraduates and high school students as part time research technicians. The level of interest and enthusiasm of all participants was extremely high, and resulted in a series of posters, promotional videos, and other outreach products.
Our pilot study revealed that 1) while large urban parks can support box turtle populations, their ecology and health may be compromised compared to rural populations, and that 2) studies of urban box turtle populations and those close to large cities and towns provide exciting opportunities for outreach to local people with potentially high conservation and societal impacts.
5) Box Turtles: Connecting People to Nature
Kimberly Burge
NC Wildlife Resources Commission, Centennial Campus Center for Wildlife Education, Raleigh, NC
The Centennial Campus Center for Wildlife Education is located in Raleigh, N.C., and is the piedmont region education center run by the N.C. Wildlife Resources Commission. Since July 2007 our program participants have been using radio telemetry to track and monitor a population of wild box turtles in Lake Raleigh Woods, a 90 + acre forest on North Carolina State University’s Centennial Campus. We became the first monitoring study site for the North Carolina Box Turtle Collaborative’s long term box turtle mark-recapture project started in 2008. We have incorporated the radio telemetry, mark-recapture studies, and mark-recapture studies with turtle tracking dogs in our regular programing at our education center with great success. Our audience is a very urban population with some fears associated with woodland exploration. Box turtles are charismatic and usually well liked which makes them excellent ambassadors during our outdoor program. The box turtles provide a connection with nature and at the same time offer the opportunity for real-life data collection, citizen science projects, and hands-on technology use. This presentation will go over our project history, how we have integrated the research into our educational programming, challenges and obstacles we’ve encountered along the way, and also the benefits and rewards.
6) Identifying Layers of Opportunity: How Box Turtles Can Help Promote Partnerships Which Include Foresters, Researchers, Developers, Educators, Conservationists, and the General Public.
John Byrd1, the CRESO Research Team, and the CRESO Education/Research Team
Clinch River Environmental Studies Organization (CRESO), Clinton, TN
As we exited our 11ha timber harvest study site, we were sternly, but politely, confronted by the developer of the subdivision adjacent to the harvest zone. He shared with us his displeasure on the difficulty of selling houses located next to an unsightly “mess of cut trees.” So there we stood at the boundary of two heavily impacted landscapes – on one site, trees would return through a slow successional process, while on the other, trees would be rapidly transformed into houses. We were struck by the fact that our Box Turtle study and its focus on research questions (survivorship, movement patterns etc.) had narrowed our thinking and resulted in overlooking important educational and partnership possibilities. It is not uncommon for government agencies to provide stakeholders a format for discussion. Unfortunately, the reality of this input design is often politically motivated. Providing “layers of opportunity” for stakeholders – landowners, developers, researchers, foresters, educators, etc. –impacted by forestry or development practices requires recognition of what the potential wins are for all parties. Ideas for using Box Turtles –the poster critter of forest and field–to develop partnerships, and promote positive landscape options and educational strategies will be presented.
7) Current Status of the Box Turtle Terrapene coahuila in the Wetlands of Cuatrocienegas Coahuila
Juan Gamaliel Castañeda Gaytán1,3, Jorge E. Becerra López2, Alejandra Cueto Mares1, Sara Valenzuela Ceballos1 and Ivonne Salas Westphal1
1Departamento de Ciencias Biológicas, Universidad Juárez del Estado de Durango, Durango, Mexico
2Laboratorio de Sistemática Molecular del Centro de Investigaciones Biológicas Universidad Autónoma del Estado de Hidalgo, Hildago, Mexico
The box turtle (Terrapene coahuila) is the flagship species of the conservation area, Cuatrocienegas, Coahuila. Its importance is that it is one of the most charismatic semi-aquatic vertebrates of the valley that are basis of numerous conservation efforts due to their association with water bodies. Thus, it is considered a potential species to assess the health of wetlands and therefore identify whether conservation practices are effective today. As part of compliance with the National Commission of Natural Protected Areas (CONANP) the Mexican government has invested resources to meet the current situation of the box turtle and then implement a simple method to maintain a constant monitoring of the populations of this seriously threatened and endangered species. From June 2010 to May 2011, we assessed an area of 30 hectares 4 or 5 days per month. In addition, we selected 143 sites distributed throughout the valley to search for live turtles or traces of turtle and evaluate their potential distribution using ecological niche models (MAXENT). For sites where live specimens were observed, data were taken as to the type of microhabitat and water (if observed within the pools). All specimens were marked with notches in the shell. The general analysis of the population describes the proportion of individuals recorded per unit area, the proportion of age classes, sex and habitat use. Additionally, radio tags were placed on the carapace of 14 male and female T. coahuila to assess the areas of activity during two different periods of the years 2011 and 2012. Preliminary results point out that the species is found in wetlands in the valley with a high degree of fragmentation. There is a high probability of box turtles occurring within an area of 1.5 km sq. (intensive sampling site). Over a larger area of slightly greater than 7 km sq, the probability of box turtles occurring drops to 51%. Within the intensive sampling site, we collected approximately 95 individuals during the first year of study with a sex ratio male: female 1:0.53. Extrapolating the abundance of individuals per unit area of search yielded an approximate abundance of 3.16 ind / ha, which is a very small population for the entire valley (potentially 461 to 2268 turtles). In most cases, turtles were observed in wetlands with a depth less than 50 cm. In the case of individuals observed on land, most were located near plants of the genus Eleocharis, Scirpus, and Distichlis (species associated with permanent water film). The months with the greatest possibility of seeing active turtles were recorded. Area of activity, has been estimated at 3 hectares for females and 5 hectares for males. These findings suggest a use of space that is higher than previously reported by Brown (1968) for this species. We are currently developing a monitoring program based on information obtained during the last two years of study and which will provide answers to major questions about why and where we should sample the box turtle population in the Valley of Cuatrocienegas to encourage its conservation
8) Impact of Railroad Tracks on Box Turtles (and Other Species of Turtles)
Christian A. d’Orgeix1,2,5, J.C. Mitchell3, and T. Laxson4
1National Evolutionary Synthesis Center, Durham, NC
2Current address: Department of Biology, Virginia State University, Petersburg, VA
3Mitchell Ecological Research Service, LLC, High Springs, FL
4USGS Gap Analysis Program, Moscow, ID
Railroads represent the largest terrestrial transportation infrastructure system in the U.S., after roads. Reports of railroad track impacts on turtles are mainly anecdotal, e.g., posing as barriers to seasonal movement between habitats, interfering with female nest site selection, and mortality of turtles trapped between the rails. We conducted weekly turtle censuses May - August 2005 and May - November 2006 using two 1.65 km sections of railroad track in Prince George County, Virginia to address the following questions: (1) What percentage of turtles encountered were box turtles? (2) Do railroad tracks contribute to box turtle mortality? (3) Are differences in sex accurate predictors of which turtles will be impacted by railroad tracks? (4) How do box turtles become trapped between the tracks? In addition, as a first step in accessing the potential global impacts of railroad tracks on turtles and the broad conservation problems associated with this transportation system, we used Geographic Information System (GIS) data to estimate how many turtles might be impacted by railroad tracks in the State of Virginia, USA over a similar period to our study. We found 38 turtles comprising seven species with eastern box turtles, Terrapene carolina carolina, composing 58% of all turtles encountered. Approximately equal numbers of turtles were found between the tracks and on the outside of the tracks, however, the mortality rate was 83% for turtles found between the tracks. There were no significant differences between male and female sex ratios in T. c. carolina. For turtles to access to our census area, tracks through railroad crossings would require moving 1-2.5 km distance, suggesting that the two primary means of accessing railroad tracks were climbing over the rails or passing under the rails through gaps. We estimate that thousands of turtles would suffer mortality or interference with movement to nesting sites or seasonal habitat shifts along Virginia's 7,450 km of railroad tracks.
9) Home Range, Habitat Use, and Mortality of Hatchling and Juvenile Ornate Box Turtles (Terrapene ornata ornata) in Iowa
 Rachel Hannah Fendrich
Cornell College, Mount Vernon, IA
 The ornate box turtle, Terrapene ornata ornata, is threatened in Iowa, as the species’ prairie habitat has mostly been converted into agricultural land, reducing and fragmenting natural habitat, increasing contact with motor vehicles, farm implements, and most likely with mesopredators like raccoons, which thrive around humans and edge habitats. Our long-term project strives to understand the ecology and natural history of the second largest population of T. ornata in Iowa and provide information to management agencies to help develop conservation plans for this species. Perhaps the largest gap in our knowledge of ornate box turtles pertains to the biology of juveniles or hatchlings, as they are both uncommon and secretive. With the advent of miniature radio telemetry transmitters, we have begun monitoring hatchling and juvenile turtles to obtain data on home range, habitat, and microhabitat. I will discuss the results from our first two years of efforts at tracking hatchlings and juveniles. We tracked 19 adult turtles, 12 hatchling turtles, and 9 juvenile turtles in the Hawkeye Wildlife Area in Johnson County, Iowa, during the summers of 2011 and 2012 using radio telemetry. We recorded each turtle’s GPS coordinates daily and calculated their home ranges using 95% fixed kernels and minimum convex polygons. In addition, we gathered data on mortality, habitat, and microhabitat used by these turtles.
Mean adult weekly and monthly home ranges were consistently larger than weekly and monthly juvenile and hatchling home ranges. Juvenile home ranges were not significantly different than hatchling home ranges.
Habitat and microhabitat preferences varied based on age. Juveniles were far more likely than either adults or hatchlings to be found in a forest habitat, while hatchlings were more likely to be found in prairie. No age group spent more time in agricultural fields, even though the nesting area was within 100 meters of agriculture. Hatchlings were far more likely to be found buried than were juveniles or adults. While our sample remains small in both number of individuals and duration of individual observations for hatchlings, it appears that compared to older turtles, hatchlings have smaller home ranges and use fewer different habitat types. This may simply reflect reduced movement compared to older turtles. If they do not move far from the nesting site in the first month after emerging, they will encounter mostly prairie habitat. The reduced movement may reflect simple size scaling of movements or a greater need to remain underground to avoid predators or high daytime temperatures.
During 2011, one hatchling died from human causes; during the summer of 2012, all seven of our hatchlings were preyed upon within two months of the time we began tracking them. While it remains unclear whether predation on nests, hatchlings and juveniles is artificially high at our site, predation on early life stages is substantial; our data suggest that the nesting site and the immediate surrounding area might be an appropriate and cost-effective focus for conservation measures.
10) A Population Study of the Eastern Box Turtle (Terrapene carolina) in the Piedmont of North Carolina
 John D. Groves1 and Jessica Foti
 North Carolina Zoological Park, Asheboro, NC
Population studies are important to understanding the dynamics of declining Box Turtle populations throughout their range, yet few recent long- term studies are available for comparison in different parts of this turtle’s extensive historical distribution throughout North America. In order to contribute population information on a basically undisturbed protected population in the Piedmont of North Carolina we conducted a mark/recapture study from 2002 - 2011 to establish baseline information. A total of 503 turtles were captured and marked and 51% of these turtles were recaptured between one and three times during the study period. We examined meristic characters, body condition, color patterns, activity patterns, and population structure. Some information on reproduction and mortality in this population will be discussed. Turtles in this population are active during all seasons with normal activity patterns between April and November. Activity patterns appear to be similar between males and females, with the height of activity in the summer months. More turtle activity is correlated with some rain activity within a 48 hr. period. Approximately one third of the population was active when no rain event occurred and two-thirds were active during rain events. Box Turtles are more active between 21-32 º C. Box Turtles are active throughout daylight hours during their active periods. Most turtles are found between 0800 until 1500 each day of activity. There is a steady decline of Box Turtles activity in the afternoon. Population size is estimated to be between 800-1000 turtles on 500 acres. Density of this population is approximately 1 turtle per acre. Age structure between the sexes appears to be similar for this population of Box turtles. Age estimates are between hatchlings and 50 years of age, with the majority of the population between 10-30 years. Home range of several turtles were investigated by radio telemetry. Home range sizes ranged from one acre in a disturbed area to 11 acres in an undisturbed area. This population had very few health concerns, with shell damage by predators being the most important concern. Information gained from this baseline study should be repeated periodically in the future to monitor trends and health of this population.
11) Experimental Challenge Study of Ranavirus Infection in Previously Infected Eastern Box Turtles (Terrapene carolina carolina) to Assess Immunity
Jennifer C. Hausmann1,8, Allison N. Wack1, Matthew C. Allender2, Michael R. Cranfield1, Kevin J. Murphy3, Kevin Barrett4, Jennell L. Romero5, James F. Wellehan6, Chris Zink7, and Ellen Bronson1
1Medical Department, The Maryland Zoo in Baltimore, Baltimore, MD
2Department of Comparative Biosciences, University of Illinois, Urbana, IL
3Animal Department, The Philadelphia Zoo, Philadelphia, PA
4Animal Department, The Maryland Zoo in Baltimore, Baltimore, MD
5Comparative Pathology, School of Medicine, Johns Hopkins University, Baltimore, MD
6Zoological Medicine Service, University of Florida College of Veterinary Medicine, Gainesville, FL
7Comparative Medicine Retrovirus Bio Laboratory, School of Medicine, Johns Hopkins University, Baltimore, MD
The Maryland Zoo in Baltimore lost 13 of 27 captive Eastern box turtles (Terrapene carolina carolina) to an outbreak of Ranavirus (100% homology to 531 bp segment of FV3 MCP) during the summer of 2011. To assess survival and shedding post-infection, an experimental challenge study was performed, in which the surviving, previously infected turtles were re-infected with the outbreak strain of Ranavirus that was harvested in terrapene heart cell lines (TH-1). Seven turtles were inoculated with a predetermined dose of infectious virus IM and four control turtles were injected with an equal volume of saline IM. The turtles were monitored for 9 weeks with blood and oral swabs collected for PCR and antibody testing. During that time only one of the seven (14.3%) inoculated turtles and none of the controls (0%) died; there was no significant difference in survival. All clinical signs seen in the inoculated turtles, except for the turtle that died, were very mild (lethargy, weight loss, oral ulcers, sublingual swelling, skin excoriations, ocular discharge and periocular swelling) when compared to the severe clinical signs shown by these turtles during the previous outbreak. The inoculated turtle that died showed intracytoplasmic inclusion bodies in the kidney, lungs, pancreas, liver, and vas deferens; vasculitis in the spleen, pancreas, lungs, and liver; nephritis; pnuemonia; esophagitis; hepatitis; and enteritis. Five of the surviving ten turtles were euthanized at the end of the study for histopathology and PCR testing. The remaining five turtles began normal brumation.
12) Landscape Use and Movement of Three-toed Box Turtles (Terrapene carolina triunguis) During Drought Conditions
Lynne W. Hooper1 and Dylan C. Kesler
Department of Fisheries and Wildlife Sciences, University of Missouri, Columbia, MO
Adult Three-toed box turtles (Terrapene carolina triunguis) were captured in Thomas S. Baskett Wildlife Research and Education Area, in Boone County, Missouri. The variable topography of the study area includes a floodplain of Brushy Creek bounded by a steep rock cliff on the south and a series of ridges to the north. Turtles were captured for study during visual surveys, and weighed, measured, sexed, and fitted with radio telemetry transmitters and temperature loggers. Temperature loggers were also distributed throughout the study area to measure ambient conditions. Four female and 6 male turtles were tracked daily for 30 days, beginning on 2 July 2012. Average regional temperatures during the month of July were the highest on record, exceeding the normal value by 4.5°C, and precipitation for July was 8.61 cm below normal. We used generalized linear mixed models to evaluate relationships between turtle temperatures, movement rates, the position of the turtle on the landscape, and ambient climate conditions. Results indicated that turtles were cooler than nearby ambient conditions (P < 0.0001), and that daily fluctuation in turtle temperatures was less than fluctuation in ambient temperatures (P < 0.0001). Results further illustrated a positive relationship between daily low relative humidity and the distance of study subjects to a nearby ephemeral stream (P = 0.0336). Further, the daily rate of movement (m/d) was positively associated with ambient relative humidity and negatively associated with daily mean ambient temperature (P < 0.0001 for both). We found no association between the elevation of turtles and temperature or humidity, and there was no difference in the movement rates of males and females. Movement rates, turtle temperatures, and proximity to the ephemeral stream were all associated with climate conditions, which supports previous findings that turtles use behavioral mechanisms to actively manage internal thermal conditions. Even during climatic conditions of high temperatures and drought, Three-toed box turtles were able to maintain lower temperatures and avoid evaporative water loss.
13) Rangewide Population Genetics of the Eastern Box Turtle Terrapene c. carolina
Kimble, Steven J.A.1,3, O.E. Rhodes Jr2, and Rod N. Williams1
1Department of Forestry and Natural Resources, Purdue University, West Lafayette, IN
2Savannah River Ecology Laboratory, Aiken, SC
The eastern box turtle Terrapene c. carolina is experiencing population declines across its range. The genetic consequences of such declines vary considerably, especially in long-lived taxa such as Testudines. For example, declines in census numbers may reduce the genetic diversity available for a species to evolve responses to novel challenges such as climate change or emerging diseases. Genetic considerations are also paramount in designing conservation management practices such as head starting or transplantation. To address these gaps in knowledge we collected samples from approximately 1550 individuals from the full range of the species and genotyped them at eleven highly polymorphic microsatellite loci. Genetic diversity is high (mean observed heterozygosity = 0.756) and our data suggested the species is defined by two genetic populations. We sampled intensely (n ≈ 600) within two contiguous state forests in Indiana and found that mean pairwise relatedness among individuals was low (mean relatedness = 0.002), suggesting high dispersal in this species and low chance of inbreeding depression. The low level of population structure also suggests high gene flow, a surprising result in light of multiple studies demonstrating low vagility among most adults. These conclusions should be interpreted cautiously, however, as the genetic signatures observed may be biased by sampling of mainly adults, and may therefore represent a historical signal rather than contemporary genetic processes.

14) Ornate Box Turtles in the Sand Hills of Colorado: Population Structure and Behavioral Ecology
Ann-Elizabeth Nash1,5, Caitlin Wilhelm2, Graham Dawson3, and Jason F. Martin4
1Department of Ecology and Evolutionary Biology, University of Colorado, Boulder, CO
2 Department of Biology, Missouri State University, Springfield, MO
3Department of Biology, Skidmore College, Saratoga Springs, NY
4Zoology Program, Colorado State University, Ft Collins, CO
For the past 6 years, beginning in 2007, the Colorado Box Turtle Project (CoBTP) has studied a population of Terrapene ornata ornata on ~50 hectares of sand hills habitat on the Eastern Plains of Colorado. The site is a human disturbed cattle ranch with numerous gas wells, all of which were hydraulically fractured in 2011. At least 160 turtles use the site (density = 3.18 turtles/ha) consisting of 44% females, 35% males, and 21% juveniles with the population skewed in favor of females. A shell damage scale was created, with females experiencing 22% less shell damage than males. Plant surveys were conducted identifying more than 100 species with two internal sites compared using the Jaccard diversity index.
 Since 2010, 10 box turtles (6 females and 4 males) have been tracked with radio telemetry, located three times per week from mid-May through August, and intermittently from September through December. GPS locations were recorded to show turtle movements and understand home range size. Ambient temperature and humidity loggers were used in 2011 to determine preferred ranges of turtles at this site. Most turtles are active mid-April through mid-September depending on rainfall, spending the winter hibernating underground, primarily using rodent burrows. Tracking data reveals high site fidelity to hibernacula location as well as preferred locations during summer months. Females move to preferred nesting locations and return to favored foraging/living areas; hibernacula sites are usually a third different location. Dietary items include various beetles, White-lined sphinx (Hyles lineata) larva, cactus (Opuntia spp.), prairie fameflower (Talinum parviflorum), and spiderwort (Tradescantia occidentalis). In contrast to the Nebraska sand hills at Crescent Lake National Wildlife Refuge, Colorado T. o. ornata habitat is waterless, making plants the likely hydration source for turtles.
Since the State of Colorado presently allows the private take of ornate turtles for personal use, the Colorado Box Turtle Project established goals of analyzing the regional ecology of this species and enhancing its protections. This study contributes to the knowledge of a widespread and once-common species, never studied before in Colorado.
15) The Georgia Sea Turtle Center: Connecting Eastern Box Turtle Rehabilitation, Population Health and Disease Monitoring, Education and Research
Terry M. Norton1, Michelle Kaylor, Steven Nelsen, Amy Hupp, Rachael Thomas, Kimberly Andrews
Georgia Sea Turtle Center, Jekyll Island Authority, Jekyll Island, GA
Over the last 5 years there have been tremendous advances in our knowledge about health and infectious disease in chelonians including T. carolina (Allender et al., 2011, Johnson et al., 2007, 2008, Feldman et al., 2006, and Alverez et al., 2012). A Ranavirus, Frog Virus 3, is known to affect Terrapene populations. Other infectious agents include a Mycoplasma sp. (distinct from other species but as yet unnamed) associated with upper respiratory tract disease, an adenovirus (a member of a novel genus) associated with enterohepatic disease, and a novel alphaherpesvirus which has been found concurrently with ranaviral disease. The agent of intranuclear coccidiosis, a significant chelonian pathogen, has recently been identified in T. carolina.
The Georgia Sea Turtle Center (GSTC) is assessing the health of resident box turtles on Jekyll Island and those brought to the GSTC for rehabilitation. This assessment is guiding future rehabilitation and management priorities in determining whether offspring of displaced and injured individuals can be established in the wild and potentially contribute to the viability of local populations. Additionally, rehabilitation and release protocols are being established based on information generated from this assessment.
Over a half a million people have had the opportunity to learn about chelonians though the interactive educational exhibits and programs at the GSTC since its opening over 5 years ago. Educators present medical updates on patients, but more importantly discuss the population effects of the various threats on turtle population survival and how the average person can help. An exhibit dedicated to box turtle conservation has been developed and is extremely popular with visitors.
Allender, M. C., Abd-eldaim, m., Schumacher, J., Mcruer, D., Christian, L.S., and Kennedy, M. 2011. PCR prevalence of Ranavirus in free-ranging eastern box turtles (Terrapene carolina carolina) in rehabilitation centers in three southeastern US states. Journal of Wildlife Diseases 47: 759–764.
Alvarez, W.A, Gibbons, P.M., Rivera, S., Archer, L.L., Childress, A.L., and Wellehan, J.F.X. 2012. Development of a quantitative PCR for rapid and sensitive diagnosis of intranuclear coccidiosis in tortoises, and identification in the critically endangered Arakan forest turtle (Heosemys depressa). Proceedings, 2012 Conference of the Association of Reptilian and Amphibian Veterinarians.
Feldman, S. H., Wimsatt, J., Marchang, R.E., Johnson, A.J., Brown, W., Mitchell, J.C., and Sleeman, J.M. 2006. A novel mycoplasma detected in association with upper respiratory disease syndrome in free-ranging eastern box turtles (Terrapene carolina carolina) in Virginia. Journal of Wildlife Diseases 42: 279–289.
Johnson, A.J., Origgi, F.C., and Wellehan, J.F.X. 2007. Molecular diagnostics. In Infectious diseases and pathology of reptiles, E. R. Jacobson (ed.). CRC Press, Boca Raton, Florida, pp. 351–380.
Johnson, A.J., pessier, a.p., wellehan,j.f.x., childress,a., norton, t.m., stedman,n.l., bloom, d.c., belzer,w., titus, v.r., wagner, r., brooks, j.w., spratt,j., and jacobson, e.r. 2008. Ranavirus infection of free-ranging and captive box turtles and tortoises in the United States. Journal of Wildlife Diseases 44: 851–863.
16) Establishment of a Reference Interval for Fibrinogen in Ornate Box Turtles (Terrepene ornata ornata)
Lily Parkinson
Colorado State University College of Veterinary Medicine and Biomedical Sciences, Fort Collins, CO
The ornate box turtle (Terrapene ornata ornata) is currently listed as “near threatened” by the International Union for Conservation of Nature. This status necessitates further study to fully understand this species’ ecology. In order to gain the fullest understanding of how to best to protect these turtle populations, researchers would be greatly aided by a method for determining the levels of inflammation in different turtle populations. This information could provide vital clues to what environmental variables most negatively affect the lives of ornate box turtles. One potential test for determining the presence of inflammatory disease in box turtles could be blood fibrinogen concentration. Blood fibrinogen levels have been shown to correlate with inflammation in several species, and could serve as a reliable indicator for inflammation in box turtles as well. Before such studies can occur, however, a normal reference interval for fibrinogen in ornate box turtles must be established. This study aims to produce a normal reference interval in box turtles through the sampling of at least 40 ornate box turtles. Currently, 24 box turtles have been enrolled in the study, and the preliminary data provides two possible interpretations. One interpretation could indicate that ornate box turtles have a relatively narrow range of fibrinogen levels, and that the few outliers observed so far were due to stress that was missed in the study’s health screening. An alternative interpretation of the data could indicate that all screened healthy box turtles provided a normal fibrinogen level for the study, but the data is not yet sufficient to confirm that the fibrinogen levels at the high end of the normal range are not outliers. The continuation of this study aims to elucidate which interpretation is most correct. The researchers also hope to provide a basis for future studies into fibrinogen as an indicator for inflammation in ornate box turtles as well as other turtle species.
 17) Under Fire: Responses of Eastern Box Turtles to Prescribed Burns
John H. Roe
University of North Carolina at Pembroke, Pembroke, NC
Prescribed fire is an essential tool for the conservation and management of longleaf ecosystems, and it is thus widely employed in protected areas of the Southeastern United States. However, such management may unintentionally overlook impacts to other non-target species that are also of conservation concern, presenting land managers with conflicts. Using radiotelemetry, temperature dataloggers, and capture-mark-recapture, we are currently studying the vital rates and seasonal behavioral responses of Eastern Box Turtles, Terrapene carolina, to prescribed fire in the North Carolina Sandhills. This study is being conducted in the Weymouth Woods Natural Area (WEWO), a primarily xeric habitat where burns have been used to manage longleaf forests for three decades, and in the nearby Lumber River State Park (LRSP), a bottomland habitat where prescribed burns are not employed. We expect that turtles inhabiting areas with frequent burns (WEWO) would largely avoid areas and habitat types subject to fire, and if not, they would suffer increased mortality, injury incidence, and other reduced vital rates relative to those where fire is not employed (LRSP). Turtle activity centers at both sites have been in more mesic microhabitats, and indeed the majority of locations have been in close proximity to watercourses or in hardwood forests outside of the burn management units. However, turtles at WEWO have used upland longleaf forests of all post-fire intervals, including one that was severely burned, with several entering burned habitats within weeks after fire where we observed foraging, thermoregulatory, and reproductive behaviors. Two turtles at WEWO are suspected to have died or been severely injured by predators, while no turtles have been lost from the LRSP. Turtles at both sites have moved extensively, exhibited similar seasonal activity, and maintained positive growth rates. While only preliminary, our findings yield insight into the behavioral and population responses of turtles to fire, thus informing burn managers of the risks to non-target animals that are also of management priority.
18) Response of the Eastern Box Turtle (Terrapene carolina carolina) to Silviculture Treatments in the Valley and Ridge Province of East Tennessee
John Rucker1, Leah Lavoie1, Kristie Fox1, Matthew Allender2, John Byrd1,3, and the CRESO Research Team1
1Clinch River Environmental Studies Organization (CRESO), Clinton, TN
2Department of Comparative Biosciences, University of Illinois, Urbana, IL
The potential impacts of different forestry management options on the Eastern Box Turtle (Terrapene carolina carolina) are uncertain. A regeneration timber harvest at the University of Tennessee Forestry Resources Research Center in Oak Ridge, Tennessee provided an opportunity to study effects of this practice on a box turtle population. Steep terrain combined with impenetrable vegetation (mainly downed tree tops) made traditional search techniques more problematic than normal for estimating turtle density. From 2006-2012, trained Boykin Spaniels were employed to find box turtles in an oak-hickory hardwood forest subject to different silviculture objectives. From 2006-2007 we captured 78 individual box turtles in an11ha pre-harvest site and 230 individual turtles in a 64 ha area adjacent to the harvest site. Thirty three of the 78 individuals (42%) were recaptured post- harvest (2008-2012), and 126 turtles (55%) were recaptured from the 64 ha. In both cases the majority of turtles (32% vs. 30%) were recaptured during 2008, the year after the timber harvest. Although there was no pre-harvest capture data for a 13 ha forest stand subject to partial harvesting techniques in 2004-05, a high density of turtles was found there during the seven year study.

19) Population Characteristics and Habitat Use of Ornate Box Turtles (Terrapene ornata) within Restored and Remnant Tallgrass Prairies
Kimberly E. Schmidt1 and Eric C. Hellgren
Cooperative Wildlife Research Laboratory, Department of Zoology, Southern Illinois University Carbondale, Carbondale, IL
The ornate box turtle (Terrapene ornata) is an integral component of tallgrass prairie ecosystems. This turtle species has been understudied in Illinois, where it is classified as state-threatened. Our objectives were to assess microhabitat selection and characterize demography of T. ornata at two sites in northwestern Illinois. We also describe critical habitat used by transmitted individuals for overwintering, aestivation, and nesting. From June 2011 to October 2012, we captured 111 individuals (34F, 46M, and 31U) via foot surveys, opportunistic encounters, road cruising, and the use of detector dogs. Capture data between sites suggests variation in sex ratios between our 2 study populations. One population was composed of approximately equal numbers of males and female turtles (17F, 16M, X2=0.03, p=0.862), and the other population contained nearly twice as many male turtles (17F, 30M, X2=3.596, p=0.058). We affixed radiotransmitters to 34 adults (14F, 20M) for monitoring in 2012, and collected microhabitat data at approximately 1000 paired turtle and random sites. These data are currently being analyzed to determine key microsite characteristics associated with habitat selection by T. ornata. Eight turtles were tracked to their overwintering locations in 2011 (3F, 5M), and 28 animals (13F, 15M) were followed to overwintering locations in 2012. Overwintering locations of 5 animals (2F, 3M) could be compared across both years. Three turtles (2F, 1M) displayed site fidelity (2-4 m from the previous overwintering location). Of the 34 transmittered turtles, there is only one suspected depredation event (1F). Five additional transmitters were lost by becoming detached from the turtles’ shells during brummation (2M) and transmitter malfunction (3M). Turtles are using remnant prairie for nesting and overwintering habitat. Early spring phenology and prolonged drought in 2012 may have encouraged early emergence, hindered turtle movements, and encouraged early aestivation. Nesting was observed in late May 2012 on a sand road on the preserve. Depredated carcasses and road-kill mortalities occurred during the late summer and fall months when turtles were active. Conservation managers may use these data to maintain and restore suitable habitat for ornate box turtles across their Midwestern range.
20) A Dollar a Turtle or How Your State Can Gather Long-term, Widespread Data on a Budget
Ann Berry Somers1,4, Gabrielle Graeter2, John D. Groves3
1Biology Department, UNC Greensboro, Greensboro, NC
2North Carolina Wildlife Resources Commission, Asheville, NC
3Animal Section, North Carolina Zoological Park, Asheboro, NC
The Box Turtle Collaborative (Collaborative) is a think tank for conservation and education centered on box turtles. The group consists of educators and biologists representing four North Carolina institutions of higher education and five state agencies. The Box Turtle Connection project (BTC) is an initiative launched by the Collaborative in 2008 to educate and engage citizens in box turtle science. The project offers opportunities for participants to become involved either by casually reporting observations to the Carolina Herp Atlas or by making a long term commitment as volunteer Project Leaders. Project Leaders are required to attend training sessions, mark turtles at their designated site using a standardized protocol, and annually enter data into a state-maintained, password-protected database. Currently there are ~ 2200 data entries with 1700 unique turtles from 30 sites, many of them state parks. Over 8,000 BTC volunteer hours were submitted by NC Wildlife Resources Commission as match for State Wildlife Grants last year. The cash outlay on the project has been minimal and the interest in the project continues to increase. Although still in the fledgling stage, the BTC can be a model for other states interested in a low-cost, recession-proof means of gathering widespread, scientifically accurate data on a presumably declining species. Analysis of turtle data begins in 2013.
21)Reintroduction Program Reveals Homing Behavior and a Previously Unknown Nesting Strategy in Terrapene ornata
Charles R. Tucker1,4,, Jeramie T. Strickland2, and Day B. Ligon3
1Department of Biology, Missouri State University, Springfield, MO
2US Fish and Wildlife Service, Thomson, IL
3Department of Biology, Missouri State University, Springfield, MO
A long term research and conservation program was initiated in 2008 at the Upper Mississippi River National Wildlife and Fish Refuge in northwestern Illinois. Because ornate box turtles (Terrapene ornata) are threatened in Illinois, our initial goals were to determine the species’ population status on seven local prairies and to assess the feasibility of a reintroduction program. Searches using trained dogs confirmed box turtle presence at six of seven prairies, one of which was not previously known to support box turtles. A reintroduction program was initiated in which eggs were collected from a donor population and head-started for one year before release at a former military depot from which box turtles had been extirpated. Each year, half of the head-started turtles were released inside an enclosure at the former military depot and half were released at the donor site. When nests were located, eggs from some nests were collected for the head-start program and temperature data loggers were placed into other nests that were allowed to incubate naturally. During nest searches, some nesting females were found to be completely underground during nesting. This behavior may allow females to oviposit deeper in the substrate, thus influencing the incubation environment. This may be important for a species that exhibits temperature-dependent sex determination. Females found to have nested underground had deeper nests, but average temperatures did not significantly differ from other nests. However, temperature profiles in deeper nests fluctuated less and had lower maximum temperatures, factors which also influence survivorship and gonadal development. Finally, some Terrapene species exhibit homing behavior, which has apparently affected spatial use of the release-site enclosure. Some translocated individuals were frequently observed at the enclosure’s edge, presumably the result of homing behavior and attempts to return to former home ranges. Monitoring revealed that translocated turtles tended to be closer to the enclosure edge than resident or head-started turtles. However, the average distance to the fence increased in successive years among translocated turtles but remained the same for resident and head-started turtles, indicating that homing declined over time.

22) The Impacts of Intraspecific Variation on Phylogenetic Resolution in Terrapene
Natasha S. Vitek and Robert W. Burroughs1
Jackson School of Geosciences, The University of Texas at Austin, Austin, TX
Levels of variation in one population that exceed interspecific levels known to be expressed between sister species can make specimen identification within a clade problematic. Box turtles of the extant genus Terrapene exemplify this problem. Historically, the clade has been divided into four extant species and ten extant subspecies based on distinct morphological characteristics. It has been noted that intraspecific variation exists within both species and subspecies, but that variation remains poorly characterized. The situation presents a circular problem; characterizing variation across the temporal and geographic range of the clade remains intractable until lineages can be separated and studied individually, but lineages remain unidentifiable, especially in the fossil record, due to a lack of understanding of variation. In systematics, this problem can translate into poor understanding of apomorphies and a lack of resolution in phylogenetic analyses. To attempt to further understand the evolutionary history of Terrapene, we chose to evaluate variation in recent specimens and Pleistocene age fossils in a phylogenetic context. Our goal was to further understand how variation can impact phylogenetic analyses overall and attempt to separate lineages present in both the modern record and fossil record for further study. We used specimen-level phylogenetic analyses to explore whether variation between specimen-level terminals still allowed for species-level resolution. We scored multiple specimens of extant species of Terrapene as well as multiple fossils from several localities. We hypothesized that specimens would cluster in polytomic assemblages by species, if variation had a minimal effect on resolution. However, in our analysis not all specimens clustered together into species assemblages. Examination of character distribution indicated that coding specimen-level, as opposed to species-level, terminals caused signal from intraspecific variation to overwhelm potential apomorphies that were traditionally used to separate species. We then collapsed recent specimens into species-level terminals and fossil specimens into locality-level terminals. That approach resulted in traditionally recognized clades. In this case, we find that currently recognized apomorphies for species of Terrapene are insufficient for specimen-level identification against the backdrop of the broad intraspecific variation that is found in the genus. In this context, reliable, apomorphy-based identification of isolated specimens of Terrapene in the fossil record is currently impossible. Adding extinct ‘species’ known from only single specimens to an analysis presents a comparable situation. Further, some currently recognized species and subspecies are not immune to this problem. We found that the hypothesized morphological distinctions between different species, such as shell shape and co-ossification of bones in the shell were not reliable indicators of species at different points in time, once variation was accounted for. Our results indicate a broader need for continued work on phylogeny of Terrapene.
23) Husbandry Techniques Used during a Ranavirus Outbreak in Eastern Box Turtles (Terrapene carolina carolina) at the Maryland Zoo in Baltimore
Allison N. Wack1 , Richard Sim2, Kevin J. Murphy3, Kevin Barrett4,5, and Ellen Bronson1
1Medical Department, The Maryland Zoo in Baltimore, Baltimore, MD
2Wildlife Center of Virginia, Waynesboro, VA
3Animal Department, The Philadelphia Zoo, Philadelphia, PA
4Animal Department, The Maryland Zoo in Baltimore, Baltimore, MD
During the summer of 2011 The Maryland Zoo in Baltimore experienced a Ranavirus outbreak in its population of 27 Eastern Box Turtles (Terrapene carolina carolina). In response to the outbreak the Animal Department, working with the zoo’s Medical Department, developed protocols in order to treat the turtles and to prevent the disease from spreading to other species in the collection. These protocols included quarantine guidelines, modified environmental parameters, intensive supportive care including nutritional support, and extensive multimodal medical treatment in conjunction with the veterinary staff. As a result 14 of 27 turtles survived the outbreak and successfully overwintered outdoors, which far exceeds previous survival numbers in this species.