Survival of cabbage stem flea beetle larvae,Psylliodes chrysocephala,exposed to low temperatures

The cabbage stem flea beetle, Psylliodes chrysocephala (L.) (Coleoptera: Chrysomelidae), is a major pest of winter oilseed rape. The larvae live throughout winter in leaf petioles and stems. Winter temperatures might play an important role in survival during winter and hence population dynamics, yet to what degree is unknown. This study investigates the effect of exposure time, cold acclimation, and larval stage on survival at ?5 and ?10 °C. Exposure time at ?5 °C was 1, 2, 4, 8, 12, 16, and 20 days and 6, 12, 24, 36, 48, 72, 96, 120, and 144 h at ?10 °C. Mortality increased with increasing exposure time and was significantly lower for cold‐acclimated larvae. Estimated time until an expected mortality of 50% (LT₅₀) and 90% (LT₉₀) of larvae exposed to ?5 °C was 7.4 and 9.6 days (non‐acclimated) and 11.0 and 15.1 days (acclimated), respectively. Estimated LT₅₀ for non‐acclimated and acclimated larvae exposed to ?10 °C was 32.6 and 70.5 h, respectively, and estimated LT₉₀ 66.8 and 132.2 h. Significant differences in mortality between larval stages were observed only at ?5 °C. When exposed to ?5 °C for 8 days, mortality of first and second instars was 81.2 and 51.3%, respectively. When exposed to ?10 °C for 2 days, mortality of first and second instars was 70.5 and 76.1%. Data on winter temperatures in Denmark from 1990 to 2013 showed that larvae were rarely exposed to a number of continuous days at ?5 or ?10 °C causing a potential larval mortality of 50–90%.     

Susceptibility of Heliothis armiger to a commercial nuclear polyhedrosis virus

The susceptibility of Heliothis armiger larvae of different ages to a commercial nuclear polyhedrosis virus (NPV), Elcar, was determined by bioassay. The median lethal dosage (LD₅₀) increased 150-fold during the first week of larval life at 25°C, i.e., during development to early fourth instar, but daily feeding rate and thus potential virus acquisition also increased. A linear relationship was determined between log LD₅₀ and larval length, indicating that larval length constitutes a useful index for estimating the susceptibility of larval populations. Median lethal times (LT₅₀s) were similar for larvae tested at ages of 0 to 7 days and ranged from 3.6 to 8.0 days at 30°C. The amount of virus produced in a single, infected neonate was equivalent to 1.4 × 10⁶ LD₅₀s for neonates, a 900,000-fold increase on the dose supplied. The data support the practice of directing the NPV against neonates, but, on the basis of larval susceptibility alone, the age of larvae at treatment may not always be critical.     

Temperature-dependent development, diapause and cold tolerance of Gratiana graminea, a potential biological control agent of Solanum viarum in Florida, USA

The objectives of this study were to examine temperature-dependent development, diapause and cold tolerance of Gratiana graminea Klug (Chrysomelidae), a candidate biological control agent of tropical soda apple, Solanum viarum Dunal (Solanaceae). Immature development was examined at six constant temperatures ranging from 15°C to 30°C. Diapause induction was determined by exposing adults to either long or short photoperiods at 20°C and cold tolerance was assessed by exposing adults to 0°C. G. graminea completed development at temperatures ranging from 20°C to 30°C. Linear regression estimated a lower temperature threshold of 11.7°C and 312 degree-days were required to complete development. Diapause was induced when adults were exposed to short photoperiods (10:14 L:D h) at 20°C. The lethal times for diapausing adults of G. graminea at 0°C (LT₅₀?=?19?days, LT₉₀?=?41?days) were two times higher compared to Gratiana boliviana Spaeth, a biological control agent already established in south and central Florida, USA. The presence of diapause and the greater cold tolerance suggest that G. graminea may establish and perform better than G. boliviana in northern Florida.     

Survival and chill-coma in the adult lesser mealworm,Alphitobius diaperinus (Coleoptera: Tenebrionidae), exposed to low temperatures

The survival of adult Alphitobius diaperinus (Coleoptera: Tenebrionidae) at low temperatures has been investigated under laboratory conditions: LT₅₀ ranges from 9.5 days at 6°C to 47.6 days at 10°C and LT₉₉ ranges from 20 days at 6°C to 102.2 days at 10°C. Mean survival shows no significant differences between sexes, but at 10°C, all the males are dead after 88 days and all the females after 107 days. Temperature of 6°C is progressively lethal for the beetles: activity ceases, all the insects are in chill-coma after 12 days and died after 22 days. At 10°C, after 1 month, 70% of the insects are still active. Chill-coma is a reversible state but insects which entered chill-coma last did not recover in most cases and also died the last. During winter sanitary disinfections, A. diaperinus is submitted to subzero temperatures: the rate of adult survival is a key factor governing the population dynamics in the poultry houses.     

Dehydrin, alcohol dehydrogenase, and central metabolite levels are associated with cold tolerance in diploid strawberry (Fragaria spp.)

The use of artificial freezing tests, identification of biomarkers linked to or directly involved in the low-temperature tolerance processes, could prove useful in applied strawberry breeding. This study was conducted to identify genotypes of diploid strawberry that differ in their tolerance to low-temperature stress and to investigate whether a set of candidate proteins and metabolites correlate with the level of tolerance. 17 Fragaria vesca, 2 F. nilgerrensis, 2 F. nubicola, and 1 F. pentaphylla genotypes were evaluated for low-temperature tolerance. Estimates of temperatures where 50 % of the plants survived (LT₅₀) ranged from ?4.7 to ?12.0 °C between the genotypes. Among the F. vesca genotypes, the LT₅₀ varied from ?7.7 °C to ?12.0 °C. Among the most tolerant were three F. vesca ssp. bracteata genotypes (FDP821, NCGR424, and NCGR502), while a F. vesca ssp. californica genotype (FDP817) was the least tolerant (LT₅₀ ?7.7 °C). Alcohol dehydrogenase (ADH), total dehydrin expression, and content of central metabolism constituents were assayed in select plants acclimated at 2 °C. The LT₅₀ estimates and the expression of ADH and total dehydrins were highly correlated (r _adh = ?0.87, r _dehyd = ?0.82). Compounds related to the citric acid cycle were quantified in the leaves during acclimation. While several sugars and acids were significantly correlated to the LT₅₀ estimates early in the acclimation period, only galactinol proved to be a good LT₅₀ predictor after 28 days of acclimation (r _galact = 0.79). It is concluded that ADH, dehydrins, and galactinol show great potential to serve as biomarkers for cold tolerance in diploid strawberry.     

Induced expression of the class II chitinase gene during cold acclimation and dehydration of bermudagrass (Cynodon sp.)

Bermudagrass cultivars vary greatly in their ability to survive freezing temperatures as a result of a differential ability to cold acclimate (CA) at temperatures slightly above 0°C. Little information exists on the genetic and physiological mechanisms associated with the cold acclimation process in bermudagrass. Experiments were conducted to study the changes in chitinase gene expression during cold acclimation of freeze-tolerant bermudagrass cultivars. A chitinase gene (CynCHT1) was isolated from ’Midiron’ bermudagrass. Because the hydrophilic protein putatively encoded by the gene lacked an N-terminal cysteine-rich domain and a hydrophobic C-terminal extension, it was classified a class II chitinase. The expression patterns of this and related chitinase genes in response to CA, drought, and ABA were investigated in freeze-tolerant ’MSU’ (LT₅₀=?11°C), Midiron (LT₅₀=?10°C) and ’Uganda’ (LT₅₀=?8°C) bermudagrasses. Northern-blot analysis indicated expression in the crown tissues induced by CA at 8°C/2°C day/night temperature cycles. Induction of gene expression was evident in tissues sampled at 2 and 28 days after initiating CA. Expression after 2-days de-acclimation at 28°C/24°C was similar to control levels. Significantly higher levels of CA-induced chitinase gene expression were observed in MSU and Midiron, compared to Uganda. Similar expression patterns were observed among the cultivars in responses to drought and ABA. These results suggest that chitinases have important roles in bermudagrass response to low temperature and dehydration stresses.     

The role of temperature in enhancing the insecticidal activity of Bacillus thuringiensis against Spodoptera littoralis larvae

The present study scrutinised how far temperature would affect the velocity of the insecticidal activity of Bacillus thuringiensis, as the rapidity of pest control achievements is of a great concern. Third instar Spodoptera littoralis larvae were treated with Bt at three concentration levels under five different temperatures (15°C, 20°C, 25°C, 30°C and 35°C). LT₅₀s were evaluated in each case. The LT₅₀ values showed various levels of reductions as temperature and/or Bt concentration increased, indicating that the velocity of mortality (1/LT₅₀) and/or the rapidity of Bt activity was almost temperature dependant. However, relatively high and low reduction percentages in the LT₅₀ values on the elevation of 5°C were obtained at lower and higher temperature ranges, respectively. The temperature coefficient, Q ₁₀ values, determined within narrow ranges (5°C) showed great reductions when temperature increased from 15°C to 20°C at all Bt concentrations. Raising temperature by 5°C above 20°C or 25°C almost caused similar Q ₁₀ values indicating constant increase in the response of Bt activity within 20–30°C temperature range. Q ₁₀ values over 30°C were comparatively very low. This proved that decrease in Q ₁₀ values due to the rise of temperature was dependant on the starting temperature.     

Effect of temperature on germination,growth, and infectivity of the mosquito pathogen Tolypocladium cylindrosporum (deuteromycotina; hyphomycetes)

The mosquito pathogen Tolypocladium cylindrosporum was examined with regard to its response to temperature. Similar temperature ranges were found for growth, germination, and infectivity of blastospores and conidia. Germination occurred at 8° and 33°C but not at 6° and 35°C. Optimal germination and growth was noted between 24° and 27°C for both spore types. Infectivity of blastospores and conidia at different temperatures was examined by exposing L₂Aedes sierrensis larvae to concentrations of 5 × 10⁵ blastospores/ml or 5 × 10⁶ conidia/ml. Larvae were incubated at 12°, 15°, 25°, and 30°C. Infection occurred at all temperatures tested with LT₅₀ values ranging from 22.7 days (12°C) to 5.6 (25°C) days for conidia and 4.7 days (12°C) to 0.6 day (25°C) for blastospores. These results confirmed earlier findings that blastospores infected and killed host larvae more rapidly than conidia and suggested that this difference is largely due to the more rapid germination rate of blastospores. These experiments demonstrated that T. cylindrosporum can be active against mosquito larvae over a broad range of temperatures encompassing both the cold-water habitat of certain temperate mosquito species as well as the habitat of tropical vector species.     

Low temperature mortality of the peach-potato aphid Myzus persicae

ABSTRACT.

• 1 The mean supercooling points of first instar and adult Myzus persicae (Sulzer) maintained at 20°C and cooled at 1°C min^?1 were ?26.6 and ?25.0°C respectively.
• 2 The LT₅₀ (temperature) of the same age groups drawn from the same population and cooled at the same rate were ?8.1 and ?6.9°C, indicating extensive pre-freeze mortality in M.persicae under laboratory conditions.
• 3 Acclimation at 10 and 5°C did not affect supercooling but depressed the LT₅₀ of both first instars and adult aphids.
• 4 Freezing of leaves during feeding did not increase mortality above that expected from the direct effects of low temperature.
• 5 The level of cold in different winters can be expressed in terms of the total number of frost days, and the frequency of abnormally cold days. Winter temperatures differ markedly in a vertical profile from the soil to the soil or grass surface, and then to the air (and foliage) above.
• 6 The time of the first record of M.persicae in suction trap samples is correlated with January and February temperatures except in the west of England and Wales. Further north December and January temperatures are relatively more important.
• 7 Winter temperatures and the resultant aphid mortality is a primary determinant of the timing of the spring migration.

     

Susceptibility of the cigarette beetle Lasioderma serricorne (Coleoptera: Anobiidae) to hypoxia

The susceptibility of the cigarette beetle Lasioderma serricorne (F.) to hypoxia was examined at three different oxygen concentrations (0.5?C0.8, 1.0?C1.3, and 2.0?C2.3?%) and four different temperature/humidity (RH) conditions: 30?°C/75?% RH, 25?°C/75?% RH, 20?°C/43?% RH, and 15?°C/43?% RH. The influence of humidity on mortality was also examined at three humidity levels (21, 43, and 75?% RH) at 1.0?C1.3?% oxygen (O₂) and 25?°C. Our results revealed that adult beetles were the most tolerant at 2.0?C2.3?% O₂ and that the larvae were the most tolerant at O₂ levels <1.0?C1.3?%. Mortality increased with increasing temperatures and decreasing O₂ concentrations. At 30?°C, 75?% RH, and 0.5?C0.8?% O₂, the 99?% lethality (LT₉₉) of larvae was 6.9?days; however, it increased to 20?days when the temperature was decreased to 25?°C or when O₂ levels were increased to 1.0?C1.3?%. Humidity also influenced mortality of both larval and adult beetles. LT₉₉ values for larvae at 25?°C and 1.0?C1.3?% O₂ were 24.0, 44.6, and 50.2?days at 21, 43, and 75?% RH, respectively. Results of this study indicate that a controlled atmosphere (CA) with reduced oxygen levels (<0.5?C0.8?% O₂) represents an effective measure for disinfesting stored tobacco as an alternative to conventional phosphine fumigation at temperatures >30?°C.     

The sea urchin Lytechinus variegatus lives close to the upper thermal limit for early development in a tropical lagoon

Thermal tolerance shapes organisms' physiological performance and limits their biogeographic ranges. Tropical terrestrial organisms are thought to live very near their upper thermal tolerance limits, and such small thermal safety factors put them at risk from global warming. However, little is known about the thermal tolerances of tropical marine invertebrates, how they vary across different life stages, and how these limits relate to environmental conditions. We tested the tolerance to acute heat stress of five life stages of the tropical sea urchin Lytechinus variegatus collected in the Bahía Almirante, Bocas del Toro, Panama. We also investigated the impact of chronic heat stress on larval development. Fertilization, cleavage, morula development, and 4‐armed larvae tolerated 2‐h exposures to elevated temperatures between 28–32°C. Average critical temperatures (LT₅₀) were lower for initiation of cleavage (33.5°C) and development to morula (32.5°C) than they were for fertilization (34.4°C) or for 4‐armed larvae (34.1°C). LT₅₀ was even higher (34.8°C) for adults exposed to similar acute thermal stress, suggesting that thermal limits measured for adults may not be directly applied to the whole life history. During chronic exposure, larvae had significantly lower survival and reduced growth when reared at temperatures above 30.5°C and did not survive chronic exposures at or above 32.3°C. Environmental monitoring at and near our collection site shows that L. variegatus may already experience temperatures at which larval growth and survival are reduced during the warmest months of the year. A published local climate model further suggests that such damaging warm temperatures will be reached throughout the Bahía Almirante by 2084. Our results highlight that tropical marine invertebrates likely have small thermal safety factors during some stages in their life cycles, and that shallow‐water populations are at particular risk of near future warming.     

Cold-induced changes of enzymes,proline, carbohydrates,and chlorophyll in wheat

Quantitative changes in total leaf soluble proteins, proline, carbohydrate content, chlorophyll fluorescence, guaiacol peroxidase (POD) and catalase (CAT) activities were determined in a less cold-hardy (LCH) spring cv. Kohdasht (LT₅₀ = −6°C), a semi cold-hardy (SCH) facultative cv. Azar 2 (LT₅₀ = −15°C), and a cold-hardy (CH) winter cv. Norstar (LT₅₀ = −26°C) of wheat (Triticum aestivum L.) exposed to 4°C for 9 weeks. Seedlings were grown in a controlled growth room for 14 days at 20°C and then transferred to 4°C (experimental day 0) for 63 days (cold treatment); otherwise they were maintained continuously at 20°C (control treatment). The samples were harvested 0, 2, 21, 28, 42, and 63 days after exposure to 4°C. The results showed significant low temperature (LT)-induced accumulation of total soluble proteins, proline, and carbohydrates and elevation in activities of CAT and POD in leaves of SCH and CH winter cultivars rather than in LCH spring cultivar. In contrast, the chlorophyll fluorescence (F _v/F _m) declined during LT treatment irrespective of cultivar. The results suggest that developmental traits such as vernalization requirement of wheat affects on cold-tolerance expression system of plants.     

Temperature and salinity tolerances of larval Californian grunion, Leuresthes tenuis (Ayres): A comparison with gulf grunion, L. Sardina (Jenkins & Evermann)

Temperature and salinity tolerances were determined for larval California grunion, Leuresthes tenuis (Ayres), and compared with previous data for Gulf of California grunion, L. sardina (Jenkins & Evermann). Larvae of similar age and acclimation history showed little interspecific difference in thermal tolerance, as measured by half-hour LT₅₀ values for 20–30 day old late postlarvae acclimated at various temperatures, and by upper and lower incipient lethal temperatures for 18°C-acclimated prolarvae. The upper incipient lethal temperature differed by 1 deg.-C (32°C for L. tenuis, 31°C for L. sardina), while the lower incipient lethal temperature for the 18°C acclimated prolarvae of both species was 7.5°C. L. tenuis larvae were much less euryhaline than L. sardina, with incipient lethal salinities of 4.2–41 %. for prolarvae and 8.6–38 %. for 20-day-old postlarvae; comparable values for L. sardina are 4–67.5 %. and 5–57.5 %. Both species show a decrease in temperature and salinity tolerance with age. The larvae of these disjunct congeners show a significant physiological divergence in euryhalinity but not in overall temperature tolerance. These tolerances are discussed in relation to the respective geographic ranges and behavioral responses of the two species.     

Effects of constant and fluctuating incubation temperatures on hatching success and hatchling traits in the diamondback terrapin (Malaclemys terrapin) in the context of the warming climate

As global temperatures continue to rise, so too will the nest temperatures of many species of turtles. Yet for most turtle species, including the estuarine diamondback terrapin (Malaclemys terrapin), there is limited information on embryonic sensitivity to elevated temperature. We incubated eggs of M. terrapin at three, mean temperatures (31, 34, 37 °C) under two thermal exposure regimes (constant or semi-naturally fluctuating temperature) and measured hatching success, developmental rate, and hatchling size. Hatching success was 100% at 31 °C and 67% at 34 °C, respectively; at 37 °C, all eggs failed early in the incubation period. These values were unaffected by exposure regime. The modeled LT₅₀ (temperature that was lethal to 50% of the test population) was 34.0 °C in the constant and 34.2 °C in the fluctuating thermal regime, reflecting a steep decline in survival between 33 and 35 °C. Hatchlings having been incubated at a constant 34 °C hatched sooner than those incubated at 31 °C under either constant or fluctuating temperature. Hatchlings were smaller in straight carapace length (CL) and width after having been incubated at 34 °C compared to 31 °C. Larger (CL) hatchlings resulted from fluctuating temperature conditions relative to constant temperature conditions, regardless of mean temperature. Based upon recent temperatures in natural nests, the M. terrapin population studied here appears to possess resiliency to several degrees of elevated mean nest temperatures, beyond which, embryonic mortality will likely sharply increase. When considered within the mosaic of challenges that Maryland's M. terrapin face as the climate warms, including ongoing habitat losses due to sea level rise and impending thermal impacts on bioenergetics and offspring sex ratios, a future increase in embryonic mortality could be a critical factor for a population already experiencing ecological and physiological challenges due to climate change.     

Chemical compositions and larvicidal effectiveness of Rhododendron ponticum essential oil on Neodiprion sertifer larvae

In many countries throughout the world, the European pine sawfly, Neodiprion sertifer (Geoffroy, 1785) (Hymenoptera: Diprionidae) is known as a significant forest pest that damages pine needles. As a result of the insect damage, young pine trees, especially, might dry up totally. Mechanical, biotechnical, biological, and chemical approaches are applied in research to keep it under control. Due to the increasing danger of chemical contamination associated with the use of synthetic pesticides to control insects, alternative control strategies have been sought, and studies on essential oils have been prioritized. In this research, in addition to determining the chemical analysis of Rhododendron ponticum L. (Ericales; Ericaceae) essential oil by GC–MS, the larvicidal effects of R. ponticum on N. setifer in the six larval stages were also investigated. The research was conducted in the Artvin Çoruh University Central Laboratory under laboratory settings (25 ± 1 °C, 65 ± 5% RH, and 14L:10D h photoperiods) during 2020–21. 1-Methyl-2-pyrrolidone (38.41%), Phenylethy alcohol (10.07%), Linalool (8.01%), Myrtenol 5.68%), β-Pinene (4.26%), Citronellol (4.05%), α-Pinene (3.02%) were found to be the major components of R. ponticum oil. The administration of R. ponticum essential oil at 10, 15, and 20 μL/Petri doses, as well as controls at the 24th, 48th, 72nd, and 96th hours, produced various rates of death (19.6–100%) on six larval stages of N. sertifer. According to LD₅₀ and LD₉₀ values, the highest toxicity was determined as L₁ (0.58 μL/larva) and the lowest toxicity was as L₆ (10.44 μL/larva). In conclusion, R. ponticum essential oil showed high toxicity against L₁ and L₂ larval stages of N. sertifer.     

Supplemental intermittent-day heat training and the lactate threshold

Heat acclimation over consecutive days has been shown to improve aerobic-based performance. Recently, it has been suggested that heat training can improve performance in a temperate environment. However, due to the multifactorial training demands of athletes, consecutive-day heat training may not be suitable. The current study aimed to investigate the effect of brief (8×30 min) intermittent (every 3–4 days) supplemental heat training on the second lactate threshold point (LT₂) in temperate and hot conditions. 21 participants undertook eight intermittent-day mixed-intensity treadmill exercise training sessions in hot (30 °C; 50% relative humidity [RH]) or temperate (18 °C; 30% RH) conditions. A pre- and post-incremental exercise test occurred in temperate (18 °C; 30% RH) and hot conditions (30 °C; 50% RH) to determine the change in LT₂. The heat training protocol did not improve LT₂ in temperate (Effect Size [ES]±90 confidence interval=0.10±0.16) or hot (ES=0.26±0.26) conditions. The primary finding was that although the intervention group had a change greater than the SWC, no statistically significant improvements were observed following an intermittent eight day supplemental heat training protocol comparable to a control group training only in temperate conditions. This is likely due to the brief length of each heat training session and/or the long duration between each heat exposure.     

Heat tolerance of early developmental stages of glacier foreland species in the growth chamber and in the field

In glacier forelands, seeds readily germinate, however, a high proportion of seedlings die shortly after their appearance. We hypothesized that besides drought, frost and missing safe sites, heat on the ground surface could be one of the major threats for seedlings. The heat strain in different ground strata was assessed from 2007 to 2010. The heat tolerance (LT₅₀) of eleven alpine species from different successional stages was tested considering imbibed (G1) and germinated seeds (G2) as well as seedlings (G3). Additionally, the heat hardening capacity of seedlings was determined in the field. Across all species, LT₅₀ decreased significantly by 9 K from G1 (55 °C) to G3 (46 °C), similarly in all species of the successional stages. Field-grown seedlings had mostly an increased LT₅₀ (2K). Intraspecifically, LT₅₀ of seedlings varied between 40.6 and 52.5 °C. Along the chronosequence, LT₅₀ in G1 was similar, but was higher in G2 and G3 of early successional species. The highest temperatures occurred at 0–0.5 cm in air (mean/absolute maximum: 42.6/54.1 °C) posing a significant heat injury risk for seedlings when under water shortage transpirational cooling is prevented. Below small stones (0–0.5 cm), maxima were 4 K lower, indicating heat safer microsites. Maxima >30 °C occurred at 32.3, >40 °C at 6.2 %. Interannually, 2010 was the hottest year with heat exceeding LT₅₀ at all microsites (0–0.5 cm). Temperature maxima on sandy surfaces were lower than on microsites with gravel (diameter <5–10 mm). The hot summer of 2010 may be a small foretaste of in future more severe and frequent heat waves. Ground surface temperature maxima at the pioneer stage are already now critical for heat survival and may partly explain the high seedling mortality recognized on recently deglaciated terrain.     

Exposure to subfreezing temperature and a freeze-thaw cycle affect freezing tolerance of winter wheat in saturated soil

Winter wheat is sown in the autumn and harvested the following summer, necessitating the ability to survive subfreezing temperatures for several months. Autumn months in wheat-growing regions typically experience significant rainfall and several days or weeks of mild subfreezing temperatures at night, followed by above-freezing temperatures in the day. Hence, the wheat plants usually are first exposed to potentially damaging subfreezing temperatures when they have high moisture content, are growing in very wet soil, and have been exposed to freeze-thaw cycles for a period of time. These conditions are conducive to freezing stresses and plant responses that are different from those that occur under lower moisture conditions without freeze-thaw cycles. This study was conducted to investigate the impact of mild subfreezing temperature and a freeze-thaw cycle on the ability of 22 winter wheat cultivars to tolerate freezing in saturated soil. Seedlings that had been acclimated at +4°C for 5 weeks in saturated soil were frozen to potentially damaging temperatures under three treatment conditions: (1) without any subzero pre-freezing treatment; (2) with a 16-h period at ?3°C prior to freezing to potentially damaging temperatures; and (3) with a freeze-thaw cycle of ?3°C for 24 h followed by +4°C for 24 h, followed by a 16-h period at ?3°C prior to freezing to potentially damaging temperatures. In general, plants that had been exposed to the freeze-thaw cycle survived significantly more frequently than plants frozen under the other two treatments. Plants that had been exposed to 16 h at ?3° (without the freeze-thaw cycle) before freezing to potentially damaging temperatures survived significantly more frequently than plants that were frozen to potentially damaging temperatures without a subzero pre-freezing treatment. These results indicated that cold-acclimated wheat plants actively acclimate to freezing stress while exposed to mild subfreezing temperatures, and further acclimate when allowed to thaw at +4°C for 24 h. The cultivar Norstar had the lowest LT₅₀ (temperature predicted to be lethal to 50% of the plants) of the 22 cultivars when frozen with either of the subzero pre-freezing treatments, but several cultivars had lower LT₅₀ scores than Norstar when frozen without a subzero pre-freezing treatment. We conclude it may be possible to improve winterhardiness of wheat grown in saturated soil by combining the ability to effectively respond to mild subzero pre-freezing temperatures with a greater ability to withstand freezing to damaging temperatures without a subzero pre-freezing exposure.