Ohio Biological Survey and Ohio Division of Wildlife: Partners in Wildlife Diversity
Ohio Amphibian Research & Conservation Conference

Amphibian Conference
Online Registration

March 10, 2012. ODOT Hilltop Conference Center. Columbus, Ohio
Keynote Presentation: "The" Cause of Amphibian Declines: A Developmental Endocrinologist's Perspective
Tyrone Hayes (tyrone[at]berkeley.edu)
Department of Integrative Biology, University of California, Berkeley, 3060 Valley Life Sciences Bldg #3140, Berkeley, California, 94720-3140

My research focuses on the role of steroid hormones in amphibian development and I conduct both laboratory and field studies in the U.S. and Africa. Dr. Tyrone HayesThe two main areas of interest are metamorphosis and sex differentiation, but I am also interested in growth (larval and adult) and hormonal regulation of reproductive behavior. My work addresses problems on several levels including ecological, organismal, and molecular questions.. Studies of metamorphosis examine the effects of temperature on developmental rates, interactions between the thyroid hormones and steroids, and hormonal regulation of skin gland development. I am also examining the effects of tadpole density on developmental rates and measuring metamorphic rates and hormone levels of tadpoles in the field and in the laboratory. My work on sex differentiation involves the African clawed frog (Xenopus laevis), the Japanese kajika (Buegeria buegeri), and the Pine Barrens treefrog (Hyla femoralis). While Xenopus serves as a good model because of its availability, the latter two species have genetically distinguishable sexes. I can therefore examine early events in gonad differentiation, steroid enzyme activities, steroid receptors, etc., knowing the genetic sex of the individual larvae. My main goal is to synthesize ecological/evolutionary, organismal/physiological, and biochemical/molecular studies to learn how an animal translates changes in its external environment to internal changes, how these internal changes are coordinated, what molecular mechanisms are involved, and in turn, how changes at the molecular level affect an animal's ability to adapt to the changes in its external environment. Most recently, my studies have been used as models to develop laboratory and field techniques to examine the effects of endocrine disrupting contaminants on amphibian development. My research now focuses on the effects of pesticides mixtures on larval development and the potential role of pesticides in amphibian declines.

Greater than 70% of the world's amphibian species are in decline. We propose that there is not likely a single cause for global amphibian declines and present a three-tiered hierarchical approach that addresses interactions among and between ultimate and proximate factors that contribute to amphibian declines. There are two immediate (proximate) causes for amphibian declines, death and decreased recruitment (reproductive failure). Although much attention has focused on death, few studies have addressed factors that contribute to declines as a result of failed recruitment. Further, a great deal of attention has focused on the role of pathogens in inducing diseases that cause death, but we suggest that pathogen success is profoundly affected by four other ultimate factors: atmospheric change, environmental pollutants, habitat modification, and invasive species. Environmental pollutants arise as likely important factors in amphibian declines because they have realized potential to affect recruitment. Further, many studies have documented immunosuppressive effects of pesticides, suggesting a role for environmental contaminants in increased pathogen virulence and disease rates. Increased attention to recruitment and ultimate factors that interct wth pathogens are important in addressing this global crisis.

Exploring Niche Divergence in Eastern Red-backed Salamanders
Carl D. Anthony (canthony[at]jcu.edu) and Cari M. Hickerson
Department of Biology, John Carroll University, 20700 North Park Blvd., University Heights, Ohio, 44118

Polymorphic species provide an opportunity to study the role of reproductive isolation in populations that may be diverging in sympatry. The Eastern Red-backed Salamander (Plethodon cinereus) is polymorphic for dorsal coloration with red-striped and all black unstriped individuals representing the most common phenotypes. Numerous studies have now established that the two common phenotypes differ in ecologically important ways that have the potential to affect when and where salamanders feed and whether or not they establish territories. Territory acquisition is a critical component of reproductive success in this species and has the potential to impact mate choice. Research in our laboratory has focused on mating behavior and on differences in diet and territorial behavior between the two phenotypes. We present a model where the striped phenotype exhibits a territorial strategy that maximizes access to high quality, prey rich territories and mates. Assortative mating by phenotype, a crucial first step in the evolution of reproductive isolation, may be one byproduct of this adaptive strategy. In the field, we observed more same color male/female pairs than expected by chance alone and striped salamanders were more apt to hold territories compared to unstriped salamanders. When salamander density peaked in spring and fall, the diets of the two phenotypes diverged, suggesting that competition for prey occurs during these seasons. In laboratory trials, striped salamanders were more aggressive and were more likely to exhibit residency effects compared to unstriped salamanders. Taken together, our results suggest that polymorphic populations of red-backed salamanders show potential for ecological divergence and that territorial behavior may play a central role in this process.

Morphological and Molecular Evidence for Widespread Hybridization Between the Northern Ravine Salamander (Plethodon electromorphus) and the Red-backed Salamander (P. cinereus)
Richard Lehtinen (rlehtinen[at]wooster.edu), Emlyne Cassagnol, Hilary Edington, Meredith Eyre, and Anthony Steratore.
Department of Biology, The College of Wooster, 931 College Mall, Wooster, Ohio, 44691

Much research in evolutionary biology has focused on how new species arise, however, relatively little work has focused on the opposite process – how species may merge back together after secondary contact. Because of the difficulty in field-identifying small Plethodon in our area, we used molecular and morphological markers to investigate the possibility that P. cinereus and P. electromorphus are currently hybridizing. We used single nucleotide polymorphisms in DNA sequences from a nuclear gene (RAG-1, 425 bp) to genotype salamanders collected from 16 localities in 7 counties in north-eastern Ohio. Of the 177 individuals genotyped, 32 (18%) were identified as hybrids and these were found at nearly all sites where both species co-occurred. Data from 21 morphological and coloration characters were consistent with the conclusions drawn from molecular data. F1 individuals as well as backcrosses to both species were identified, indicating that hybrids are both viable and fertile. Further, mitochondrial DNA sequence data from hybrids indicated that hybridization was symmetrical (occurred in both directions). Museum specimens (collected as far back as 1965) were confirmed as hybrids and indicate that gene flow between these species is not very recent. Together these data suggest that hybridization between these species is both common and geographically widespread. Future work will focus on assessing the efficacy of pre-zygotic isolation and the fitness of hybrids compared with pure parental forms. These additional data will help assess whether these species will likely remain independent units over evolutionary time.

Phylogeography of the Spring Salamander, Gyrinophilus porphyriticus
Michael Haughey (mh227810[at]ohio.edu) and Shawn Kutcha
Department of Biological Sciences, Ohio University, 107 Irwin Hall, Athens, Ohio, 45701

Climatic oscillations during the Pleistocene repeatedly formed glacial boundaries, shifted suitable habitat, and caused significant geological changes in the major drainage systems of Eastern North America. Contemporary drainage systems are therefore a result of these cycles of glacial advances and retreats causing the fragmentation and fusion of many major pre-Pleistocene drainage systems. However, few studies have looked at the impact of changes in historical drainage systems in shaping patterns of genetic diversity in stream-dwelling terrestrial vertebrates. The main goal of our study is to investigate the importance of drainage system alterations on shaping the phylogenetic patterns of genetic diversity in the stream dwelling salamander Gyrinophilus porphyriticus. The state of Ohio, located on the edge of the glacial maximum, experienced some of the most dramatic changes, making it an ideal area to investigate the influence of historical vs. contemporary drainage systems on the distribution of genetic diversity. In addition we are also investigating the hypothesis that the Hamilton County population of G. porphyriticus (near Cincinnati) is a Pleistocene refuge from Northeastern Ohio. To conduct these analyses we are collecting sequence data from three mitochondrial loci and one nuclear locus from approximately 300 individuals located across the entire range of the species. In Ohio we currently have 75 individuals from 12 localities, however, our analyses are range wide.

Fluctuation and Stability: A Seven Year Study of Spotted Salamander (Ambystoma maculatum) Demography and Migratory Orientations
Rebecca Homan (homanr[at]denison.edu)
Department of Biology, Dennison University, Talbot Hall, Granville, Ohio, 43023

Long-term studies of undisturbed amphibian populations are critical to understanding natural population fluctuations and distinguishing them from human induced fluctuations. To that end, I initiated a long-term demographic study of Spotted Salamanders (Ambystoma maculatum) at a relatively undisturbed breeding site in central Ohio. For the last seven years, I used a drift fence system to monitor the annual breeding population sizes, recruitment rates, and migration orientations for Spotted Salamanders. Additionally, a subset of the adult population was PIT tagged annually to estimate annual survival and recapture probabilities. Between 2005 and 2011, Spotted Salamander breeding population sizes ranged from 487 to 1,061. Adults entered the pond in significantly higher frequencies from the East in most years and in significantly lower frequencies from the West all years. We found that sex ratios (M:F) ranged from 1.5 to 3.8. Numbers of emerging juveniles were highly variable, with a range of 0 to 304, and newly metamorphosed juveniles tended to initially orient South when leaving the pond. Finally, from data up to 2010, we estimate that annual survival rate for Spotted Salamanders (77.0%) is not dependent on sex or year, but that the probability of recapture is time dependent. Throughout the last six years, we have seen dramatic demographic fluctuations in several metrics. The nature of these results at our undisturbed breeding site reinforces the need for long-term studies on amphibian populations if we intend to use demographic metrics to evaluate population health and/or stability.

Abundance, Distribution, and Habitat Associations of Unisexual Mole Salamanders (Ambystoma) in Ohio
Katherine Greenwald1 (katherine.greenwald[at]emich.edu) and H. Lisle Gibbs2
1Department of Biology, Eastern Michigan University, 391 Mark Jefferson, Ypsilanti, Michigan, 48197. 2Department of Evolution, Ecology, and Organismal Biology, Ohio State University, 318 W 12th Avenue, Columbus, Ohio, 43210-1235.

Unisexual salamanders in the genus Ambystoma are an abundant yet difficult to identify component of Ohio’s biodiversity that have both fascinated and frustrated herpetologists interested in understanding the distribution of salamanders in the Great Lakes region. These all-female salamanders reproduce in a unique way that involves "stealing" genomes from the sexual species that they coexist with. To date, scientists have documented genome incorporation by unisexuals from five different sexual species: the blue-spotted (A. laterale), Jefferson (A. jeffersonianum), smallmouth (A. texanum), tiger (A. tigrinum) and streamside (A. barbouri) salamanders. This phenomenon, along with the fact that the unisexuals are frequently polyploid (having more than two copies of the genome), means that there are a large number of possible genome combinations (biotypes) in these salamanders. In many cases the unisexuals closely resemble the sexual species whose genomes they have incorporated, making morphological identification problematic. We have developed and used a genetic method to identify unisexual biotypes for over 2000 salamanders from over 60 sites across Ohio. Analysis of locality data reveals a strong latitudinal gradient in the frequency of unisexuals, with frequency decreasing from north to south. Furthermore, habitat data collected at each site suggest that there is niche partitioning among the various species and biotypes found in Ohio, with unisexuals more common in higher quality habitat. These data will be used to explore the ecological and evolutionary factors that determine the range, abundance and habitat usage of sexual species and unisexual biotypes present in Ohio, as well as to develop conservation plans for the state-listed blue-spotted salamander.

Peter Pan May be Down, But Not Out! A Long-term Look at the Impact of the Lampracide TFM upon the Common Mudpuppy in the Grand River
Tim Matson (tmatson[at]cmnh.org)
Cleveland Museum of Natural History, 1 Wade Oval Drive University Circle, Cleveland, Ohio, 44106

The lampricide TFM has been used as the primary method in controlling the sea lamprey (Petromyzon marinus) in the Great Lakes since 1958 and more recently in tributaries to Lake Champlain. TFM has long been known to cause mortality in the non-targeted mudpuppy (Necturus maculosus). Applications of TFM in Ohio streams which began in 1986 have caused high mortality in mudpuppies within Conneaut Creek (1986, 1995) and within stretches of the Grand River (1987, 2003). Quantification of the impact of TFM upon the mudpuppy in the Grand River was estimated using Schnabel estimation for one site by Matson (1990). Severe time constraints did not allow inclusion of a control site prior to the first application of TFM to the Grand River on 26 April 1987. Results of that study indicated a minimum decrease in population size of 29%. Since that time both Conneaut Creek and the Grand River have undergone numerous lampricde applications. Recently we have conducted two year Schnabel estimates at the same site in the Grand River and at a control site in the Ashtabula River. Results of that project will be compared with the earlier study and additional data pertinent to the status of the Common Mudpuppy in the Grand River will be presented.

They're Declining; Now What? A Conservation Plan for the Eastern Hellbender in Ohio
Gregory J. Lipps (GregLipps[at]gmail.com)
1473 County Road 5 2, Delta, Ohio, 43515

The Eastern Hellbender (Cryptobranchus alleganiensis alleganiensis) is Ohio’s largest amphibian, a fully aquatic salamander that was once found throughout much of the Ohio River drainage. A statewide survey conducted from 2006-2009 found an 82% decline in relative abundance of hellbenders in waterways where they were found during surveys of the mid-1980s. Most remaining populations in Ohio are apparently not viable, as there is no evidence of recent successful recruitment. While all of the factors responsible for the decline are not fully understood, failing to take action now would likely result in the extirpation of most of the remaining populations in the state. The Conservation Plan for the Eastern Hellbender in Ohio lays out a vision for the recovery of the species in Ohio and strategies and methods for accomplishing this goal. Central to this plan is a head-starting/repatriation program to establish populations where they have been extirpated and augment populations that are not viable, beginning with eggs collected from the wild and reared at partner zoos. The plan also addresses other issues, ranging from measuring habitat quality to responses to pollution events in hellbender occupied habitats. In 2011, hellbender eggs were successfully collected in Ohio and the larvae are currently being reared at The Columbus Zoo and Aquarium. Four-year old individuals raised from eggs collected in neighboring West Virginia are being screened for potential pathogens prior to our first planned release. The partnership that has formed to aid in the recovery of the Eastern Hellbender in Ohio consists of government agencies, zoos, soil and water conservation districts, a land trust, and educational institutions. The expertise, resources, and cooperation of the partners are fundamental to the successful implementation of the plan.

Some Times are Better than Others: How Time of Insecticide Exposure can Influence Anuran Metamorphosis and Development
Michelle D. Boone1 (boonemd[at]muohio.edu), Melissa Youngquist1, Caren C. Helbing2, and Nick Veldhoen2
1Department of Zoology, Miami University of Ohio, Dept. of Zoology, 212 Pearson Hall, Oxford, Ohio, 45056. 2Department of Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia V8W 3P6 Canada

The orchestration of metamorphosis is initiated and integrated by thyroid hormones, which change dynamically during larval development. The impact of pesticides may vary depending on developmental stage of an amphibian, which may be influenced by physiological changes during development. Previous studies have found that some anurans reach metamorphosis earlier or are more developed when exposure to the insecticide carbaryl occurs later in development, suggesting that carbaryl could affect the thyroid hormone axis as well as alter the food web. We examined the effects of carbaryl exposure from early to late in tadpole development on green frogs (Rana clamitans) and northern leopard frogs (Rana pipiens) in the lab and/or the field. We measured effects on survival, time and size at metamorphosis, feeding and jumping performance of metamorphs, and thryoid response genes in the brain of metamorphs. We found that carbaryl did not impact survival at metamorphosis, mass at or time to metamorphosis, or feeding performance for lab reared green frogs, but it did significantly increase the abundance of some thyroid responsive genes (i.e., TR-a) in brain tissue, suggesting that carbaryl can act as an endocrine disruptor at some times in anuran development. We have also found that exposure to carbaryl in the field resulted in greater mass and increased development of green frog tadpoles when exposure occurred late, suggesting that timing of exposure can also influence traits correlated with fitness; analyses of brain tissue are currently under way. Timing of exposure also appears to influence impacts on time and size at metamorphosis on northern leopard frogs in field studies; analyses of brain tissue are currently under way for this species. Our studies suggest that physiological changes associated with development may make amphibians more susceptible to effects of an insecticide and may have disparate effects on population-level outcomes.

Seasonal and Habitat-Associated Patterns of Bd Infection in Green Frogs and Bullfrogs in Central Ohio
Chelsea Korfel (chelsea.korfel[at]gmail.com) and Thomas Hetherington
Museum of Biological Diversity, Ohio State University, 1315 Kinnear Road, Columbus, Ohio, 43212

The amphibian chytrid, Batrachochytrium dendrobatidis (Bd), is widespread throughout central Ohio. Green frogs (Lithobates clamitans) and bullfrogs (L. catesbieanus) are two generalist amphibian species that are active from spring through autumn in Ohio and inhabit a broad range of aquatic habitats: streams, emergent wetlands, and vernal pools. We collected samples of these species from March – November for two seasons (2010, 2011) and in different habitats. We found that patterns of occurrence of Bd are associated with both season and habitat, but that patterns of occurrence are not correlated with infection intensity. On multiple occasions individuals were sampled in the same habitat months apart, and data from these recaptured animals showed that frogs can lose, gain, and maintain low intensity infections throughout the season of amphibian activity. Both asymptomatic bullfrogs and green frogs were found with infection intensities above the reported critical threshold of 10,000; however, most infections were low intensity. We found that frogs displayed higher infection rates but lower infection intensities in cooler conditions (spring, ravine streams) compared to warmer conditions (summer, emergent wetlands). These findings provide valuable insights into the dynamic between amphibian hosts and this fungal pathogen in a temperate environment and contributes to a better understanding on potential impacts of Bd on temperate amphibian species worldwide.

Influence of the Invasive Mosquitofish (Gambusia affinis) on Oviposition and Colonization Success of Gray Treefrogs (Hyla versicolor) in experimental mesocosms.
Geoffrey R. Smith (smithg[at]denison.edu) and Johanna J. Harmon
Department of Biology, Denison University, Granville, Ohio, 43203

Mosquitofish (Gambusia) are an invasive species that have been introduced around the world, including Ohio. Previous studies have demonstrated that the western mosquitofish (Gambusia affinis) can have negative effects on some Ohio amphibians. Here we examine the effects of the lethal and non-lethal presence of mosquitofish on the oviposition and colonization success of Gray Treefrogs (Hyla versicolor) and macroinvertebrates in experimental ponds. Gray Treefrogs laid fewer eggs and produced fewer tadpoles at the end of the experiment in experimental ponds with lethal (uncaged) mosquitofish than in experimental ponds with non-lethal (caged) mosquitofish or no mosquitofish. In addition, the presence of mosquitofish affected the successful colonization of these ponds by macroinvertebrates, including several that may be predators on anuran tadpoles. Our results suggest that Gray Treefrogs colonization success may be influenced directly by the presence of mosquitofish, but also by the potential for indirect effects due to changes in the abundance and composition of the macroinvertebrate community.

Identifying Potential Vernal Pool Restoration Sites in Ohio Using GIS
Brian Gara1 (brian.gara[at]epa.state.oh.us) and Mick Miccachion1,2
1Division of Surface Water, Ohio Environmental Protection Agency, 4675 Homer Ohio Lane, Groveport, Ohio, 43215. 2Currrent address: Midwest Biodiversity Institute, 5530 Olentangy River Road, Columbus, Ohio, 43235.

Wetland mitigation projects frequently result in habitat that is inferior in quality to the natural wetlands being impacted via permitted development activities. Additionally, the replacement of ecological services provided vernal pools are rarely targeted as a component of wetland mitigation. These forested depressions represent critical breeding habitat for several species of sensitive amphibians, including Wood Frogs (Lithobates sylvaticus) and Spotted Salamanders (Ambystoma maculatum). Without adequate replacement of these resources, Ohio continues to experience a “net loss” of amphibian habitat. A GIS-based model was developed to provide a tool for targeting sites specifically for vernal pool restoration. As most pond-breeding amphibian species are known to travel short distances from the vernal pools where they originated, one of the primary factors considered was the proximity of the restoration site to existing amphibian breeding locations. A GIS layer of potential “high quality” vernal pools was developed using digital National Wetland Inventory data. Several predictors of landscape disturbance were considered within the area surrounding the wetland, including the quantities of impervious surface and other developed land. Wetlands were then scored depending on the overall intensity of landscape disturbance affecting each of these vernal pools. A GIS procedure was then used to identify potential vernal pool restoration sites meeting the following criteria:
1) within 500 meters of existing high quality vernal pools;
2) containing a significant amount of agricultural land mapped as having hydric soil.
The final product of this study was a Google map displaying the locations of these sites (http://wwwapp.epa.ohio.gov/dsw/gis/vernal/).

2012 Amphibian Conference Online Registration

The Ohio Amphibian Research and Conservation Conference is sponsored by the Ohio Division of Wildlife and The Columbus Zoo & Aquarium .