Pre-freeze mortality in three species of aphids from sub-Antarctic Marion Island

Understanding the mechanisms by which aphids survive low temperature is fundamental in forecasting the risk of pest outbreaks. Aphids are chill susceptible and die at a temperature close to that at which a small exothermal event is produced. This event, which can be identified using differential scanning calorimetry (DSC), normally occurs at a higher temperature than the supercooling point (SCP) and has been termed a pre-freeze event (PFE). However, it is not known what causes the PFE or whether it signifies the death of the aphid. These questions are addressed here by using a sensitive DSC to quantify the PFE and SCP and to relate these thermal events to the lower lethal temperature (LT₅₀) of sub-Antarctic aphids acclimated to low temperatures. PFEs were observed in each of the 3 species of aphids examined. They occurred over a narrower temperature range and at a higher temperature range than the SCP (−8.2 to −13.8 and −5.6 to −29.8 °C, respectively). Increased acclimation temperature resulted in increased SCPs in Myzus ascalonicus but not in Rhopalosiphum padi. The LT₅₀ reduced by approximately 1 °C from −9.3 to −10.5 °C with reduced acclimation temperature (10–0 °C). The LT₅₀ was close to the temperature at which the PFE occurred but statistically significantly higher than either the PFE or the SCP. In the majority of cases the PFE exotherm occurred well before the main exotherm produced by the bulk of the insect’s body water freezing (SCP). However, in a few cases it occurred at the same temperature or before the super-cooling point making the term, pre-freeze event (PFE), rather misleading. The possible origins of the PFE are discussed.     

Effects of acclimation and diapause on the thermal tolerance of the two-spotted spider mite Tetranychus urticae

The two-spotted spider mite, Tetranychus urticae, is a worldwide pest species that overwinters as diapausing females. Cold hardening is presumed to start during diapause development to ensure the successful overwintering of this species. To address this hypothesis, we compared cold tolerance between non-diapausing and diapausing females. We measured supercooling point (SCP) and survival to acute cold stress by exposing the mites at a range of sub-zero temperatures (from −4 to −28 °C for 2 h). The mean SCPs of non-diapausing and diapausing females were −19.6±0.5 and −24.7±0.3 °C respectively, and freezing killed the mites. Diapausing females were significantly more cold tolerant than non-diapausing ones, with LT₅₀ of −19.7 and −13.3 °C, respectively. Further, we also examined the effects of cold acclimation (10 d at 0 or 5 °C) in non-diapausing and diapausing females. Our findings indicated that diapause decreased SCP significantly, while cold acclimation had no effect on the SCP except for non-diapausing females that were acclimated at 5 °C. Acclimation at 5 °C enhanced survival to acute cold stress in diapausing and non-diapausing females, with LT₅₀ of −22.0 and −17.1 °C, respectively. Altogether, our results indicate that T. urticae is a chill tolerant species, and that diapause and cold acclimation elevate cold hardiness in this species.     

The maximum nitrate reductase activity of the seagrass Zostera noltii (Hornem.) varies along its vertical distribution

Maximum nitrate reductase (NR) activity was measured in two intertidal morphotypes of Zostera noltii (Hornem.) in Ria Formosa tidal lagoon, southern Portugal. The two morphotypes develop in the upper and lower limits of the intertidal meadows. The NR activity was measured using an in vivo method, without cell disruption. NR activity was 30-40 fold higher in leaves than in roots, which indicates that nitrate reduction is essentially made through the aerial part of the plant. The effects of assay temperature (5 °C steps, from 5 to 45 °C), pH (7, 8 and 9) and elevation (upper and lower intertidal) on leaf NR activity were tested in a factorial design (n=5). Both elevation and assay temperature had a significant effect on NR activity, but not pH. NR activity was always higher in the upper intertidal plants, at all temperatures. Activity peaks for upper and lower plants were, respectively, 6.12 μmol NO₂⁻ g⁻¹ DW 0.5 h⁻¹ at 25 °C, and 3.30 μmol NO₂⁻ g⁻¹ DW 0.5h⁻¹ at 35 °C. Further investigation on environmental factors concerning the intertidal environment must be developed, as they are probably responsible for the significant differences found between the values of NR activity in the upper and lower morphotype.     

Enhancement of frost tolerance in olive shoots in vitro by cold acclimation and sucrose increase in the culture medium

The evaluation of frost tolerance in olive shoots in vitro has been successfully accomplished. The behavior of in vitro shoots at freezing temperatures was comparable to that of intact plants. Cold acclimation was found to increase frost tolerance in cv. Moraiolo and the LT₅₀ was about 4 °C lower compared to nonacclimated shoots. Damage in acclimated shoots occurred at –15 °C, whereas control shoots were damaged at –10 °C. Olive shoots were unable to withstand freezing temperatures of –20 °C, even when acclimated. The effects of sucrose were also determined. 6% (w/v) sucrose in the medium conferred the highest frost tolerance in both acclimated and nonacclimated plants.     

The effects of long-term elevation of air temperature and CO on the frost hardiness of Scots pine

The frost hardiness of 20 to 25-year-old Scots pine (Pinus sylvestris L.) saplings was followed for 2 years in an experiment that attempted to simulate the predicted climatic conditions of the future, i.e. increased atmospheric CO₂ concentration and/or elevated air temperature. Frost hardiness was determined by an electrolyte leakage method and visual damage scoring on needles. Elevated temperatures caused needles to harden later and deharden earlier than the controls. In the first year, elevated CO₂ enhanced hardening at elevated temperatures, but this effect disappeared the next year. Dehardening was hastened by elevating CO₂ in both springs. The frost hardiness was high (相似文献   

Comparing image (fractal analysis) and electrochemical (impedance spectroscopy and electrolyte leakage) techniques for the assessment of the freezing tolerance in olive

Olive growth and productivity are limited by low temperatures mainly during winter, but sometimes also in spring and fall. The most effective way to avoid these damages in areas subjected to these climatic conditions is to select least susceptible varieties, but the choice of the right method to determine cold hardiness is extremely difficult. The aims of the work were (1) to assess LT₅₀ (lethal temperature at which 50% of damage in plants subjected to low temperatures occurs) of some olive varieties in two seasons (summer and winter) and (2) to assess the reliability of different methods to evaluate cold hardiness. LT₅₀ was determined on 21 different olive (Olea europaea L.) Italian varieties by leaf and shoot electrolyte leakage, shoot impedance spectroscopy and leaf color determination of fractal spectrum. All the experiments were conducted on non-acclimated and cold-acclimated plants. Our results showed that all the three methods were able to detect damages on olive plants after exposure to low temperatures, with leaves appearing more sensitive to cold stress than shoots. Among these methods, fractal analysis could be very useful in assessing cold hardiness of plants on the basis of visible injury, without sophisticated or expensive instruments and in a reliable and cost-effective way, using only a scanning device, a personal computer and dedicated freeware software.     

The timing of leaf fall affects cold acclimation by interactions with air temperature through water and carbohydrate contents

Three parameters (i.e. the water content, soluble sugar content and minimal air temperature) can be used to predict the cold acclimation process of walnut trees. In order to test this assumption, two-year-old walnuts were defoliated at two different dates, i.e. mechanical defoliation in early October (early leaf fall, EF) or natural defoliation in early November (natural leaf fall, NF) and conditioned in either outdoor freeze-deprived or cold-deprived (T_min > 13 °C) greenhouses over winter. Even if early defoliation date could have affected short day signal perception (SDSP), water balance and carbohydrate metabolism were more altered. EF treatment, by stopping transpiration, significantly increased tree's water content and at warm temperature high root activity stopped normal winter dehydration. Starch content decreased in all treatments, but there was only a significant increase in soluble sugar content when water content had sufficiently decreased. Thus, depending on date of defoliation, cold-deprived trees were or were not able to acclimate to frost (minimal frost hardiness = −21.8 °C vs. −22.1 °C in controls (freeze-deprived) for NF and −13.7 °C vs. −25.3 °C in controls for EF). Different treatments showed the relationship between minimal water content observed during winter and maximal soluble sugars synthesized. Thus, the cold acclimation process appeared dependent on these physiological parameters (water and soluble sugar contents) through the interaction between air temperature and timing of leaf fall.     

Cold tolerance of the overwintering larval instars of light brown apple moth Epiphyas postvittana

The light brown apple moth, Epiphyas postvittana, a leafroller native to southeastern Australia was discovered in California in 2006. The highly polyphagous nature of this pest adds to the importance of being able to predict the potential distribution of this invader across the North American continent. The spread of ectothermic species that lack winter diapause, such as E. postvittana, can be limited by their ability to tolerate cold temperature extremes. In this study we examined the cold hardiness of 4th to 6th instar E. postvittana, the only life stages known to overwinter in California, through a combination of supercooling point (SCP) and mortality at low temperatures. Our results showed that the mean SCP for E. postvittana ranged from −14.1 °C for 6th instars to −16.0 °C for 4th instars. Lethal time leading to 50% mortality (LT₅₀) for the three instars combined were 2.5 h at −10.5 °C, 41 h at −6.5 °C and 198 h at −0.9 °C. At 3 °C, the LT₅₀ of 4th instars was significantly lower at 775 h than that for 5th and 6th instars combined at 1029 h. The cold hardiness characteristics of later-instar E. postvittana larvae were comparable to those of pink bollworm, Pectinophora gossypiella, a diapausing invasive with a geographic distribution restricted to southern California. Slightly greater cold hardiness is shown by the indigenous non-diapausing leafroller Argyrotaenia franciscana, which is restricted to the Pacific Coast of North America. We therefore conclude that the moderate cold hardiness of E. postvittana will substantially limit its spread into northern temperate regions of North America.     

Ultrastructural and protein changes in cell suspension cultures of peach associated with low temperature-induced cold acclimation and abscisic acid treatment

Cell suspension cultures were initiated from callus derived from xylem tissues of peach [Prunus persica (L.) Batsch]. Cold acclimation was induced (LT₅₀ of-13°C) in cell suspensions at 3°C in the dark for 10 days. Freezing tolerance returned to the level of nonacclimated cells (LT₅₀ of –4.5°C) when cold-acclimated cells were transferred to 24°C (in dark) for 3 days. Addition of 75 M abscisic acid (ABA) to the growth medium failed to induce cold acclimation after cells were cultured for 5 days at 24°C. Microvacuolation, cytoplasmic augmentation and disappearance of starch grains were observed in cells that were cold-acclimated by exposure to low temperature. Similar ultrastructural alterations were not observed in ABA-treated cells. Several qualitative and quantitative changes in proteins were noted during both cold acclimation and ABA treatment. Both the ultrastructural and protein changes observed during cold acclimation were reversed during deacclimation. The relationship of these changes to cold acclimation in peach cell-cultures is discussed.Abbreviations ABA abscisic acid - 2,4-d 2,4-dichlorophenoxyacetic acid - IBA indole-3-butyric acid - Ms Murashige & Skoog - PMSF phenylmethylsulfonyl fluoride - LT₅₀ or Freezing Tolerance temperature that resulted in 50% decrease in TTC reduction - TTC 2,3,5-triphenyltetrazolium chloride     

The use of small subcutaneous transponders for quantifying thermal biology and torpor in small mammals

Remote measurements of body temperature (T_b) in animals require implantation of relatively large temperature-sensitive radio-transmitters or data loggers, whereas rectal temperature (T_rec) measurements require handling and therefore may bias the results. We investigated whether ∼0.1 g temperature-sensitive subcutaneously implanted transponders can be reliably used to quantify thermal biology and torpor use in small mammals. We examined (i) the precision of transponder readings as a function of temperature and (ii) whether subcutaneous transponders can be used to remotely record subcutaneous temperature (T_sub). Five adult male dunnarts (Sminthopsis macroura, body mass 24 g) were implanted with subcutaneous transponders to determine T_sub as a function of time and ambient temperature (T_a), and in comparison to thermocouple readings of T_rec. Transponder temperature was highly correlated with water bath temperature (r²=0.96–0.99) over a range of approximately 10.0–40.0 °C. Transponders provided reliable data (±0.6 °C) over the T_sub of 21.4–36.9 °C and could be read from a distance of up to 5 cm. Below 21.4 °C, accuracy was reduced to ±2.8 °C, but individual transponder accuracy varied. Consequently, small subcutaneous transponders are useful to remotely quantify thermal physiology and torpor patterns without having to disturb the animal and disrupt torpor. Even at T_sub<21.4 °C where the accuracy of the temperature readings was reduced, transponders do provide reliable data on whether and when torpor is used.     

Cold tolerance of the harlequin ladybird Harmonia axyridis in Europe

As an essential aspect of its invasive character in Europe, this study examined the cold hardiness of the harlequin ladybird Harmonia axyridis. This was done for field-collected populations in Belgium overwintering either in an unheated indoor or an outdoor hibernaculum. The supercooling point, lower lethal temperature and lower lethal time at 0 and −5 °C were determined. Possible seasonal changes were taken into account by monitoring the populations during each winter month. The supercooling point and lower lethal temperature remained relatively constant for the overwintering populations in the outdoor hibernaculum, ranging from −17.5 to −16.5 °C and −17.1 to −16.3 °C, respectively. In contrast, the supercooling point and lower lethal temperature of the population overwintering indoors clearly increased as the winter progressed, from −18.5 to −13.2 °C and −16.7 to −14.1 °C, respectively. A proportion of the individuals overwintering indoors could thus encounter problems surviving the winter due to premature activation at times when food is not available. The lower lethal time of field populations at 0 and −5 °C varied from 18 to 24 weeks and from 12 to 22 weeks, respectively. Morph type and sex had no influence on the cold hardiness of the overwintering adults. In addition, all cold tolerance parameters differed greatly between the laboratory population and field populations, implying that cold tolerance research based solely on laboratory populations may not be representative of field situations. We conclude from this study that the strong cold hardiness of H. axyridis in Europe may enable the species to establish in large parts of the continent.     

A comparison of low temperature tolerance traits between closely related aphids from the tropics, temperate zone, and Arctic

The survival of aphids exposed to low temperatures is strongly influenced by their ability to move within and between plants and to survive exposure to potentially lethal low temperatures. Little is known about the physiological and behavioural limitations on aphid movement at low temperatures or how they may relate to lethal temperature thresholds. These questions are addressed here through an analysis of the thermal ecology of three closely related aphid species: Myzus persicae, a ubiquitous temperate zone pest, Myzus polaris, an arctic species, and Myzus ornatus, a sub-tropical species. Lower lethal temperatures (LLT₅₀) of aphids reared at 15 °C were similar for M. persicae and M. polaris (range: −12.7 to −13.9 °C), but significantly higher for M. ornatus (−6.6 °C). The temperature thresholds for activity and chill coma increased with rearing temperature (10, 15, 20, and 25 °C) for all clones. For M. polaris and M. ornatus the slopes of these relationships were approximately parallel; by contrast, for M. persicae the difference in slopes meant that the difference between the temperatures at which aphids cease walking and enter coma increased by approximately 0.5 °C per 1 °C increase in rearing temperature. The data suggest that all three species have the potential to increase population sizes and expand their ranges if low temperature limitation is relaxed.     

Use of light and inorganic carbon acquisition by two morphotypes of Zostera noltii Hornem

The affinity for dissolved inorganic carbon (DIC) and the mechanisms to use HCO₃⁻ as a source of DIC for photosynthesis were investigated in two morphotypes of Zostera noltii Hornem. Both morphotypes were collected at Ria Formosa lagoon (Southern Portugal) at two different levels in the intertidal. Affinity for DIC at saturating photon fluence rate (PFR), estimated as photosynthetic conductance for CO₂ (g_p(CO2)), was reduced by 75% in the Z. noltii plants adapted to shade conditions (lower intertidal) in comparison to the sun morphotype (45×10⁻⁶ and 182×10⁻⁶ m s⁻¹, respectively), indicating that the plants acclimated to sun conditions (higher intertidal) had a higher capacity to use HCO₃⁻ as DIC source for photosynthesis. Since external carbonic anhydrase activity was negligible and a large inhibitory effect was produced by Tris buffer addition, this HCO₃⁻ use was attributed to the operation of H⁺ ATPases creating low pH zones in periplasmic space. The photosynthetic CO₂-flux supported for this mechanism was calculated to be 53 μmol O₂ m⁻² s⁻¹ in sun morphotype, about 80% out of maximum photosynthesis rate. In order to determine the possible photosynthetic energy cost of the HCO₃⁻ use, the effect of decreasing light on photosynthetic rates and g_p(CO2) was estimated. Photosynthetic conductance decreased in both morphotypes at non-saturating PFR. This dependence of g_p(CO2) on PFR indicated the existence of a positive interactive effect between DIC and PFR which was more pronounced in the shade morphotype since the ascending slope of O₂ evolution vs. PFR curves at limiting PFRs was reduced from 7.2 to 2.3 mmol O₂ mol photon⁻¹ at 4 and 0.5 mol m⁻³ of DIC, respectively.     

Thermal tolerance,net CO2 exchange and growth of a tropical tree species, Ficus insipida, cultivated at elevated daytime and nighttime temperatures

Global warming and associated increases in the frequency and amplitude of extreme weather events, such as heat waves, may adversely affect tropical rainforest plants via significantly increased tissue temperatures. In this study, the response to two temperature regimes was assessed in seedlings of the neotropical pioneer tree species, Ficus insipida. Plants were cultivated in growth chambers at strongly elevated daytime temperature (39 °C), combined with either close to natural (22 °C) or elevated (32 °C) nighttime temperatures. Under both growth regimes, the critical temperature for irreversible leaf damage, determined by changes in chlorophyll a fluorescence, was approximately 51 °C. This is comparable to values found in F. insipida growing under natural ambient conditions and indicates a limited potential for heat tolerance acclimation of this tropical forest tree species. Yet, under high nighttime temperature, growth was strongly enhanced, accompanied by increased rates of net photosynthetic CO₂ uptake and diminished temperature dependence of leaf-level dark respiration, consistent with thermal acclimation of these key physiological parameters.     

Effects of salinity and temperature on the survival and byssal attachment of the lion's paw scallop Nodipecten nodosus at its southern distribution limit

The lion's paw scallop, Nodipecten nodosus, is subject to wide temperature variations on seasonal and short-term scales, and may be exposed to low-salinity events, caused by oceanographic and meteorological processes at its southern distribution limit (Santa Catarina State, Brazil). Such variations may have important implications on the distribution and on aquaculture site selection. The upper and lower temperature tolerances and the percentage of byssal attachment at different temperatures (11 to 35 °C) were studied for spat, juvenile and adult scallops. The lethal and sublethal effects of reduced salinity (13‰ to 33‰) on spat, juvenile and adult scallops were studied at ambient temperature (23.5 °C) and on spat also at low (16 °C) and high (28 °C) temperatures during 96-h bioassays. In addition, the influences of short exposure (1 h) to low salinity (13‰ and 17‰) at different temperatures (16 and 28 °C), and the effects of exposure (2 and 4 h) to high temperature (33 °C) at ambient salinity (33‰) were studied. N. nodosus is a moderately eurythermal but stenohaline tropical species, adults having lower tolerance to high temperature and low salinity than spat. Lethal temperatures for a 48-h exposure (LT₅₀) were 29.8 °C for adult and juveniles, and 31.8 °C for spat. Maximum rate of byssal attachment occurred in a narrower temperature range for juveniles and adults (23 to 27 °C) than for spat (19 to 27 °C), which are suggested as the optimum ranges of temperatures for growth. Lethal salinities (LC₅₀) for a 48-h exposure at ambient temperature were 23.2‰, 23.6‰ and 20.1‰ for adults, juveniles and spat, respectively, but the percent byssal attachment was significantly reduced below salinities of 29‰ indicating that scallops were physiologically stressed. A 1-h exposure to 17‰ was lethal to spat at 28 °C, but at 16 °C there was a 28.5% survival, 96 h after the exposure. Temperatures and salinity in coastal areas of southern Brazil can reach levels leading to sublethal effects, and in some sites, it may surpass the limits of tolerance for the survival of the species.     

Genetic analysis of frost hardiness traits in tuber-bearingSolanum species

The inheritance of frost hardiness and cold acclimation potential traits was studied in three segregating populations derived from a cross betweenSolanum commersonii Dun. PI 243503 (cmm) andSolanum cardiophyllum Lindl., PI 184762 (cph), two parental genotypes with contrasting frost hardiness and cold acclimation potential. The levels of frost hardiness and cold acclimation potential were expressed as the LT₅₀, the temperature at which 50% of the cells in leaf discs were killed, as measured by the ion leakage method, following a controlled freeze test There was considerable variation in both frost hardiness and cold acclimation potential in all three segregating populations (F₁ F₁ xcmm, and F₁ xcph). Frost hardiness and cold acclimation potential were not correlated, suggesting that these two traits are under independent genetic control. The analysis of generation means indicated that the variation for both traits could be best explained by an additive-dominance model, with additive gene effects the most important Broad-sense heritability was 0.73 and 0.74 in the F₁ population, for frost hardiness and cold acclimation potential, respectively, and was 0.85 for either trait in the F₁ xcmm population, indicating that these two traits are highly inheritable. Our results suggest that it should be possible to incorporate the frost hardiness and cold acclimation traits from S.commersonii into cultivated potato species.     

Ambient temperature-dependent thermoregulatory role of REM sleep

Changes in ambient temperature produce complex effects on sleep–wakefulness. In order to find out the mechanisms involved in temperature-sensitive changes in sleep in rats, their thermal preference, body temperature and sleep were studied before and after the destruction of both peripheral and central warm receptors, by systemic administration of 375 mg/kg capsaicin. Though the pre-treated rats preferred to stay mostly at the ambient temperature of 27 °C, post-treated rats strayed freely into chambers having ambient temperature of 30 °C and 33 °C. Sleep and body temperature of these rats were studied for six hours each, when they were kept at an ambient temperature of 18–36 °C. Total sleep time, especially REM sleep, was maximum at 30 °C in pre-treated rats, but this REM sleep peak at 30 °C disappeared after capsaicin administration. Body temperature increased sharply in post-treated rats, at ambient temperatures above 30 °C. Apart from the ability to defend body temperature at high ambient temperature, avoidance of warm ambient temperature and increase in REM sleep are the behavioral measures which are lost in post-treated rats. Results of this study suggest that the ambient temperature-related increase in REM sleep at 30 °C could be part of the thermoregulatory measures.     

Induction of freezing tolerance in alfalfa cell suspension cultures

The effects of ABA, 2,4-D, kinetin and cold exposure on the cold hardiness of Medicago sativa L. cell suspensions were investigated. Cultures treated with 5×10^–5 M ABA at 2°C for 4 weeks in the absence of kinetin showed a 50% survival after freezing to –12.5°C, whereas cultures grown at 25°C under normal conditions tolerated freezing to only –3°C. The optimum ABA treatment of 5×10^–5 M for 4 weeks was effective only in combination with cold exposure. Of six cell lines tested, all showed different degrees of induced cold hardiness. The results suggest that ABA alone cannot induce freezing tolerance on alfalfa cell suspension cultures and that the deletion of kinetin and combination of low temperature and ABA is critical for the induction of cold hardiness in alfalfa cell suspension cultures.Abbreviations ABA abscisic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - LT₅₀ 50% killing temperature     

Inheritance of autumn frost hardiness in Pinus sylvestris L. seedlings

Summary Inheritance of frost hardiness was analysed making use of a 12×12 incomplete factorial mating design. Owing to space limitations only 59 families could be tested in four experiments. To link the four experiments, some families were common to two or more experiments. The seedlings were grown in climate chambers under conditions inducing autumn hardening. The plants were exposed to a freezing temperature of –10 °C for three hours at night lengths of 11–13 h. A statistical model was developed for analyses of variance of our data. The genetic variation and the variation due to the cultivation regimes during autumn hardening were of the same magnitude. The additive effects were the most important ones for induction of frost damage. No interaction following long-distance crossing was noted. Mixed model equations were used for ranking of the parents. The results obtained support a polygenic inheritance of frost hardiness. The large within-population variation offers good opportunities for hardiness breeding.     

Multigenerational analysis of temperature and salinity variability affects on metabolic rate,generation time,and acute thermal and salinity tolerance in Daphnia pulex

Climate change, sea level rise, and human freshwater demands are predicted to result in elevated temperature and salinity variability in upper estuarine ecosystems. Increasing levels of environmental stresses are known to induce the cellular stress response (CSR). Energy for the CSR may be provided by an elevated overall metabolic rate. However, if metabolic rate is constant or lower under elevated stress, energy for the CSR is taken from other physiological processes, such as growth or reproduction. This study investigated the examined energetic responses to the combination of temperature and salinity variability during a multigenerational exposure of partheogenetically reproducing Daphnia pulex. We raised D. pulex in an orthogonal combination of daily fluctuations in temperature (15, 15–25, 15–30 °C) and salinity (0, 0–2, 0–5). Initially metabolic rates were lower under all variable temperature and variable salinity treatments. By the 6th generation there was little metabolic variation among low and intermediate temperature and salinity treatments, but metabolic suppression persisted at the most extreme salinity. When grown in the control condition for the 6th generation, metabolic suppression was only observed in D. pulex from the most extreme condition (15–30 °C, 0–5 salinity). Generation time was influenced by acclimation temperature but not salinity and was quickest in specimens reared at 15–25 °C, likely due to Q₁₀ effects at temperatures closer to the optima for D. pulex, and slowest in specimens reared at 15–30 °C, which may have reflected elevated CSR. Acute tolerance to temperature (LT₅₀) and salinity (LC₅₀) were both highest in D. pulex acclimated to 15–30 °C and salinity 0. LT₅₀ and LC₅₀ increased with increasing salinity in specimens raised at 15 °C and 15–25 °C, but decreased with increasing salinity in specimens raised at 15–30 °C. Thus, increasing temperature confers cross-tolerance to salinity stress, but the directionality of synergistic effects of temperature and salinity depend on the degree of environmental variability. Overall, the results of our study suggest that temperature is a stronger determinant of metabolism, growth, and tolerance thresholds, and assessment of the ecological impacts of environmental change requires explicit information regarding the degree of environmental variability.