ECOLOGY AND PHYSIOLOGY OF HIBERNATION AND OVERWINTERING AMONG FRESHWATER FISHES, TURTLES, AND SNAKES

1. Freshwater fishes are the most northerly of freshwater ectotherms, followed by frogs. North American freshwater snakes, turtles, and salamanders do not range farther north than southernmost Canada. 2. Freezing and desiccation are the main challenges during terrestrial hibernation of ectotherms. Oxygen depletion, water balance, and ionic balance are the major problems for air breathing ectotherms that hibernate underwater. 3. The importance of accumulation of energy stores for overwintering among fishes depends upon the length and severity of the winters, whether or not there is springtime reproduction, body size, latitude, and the availability and use of food during overwintering. 4. Fishes can decrease energy demands during the winter by reductions in activity, metabolic depression, and entrance in semi-torpidity. 5. Adaptations for coping with hypoxia and anoxia among overwintering freshwater fishes may include metabolic depression, a decrease in blood O₂ affinity, microhabitat selection, air breathing, short-distance migration, biochemical modifications aimed at adjusting glycolytic rates, and alcoholic fermentation. 6. Freshwater turtles have a worldwide northern limit of approximately 50° N, which means that some species spend about half of their lives hibernating. 7. Aquatic turtles normally hibernate underwater, although occasionally they hibernate on land. In water they usually hibernate in a hypoxic or anoxic (mud) environment and in relatively shallow water. Wintertime movements of unknown frequency occur in some species. 8. The hatchlings of many turtle species can overwinter in the nest. Among northern species this behaviour is most common among painted turtles, whose hatchlings can withstand freezing. 9. Mortality among adult turtles is probably highest during the hibernation cycle. 10. Temperature appears to the most important cue for entry and exit from hibernation among freshwater turtles. 11. Little is known of the energetics of overwintering turtles. Energy stores for overwintering may be more important at lower latitudes than at higher ones, due to the higher metabolic rates of overwintering, but non-feeding, southern turtles. 12. The ability of turtles to tolerate submergence is a function of temperature, degree of water oxygenation, latitude of origin, efficacy of extrapulmonary respiratory pathways, and metabolic rate. 13. For turtles that hibernate in an anoxic hibernaculum, and for those without sufficient extrapulmonary uptake of O₂ to allow metabolism to be completely aerobic, the most important physiological perturbation is an acidosis developed from a continuing production of lactate. If sufficient O₂ can be obtained, the most likely factors limiting hibernation time are water balance and ion balance. 14. Mechanisms of turtles for coping with acidosis include metabolic depression, integumental CO₂ loss, bicarbonate buffering, and changes in ion concentrations that minimize the decrease in SID (strong ion difference). The most important among the latter are a decrease in plasma [Cl^-] and large increases in plasma calcium and magnesium. 15. Turtles are unique among reptiles in their ability to maintain both cardiovascular and nervous system function during prolonged anoxia. 16. Turtles gain weight from water uptake during submerged hibernation, but apparently maintain some kidney function; however, osmoregulation is one of the least known areas of the physiology of hibernation. 17. Recovery of turtles upon emergence commences with a rapid hyperventilatory compensation of pH, followed by a slower adjustment of ion levels. Basking speeds recovery greatly. 18. While hibernation of turtles in the northern parts of their ranges is most likely very stressful physiologically, northern range limits are more likely to be determined by reproductive restraints than by the rigors of extended hibernation. 19. The superior ability of turtles to tolerate anoxia may be more the result of an annual hibernation than of their diving habits during active periods of the year. 20. Freshwater snakes usually hibernate on land. However, they appear to be capable of aquatic hibernation and may not do so because of the risk of death from anoxia. 21. Some species of terrestrial snakes are known to hibernate underwater, and are able to do so in the laboratory for months. In the field, this behaviour is considered opportunistic, as there is no evidence to suggest that any snakes can tolerate extended anoxia.     

Physiological ecology of overwintering in hatchling turtles

Temperate species of turtles hatch from eggs in late summer. The hatchlings of some species leave their natal nest to hibernate elsewhere on land or under water, whereas others usually remain inside the nest until spring; thus, post-hatching behavior strongly influences the hibernation ecology and physiology of this age class. Little is known about the habitats of and environmental conditions affecting aquatic hibernators, although laboratory studies suggest that chronically hypoxic sites are inhospitable to hatchlings. Field biologists have long been intrigued by the environmental conditions survived by hatchlings using terrestrial hibernacula, especially nests that ultimately serve as winter refugia. Hatchlings are unable to feed, although as metabolism is greatly reduced in hibernation, they are not at risk of starvation. Dehydration and injury from cold are more formidable challenges. Differential tolerances to these stressors may explain variation in hatchling overwintering habits among turtle taxa. Much study has been devoted to the cold-hardiness adaptations exhibited by terrestrial hibernators. All tolerate a degree of chilling, but survival of frost exposure depends on either freeze avoidance through supercooling or freeze tolerance. Freeze avoidance is promoted by behavioral, anatomical, and physiological features that minimize risk of inoculation by ice and ice-nucleating agents. Freeze tolerance is promoted by a complex suite of molecular, biochemical, and physiological responses enabling certain organisms to survive the freezing and thawing of extracellular fluids. Some species apparently can switch between freeze avoidance or freeze tolerance, the mode utilized in a particular instance of chilling depending on prevailing physiological and environmental conditions.     

Frogs and turtles: different ectotherm overwintering strategies

The ability of frogs and turtles to overwinter and to survive hypoxia and anoxia has long been a topic of interest. While data remains scant, the emerging picture shows fundamentally different approaches to overwintering in these two groups of ectotherms. Frogs are far more limited by availability of oxygen than are turtles, even at near-freezing ambient temperatures. The reasons for this probably involve the vastly greater cutaneous permeability of the former. With their extreme tolerance of anoxia and profound suppression of metabolism, overwintering in turtles should not be viewed as simply prolonged diving but rather as ectotherm hibernation. Their incredible diving capabilities are merely a spin-off of a successful overwintering strategy. The following discussion reviews the major physiological mechanisms involved in the overwintering strategies of these two ectotherm groups.     

First records of dive durations for a hibernating sea turtle

The first published record, from the early 1970s, of hibernation in sea turtles is based on the reports of the indigenous Indians and fishermen from Mexico, who hunted dormant green turtles (Chelonia mydas) in the Gulf of California. However, there were no successful attempts to investigate the biology of this particular behaviour further. Hence, data such as the exact duration and energetic requirements of dormant winter submergences are lacking. We used new satellite relay data loggers to obtain the first records of up to 7h long dives of a loggerhead turtle (Caretta caretta) overwintering in Greek waters. These represent the longest dives ever reported for a diving marine vertebrate. There is strong evidence that the dives were aerobic, because the turtle surfaced only for short intervals and before the calculated oxygen stores were depleted. This evidence suggests that the common belief that sea turtles hibernate underwater, as some freshwater turtles do, is incorrect.     

A novel hypothesis for the adaptive maintenance of environmental sex determination in a turtle

Temperature-dependent sex determination (TSD) is widespread in reptiles, yet its adaptive significance and mechanisms for its maintenance remain obscure and controversial. Comparative analyses identify an ancient origin of TSD in turtles, crocodiles and tuatara, suggesting that this trait should be advantageous in order to persist. Based on this assumption, researchers primarily, and with minimal success, have employed a model to examine sex-specific variation in hatchling phenotypes and fitness generated by different incubation conditions. The unwavering focus on different incubation conditions may be misplaced at least in the many turtle species in which hatchlings overwinter in the natal nest. If overwintering temperatures differentially affect fitness of male and female hatchlings, TSD might be maintained adaptively by enabling embryos to develop as the sex best suited to those overwintering conditions. We test this novel hypothesis using the painted turtle (Chrysemys picta), a species with TSD in which eggs hatch in late summer and hatchlings remain within nests until the following spring. We used a split-clutch design to expose field-incubated hatchlings to warm and cool overwintering (autumn–winter–spring) regimes in the laboratory and measured metabolic rates, energy use, body size and mortality of male and female hatchlings. While overall mortality rates were low, males exposed to warmer overwintering regimes had significantly higher metabolic rates and used more residual yolk than females, whereas the reverse occurred in the cool temperature regime. Hatchlings from mixed-sex nests exhibited similar sex-specific trends and, crucially, they were less energy efficient and grew less than same-sex hatchlings that originated from single-sex clutches. Such sex- and incubation-specific physiological adaptation to winter temperatures may enhance fitness and even extend the northern range of many species that overwinter terrestrially.     

Survival and physiological responses of hatchling blanding's turtles (Emydoidea blandingii) to submergence in normoxic and hypoxic water under simulated winter conditions

Overwintering habits of hatchling Blanding's turtles (Emydoidea blandingii) are unknown. To determine whether these turtles are able to survive winter in aquatic habitats, we submerged hatchlings in normoxic (155 mmHg Po2) and hypoxic (6 mmHg Po2) water at 4 degrees C, recording survival times and measuring changes in key physiological variables. For comparison, we simultaneously studied hatchling softshell (Apalone spinifera) and snapping (Chelydra serpentina) turtles, which are known to overwinter in aquatic habitats. In normoxic water, C. serpentina and A. spinifera survived to the termination of the experiment (76 and 77 d, respectively). Approximately one-third of the E. blandingii died during 75 d of normoxic submergence, but the cause of mortality was unclear. In hypoxic water, average survival times were 6 d for A. spinifera, 13 d for E. blandingii, and 19 d for C. serpentina. Mortality during hypoxic submergence was probably caused by metabolic acidosis, which resulted from accumulated lactate. Unlike the case with adult turtles, our hatchlings did not increase plasma calcium and magnesium, nor did they sequester lactate within the shell. Our results suggest that hatchling E. blandingii are not particularly well suited to hibernation in hypoxic aquatic habitats.     

Hibernal thermal ecology of eastern box turtles within a managed forest landscape

Box turtles are being extirpated from much of their former range and remaining populations often live in association with anthropogenically altered habitats. This is particularly evident at the northern distributional limit of eastern box turtles (Terrapene carolina carolina) and is an important factor to consider during the winter months when their ability to respond to microclimatic change is limited. Using temperature dataloggers, we studied the hibernal microclimate of box turtles and associated habitat following timber harvests. We monitored the body temperatures of 38 eastern box turtles and collected detailed air and soil profile temperatures of 12 box turtle hibernacula, 6 clearcuts, and 6 adjacent forested areas during the hibernal season (winter 2009–2010). We partitioned the hibernal season into 2 biologically significant thermal periods: hibernation and emergence. The mean hibernation body temperature averaged (3.28° C, SE = 0.09) and corresponded to an average depth of 10 cm. Clearcuts were consistently colder ( = 1.91° C) than forests ( = 2.68° C) and hibernacula ( = 2.77° C) during hibernation, but became the warmest areas during emergence ( = 9.96° C). We found that in the average clearcut, turtles could burrow to approximately 20 cm to attain the average hibernation body temperature or to approximately 15 cm to attain a body temperature no different than those overwintering on colder, northeast-facing slopes in the forest ( = 2.83° C). Alternatively, we found that southwest-facing slopes were warmer and if turtles chose to overwinter only in clearcuts on those slopes, they could remain shallower. All but 1 turtle overwintered in forested areas; however, our study suggests that some timber harvested areas offer various microhabitats exploitable by hibernating box turtles based on soil profile temperatures, slope aspect, and depth of hibernation. © 2012 The Wildlife Society.     

The 'lost years' of green turtles: using stable isotopes to study cryptic lifestages

Ignorance of the location or inaccessible locations of lifestages can impede the study and management of species. We used stable isotopes of carbon and nitrogen to identify the habitats and diets and to estimate the duration of a 'missing' lifestage: the early juvenile stage of the green turtle, Chelonia mydas. Stable isotopes in scute from young herbivorous green turtles in shallow-water habitats revealed that they spend 3-5 years as carnivores in oceanic habitats before making a rapid ontogenetic shift in diet and habitat. Stable isotopes in persistent and continuously growing tissues, such as some fish scales, bird bills and claws and mammal hair and claws, can be used to evaluate the ecology of inaccessible lifestages.     

Energy use and management of energy reserves in hatchling turtles (Chrysemys picta) exposed to variable winter conditions

The painted turtle (Chrysemys picta) is an especially useful organism in the study of metabolic regulation during dormancy because it is sustained by finite energy reserves from hatching until emerging from its nest, about nine months later. In this study we subjected overwintering C. picta hatchlings to 4, 10, or 15 °C, temperatures simulating cold, mild, and warm winters, respectively, to investigate how various energy reserves are impacted by differential metabolic demands. An energy budget based on seasonal changes in caloric content showed that these turtles consumed an average of 0.39, 0.75, or 1.21 kJ g⁻¹, respectively, during the 6-month period of simulated hibernation. These estimates of energy use agreed reasonably well with estimates based solely on respirometric data. Unexpectedly, turtles in autumn contained little residual yolk, none of which was consumed by turtles in the cold- and mild-winter groups, this finding contradicting the widely held belief that residual yolk plays an important, direct role in the survival of turtles that overwinter inside their natal nest. By contrast, a marked reduction in dry mass of both liver and carcass attested to their importance in fueling metabolism and, indeed, catabolism of substrates from these components accounted for 31–52 and 35–63%, respectively, of the energetic cost of overwintering. The greater dependence on carcass reserves and relatively poor physiological condition of turtles in the mild- and warm-winter groups implies that metabolic demands imposed by high environmental temperatures would likely constrain post-emergence fitness.     

Does maternal oviposition site influence offspring dispersal to suitable habitat?

Orientation and dispersal to suitable habitat affects fitness in many animals, but the factors that govern these behaviors are poorly understood. In many turtle species, hatchlings must orient and disperse to suitable aquatic habitat immediately after emergence from subterranean nests. Thus, the location of nest sites relative to aquatic habitats ideally should be associated with the direction of hatchling dispersal. At our study site, painted turtles (Chrysemys picta) nest to the west (on an island) and east (on the mainland) of a wetland, which determines the direction that hatchlings must travel to reach suitable aquatic habitat. To determine if hatchling orientation is intrinsically influenced by the location where their mothers nest, we employed a two-part cross-fostering experiment in the field, whereby half the eggs laid in mainland nests were swapped with half the eggs laid in island nests. Moreover, because C. picta hatchlings overwinter inside their nests, we performed a second cross-fostering experiment to fully decouple the effects of (1) the maternally chosen nest location, (2) the embryonic developmental location, and (3) the overwinter location. We released hatchlings into a circular arena in the field and found that turtles generally dispersed in a westerly direction, regardless of the maternally chosen nest location and independent of the locations of embryonic development and overwintering. Although this westerly direction was towards suitable aquatic habitat, we could not distinguish whether naïve hatchling turtles (i) use environmental cues/stimuli to orient their movement, or (ii) have an intrinsic bias to orient west in the absence of stimuli. Nevertheless, these findings suggest that the orientation behavior of naïve hatchling turtles during terrestrial dispersal is not dependent upon the location of maternally-chosen nest sites.     

Retinal Anatomy of hatchling sea turtles: Anatomical specializations and behavioral correlates

The eyes of three species of sea turtle hatchlings (loggerheads, green turtles, and leatherbacks) possess visual streaks, areas of densely packed ganglion cells running along the antero‐posterior retinal axis. These probably function to provide heightened visual acuity along the horizon. The vertical extent and absolute concentration of cells within the streak, compared to the rest of the retina, differ among the species. Leatherbacks have an additional specialized region (area temporalis) that might enhance their ability to detect prey below them in the water column. Green turtles and loggerheads, but not leatherbacks, show compensatory eye reflexes that keep the visual streak horizontal. Species differences in retinal structure and eye reflexes probably reflect their unique specializations in visual ecology and behaviour.     

Effects of wetland connectivity on overwintering and movement behaviours of Australian freshwater turtles

Freshwater turtles are important consumers in Australian freshwater ecosystems. They serve as scavengers, nutrient regulators, and as food sources and Totems for Traditional Owners throughout Australia. Despite their importance, most Australian freshwater turtle species are declining. The impact of winter wetland drying on turtle populations remains unknown, and winter exposure of hibernating turtles may be an important additional source of mortality. We aimed to examine turtle responses to seasonal and episodic wetland drying in wetlands using acoustic telemetry and active trapping. Wetlands were chosen that spanned a range of hydrological connectivity to the adjacent Edward/Kolety-Wakool River. We found that tagged Emydura macquarii typically exit wetlands disconnected from the adjacent permanent river prior to winter, and overwinter in the river. Female E. macquarii rapidly re-entered ‘home’ wetlands (wetlands in which they were initially tagged) the following spring, whereas males tended to leave the study area, returning occasionally. Although we were not able to evaluate a winter drying event, one of the wetlands experienced partial summer drying. All three local turtle species (E. macquarii, Chelodina expansa, C. longicollis) exited the wetland long before winter drying would have become a potential threat. Our results suggest that turtles in this system may be protected from winter wetland drying because they move to the adjacent permanent river prior to winter. Spending the winter in the river channel reduces the risks of being trapped in a drying wetland as temperatures drop in winter.     

Population genetics and phylogeography of sea turtles

The seven species of sea turtles occupy a diversity of niches, and have a history tracing back over 100 million years, yet all share basic life-history features, including exceptional navigation skills and periodic migrations from feeding to breeding habitats. Here, we review the biogeographic, behavioural, and ecological factors that shape the distribution of genetic diversity in sea turtles. Natal homing, wherein turtles return to their region of origin for mating and nesting, has been demonstrated with mtDNA sequences. These maternally inherited markers show strong population structure among nesting colonies while nuclear loci reveal a contrasting pattern of male-mediated gene flow, a phenomenon termed 'complex population structure'. Mixed-stock analyses indicate that multiple nesting colonies can contribute to feeding aggregates, such that exploitation of turtles in these habitats can reduce breeding populations across the region. The mtDNA data also demonstrate migrations across entire ocean basins, some of the longest movements of marine vertebrates. Multiple paternity occurs at reported rates of 0-100%, and can vary by as much as 9-100% within species. Hybridization in almost every combination among members of the Cheloniidae has been documented but the frequency and ultimate ramifications of hybridization are not clear. The global phylogeography of sea turtles reveals a gradient based on habitat preference and thermal regime. The cold-tolerant leatherback turtle (Dermochelys coriacea) shows no evolutionary partitions between Indo-Pacific and Atlantic populations, while the tropical green (Chelonia mydas), hawksbill (Eretmochelys imbricata), and ridleys (Lepidochelys olivacea vs. L. kempi) have ancient separations between oceans. Ridleys and loggerhead (Caretta caretta) also show more recent colonization between ocean basins, probably mediated by warm-water gyres that occasionally traverse the frigid upwelling zone in southern Africa. These rare events may be sufficient to prevent allopatric speciation under contemporary geographic and climatic conditions. Genetic studies have advanced our understanding of marine turtle biology and evolution, but significant gaps persist and provide challenges for the next generation of sea turtle geneticists.     

Adaptations to terrestrial overwintering of hatchling northern map turtles,<Emphasis Type="Italic"> Graptemys geographica</Emphasis>

We conducted a 3-year field and laboratory study of winter biology in hatchlings of the northern map turtle (Graptemys geographica). At our study area in northern Indiana, hatchlings routinely overwintered in their natal nests, emerging after the weather warmed in spring. Winter survival was excellent despite the fact that hatchlings were exposed frequently to subfreezing temperatures (to –5.4 °C). In the laboratory, cold-acclimated hatchlings exhibited low rates of evaporative water loss (mean=2.0 mg g^–1 day^–1), which would enable them to conserve body water during winter. Laboratory-reared hatchlings were intolerant of freezing at –2.5 °C for 24 h, conditions that are readily survived by freeze-tolerant species of turtles. Winter survival of hatchling G. geographica probably depended on their extensive capacity for supercooling (to –14.8 °C) and their well-developed resistance to inoculative freezing, which may occur when hatchlings contact ice and ice-nucleating agents present in nesting soil. Supercooled hatchlings survived a brief exposure to –8 °C. Others, held at –6 °C for 5 days, maintained ATP concentrations at control levels, although they did accumulate lactate and glucose, probably in response to tissue hypoxia. Therefore, anoxia tolerance, as evidenced by the viability of hatchlings exposed to N₂ gas for 8 days, may promote survival during exposure to subfreezing temperatures.Abbreviations EWL evaporative water loss - FP_eq equilibrium freezing point - INA ice-nucleating agents - T_c temperature of crystallizationCommunicated by L.C.-H. Wang     

Natural freeze-tolerance in hatchling painted turtles?

Hatchlings of the North American painted turtle (Family Emydidae: Chrysemys picta) typically spend their first winter of life inside a shallow, subterranean hibernaculum (the natal nest) where life-threatening conditions of ice and cold commonly occur. Although a popular opinion holds that neonates exploit a tolerance for freezing to survive the rigors of winter, hatchlings are more likely to withstand exposure to ice and cold by avoiding freezing altogether-and to do so without the benefit of an antifreeze. In the interval between hatching by turtles in late summer and the onset of wintery weather in November or December, the integument of the animals becomes highly resistant to the penetration of ice into body compartments from surrounding soil, and the turtles also purge their bodies of catalysts for the formation of ice. These two adjustments, taken together, enable the animals to supercool to temperatures below those that they routinely experience in nature. However, cardiac function in hatchlings is diminished at subzero temperatures, thereby compromising the delivery of oxygen to peripheral tissues and eliciting an increase in reliance by those tissues on anaerobic metabolism for the provision of ATP. The resulting increase in production of lactic acid may disrupt acid/base balance and lead to death even in animals that remain unfrozen. Although an ability to undergo supercooling may be key to survival by overwintering turtles in northerly populations, a similar capacity to resist inoculation and undergo supercooling characterizes animals from a population near the southern limit of distribution, where winters are relatively benign. Thus, the suite of characters enabling hatchlings to withstand exposure to ice and cold may have been acquired prior to the northward dispersal of the species at the end of the Pleistocene, and the characters may not have originated as adaptations specifically to the challenges of winter.     

The ontogeny of morphological defenses in Kemp's ridley (Lepidochelys kempii) and loggerhead (Caretta caretta) sea turtles

Marine turtles are large reptiles that compensate for high juvenile mortality by producing hundreds of hatchlings during a long reproductive lifespan. Most hatchlings are taken by predators during their migration to, and while resident in, the open ocean. Their survival depends upon crypticity, minimizing movement to avoid detection, and foraging efficiently to grow to a size too difficult for predators to either handle or swallow. While these behavioral antipredator tactics are known, changes in morphology accompanying growth may also improve survival prospects. These have been only superficially described in the literature. Here, we compare the similarities and differences in presumed morphological defenses of growing loggerhead (Caretta caretta) and Kemp's ridley (Lepidochelys kempii) posthatchlings, related species that differ in growth rate, timing of habitat shift (the return from oceanic to neritic locations), and size at maturity. In both species, vertebral spination and carapace widening increase disproportionally as small turtles grow, but later in ontogeny, the spines regress, sooner in ridley than in loggerhead turtles. Carapace widening occurs in both species but loggerheads are always longer than they are wide whereas in Kemp's ridley turtles, the carapace becomes as wide as long. Our analysis indicates that these changes are unrelated to when each species shifts habitat but are related to turtle size. We hypothesize that the spines function in small turtles as an early defense against gape‐limited predators, but changes in body shape function throughout ontogeny—initially to make small turtles too wide to swallow and later by presenting an almost flat and hardened surface that large predators (such as a sharks) are unable to grasp. The extremely wide carapace of the Kemp's ridley may compensate for its smaller adult size (and presumed greater vulnerability) than the loggerhead. J. Morphol. 276:929–940, 2015. © 2015 Wiley Periodicals, Inc.