Concurrent threats and extinction risk in a long-lived, highly fecund vertebrate with parental care.

Detecting declines and quantifying extinction risk of long-lived, highly fecund vertebrates, including fishes, reptiles, and amphibians, can be challenging. In addition to the false notion that large clutches always buffer against population declines, the imperiled status of long-lived species can often be masked by extinction debt, wherein adults persist on the landscape for several years after populations cease to be viable. Here we develop a demographic model for the eastern hellbender (Cryptobranchus alleganiensis), an imperiled aquatic salamander with paternal care. We examined the individual and interactive effects of three of the leading threats hypothesized to contribute to the species' demise: habitat loss due to siltation, high rates of nest failure, and excess adult mortality caused by fishing and harvest. We parameterized the model using data on their life history and reproductive ecology to model the fates of individual nests and address multiple sources of density-dependent mortality under both deterministic and stochastic environmental conditions. Our model suggests that high rates of nest failure observed in the field are sufficient to drive hellbender populations toward a geriatric age distribution and eventually to localized extinction but that this process takes decades. Moreover, the combination of limited nest site availability due to siltation, nest failure, and stochastic adult mortality can interact to increase the likelihood and pace of extinction, which was particularly evident under stochastic scenarios. Density dependence in larval survival and recruitment can severely hamper a population's ability to recover from declines. Our model helps to identify tipping points beyond which extinction becomes certain and management interventions become necessary. Our approach can be generalized to understand the interactive effects of various threats to the extinction risk of other long-lived vertebrates. As we face unprecedented rates of environmental change, holistic approaches incorporating multiple concurrent threats and their impacts on different aspects of life history will be necessary to proactively conserve long-lived species.

The habitat quality paradox: loss of riparian forest cover leads to decreased risk of parasitism and improved body condition in an imperiled amphibian.

Amphibian declines are a global phenomenon but responses of populations to specific threats are often context dependent and mediated by individual physiological condition. Habitat degradation due to reduced riparian forest cover and parasitism are two threats facing the hellbender salamander (Cryptobranchus alleganiensis), but their potential to interact in nature remains largely unexplored. We investigated associations between forest cover, parasitic infection and physiology of hellbenders to test the hypotheses that physiological condition responds to infection and/or habitat degradation. We sampled 17 stream reaches in southwest Virginia, USA, on a year-round basis from 2013 to 2016 and recorded 841 captures of 405 unique hellbenders. At each capture we documented prevalence of two blood-associated parasites (a leech and trypanosome) and quantified up to three physiological condition indices (body condition, hematocrit, white blood cell [WBC] differentials). We used generalized linear mixed models to describe spatiotemporal variation in parasitic infection and each condition index. In general, living in the most heavily forested stream reaches, where hellbender density was highest, was associated with the greatest risk of parasitism, elevated neutrophil-to-lymphocyte (N:L) ratios and eosinophils, slightly lower hematocrit and lower mean body condition in hellbenders. All condition indices fluctuated temporally in a manner consistent with seasonal variation in hellbender metabolic demands and breeding phenology and were associated with land use during at least part of the year. Paradoxically, relatively low levels of forest cover appeared to confer a potential advantage to individuals in the form of release from parasites and improved body condition. Despite improved body condition, individuals from less forested areas failed to exhibit fluctuating body condition in response to spawning, which was typical in hellbenders from more forested habitats. We postulate this lack of fluctuation could be due to reduced conspecific competition or reproductive investment and/or high rates of filial cannibalism in response to declining forest cover.

Filial Cannibalism Leads to Chronic Nest Failure of Eastern Hellbender Salamanders (Cryptobranchus alleganiensis).

AbstractIn species that provide parental care, parents will sometimes cannibalize their own young (i.e., filial cannibalism). Here, we quantified the frequency of whole-clutch filial cannibalism in a species of giant salamander (eastern hellbender; Cryptobranchus alleganiensis) that has experienced precipitous population declines with unknown causes. We used underwater artificial nesting shelters deployed across a gradient of upstream forest cover to assess the fates of 182 nests at 10 sites over 8 years. We found strong evidence that nest failure rates increased at sites with low riparian forest cover in the upstream catchment. At several sites, reproductive failure was 100%, mainly due to cannibalism by the caring male. The high incidence of filial cannibalism at degraded sites was not explained by evolutionary hypotheses for filial cannibalism based on poor adult body condition or low reproductive value of small clutches. Instead, larger clutches at degraded sites were most vulnerable to cannibalism. We hypothesize that high frequencies of filial cannibalism of large clutches in areas with low forest cover could be related to changes in water chemistry or siltation that influence parental physiology or that reduce the viability of eggs. Importantly, our results identify chronic nest failure as a possible mechanism contributing to population declines and observed geriatric age structure in this imperiled species.

Warmer incubation temperatures and later lay-orders lead to shorter telomere lengths in wood duck (Aix sponsa) ducklings.

The environment that animals experience during development shapes phenotypic expression. In birds, two important aspects of the early-developmental environment are lay-order sequence and incubation. Later-laid eggs tend to produce weaker offspring, sometimes with compensatory mechanisms to accelerate their growth rate to catch-up to their siblings. Further, small decreases in incubation temperature slow down embryonic growth rates and lead to wide-ranging negative effects on many posthatch traits. Recently, telomeres, noncoding DNA sequences at the end of chromosomes, have been recognized as a potential proxy for fitness because longer telomeres are positively related to lifespan and individual quality in many animals, including birds. Although telomeres appear to be mechanistically linked to growth rate, little is known about how incubation temperature and lay-order may influence telomere length. We incubated wood duck (Aix sponsa) eggs at two ecologically-relevant temperatures (34.9°C and 36.2°C) and measured telomere length at hatch and 1 week after. We found that ducklings incubated at the lower temperature had longer telomeres than those incubated at the higher temperature both at hatch and 1 week later. Further, we found that later-laid eggs produced ducklings with shorter telomeres than those laid early in the lay-sequence, although lay-order was not related to embryonic developmental rate. This study contributes to our broader understanding of how parental effects can affect telomere length early in life. More work is needed to determine if these effects on telomere length persist until adulthood, and if they are associated with effects on fitness in this precocial species.

Hematological and histological changes from ingestion of Deepwater Horizon crude oil in zebra finches (Taeniopygia guttata).

Exposure to crude oil during spill events causes a variety of pathologic effects in birds, including oxidative injury to erythrocytes, which is characterized in some species by the formation of Heinz bodies and subsequent anemia. However, not all species appear to develop Heinz bodies or anemia when exposed to oil, and there are limited controlled experiments that use both light and electron microscopy to evaluate structural changes within erythrocytes following oil exposure. In this study, we orally dosed zebra finches (Taeniopygia guttata) with 3.3 or 10 mL/kg of artificially weathered Deepwater Horizon crude oil or 10 mL/kg of peanut oil (vehicle control) daily for 15 days. We found that birds receiving the highest dosage experienced a significant increase in reticulocyte percentage, mean corpuscular hemoglobin concentration, and liver mass, as well as inflammation of the gastrointestinal tract and lymphocyte proliferation in the spleen. However, we found no evidence of Heinz body formation based on both light and transmission electron microscopy. Although there was a tendency for packed cell volume and hemoglobin to decrease in birds from the high dose group compared to control and low dose groups, the changes were not statistically significant. Our results indicate that additional experimental dosing studies are needed to understand factors (e.g., dose- and species-specific sensitivity) and confounding variables (e.g., dispersants) that contribute to the presence and severity of anemia resulting from oil exposure in birds.

Pre-breeding androgen and glucocorticoid profiles in the eastern hellbender salamander (Cryptobranchus alleganiensis alleganiensis).

Seasonally breeding species exhibit cyclical changes in circulating steroid hormone profiles that correspond with changes to their reproductive behavior and ecology. Such information is critical to the conservation of imperiled and data-deficient species, such as the eastern hellbender salamander (Cryptobranchus alleganiensis alleganiensis). We determined changes in plasma testosterone (T), dihydrotestosterone (DHT), 11-ketotestosterone (11-KT), 11-ketoandrostenedione (11-KA), dehydroepiandrosterone (DHEA), cortisol, corticosterone, and progesterone (P4) during a four-month period preceding breeding in adult male and female eastern hellbenders. This pre-breeding period is characterized by increased diel movement and aggression by both sexes, follicular development and yolk production in females, and sperm production, territoriality, and nest site establishment in males. In both males and females, we observed a progressive increase in circulating T and DHT during the pre-reproductive season, both peaking in August (17 days before breeding), but concentrations of both hormones were higher in males. Conversely, 11-KT was higher in females, but did not vary significantly by date. These results suggest that T and DHT are the predominant androgens in eastern hellbenders and are likely important regulators of reproductive processes in both males and females. The detection of significant quantities of DHT and 11-KT in females is particularly interesting, considering that unlike T, neither of these androgens can be converted to estrogens. Therefore, it seems possible that aggression or some aspect of reproduction in the female eastern hellbender may be directly mediated by androgen signaling. Baseline cortisol did not vary throughout the pre-breeding period but was higher in females than males, and also became highly variable in females leading up to breeding. Progesterone, 11-KA, DHEA, and corticosterone were rarely or never detected, and thus, do not appear to be important during the pre-reproductive season. This study provides a physiological framework for future studies of hellbender reproductive biology, which could ultimately be important for their conservation.

Ultraviolet-assisted oiling assessment improves detection of oiled birds experiencing clinical signs of hemolytic anemia after exposure to the Deepwater Horizon oil spill.

While large-scale oil spills can cause acute mortality events in birds, there is increasing evidence that sublethal oil exposure can trigger physiological changes that have implications for individual performance and survival. Therefore, improved methods for identifying small amounts of oil on birds are needed. Because ultraviolet (UV) light can be used to identify thin crude oil films in water and on substrate that are not visually apparent under normal lighting conditions, we hypothesized that UV light could be useful for detecting small amounts of oil present on the plumage of birds. We evaluated black skimmers (Rynchops niger), brown pelicans (Pelecanus occidentalis), clapper rails (Rallus crepitans), great egrets (Ardea alba), and seaside sparrows (Ammodramus maritimus) exposed to areas affected by the Deepwater Horizon oil spill in the Gulf of Mexico as well as from reference areas from 20 June, 2010 to 23 February, 2011. When visually assessed without UV light, 19.6% of birds evaluated from areas affected by the spill were determined to be oiled (previously published data), whereas when examined under UV light, 56.3% of the same birds were determined to have oil exposure. Of 705 individuals examined in areas potentially impacted by the spill, we found that fluorescence under UV light assessment identified 259 oiled birds that appeared to be oil-free on visual exam, supporting its utility as a simple tool for improving detection of modestly oiled birds in the field. Further, UV assessment revealed an increase in qualitative severity of oiling (approximate % of body surface oiled) in 40% of birds compared to what was determined on visual exam. Additionally, black skimmers, brown pelicans, and great egrets exposed to oil as determined using UV light experienced oxidative injury to erythrocytes, had decreased numbers of circulating erythrocytes, and showed evidence of a regenerative hematological response in the form of increased reticulocytes. This evidence of adverse effects was similar to changes identified in birds with oil exposure as determined by visual examination without UV light, and is consistent with hemolytic anemia likely caused by oil exposure. Thus, UV assessment proved useful for enhancing detection of birds exposed to oil, but did not increase detection of birds experiencing clinical signs of anemia compared to standard visual oiling assessment. We conclude that UV light evaluation can help identify oil exposure in many birds that would otherwise be identified visually as unexposed during oil spill events.

Prolactin is related to incubation constancy and egg temperature following a disturbance in a precocial bird.

To maximize fitness, parents may trade-off time and energy between parental care and self-maintenance. In vertebrates, prolactin and corticosterone are two important hormones that regulate parental investment because they stimulate parental care and mobilize energy, respectively. Further, concentrations of both hormones change in response to disturbances. One of the most important parental care behaviors in birds is incubation, since small changes in egg temperature have large effects on offspring. We investigated how prolactin and corticosterone may mediate parental incubation constancy (i.e., the daily amount of time spent incubating eggs) and regulation of egg temperature. We collected blood samples from female wood ducks (Aix sponsa) near the start and end of the incubation period to measure baseline and stress-induced (30 min after capture and restraint) hormone concentrations. We also quantified incubation constancy and egg temperature using artificial egg temperature loggers. As expected, prolactin decreased and corticosterone increased after 30 min of capture and restraint. Corticosterone concentrations (baseline and stress-induced) were negatively related to body mass, but were not related to incubation constancy. In contrast, prolactin concentrations (baseline and stress-induced) were higher at the end than the start of the incubation period, and stress-induced prolactin concentrations were positively related to incubation constancy following a nest disturbance (i.e., capture). Further, prolactin (baseline and stress-induced) concentrations were positively related to egg temperatures, but only after the disturbance. These results suggest that prolactin may be associated with the regulation of parental incubation constancy and resulting heat-transfer after a disturbance, which may ultimately affect offspring development.

Cortisol is the predominant glucocorticoid in the giant paedomorphic hellbender salamander (Cryptobranchus alleganiensis).

Corticosterone is widely regarded to be the predominant glucocorticoid produced in amphibians. However, we recently described unusually low baseline and stress-induced corticosterone profiles in eastern hellbenders (Cryptobranchus alleganiensis alleganiensis), a giant, fully aquatic salamander. Here, we hypothesized that hellbenders might also produce cortisol, the predominant glucocorticoid used by fishes and non-rodent mammals. To test our hypothesis, we collected plasma samples in two field experiments and analyzed them using multiple analytical techniques to determine how plasma concentrations of cortisol and corticosterone co-varied after 1) physical restraint and 2) injection with adrenocorticotropic hormone (ACTH), the pituitary hormone responsible for triggering the release of glucocorticoids from amphibian interrenal glands. Using liquid chromatography-mass spectrometry, we found that baseline and restraint-induced plasma concentrations of cortisol were more than five times those of corticosterone. We then demonstrated that plasma concentrations of both glucocorticoids increased in response to ACTH in a dose-dependent manner, but cortisol concentrations were consistently higher (up to 10-fold) than corticosterone. Cortisol and corticosterone concentrations were not correlated with one another at basal or induced conditions. The extremely low plasma concentrations of corticosterone in hellbenders suggests that corticosterone could simply be a byproduct of cortisol production, and raises questions as to whether corticosterone has any distinct physiological function in hellbenders. Our results indicate that hellbenders produce cortisol as their predominant glucocorticoid, supporting a small and inconclusive body of literature indicating that some other amphibians may produce appreciable quantities of cortisol. We hypothesize that the use of cortisol by hellbenders could be an adaptation to their fully aquatic life history due to cortisol's ability to fulfill both mineralocorticoid and glucocorticoid functions, similar to its functions in fishes. Given the large number of amphibian species that are fully aquatic or have aquatic life stages, we suggest that the broadly held assumption that corticosterone is the predominant glucocorticoid in all amphibians requires further scrutiny. Ultimately, multi-species tests of this assumption will reveal the ecological factors that influenced the evolution of endocrine adaptations among amphibian lineages, and may provide insight into convergent evolution of endocrine traits in paedomorphic species.

Limited Support for Thyroid Hormone or Corticosterone Related Gene Expression as a Proximate Mechanism of Incubation Temperature-Dependent Phenotypes in Birds.

The conditions that animals experience during early development can have profound consequences for health and fitness. In birds, one of the most important aspects of development is egg incubation temperature. A small decrease in average temperature leads to various impacts on offspring phenotype, such as smaller body sizes, slower growth rates, and less efficient metabolic activity. Little is known, however, about the proximate mechanisms underlying these incubation temperature-induced phenotypic changes. Two important hormones which could play a proximate role are thyroid hormone and corticosterone, which mobilize stored energy reserves and coordinate the normal growth of tissues, particularly in the brain. Previous research shows that circulating blood concentrations of both hormones are influenced by incubation temperature, but the mechanism by which incubation temperature may lead to these changes is unknown. We hypothesized that incubation temperature induces changes in thyroid hormone and corticosterone regulation, leading to changes in expression of hormone-sensitive genes in the brain. To test this, we incubated wood duck (Aix sponsa) eggs at three different temperatures within the natural range (35.0, 35.8, and 37.0°C). We measured mRNA expression of thyroid hormone-related neuroendocrine endpoints (deiodinase 2/3, thyroid hormone receptor α/ß, neural regeneration related protein, and Krueppel-like factor 9) in newly hatched ducklings and corticosterone-related neuroendocrine endpoints (mineralocorticoid receptor, glucocorticoid receptor, cholecystokinin, and brain-derived neurotrophic factor) in 15 day-old ducklings using qPCR on brain tissue from the hippocampus and hypothalamus. Contrary to our predictions, we found that mRNA expression of thyroid hormone-related endpoints in both brain areas were largely unaffected by incubation temperature, although there was a trend for an inverse relationship between mRNA expression and incubation temperature for several genes in the hypothalamus. We also found that mineralocorticoid receptor mRNA expression in the hypothalamus was lower in ducklings incubated at the low relative to the high temperatures. This study is the first to evaluate the effects of incubation temperature on mRNA expression of neuroendocrine endpoints in the developing avian brain and suggests that these particular endpoints may be largely resistant to changes in incubation temperature. Thus, further research into the proximate mechanisms for incubation temperature-induced developmental plasticity is needed.

Selenium dynamics in headwater streams of the central Appalachian coalfield.

Coal mining can cause selenium (Se) contamination in US Appalachian streams, but linkages between water-column Se concentrations and Se bioaccumulation within Appalachian headwater streams have rarely been quantified. Using elevated specific conductance (SC) in stream water as an indicator of mining influence, we evaluated relationships between SC and Se concentrations in macroinvertebrates and examined dynamics of Se bioaccumulation in headwater streams. Twenty-three Appalachian streams were categorized into 3 stream types based on SC measurements: 1) reference streams with no coal-mining history; 2) mining-influenced, high-SC streams; and 3) mining-influenced, low-SC streams. Selenium concentrations in macroinvertebrates exhibited strong positive associations with both SC and dissolved Se concentrations in stream water. At 3 streams of each type, we further collected water, particulate matter (sediment, biofilm, leaf detritus), and macroinvertebrates and analyzed them for Se during 2 seasons. Enrichment, trophic transfer, and bioaccumulation factors were calculated and compared among stream types. Particulate matter and macroinvertebrates in mining-influenced streams accumulated high Se concentrations relative to reference streams. Concentrations were found at levels indicating Se to be a potential environmental stressor to aquatic life. Most Se enrichment, trophic transfer, and bioaccumulation factors were independent of season. Enrichment factors for biofilm and sediments and bioaccumulation factors for macroinvertebrate predators varied negatively with water-column Se. Our results increase scientific understanding of Se bioaccumulation processes in Appalachian headwater streams. Environ Toxicol Chem 2018;37:2714-2726. © 2018 SETAC.

Agricultural land use creates evolutionary traps for nesting turtles and is exacerbated by mercury pollution.

The rate of anthropogenic habitat conversion often exceeds the rate of natural ecological and evolutionary processes, which sometimes creates mismatches between environmental cues and adaptive behaviors. In numerous species of aquatic turtles, nest site selection is primarily based on cues related to high solar exposure, which leads females to select sites where humans have disturbed habitat. These disturbed sites are often contaminated by pollutants, such as mercury. Despite the fact that anthropogenic disturbances often co-occur, few studies have examined the interactive influence of major global changes on animal development. Using Chelydra serpentina, we investigated the individual and interactive effects of crop agriculture and mercury pollution on hatch success and offspring phenotype. We hypothesized that following nesting, rapid crop growth would shade and cool nests in agricultural fields and subsequently negatively impact embryonic development. Agricultural and control nests were similar in temperature at the time of oviposition, but temperatures diverged as crops grew: agricultural nests averaged 2.5°C cooler than control nests over the course of incubation. In laboratory and field experiments, we found that turtles incubated under agricultural thermal regimens took longer to hatch, hatched at smaller body sizes, lost more mass, and had lower posthatching growth rates. Additionally, thermal conditions associated with agricultural land use interacted with mercury contamination to decrease hatching success. To our knowledge, this is the first documentation of negative interactive effects of mercury pollution and habitat quality on early vertebrate development and highlights the importance of examining the combined influence of anthropogenic global changes on organisms.

Incubation temperature influences the behavioral traits of a young precocial bird.

The environment in which animals develop can have important consequences for their phenotype. In reptiles, incubation temperature is a critical aspect of the early developmental environment. Incubation temperature influences morphology, physiology, and behavior of non-avian reptiles, however, little is known about how incubation temperature influences offspring phenotype and behaviors important to avian survival. To investigate whether incubation temperature influences avian behaviors, we collected wood duck (Aix sponsa) eggs from the field and incubated them at three naturally occurring incubation temperatures (35.0, 35.8, and 37.0°C). We conducted multiple repeated behavioral trials on individual ducklings between 5 and 15 days post-hatch to assess activity, exploratory, and boldness behaviors, classified along a proactive-reactive continuum. We measured growth rates and circulating levels of baseline and stress-induced corticosterone levels to investigate possible physiological correlates of behavior. Ducklings incubated at the lowest temperature displayed more proactive behaviors than those incubated at the two higher temperatures. We also found that younger ducklings exhibited more proactive behavior than older ducklings and males exhibited more proactive behavior than females. Further, duckling behaviors were repeatable across time and contexts, indicative of a proactive-reactive continuum of behavioral tendencies. However, neither corticosterone levels nor growth rates were related to behavior. This provides some of the first evidence that incubation temperature, a critical parental effect, influences avian offspring behaviors that may be important for survival. Our results identify incubation temperature as a mechanism that contributes to the development of behavioral traits and, in part, explains how multiple behavioral types may be maintained within populations.

Modulators of mercury risk to wildlife and humans in the context of rapid global change.

Environmental mercury (Hg) contamination is an urgent global health threat. The complexity of Hg in the environment can hinder accurate determination of ecological and human health risks, particularly within the context of the rapid global changes that are altering many ecological processes, socioeconomic patterns, and other factors like infectious disease incidence, which can affect Hg exposures and health outcomes. However, the success of global Hg-reduction efforts depends on accurate assessments of their effectiveness in reducing health risks. In this paper, we examine the role that key extrinsic and intrinsic drivers play on several aspects of Hg risk to humans and organisms in the environment. We do so within three key domains of ecological and human health risk. First, we examine how extrinsic global change drivers influence pathways of Hg bioaccumulation and biomagnification through food webs. Next, we describe how extrinsic socioeconomic drivers at a global scale, and intrinsic individual-level drivers, influence human Hg exposure. Finally, we address how the adverse health effects of Hg in humans and wildlife are modulated by a range of extrinsic and intrinsic drivers within the context of rapid global change. Incorporating components of these three domains into research and monitoring will facilitate a more holistic understanding of how ecological and societal drivers interact to influence Hg health risks.

Hematological indices of injury to lightly oiled birds from the Deepwater Horizon oil spill.

Avian mortality events are common following large-scale oil spills. However, the sublethal effects of oil on birds exposed to light external oiling are not clearly understood. We found that American oystercatchers (area of potential impact n = 42, reference n = 21), black skimmers (area of potential impact n = 121, reference n = 88), brown pelicans (area of potential impact n = 91, reference n = 48), and great egrets (area of potential impact n = 57, reference n = 47) captured between 20 June 2010 and 23 February 2011 following the Deepwater Horizon oil spill experienced oxidative injury to erythrocytes, had decreased volume of circulating erythrocytes, and showed evidence of a regenerative hematological response in the form of increased reticulocytes compared with reference populations. Erythrocytic inclusions consistent with Heinz bodies were present almost exclusively in birds from sites impacted with oil, a finding pathognomonic for oxidative injury to erythrocytes. Average packed cell volumes were 4 to 19% lower and average reticulocyte counts were 27 to 40% higher in birds with visible external oil than birds from reference sites. These findings provide evidence that small amounts of external oil exposure are associated with hemolytic anemia. Furthermore, we found that some birds captured from the area impacted by the spill but with no visible oiling also had erythrocytic inclusion bodies, increased reticulocytes, and reduced packed cell volumes when compared with birds from reference sites. Thus, birds suffered hematologic injury despite no visible oil at the time of capture. Together, these findings suggest that adverse effects of oil spills on birds may be more widespread than estimates based on avian mortality or severe visible oiling. Environ Toxicol Chem 2018;37:451-461. © 2017 SETAC.

Mercury alters initiation and construction of nests by zebra finches, but not incubation or provisioning behaviors.

Mercury is an environmental contaminant that impairs avian reproduction, but the behavioral and physiological mechanisms underlying this effect are poorly understood. The objective of this study was to determine whether lifetime dietary exposure to mercury (1.2 µg/g wet weight in food) impacted avian parental behaviors, and how this might influence reproductive success. To distinguish between the direct effects of mercury on parents and offspring, we created four treatment groups of captive-bred zebra finches (Taeniopygia guttata), with control and mercury-exposed adults raising cross-fostered control or mercury-exposed eggs (from maternal transfer). Control parents were 23% more likely to fledge young than parents exposed to mercury, regardless of egg exposure. Mercury-exposed parents were less likely to initiate nests than controls and spent less time constructing them. Nests of mercury-exposed pairs were lighter, possibly due to an impaired ability to bring nest material into the nestbox. However, nest temperature, incubation behavior, and provisioning rate did not differ between parental treatments. Unexposed control eggs tended to have shorter incubation periods and higher hatching success than mercury-exposed eggs, but there was no effect of parental exposure on these parameters. We accidentally discovered that parent finches transfer some of their body burden of mercury to nestlings during feeding through secretion in the crop. These results suggest that, in mercury-exposed songbirds, pre-laying parental behaviors, combined with direct exposure of embryos to mercury, likely contribute to reduced reproductive success and should be considered in future studies. Further research is warranted in field settings, where parents are exposed to greater environmental challenges and subtle behavioral differences might have more serious consequences than were observed in captivity.

High levels of maternally transferred mercury disrupt magnetic responses of snapping turtle hatchlings (Chelydra serpentina).

The Earth's magnetic field is involved in spatial behaviours ranging from long-distance migration to non-goal directed behaviours, such as spontaneous magnetic alignment (SMA). Mercury is a harmful pollutant most often generated from anthropogenic sources that can bio-accumulate in animal tissue over a lifetime. We compared SMA of hatchling snapping turtles from mothers captured at reference (i.e., low mercury) and mercury contaminated sites. Reference turtles showed radio frequency-dependent SMA along the north-south axis, consistent with previous studies of SMA, while turtles with high levels of maternally inherited mercury failed to show consistent magnetic alignment. In contrast, there was no difference between reference and mercury exposed turtles on standard performance measures. The magnetic field plays an important role in animal orientation behaviour and may also help to integrate spatial information from a variety of sensory modalities. As a consequence, mercury may compromise the performance of turtles in a wide variety of spatial tasks. Future research is needed to determine the threshold for mercury effects on snapping turtles, whether mercury exposure compromises spatial behaviour of adult turtles, and whether mercury has a direct effect on the magnetoreception mechanism(s) that mediate SMA or a more general effect on the nervous system.

Effects of Echinostoma trivolvis metacercariae infection during development and metamorphosis of the wood frog (Lithobates sylvaticus).

Many organisms face energetic trade-offs between defense against parasites and other host processes that may determine overall consequences of infection. These trade-offs may be particularly evident during unfavorable environmental conditions or energetically demanding life history stages. Amphibian metamorphosis, an ecologically important developmental period, is associated with drastic morphological and physiological changes and substantial energetic costs. Effects of the trematode parasite Echinostoma trivolvis have been documented during early amphibian development, but effects during later development and metamorphosis are largely unknown. Using a laboratory experiment, we examined the energetic costs of late development and metamorphosis coupled with E. trivolvis infection in wood frogs, Lithobates [=Rana] sylvaticus. Echinostoma infection intensity did not differ between tadpoles examined prior to and after completing metamorphosis, suggesting that metacercariae were retained through metamorphosis. Infection with E. trivolvis contributed to a slower growth rate and longer development period prior to the initiation of metamorphosis. In contrast, E. trivolvis infection did not affect energy expenditure during late development or metamorphosis. Possible explanations for these results include the presence of parasites not interfering with pronephros degradation during metamorphosis or the mesonephros compensating for any parasite damage. Overall, the energetic costs of metamorphosis for wood frogs were comparable to other species with similar life history traits, but differed from a species with a much shorter duration of metamorphic climax. Our findings contribute to understanding the possible role of energetic trade-offs between parasite defense and host processes by considering parasite infection with simultaneous energetic demands during a sensitive period of development.

Spatial differences in trace element bioaccumulation in turtles exposed to a partially remediated coal fly ash spill.

Large-scale releases of environmental contaminants from industrial facilities can cause considerable damage to surrounding ecosystems and require remediation. The expense and/or undesirable environmental side effects of physical removal may constrain remediation efforts. In 2008, approximately 4.1 million m3 of fly ash were released into the Emory River at a coal-burning power plant in Kingston, Tennessee, USA. Approximately 390 000 m3 of fly ash were not removed (hereafter "residual ash"), to avoid disturbing underlying legacy contamination from unrelated historical industrial activity. In 2011 and 2012, the authors measured trace element concentrations in an assemblage of freshwater turtles in 2 rivers impacted by the spill and in a third river that was unaffected. Concentrations of arsenic, copper, iron, mercury, manganese, selenium, and zinc were higher in turtles from rivers affected by the spill but low relative to concentrations known to be toxic to other vertebrates. Concentrations of some trace elements decreased with distance from the original spill site but were not strongly affected by nearby volumes of residual ash. Among-species differences in trace element bioaccumulation and/or the relatively low spatial resolution of available data on residual ash volumes may have obscured this effect. The results suggest that the spill influenced turtle bioaccumulation of trace elements but that distance from the spill site may be a more important factor than residual ash in influencing postremediation bioaccumulation. Environ Toxicol Chem 2017;36:201-211. © 2016 SETAC.

Repeatability and sources of variation of the bacteria-killing assay in the common snapping turtle.

Research on reptile ecoimmunology lags behind that on other vertebrates, despite the importance of such studies for conservation and evolution. Because the innate immune system is highly conserved across vertebrate lineages, assessments of its performance may be particularly useful in reptiles. The bacteria-killing assay requires a single, small blood sample and quantifies an individual's ability to kill microorganisms. The assay's construct validity and interpretability make it an attractive measure of innate immunity, but it requires proper optimization and sample storage. We optimized this assay for the common snapping turtle (Chelydra serpentina) to assess the repeatability of the assay and the effects of freezing and thawing on bactericidal capacity. We determined whether age (adult female and hatchlings) or incubation temperature influenced bactericidal capacity. We found that the assay was repeatable and that freezing plasma samples for 6 weeks at -80°C did not decrease bactericidal capacity nor did a single 30-min thaw and subsequent refreezing. However, we detected subtle interassay variation and results from one assay were 5-6% greater than those from the other two. Adult females had significantly greater bactericidal ability than hatchlings and we found no relationship between incubation temperature and bactericidal capacity. This assay is a useful tool in snapping turtles and may have applicability in other reptiles. However, species-specific optimization is required to ensure that variation among individuals exceeds interassay variation. Consideration should be given to optimization conditions that facilitate comparisons between or within groups, particularly groups that differ considerably in bactericidal capacity.