Cold tolerance in sealworm (Pseudoterranova decipiens) due to heat-shock adaptations

SUMMARYThird-stage larvae of Pseudoterranova decipiens commonly infect whitefish such as cod, and the parasite can be transferred to humans through lightly prepared (sushi) meals. Because little is known about the nematode's cold tolerance capacity, we examined the nematode's ability to supercool, and whether or not cold acclimation could induce physiological changes that might increase its ability to tolerate freezing conditions. Even if third-stage Pseudoterranova decipiens larvae have some supercooling ability, they show no potential for freezing avoidance because they are not able to withstand inoculative freezing. Still, they have the ability to survive freezing at high subzero temperatures, something which suggests that these nematodes have a moderate freeze tolerance. We also show that acclimation to high temperatures triggers trehalose accumulation to an even greater extent than cold acclimation. Trehalose is a potential cryoprotectant which has been shown to play a vital role in the freeze tolerance of nematodes. We suggest that the trehalose accumulation observed for the cold acclimation is a general response to thermal stress, and that the nematode's moderate freeze tolerance may be acquired through adaptation to heat rather than coldness.

Inoculative freezing promotes winter survival in hatchling diamondback terrapin, Malaclemys terrapin

We investigated the hibernation ecology and cold hardiness of hatchling diamondback terrapins, Malaclemys terrapin (Schoepf, 1793), an estuarine species that reaches 42°N along the Atlantic Ocean. During 3 years of study, about 50% of the nests we monitored harboured hatchlings during winter, and the majority (87%) of these individuals survived despite being intermittently exposed to subfreezing temperatures. Most such exposures were brief (ca. 12 h) and mild (minimum temperature: ca. –1.2 °C); however, turtles were occasionally subjected to longer chilling episodes and lower temperatures. In laboratory experiments, hatchlings supercooled extensively, attaining ca. –15 °C before spontaneously freezing. However, they were highly susceptible to inoculative freezing through contact with external ice and (or) ice-nucleating agents, which occur in nesting soil. Therefore, freeze avoidance through supercooling does not appear to be a viable cold-hardiness strategy in these turtles. Hatchlings subjected to experimental freezing survived exposure to temperatures as low as –3.0 °C, suggesting that freeze tolerance may account for the high winter survival observed in natural nests. We conclude that freeze tolerance in hatchling M. terrapin is promoted by high susceptibility to inoculation, which is known to moderate freezing, allowing cells time to adapt to the attendant physical and osmotic stresses.

Cold-hardiness and dehydration resistance of hatchling Blanding's turtles (Emydoidea blandingii): implications for overwintering in a terrestrial habitat

The overwintering habits of hatchling Blanding's turtles, Emydoidea blandingii (Holbrook, 1838), are not well understood. To ascertain whether these turtles are well suited to hibernation on land, we examined susceptibility to dehydration, supercooling capacity, resistance to inoculative freezing, capacity for freeze tolerance, and physiological responses to somatic freezing in laboratory-reared, hatchling E. blandingii. Rates of evaporative water loss (mean ± SE = 4.1 ± 0.2 mg·g–1·d–1) were intermediate to rates previously reported for the hatchlings of species known to hibernate on land and in water. Supercooled hatchlings recovered from a 1-h exposure to –8 °C or a 7-d exposure to –4 °C. Additional turtles supercooled to –14.3 ± 1.2 °C (mean ± SE) before spontaneously freezing. However, when immersed in frozen soil, their capacity to supercool was severely limited by an inability to resist inoculative freezing following contact with external ice and ice nuclei. Therefore, hatchlings likely do not use supercooling as a winter survival strategy. Hatchlings tolerated a 72-h period of somatic freezing to –3.5 °C and responded to somatic freezing by increasing plasma concentrations of the putative cryoprotectants lactate and glucose. Our results suggest that hatchling E. blandingii could overwinter in moist, terrestrial hibernacula where risk of dehydration is reduced and freeze tolerance is promoted.

The response of Anisakis larvae to freezing

Anisakis third stage larvae utilize a variety of fish as intermediate hosts. Uncooked fish are rendered safe for human consumption by freezing. Larvae freeze by inoculative freezing from the surrounding medium but can survive freezing at temperatures down to -10°C. This ability may be aided by the production of trehalose, which can act as a cryoprotectant, but does not involve recrystallization inhibition. Monitoring of fish freezing in commercial blast freezers and under conditions which simulate those of a domestic freezer, indicate that it can take a long time for all parts of the fish to reach a temperature that will kill the larvae. This, and the moderate freezing tolerance of larvae, emphasizes the need for fish to be frozen at a low enough temperature and for a sufficient time to ensure that fish are safe for consumption.

Seasonal changes in physiology and development of cold hardiness in the hatchling painted turtle Chrysemys picta

Hatchling painted turtles (Chrysemys picta) commonly hibernate in shallow, natal nests where winter temperatures may fall below −10 degrees C. Although hatchlings are moderately freeze-tolerant, they apparently rely on supercooling to survive exposure to severe cold. We investigated seasonal changes in physiology and in the development of supercooling capacity and resistance to inoculative freezing in hatchling Chrysemys picta exposed in the laboratory to temperatures that decreased from 22 to 4 degrees C over a 5.5 month period. For comparison, we also studied hatchling snapping turtles (Chelydra serpentina), a less cold-hardy species that usually overwinters under water. Although Chrysemys picta and Chelydra serpentina differed in some physiological responses, both species lost dry mass, catabolized lipid and tended to gain body water during the acclimation regimen. Recently hatched, 22 degrees C-acclimated Chrysemys picta supercooled only modestly (mean temperature of crystallization −6.3+/−0.2 degrees C; N=6) and were susceptible to inoculation by ice nuclei in a frozen substratum (mean temperature of crystallization −1.1+/−0.1 degrees C; N=6) (means +/− s.e.m.). In contrast, cold-acclimated turtles exhibited pronounced capacities for supercooling and resistance to inoculative freezing. The development of cold hardiness reflected the elimination or deactivation of potent endogenous ice nuclei and an elevation of blood osmolality that was due primarily to the retention of urea, but was not associated with accumulation of the polyols, sugars or amino acids commonly found in the cryoprotection systems of other animals. Also, Chrysemys picta (and Chelydra serpentina) lacked both antifreeze proteins and ice-nucleating proteins, which are used by some animals to promote supercooling and to initiate freezing at the high temperatures conducive to freezing survival, respectively.