Metabolism and bound water in overwintering insects

The freezing-tolerant gall fly larva, Eurosta solidaginis, provides an excellent model system for the study of metabolic adaptation and metabolic control for lowtemperature survival during overwintering. Low-temperature acclimation of the larvae results in dramatic alterations in metabolic flux producing a sequential synthesis of two cryoprotectants, glycerol at warmer temperatures followed by sorbitol when larvae are exposed to 5 °C. Regulation of metabolism in the larvae appears to exploit temperature change, temperature effects on enzyme kinetics, and temperature/modulator interactions with enzymes producing the alterations in metabolic flux leading to differential polyol synthesis. For instance, temperature/modulator effects on phosphofructokinase appear to be the major factor halting carbon flow into glycerol synthesis at low temperatures and diverting flux instead into the pathway of sorbitol synthesis. Alterations in the cellular content of bound water and the metabolic pools of free versus bound soluble metabolites may also have important regulatory consequences for low-temperature metabolism. Bound water content of the larvae increases with low-temperature acclimation and is attributable to changes in water binding by both low-molecular-weight (polyols) and highmolecular-weight (proteins, glycogen) subcellular components. A restrictive effect of high bound water content may be one factor causing the strong depression of metabolic activity seen in the larvae as a result of extracellular freezing. In addition, bound water may have a more subtle effect in determining the relative pool sizes of bound versus free metabolites in the cell. ³¹P-NMR studies of whole larvae show that the content of free phosphorylated intermediates in the cell diminishes with decreasing temperatures despite a measured constancy in the total pool size of these intermediates. An increase in the content of bound metabolites with low temperature may restrict metabolism by limiting the availability of substrates and effectors of enzyme reactions.     

Temperature acclimation and seasonal responses by enzymes in cold-hardy gall insects

Changes in the activity of over 20 enzymes of intermediary metabolism in 15°C or ?4°C acclimated goldenrod gall moth (Epiblema scudderiana) and gall fly (Eurosta solidaginis) larvae were measured. Increased activities of glyco-genolytic and hexose monophosphate shunt enzymes in cold-acclimated Epiblema scudderiana suggest a role for coarse control in the conversion of glycogen reserves into glycerol cryoprotectant synthesis. In Eurosta solidaginis, high glycogen phosphorylase activity with decreased activities of glycolytic enzymes may account in part for the temperature-dependent switch from glycerol to sorbitol synthesis in these larvae upon cold acclimation. Isoelectric focusing analyses of five enzymes in overwintering Epiblema scudderiana revealed transient mid-winter changes in the isoelectric points of phosphofructokinase and pyruvate kinase, suggesting seasonal changes in the phosphorylation state of these enzymes. A distinct developmental pattern of aldolase isozymes suggests a role for a new isozyme during overwintering or upon spring emergence. Regulation of metabolism by changes in enzyme activities is indicated for both larvae. © 1995 Wiley-Liss, Inc.     

The fate of [14C]glucose during cold-hardening in Eurosta solidaginis (Fitch)

Glucose catabolism in overwintering larvae Eurosta solidaginis was examined to determine the relative contributions of glycolysis and the pentose phosphate pathway to polyol synthesis at different temperatures. Rates of ¹⁴CO₂ evolution were determined after injection of [¹⁴C]1-glucose, [¹⁴C]6-glucose, and [¹⁴C]3,4-glucose. In addition incorporation of label from each isotope into sorbitol and glycerol was monitored. The respirometric studies showed a relative increase in pentose phosphate activity between 10 and 5°C. Similar results were obtained from the changes of radioactivities incorporated into glycerol, although the activation of the pentose phosphate pathway was low. The conversion of [¹⁴C]glucose to glycerol was highest at 10°C, suggesting that maximum glycerol synthesis may occur at this temperature. Radioactivity appeared in the sorbitol fraction of larvae incubated at temperatures below 5°C. Late autumn larvae converted more [¹⁴C]glucose than did early autumn larvae.     

Cold-hardiness of the arctic beetle, Pytho americanus Kirby Coleoptera,Pythidae (Salpingidae)

The arctic beetle, Pytho americanus Kirby, is frost tolerant in both larval and adult stages. This is the first demonstration that an insect can tolerate freezing in more than one life stage, a situation which would be congruous with its northern distribution and allow it to spread its life cycle over a number of growing seasons. The main biochemical correlates during the cold hardening process of low temperature acclimation are increasing glycerol and decreasing glycogen concentrations. Glycerol is the only polyol to be synthesized during acclimation, and it accumulates to a maximum of 8.2 and 12.2% of the fresh body weight in larvae and adults respectively. This coincides with the peak of frost tolerance. In addition to its normally assumed roles in cryoprotection it is suggested that glycerol may further serve to minimize dehydration in the overwintering insect by increasing the level of ‘bound’ water. Evidence is presented that indicates that glycerol is synthesized mainly from carbohydrate reserves, especially glycogen, but it does not rule out the possibility that a proportion of free glycerol comes from glyceride sources.P. americanus larvae and adults have low supercooling potential and maintain their supercooling points in the region of ?4° to ?8°C. It is hypothesized that these elevated supercooling points are a result of the presence in the haemolymph of nucleating agents which ensure ice formation at high sub-zero temperatures. It is believed that this beetle overwinters in a frozen state within its microhabitat, which is under bark of fallen spruce which is, in turn, covered by an insulating blanket of snow. The advantages of this overwintering strategy are discussed.     

Adaptations to cold in Canadian arctic insects

The survival of insects that inhabit Canadian arctic regions depends on a number of factors which have important ecological, behavioral, physiological, and biochemical components. The ability to withstand low winter temperatures is one of the most conspicuous adaptations of northern insects and the one most closely studied in the laboratory. Most species studied so far conform to one or other of the two major overwintering strategies, namely, frost susceptibility, the ability to avoid freezing by supercooling to a considerable degree, or frost tolerance, the survival of actual ice formation within the body. The Arctic beetle, Pytho americanus Kirby, is frost tolerant in both larval and adult stages, a situation which would be congruous with its northern distribution and allow it to spread its life cycle over a number of growing seasons. The main biochemical correlates during the cold-hardening process in this species are increasing glycerol and decreasing glycogen concentrations. In addition to its normally assumed roles in cryoprotection there is evidence to suggest that glycerol may further serve to minimize dehydration in the overwintering insect by increasing the level of bound water. P. americanus larvae and adults have narrow supercooling ranges and maintain their supercooling points in the region of ?4 to ?8 °C. It is hypothesized that these elevated supercooling points are a result of the presence in the hemolymph of nucleating agents which ensure ice formation at high subzero temperatures.Low temperature tolerance strategies of some other arctic and alpine species have been examined and compared with those of relatives from more southerly latitudes. P. americanus has been collected in the Canadian Rockies at elevations of over 6000′, and its frost-tolerant attributes are identical to those of the population collected in the Arctic. A closely related species, P. deplanatus, from the Rockies, however, although it too exhibits frost tolerance in the larval stage, differs markedly from P. americanus in its ability to depress its supercooling range to ?54 °C. It appears that P. deplanatus does not have the ability to synthesize ice-nucleating agents and, therefore, can overwinter in a supercooled condition. Two congeneric species of willow leaf gall sawflies (Pontania spp.), one from Tuktoyaktuk, N.W.T., and the other from southern Vancouver Island have also been compared and contrasted. Pontania sp. on Salix glauca (Tuk., ca. 70 °N) is frost tolerant in its larval stage, has relatively high supercooling points (ca. ?9.0 °C), but does not accumulate glycerol. Pontania sp. from Salix lasiandra (Victoria, ca. 48 °N) has almost identical overwintering properties, indicating the close phylogenetic affinities of cold tolerance in this genus rather than independent adaptation to widely different climatic conditions. Some of the lowest supercooling points ever recorded are from willow stem gall forming insects. Rhabdophaga sp. (Cecidomyiidae) forms potato galls on the stems of Salix lanata in the Inuvik area, N.W.T. After low temperature acclimation, supercooling points down to ?66 °C have been recorded from individual larvae. This is a record, and it indicates that we may be dealing with a system in which most water is in a metabolically bound state. Glycerol levels reach 20% of the fresh body weight during this period. Diastrophus kincaidii Cynipidae) forms stem galls on Thimble Berry (Rubus parviflorus) on southern Vancouver Island. Both of the forementioned species overwinter as larvae in their galls and are, therefore, exposed to ambient air temperatures. A more benign winter climate on Vancouver Island is reflected in the fact that D. kincaidii has supercooling points only in the ?30 to ?33 °C range at the peak of low temperature acclimation, and glycerol levels just below 4% of fresh body weight. Both species are frost susceptible and depend on their supercooling abilities to survive low winter temperatures.     

Environmental triggers to cryoprotectant modulation in separate populations of the gall fly, Eurost a solidaginis (Fitch)

Northern and southern populations of the gall fly Eurosta solidaginis utilize quantitatively distinct adaptive strategies when exposed to a laboratory simulation of winter temperatures. Both populations are freezing tolerant and rely in part on the temperature-dependent accumulation of glycerol and sorbitol, and static but elevated levels of trehalose for protection. The accumulation triggers are time-temperature dependent. For northern and southern populations, exposure to 5°C for periods exceeding 24 hr, but less than 5 days or 5°C with a gradual reduction (1°C/day) results in the accumulation of sorbitol at 1.5 μg/mg/day. Glycerol levels remain essentially constant between 10° and ?25°C in each population. However, the concentration of glycerol in the Minnesota population is 3–4 times greater that that of the Texas specimens. Haemolymph melting points varied quantitatively with changing cryoprotectant levels. No significant difference was noted between the supercooling points of each population. This suggests that ice-nucleator levels were comparable throughout the exposure period.     

RAPID COLD HARDENING PROVIDING HIGHER COLD TOLERANCE THAN COLD ACCLIMATION IN THE PINE NEEDLE GALL MIDGE THECODIPLOSIS JAPO‐NENSIS LARVAE*

Abstract The responses of overwintering larvae of the pine needle gall midge Thecodiplosis japonensis Uchida et Inouye to rapid cold hardening and cold acclimation were studied. A rapid cold hardening response is found in the 3rd instar larvae of T. japonensis. When overwintering larvae are transferred directly from 27°C to ‐ 15°C for 3 h, there is only 17.9% survival, whereas exposure to 4°C for 2 h prior to transfer to ‐ 15°C increases survival to 40.0%. The acquired cold tolerance is transient and is rapidly lost (after 15 min at 27°C). Rapid cold hardening is more effective in maintaining larval survival than cold acclimation. Different mechanisms are suggested to regulate the insect's cold hardiness under rapid cold hardening and cold acclimation.     

Juvenile hormone modulation of insect cold hardening: Ice-nucleating activity

Data are presented offering the first evidence for probable endocrine involvement in the control of cold hardening in Eurosta solidaginis. Juvenile hormone (JH) deprivation experiments in which the corpora allata were removed by head ligation resulted in a loss of supercooling (SC) capacity in larvae collected over 2 years. This loss of supercooling capacity is indicative of synthesis of organismal pools of ice-nucleating agents (INA). Larval sensitivity to JH removal (ligation) on SC is seasonally dependent. For example, in 1983, larvae were most sensitive in October, secondarily so in September, and relatively insensitive in December regardless of acclimation temperature. While in 1984, larvae acclimated to +5 °C were most sensitive in November, secondarily so in October, and relatively insensitive in December, and larvae acclimated to +15 °C were most sensitive in October and December and least so in November. Supercooling point elevations as great as 7 °C over controls were observed with maximal responses occurring within 1 day following ligation. In 1983, juvenile hormone replacement following ligation generally resulted in an expansion of supercooling capacity when compared to controls. As with ligation, the sensitivity to JH replacement on SC was seasonally dependent: September and October larvae being the most sensitive with December larvae being insensitive. Larvae collected in 1984 were given a greater dose of JH than those in the previous year and showed no significant change in SC over ligated-acetone controls. Hormone analog potentiation experiments in which unligated larvae collected in 1983 and acclimated to +5 °C were given methoprene resulted in depression of supercooling points for September and October larvae. JH titres appear to play an important role in the regulation of SC capacity in E. solidaginis larvae.     

Changes in abundance of aquaporin-like proteins occurs concomitantly with seasonal acquisition of freeze tolerance in the goldenrod gall fly, Eurosta solidaginis

The accumulation of cryoprotectants and the redistribution of water between body compartments play central roles in the capacity of insects to survive freezing. Aquaporins (AQPs) allow for rapid redistribution of water and small solutes (e.g. glycerol) across the cell membrane and were recently implicated in promoting freeze tolerance. Here, we examined whether aquaporin-like protein abundance correlated with the seasonal acquisition of freezing tolerance in the goldenrod gall fly, Eurosta solidaginis (Diptera: Tephritidae). Through the autumn, larvae became tolerant of freezing at progressively lower temperatures and accumulated the cryoprotectant glycerol. Furthermore, larvae significantly increased the abundance of membrane-bound aquaporin and aquaglyceroporin-like proteins from July through January. Acute exposure of larvae to cold and desiccation resulted in upregulation of the AQP3-like proteins in October, suggesting that their abundance is regulated by environmental cues. The seasonal increase in abundance of both putative aquaporins and aquaglyceroporins supports the hypothesis that these proteins are closely tied to the seasonal acquisition of freeze tolerance, functioning to permit cells to quickly lose water and take-up glycerol during extracellular ice formation, as well as reestablish water and glycerol concentrations upon thawing.     

Response diversity of wild bees to overwintering temperatures

Biodiversity can provide insurance against environmental change, but only if species differ in their response to environmental conditions (response diversity). Wild bees provide pollination services to wild and crop plants, and response diversity might insure this function against changing climate. To experimentally test the hypothesis that bee species differ in their response to increasing winter temperature, we stored cocoons of nine bee species at different temperatures during the winter (1.5–9.5 °C). Bee species differed significantly in their responses (weight loss, weight at emergence and emergence date). The developmental stage during the winter explained some of these differences. Bee species overwintering as adults generally showed decreased weight and earlier emergence with increasing temperature, whereas bee species overwintering in pre-imaginal stages showed weaker or even opposite responses. This means that winter warming will likely affect some bee species negatively by increasing energy expenditure, while others are less sensitive presumably due to different physiology. Likewise, species phenologies will respond differently to winter warming, potentially affecting plant–pollinator interactions. Responses are not independent of current flight periods: bees active in spring will likely show the strongest phenological advances. Taken together, wild bee diversity provides response diversity to climate change, which may be the basis for an insurance effect.     

Subtropical freshwater phytoplankton show a greater response to increased temperature than to increased pCO2

Global increases in atmospheric CO₂ and temperatures will impact aquatic systems, with freshwater habitats being affected. Some studies suggest that these conditions will promote cyanobacterial dominance. There is a need for a clearer picture of how algal species and strains within species will respond to higher temperatures and CO₂, especially in combination. This study examined two chlorophytes (Monoraphidium and Staurastrum), and two strains of the cyanobacterium Raphidiopsis raciborskii (straight S07 and coiled C03), to determine how the combination of higher temperature and CO₂ levels will affect their growth and maximum cell concentrations. Continuous cultures were used to compare the steady state cell concentrations at 28 °C and 30 °C, and CO₂ partial pressures (pCO₂), 400 and 750 ppm for all cultures, and in addition 1000 ppm at 28 °C for R. raciborskii strains. This study showed that, for all species, water temperature had a greater effect than higher pCO₂ on cell concentrations. There were clear differences in response between the chlorophyte species, with Monoraphidium preferring 28 °C and Staurastrum preferring 30 °C. There were also differences in response of the R. raciborskii strains to increasing temperature and pCO₂, with S07 having a greater increase in cell concentration. Genome analysis of R. raciborskii showed that the straight strain has five additional carbon acquisition genes (β-CA, chpY, cmpB, cmpD and NdhD4), indicative of increased carbon metabolism. These differences in the strains’ response to elevated pCO₂ will lead to changes in the species population structure and distribution in the water column. This study shows that it is important to examine the effects of both pCO₂ and temperature, and to consider strain variation, to understand how species composition of natural systems may change under future climate conditions.     

水稻二化螟越冬幼虫耐寒性物质的动态变化

为从生理生化水平上探讨二化螟越冬幼虫耐寒机制,分别对不同时期采集的二化螟越冬幼虫过冷却点及水、灰分、元素、脂肪、脂肪酸、甘油、总糖和蛋白质含量进行了测定.结果表明：越冬期幼虫过冷却点及游离水、游离脂肪、总糖和蛋白质含量变化均呈先减后增,而结合水、灰分、高含量元素（K、Na、Mg、Fe、Ni和Cr）、结合脂肪和甘油含量则呈相反变化;越冬期幼虫脂肪酸组分出现变化,但主要成分均为9-十六碳烯酸、9-十六烷酸和9-十八碳烯酸,其中前二者含量呈先减后增变化而后者则相反.越冬幼虫体内理化成分含量的动态变化可以反映其耐寒性强弱.     

The tissue water relationships of Callitris columellaris,Eucalyptus melliodora and Eucalyptus microcarpa investigated using the pressure-volume technique

Using the pressure-bomb to construct pressure-volume curves, a cellular basis of differential drought resistance was found between Callitris columellaris (F. Muell), Eucalyptus melliodora A. Cunn. ex Schauer, and Eucalyptus microcarpa Maiden. Between these three species differences were found in bound water, relative water content and water potential at zero turgor, osmotic potential at full turgor and bulk modulus of elasticity. It is suggested that these parameters showed C. columellaris to be the most, and E. melliodora the least drought resistant of the three species. Preliminary studies also showed that drought hardening may involve an increase in bound water content, dry weight: turgid weight ratio and a decrease in osmotic potential at full turgor and water potential at zero turgor.     

Thermal factors affecting egg development in the wandering glider dragonfly, <Emphasis Type="Italic">Pantala flavescens</Emphasis> (Odonata: Libellulidae)

The wandering glider dragonfly, Pantala flavescens (Fabricius), arrives in Japan from tropical regions every spring. The offspring colonize areas throughout Japan, with rapid increases in populations in the autumn, but all individuals die in the winter, suggesting low tolerance to low temperatures. However, few quantitative data on egg development and water temperature have been reported for this species. Females at the reproductive stage were collected from fields throughout the flying season and their eggs released using an artificial oviposition technique. Almost all of the eggs were fertilized. Egg size was stable throughout the seasons. Most eggs hatched within a period of 5 days at high water temperatures (35 and 30 °C), which were recorded in the shallow ponds and rice paddy fields from summer to early autumn. However, the egg-stage duration increased with declining water temperature. All eggs in water at 15 °C had failed to hatch by 90 days. The calculated critical temperature of water was determined to be approximately 14.3 °C; the total effective temperature for the egg stage was about 80 degree-days. Thus, low water temperatures in winter may prevent P. flavescens overwintering in Japan.     

Kinetic properties and regulation of glycerol-3-phosphate dehydrogenase from the overwintering, freezing-tolerant gall fly larva, Eurosta solidagenis

The kinetic properties of cytoplasmic glycerol-3-P dehydrogenase from the third instar larva of the gall fly, Eurosta solidaginis, were studied with emphasis on temperature effects on the enzyme and the regulation of enzyme activity during the synthesis of the cryoprotectant, glycerol. Isoelectrofocusing revealed one major and two minor forms of the enzyme with no alteration in the pI's or relative activities of the forms in larvae acclimated to 24 versus ?30 °C. Kinetic properties of the enzyme were also the same in larvae acclimated to high and low temperatures. Arrhenius plots were linear over a 30 to 0 °C range with an activation energy of 12,630 ± 185 cal/mol and a Q₁₀ of 2.16. The K_m for dihydroxyacetone-P was constant, at 50 μM, between 30 and 10 °C but increased by 75% at 0 °C; this increase may be a factor in the cessation of glycerol synthesis which occurs below 5 °C in this species. The K_m(NADH), by contrast, was higher (5–6 μM) at 30 °C but decreased (3 μM) at lower temperatures. In the reverse direction, K_m's were 340 μM for glycerol-3-P and 12 μM for NAD⁺. Effects of most inhibitors (of the forward reaction), glycerol-3-P (K_i = 2.4 mM), NAD⁺ (K_i = 0.2 mM), ATP, Mg·ATP, and P_i, were unaltered by assay temperature but ADP effects were potentiated by low temperature while citrate inhibition was greatest at high temperatures. Glycerol and sorbitol, which accumulate as cryoprotectants in E. solidaginis, had no significant effects on kinetic constants at any temperature but decreased the V_max activity of the enzyme. Thermal inactivation studies showed an increased thermal stability of the larval enzyme compared to the homologous enzyme from rabbit muscle while added polyols stabilized enzyme activity, decreasing the rate of enzyme inactivation at 50 °C.     

The effect of overwintering temperature on the body energy reserves and phenoloxidase activity of bumblebee Bombus lucorum queens

Warming winters and changes in species composition related to the estimated global warming may cause a threat to bumblebees adapted to cold winters. During the overwintering period, their intermediary and respiratory metabolism decreases but metabolism remains responsive to temperature. The effect of temperature on diapause survival, phenoloxidase (PO) activity, and energy expenditure of the white-tailed bumblebee (Bombus lucorum) after a 4-month diapause were studied by manipulating the diapause temperature. Two overwintering temperatures were used, cold (1.8 °C) and warm (9 °C). Body fat content was used as an estimate of the remaining energy resources and PO activity as an immune function parameter of overwintering bumblebee queens. The baseline levels of PO activity were used to measure the differences in B. lucorum queen responses after overwintering in either temperature. We found a 0.4 g pre-diapause threshold weight of survival in B. lucorum. Large queens had more fat left and a higher PO activity compared to small ones after overwintering in warm conditions, but in the cold there was no effect of size on the remaining fat in the fat body of queens or their PO activity. The observed difference in energy usage appears to relate to normal size-dependent metabolism and variation in energy allocation between basic metabolism and immune functions.     

Indications that mutagenesis in Salmonella may be subject to catabolite repression

Spontaneous reversion of the base-pair substitution trpE8 marker in the LT2 sub-line of Salmonella typhimurium is significantly increased in the presence of the ultraviolet light-protecting and mutation-enhancing plasmid pKM101. The numbers of Trp⁺ revertants arising on plates of defined medium supplemented with trace amounts of nutrient broth have been found to depend upon the nature of the carbon source provided to support growth of both the background lawn and any revertants which may arise. For example, the yield of Trp⁺ revertants can be some 5–8 times greater when glycerol is the carbon source as compared to when glucose is the carbon source. S. typhimurium strain TA100, which carries the base-pair substitution hisG46 marker and pKM101, shows a similar response, although the difference is much smaller. Time-course experiments using both carbon sources indicate that the final trpE8 → Trp⁺ mutation yield is depressed by glucose rather than enhanced by a ‘mutagenic’ effect of glycerol. These results are consistent with the idea that a glucose-repressible function responsible for generating mutations can be switched on by growth on glycerol as sole carbon source. Evidence is also presented that many more mutational events occur in response to a mild temperature stress (42°) in populations growing on glycerol as carbon source than occur in populations growing on glucose.     

Effect of mating on survival at low temperature in females of the Japanese common grass yellow,Eurema mandarina

1. In some insects that overwinter as adults, mating occurs both before and after overwintering. Two hypotheses have been proposed to explain the adaptive significance of pre‐overwintering copulation of females. One is the bet‐hedging hypothesis, which explains pre‐overwintering copulation as a preparation for less chance of mating in the following spring. The other is the nuptial gift hypothesis, which states that secretions derived from males increase overwintering success of females. 2. In Eurema mandarina, both diapause autumn‐ and non‐diapause summer‐form male adults emerge with autumn‐form female adults in the last generation in a year. Most autumn‐form females mate with summer‐form males before winter, and re‐mate with autumn‐form males in the following spring. Because autumn‐form females have sufficient chances for mating after overwintering, the nuptial gift hypothesis has been regarded as the more probable hypothesis. 3. To test the nuptial gift hypothesis, the survival period was compared under short‐day conditions at 10 °C between mated and unmated females that had been reared on sucrose solution at 25 °C for 15–21 days. The mated females had significantly greater longevity than the unmated females, supporting the nuptial gift hypothesis. Body size also affected the survival period. 4. The results suggest that the nuptial gift is an important factor for the evolution of pre‐overwintering copulation in species in which females mate both before and after overwintering.     

Evidence for lower plasticity in CTMAX at warmer developmental temperatures

Understanding the capacity for different species to reduce their susceptibility to climate change via phenotypic plasticity is essential for accurately predicting species extinction risk. The climatic variability hypothesis suggests that spatial and temporal variation in climatic variables should select for more plastic phenotypes. However, empirical support for this hypothesis is limited. Here, we examine the capacity for ten Drosophila species to increase their critical thermal maxima (CT_MAX) through developmental acclimation and/or adult heat hardening. Using four fluctuating developmental temperature regimes, ranging from 13 to 33 °C, we find that most species can increase their CT_MAX via developmental acclimation and adult hardening, but found no relationship between climatic variables and absolute measures of plasticity. However, when plasticity was dissected across developmental temperatures, a positive association between plasticity and one measure of climatic variability (temperature seasonality) was found when development took place between 26 and 28 °C, whereas a negative relationship was found when development took place between 20 and 23 °C. In addition, a decline in CT_MAX and egg‐to‐adult viability, a proxy for fitness, was observed in tropical species at the warmer developmental temperatures (26–28 °C); this suggests that tropical species may be at even greater risk from climate change than currently predicted. The combined effects of developmental acclimation and adult hardening on CT_MAX were small, contributing to a <0.60 °C shift in CT_MAX. Although small shifts in CT_MAX may increase population persistence in the shorter term, the degree to which they can contribute to meaningful responses in the long term is unclear.