Hyla Versicolor Research Articles

An extension of gray treefrog range in Manitoba and into Saskatchewan

An extension of gray treefrog range in Manitoba and into Saskatchewan

Impacts of Batrachochytrium dendrobatidis Infection on Tadpole Foraging Performance

Pathogen-induced modifications in host behavior, including alterations in foraging behavior or foraging efficiency, can compromise host fitness by reducing growth and development. Chytridiomycosis is an infectious disease of amphibians caused by the fungus Batrachochytrium dendrobatidis (Bd), and it has played an important role in the worldwide decline of amphibians. In larval anurans, Bd infections commonly result in reduced developmental rates, however, the mechanism(s) responsible are untested. We conducted laboratory experiments to test whether Bd infections reduced foraging performance of Grey Treefrog (Hyla chrysoscelis) and Fowler's Toad (Anaxyrus [= Bufo] fowleri) tadpoles. In the first experiment, we observed foraging behavior of Bd-infected and uninfected tadpoles to test for differences in foraging activity. In a second experiment, we tested for differences in the ingestion rates of tadpoles by examining the amount of food in their alimentary track after a 3-hour foraging period. We hypothesized that Bd-infected tadpoles would forage less often and less efficiently than uninfected tadpoles. As predicted, Bd-infected larvae forage less often and were less efficient at obtaining food than uninfected larvae. Our results show that Bd infections reduce foraging efficiency in Anaxyrus and Hyla tadpoles, and that Bd differentially affects foraging behavior in these species. Thus, our results provide a potential mechanism of decreased developmental rates of Bd-infected tadpoles.

Processing of clustered sensory signals in gray treefrogs (Hyla versicolor): Behavioral responses in the laboratory and field.

Anuran mating choruses may consist of hundreds of sympatric and conspecific males calling in addition to other biotic and abiotic noise, representing a complex auditory scene whose characteristics change with distance. Detecting and localizing individual males in the chorus poses a challenge to the female, especially at long ranges. We carried out a series of recordings and behavioral experiments that are aimed at determining the distance at which female treefrogs (Hyla versicolor) use the chorus as an acoustic cue for their orienting responses. We analyzed natural choruses recorded at distances of 1–100 m and used these to create stimuli for phonotaxis experiments evaluating how the chorus attracts and provides cues for localization by females. In addition, we performed phonotaxis tests in the field near natural choruses to compare the frog’s behavior under laboratory and field conditions. We found that females utilize acoustic cues for orientation toward choruses at a distance up to 50 m, but additional cues may be used to ensure orientation in the field at distances >50 m. Additional experiments revealed that temporal coherence of the male call, as opposed to a general reduction in stimulus intensity, was the most important factor in female phonotactic behavior.

Apparent Continuing Expansion in the Range of the Gray Treefrog, <em>Hyla versicolor</em>, in New Brunswick

Recent observations of the Gray Treefrog, Hyla versicolor, in the Maritimes suggest that this species may be expanding its range and abundance, perhaps partly in response to human activities. However, during the breeding season the species characteristically calls in very small choruses in the region and may be present but not call, or call for very short periods. This has made it difficult to determine distribution and to make an assessment of unconfirmed historic records that suggest presence beyond the current range more than 50 years ago. The inadvertent introduction of frogs may also be playing a role in the apparent recent range expansion of this species.

Extension of the Known Range of the Gray Treefrog, <em>Hyla versicolor</em>, in Northwestern Ontario

I present the first documented records of Gray Treefrog, Hyla versicolor, from the Red Lake area of Kenora District in northwestern Ontario. A photograph of H. versicolor was taken at a locality west-northwest of the town of Red Lake in 2007 and another east of Red Lake in 2009. H. versicolor choruses were heard at 11 localities east and southeast of Red Lake in June 2008, and were tape recorded at 5. These seven documented records are 46-56 km north-northwest of the previous most northern voucher, west of the town of Ear Falls, and they represent the most northern locations known for H. versicolor in Ontario.

PCR Detection of Ranavirus in Adult Anurans from the Louisville Zoological Garden

Ranaviruses are known to cause mortality in a variety of anuran species and have the potential to significantly impact wild and captive frog populations. In this study, 16 captive frogs and toads from the Louisville Zoological Garden were examined for the presence of ranavirus; this group included 14 Cope's grey tree frogs (Hyla chrysoscelis), an American toad (Bufo americanus), and a southern toad (Bufo terrestris). All animals were wild caught and were evaluated via polymerase chain reaction (PCR), while animals that died were also assessed via histologic study to understand the role of ranaviral disease in these specimens. Of the animals that died, 82% were positive for ranavirus via PCR. Multiple swab samples collected over time from live tree frogs were positive for ranavirus via PCR. These findings reveal that ranaviral infection in captive adult anurans may occur without clinical signs or consistent histopathologic lesions.

The toxicity of Roundup Original Max® to 13 species of larval amphibians

With the increased use of glyphosate-based herbicides (marketed under several names, including Roundup and Vision), there has been a concomitant increased concern about the unintended impacts that particular formulations containing the popular surfactant polyethoxylated tallowamine (POEA) might have on amphibians. Published studies have examined a relatively small number of anuran species (primarily from Australia and eastern North America) and, surprisingly, no species of salamanders. Using a popular formulation of glyphosate (Roundup Original Max), the goal of the present study was to conduct tests of lethal concentrations estimated to kill 50% of a population after 96 h (LC50(96-h)) on a wider diversity of species from both eastern and western North America. Tests were conducted on nine species of stage 25, larval anurans from three families (Ranidae: Rana pipiens, R. clamitans, R. sylvatica, R. catesbeiana, R. cascadae; Bufonidae: Bufo americanus, B. boreas; and Hylidae: Hyla versicolor, Pseudacris crucifer) and four species of larval salamanders from two families (Ambystomatidae: Ambystoma gracile, A. maculatum, A. laterale; and Salamandridae: Notophthalmus viridescens). For the nine species of larval anurans, LC50(96-h) values ranged from 0.8- to 2.0-mg acid equivalents per liter with relatively little pattern in differential sensitivity among the species or families. The four species of larval salamanders were less sensitive than the anurans, with LC50(96-h) values ranging from 2.7- to 3.2-mg acid equivalents per liter and no substantial differences among the species of salamanders. This work substantially increases the available data on amphibian sensitivity to glyphosate formulations that include either POEA surfactants or the equally moderately to highly toxic surfactants of Roundup Original Max and should be useful for improving future risk assessments.

Very highly toxic effects of endosulfan across nine species of tadpoles: Lag effects and family‐level sensitivity

Pesticides are commonly used for health and economic benefits worldwide, but increased use has led to increased contamination of aquatic habitats. To understand potential impacts on nontarget organisms in these habitats, toxicologists generally use short-term (4-d) toxicity tests on model organisms. For most pesticides, few amphibian tests have been conducted, but there is growing concern about the potential impact of pesticides to amphibian populations. For the insecticide endosulfan, previous studies have found that low concentrations can be very highly toxic to amphibians and have suggested that this mortality may exhibit important lag effects. To estimate the lethal concentration of endosulfan that would cause 50% mortality after 4 d (LC50(4-d)) across a diversity of amphibians and the presence of lag effects, LC50(4-d) experiments were conducted on nine species of tadpoles from three families (Bufonidae: Bufo americanus, B. boreas; Hylidae: Pseudacris crucifer, P. regilla, Hyla versicolor; and Ranidae: Rana pipiens, R. clamitans, R. cascadae, R. catesbeiana) and then held the animals for an additional 4 d in clean water. The LC50(4-d) values for endosulfan ranged from 1.3 to 120 ppb, which classifies endosulfan as highly toxic to very highly toxic. Moreover, holding the animals for an additional 4 d in clean water revealed significant additional mortality in three of the nine species. Leopard frogs, for example, experienced no significant death during the initial 4-d exposure to 60 ppb but 97% death after an additional 4 d in clean water. A phylogenetic pattern also appears to exist among families, with Bufonidae being least susceptible, Hylidae being moderately susceptible, and Ranidae being most susceptible. Results from the present study provide valuable data to assess the impact of endosulfan on a globally declining group of vertebrates.

Lethal and Sublethal Effects of Embryonic and Larval Exposure of Hyla versicolor to Stormwater Pond Sediments

Stormwater ponds are common features of modern stormwater management practices. Stormwater ponds often retain standing water for extended periods of time, develop vegetative characteristics similar to natural wetlands, and attract wildlife. However, because stormwater ponds are designed to capture pollutants, wildlife that utilize ponds might be exposed to pollutants and suffer toxicological effects. To investigate the toxicity of stormwater pond sediments to Hyla versicolor, an anuran commonly found using retention ponds for breeding, we exposed embryos and larvae to sediments in laboratory microcosms. Exposure to pond sediments reduced survival of embryos by approximately 50% but did not affect larval survival. Larvae exposed to stormwater pond sediment developed significantly faster (x = 39 days compared to 42 days; p = 0.005) and were significantly larger at metamorphosis (x = 0.49 g compared to 0.36 g; p < 0.001) than controls that were exposed to clean sand. Substantial amounts (712-2215 mg/l) of chloride leached from pond sediments into the water column of treatment microcosms; subsequently, survival of embryos was negatively correlated (r (2) = 0.50; p < 0.001) with water conductivity during development. Our results, along with the limited number of other toxicological studies of stormwater ponds, suggest that road salt contributes to the degradation of stormwater pond habitat quality for amphibian reproduction and that future research should focus on understanding interactions among road salts and other pollutants and stressors characteristic of urban environments.

Predation risk assessment by green frog (Rana clamitans) tadpoles through chemical cues produced by multiple prey

Many prey assess predation risk through predator chemical cues. Numerous studies have shown that (1) prey sometimes respond to chemical cues produced by heterospecifics and (2) that many species are capable of associative learning. This study extends this research by focusing on predation risk assessment and antipredator behavior in environments containing chemical cues produced by multiple prey species. The results show that green frog (Rana clamitans) tadpoles (1) assess risk from the chemical cue produced during predation by a heterospecific (gray tree frog, Hyla versicolor, tadpoles) and (2) can exhibit similarly strong behavioral responses to a mix of conspecific and heterospecific cues compared to conspecific cue alone, depending on their conditioning environment. I then discuss how the prey choice of the predators and the relative abundances of the prey species should influence the informational value of heterospecific cues.

EXPERIMENTAL ANALYSIS OF MULTIVARIATE FEMALE CHOICE IN GRAY TREEFROGS (HYLA VERSICOLOR): EVIDENCE FOR DIRECTIONAL AND STABILIZING SELECTION

Even simple biological signals vary in several measurable dimensions. Understanding their evolution requires, therefore, a multivariate understanding of selection, including how different properties interact to determine the effectiveness of the signal. We combined experimental manipulation with multivariate selection analysis to assess female mate choice on the simple trilled calls of male gray treefrogs. We independently and randomly varied five behaviorally relevant acoustic properties in 154 synthetic calls. We compared response times of each of 154 females to one of these calls with its response to a standard call that had mean values of the five properties. We found directional and quadratic selection on two properties indicative of the amount of signaling, pulse number, and call rate. Canonical rotation of the fitness surface showed that these properties, along with pulse rate, contributed heavily to a major axis of stabilizing selection, a result consistent with univariate studies showing diminishing effects of increasing pulse number well beyond the mean. Spectral properties contributed to a second major axis of stabilizing selection. The single major axis of disruptive selection suggested that a combination of two temporal and two spectral properties with values differing from the mean should be especially attractive.

Elucidation of the role of aquaporins and glyceroporins in freeze tolerance of Hyla chrysoscelis

Cope's gray tree frog (Hyla chrysoscelis) can tolerate freezing of its extracellular water. While the exact mechanism of freeze tolerance is unknown, aquaporins may play a role by allowing water and solutes like glycerol to move between compartments prior to and during freezing. Previously, two aquaporins (HC1, HC2) and one glyceroporin (HC3) were identified in Hyla chrysoscelis in a tissue and temperature dependent manner. To further elucidate the role of aquaporins in freeze tolerance, three putative aquaporins (HC7, HC8 and HC0) have been identified. Degenerate PCR primers were developed using conserved amino acid regions from Xenopus tropicalis and compared to other species with high sequence homology. For the putative aquaporins, mRNA was isolated from the fat, large intestines, and the eye for HC7, HC8 and HC0 respectively. The templates were amplified using RT‐PCR and products (326 bp, 377 bp and 400 bp respectively) were examined by electrophoresis. Further experiments are underway to confirm these putative aquaporins, and to examine their tissue and temperature dependence. Supported by NSF IOB‐0517301.

Single and interactive effects of malathion, overwintered green frog tadpoles, and cyanobacteria on gray treefrog tadpoles

Amphibian population declines around the world are associated with invasive species, pesticides, pathogens, habitat destruction, or a combination of factors. Because contamination is widespread, it represents a relevant environmental stress that can affect the ability of organisms to deal with other factors present in the environment. We examined the effects of the insecticide malathion, larger tadpole competitors (green frogs, Rana clamitans), and a toxic cyanobacteria (Anabaena spp.) on tadpoles of Cope's gray treefrog (Hyla chrysoscelis) reared from hatching through metamorphosis in outdoor mesocosms. The response of mass at metamorphosis and time to metamorphosis was significantly affected by exposure to malathion and presence of overwintered green frog tadpoles. Malathion generally led to increased mass at metamorphosis, earlier time to metamorphosis, and increased activity during larval development. These results likely stem from short-term increases in periphyton associated with malathion exposure (although these effects were nonsignificant). Exposure of gray treefrogs to overwintered green frog tadpoles led to an earlier time to metamorphosis without differences in mass at metamorphosis and was associated with increased activity in gray treefrogs. Survival of gray treefrogs was significantly affected by an interaction of green frog and malathion, indicating nonadditive effects of these treatments. Exposure to cyanobacteria had a significant negative effect on green frogs but no effect on treefrogs. Malathion had the strongest effect on the community, but our results indicated that some factors can interact in ways not predicted by single factors alone.

Amphibian Community Response to Variation in Habitat Structure and Competitor Density

Habitat destruction and alteration is a major cause of population declines; thus, habitat restoration is essential for preventing species extinctions. Habitat restoration, however, may be hindered by an inadequate understanding of species' habitat requirements. Habitat structure may play a major role in influencing the biotic interactions among species. Pond-breeding amphibians are one group that has experienced loss and alteration of critical habitats. Vegetation structure in wetland environments may be important to amphibian larvae if its presence mediates biotic interactions, such as competition, which can affect growth and survival to metamorphosis. Despite the potential importance of habitat structure and competition, few studies have examined these factors simultaneously. In a full-factorial pond mesocosm experiment, we tested for the effects of variation in habitat structure and density of a larger tadpole competitor (Rana pipiens, northern leopard frog) on development and survival of three species of larval amphibians: Bufo americanus (American toad), Hyla versicolor (gray treefrog), and Ambystoma maculatum (spotted salamander). Survival of Bufo was greater in mesocosms with live or artificial cattails than in those lacking vegetation. Although responses varied among species, our data suggest that time to metamorphosis, survival, and species evenness can be influenced by both aquatic habitat structure and competition, whereas size at metamorphosis is affected primarily by competition. Our study is the first to demonstrate that vegetation structure alone can affect the expression of metamorphic traits for some anuran species.

Pesticide alters habitat selection and aquatic community composition

Anthropogenic chemical contamination is an important issue for conservation of aquatic ecosystems. While recent research highlights that community context can mediate the consequences of contaminant exposure, little is known about how contaminants themselves might determine this context by altering habitat selection and thus initial community composition. Here we show that the insecticide carbaryl and its commercial counterpart Sevin can affect aquatic community composition by differentially altering oviposition and colonization of experimental pools by amphibians and insects. On average, contaminated pools received 20-fold more adult beetle and heteropteran colonists and 12-fold more Culex mosquito and chironomid midge egg masses. On the other hand, ovipositing Anopheles mosquitoes and cricket frogs showed no preference and we have shown previously that gray treefrogs strongly avoid contaminated pools. Overall, initial richness doubled in contaminated pools compared with controls. By affecting colonizing taxa differently and increasing richness, the contaminant may alter the ecological context under which subsequent effects of exposure will unfold. Given that community context is important for evaluating toxicity effects, understanding the net effects of contaminants in natural systems requires an understanding of their effects on community assembly via shifts in habitat selection.

When Should Prey Respond to Consumed Heterospecifics? Testing Hypotheses of Perceived Risk

In aquatic systems, a long-standing question is why chemical cues from some diets consumed by a predator induce strong anti-predator responses in prey while other diets induce weak or no responses. We performed an experiment to determine if strong prey responses to particular predator diets are due to prey being closely related to the predator's diet (i.e., phylogenetic relatedness) or due to prey coexisting with the predator's diet and thereby sharing a risk of predation. We compared the behavior of Gray Treefrog tadpoles (Hyla versicolor) to cues from a dragonfly nymph (Anax junius) that consumed either conspecific Gray Treefrogs, one of six diets that commonly coexist with Gray Treefrogs (spanning a wide range of phylogenetic relatedness), or one diet that is closely related to Gray Treefrogs but has an allopatric range that has not overlapped for at least 20,000 yrs. We found that tadpoles could discriminate among the diets and that the magnitude of behavioral response supported the hypothesis of diet phylogenetic relatedness and refuted the hypothesis of diet coexistence.

All hosts are not equal: explaining differential patterns of malformations in an amphibian community

1. Within a community, different host species often exhibit broad variation in sensitivity to infection and disease. Because such differences can influence the strength and outcome of community interactions, it is essential to understand differential disease patterns and identify the mechanisms responsible. 2. In North American wetlands, amphibian species often exhibit extraordinary differences in the frequency of limb malformations induced by the digenetic trematode, Ribeiroia ondatrae. By coupling field studies with parasite exposure experiments, we evaluated whether such patterns were due to differences in (i) parasite encounter rate, (ii) infection establishment, or (iii) parasite persistence within hosts. 3. Field results underscored the broad variation in malformations and infection between host species; while nearly 60% (n = 618) of emerging American toads exhibited severe limb deformities such as bony triangles, skin webbings and missing limbs, fewer than 4% (n = 251) of Eastern gray treefrogs from the same pond were abnormal. Despite similarities in the phenology and larval development period of these species, they differed sharply in Ribeiroia infection. On average, toads supported 75x more metacercariae than did metamorphic treefrogs. 4. Experimental exposures of larval toads and treefrogs to a realistic range of Ribeiroia cercariae revealed strong differences in the sensitivity of these species to infection; exposed toads suffered elevated mortality (up to 95%), delayed metamorphosis, and severe limb malformations consistent with field observations. Treefrogs, in contrast, exhibited limited mortality and no malformations, regardless of exposure level. Ribeiroia cercariae were substantially less successful in locating and infecting Hyla versicolor larvae. 5. Our results indicate that the observed differences in infection and malformations owe to a lower ability of Ribeiroia cercariae to both find and establish within larval treefrogs, possibly stemming from a heightened immune response to infection. Because Ribeiroia is a highly pathogenic parasite with negative effects on larval and metamorphic amphibian survival, variation in infection resistance among species could have important implications for understanding patterns of species co-occurrence, competition, and community diversity.

A cocktail of contaminants: how mixtures of pesticides at low concentrations affect aquatic communities

The ubiquity of anthropogenic chemicals in nature poses a challenge to understanding how ecological communities are impacted by them. While we are rapidly gaining an understanding of how individual contaminants affect communities, communities are exposed to suites of contaminants yet investigations of the effects of diverse contaminant mixtures in aquatic communities are rare. I examined how a single application of five insecticides (malathion, carbaryl, chlorpyrifos, diazinon, and endosulfan) and five herbicides (glyphosate, atrazine, acetochlor, metolachlor, and 2,4-D) at low concentrations (2-16 p.p.b.) affected aquatic communities composed of zooplankton, phytoplankton, periphyton, and larval amphibians (gray tree frogs, Hyla versicolor, and leopard frogs, Rana pipiens). Using outdoor mesocosms, I examined each pesticide alone, a mix of insecticides, a mix of herbicides, and a mix of all ten pesticides. Individual pesticides had a wide range of direct and indirect effects on all trophic groups. For some taxa (i.e., zooplankton and algae), the impact of pesticide mixtures could largely be predicted from the impacts of individual pesticides; for other taxa (i.e., amphibians) it could not. For amphibians, there was an apparent direct toxic effect of endosulfan that caused 84% mortality of leopard frogs and an indirect effect induced by diazinon that caused 24% mortality of leopard frogs. When pesticides were combined, the mix of herbicides had no negative effects on the survival and metamorphosis of amphibians, but the mix of insecticides and the mix of all ten pesticides eliminated 99% of leopard frogs. Interestingly, these mixtures did not cause mortality in the gray tree frogs and, as a result, the gray tree frogs grew nearly twice as large due to reduced competition with leopard frogs. In short, wetland communities can be dramatically impacted by low concentrations of pesticides (both separate and combined) and these results offer important insights for the conservation of wetland communities.

Pesticide alters oviposition site selection in gray treefrogs

Pesticide alters oviposition site selection in gray treefrogs

Hormonal modulation of phonotaxis and advertisement-call preferences in the gray treefrog ( Hyla versicolor)

Hormonal levels fluctuate during the breeding season in many anurans, but the identity of the hormones that modulate breeding behavior and their effects remain unclear. We tested the influence of a combined treatment of progesterone and prostaglandin on phonotaxis, the key proceptive reproductive behavior of female anurans. First, we found that female gray treefrogs ( Hyla versicolor) treated with progesterone and prostaglandin exhibited phonotaxis to synthetic male advertisement signals significantly more often than animals treated with ringers vehicle or uninjected controls. Responsive females had greater levels of plasma progesterone and estradiol compared to both control groups, suggesting that these steroids may be promoting phonotaxis. Second, we found that the selectivity of hormonally-induced phonotaxis in H. versicolor was similar to that observed in freshly captured breeding animals. Females made the same choices between acoustic signals after hormone treatments in tests of frequency, call rate and pulse rate, compared to their responses without treatment immediately after collection from the breeding chorus. The preference for a longer call was, however, significantly weaker after hormonal induction of phonotaxis. Hormonally primed females were also less likely to respond in any test and took longer to respond than did freshly collected females. Consequently, our study shows how progesterone–prostaglandin induced phonotaxis in female treefrogs influences both the quality and quantity of phonotaxis, relative to that exhibited by naturally breeding females.