Alfalfa water status and cold hardiness as influenced by cold acclimation and water stress

Abstract Water stress at a nonacclimating temperature (18–20°C) increased the cold hardiness of Medicagosativa L. (alfalfa) plants. This increased cold hardiness was retained when the previously water-stressed plants were cold acclimated (2–9°C) in the absence of water stress. Water stress during cold acclimation also increased cold hardiness. Alfalfa was demonstrated to suffer injury, measured as decreased growth following freezing, at sub-lethal temperatures. During cold acclimation the turgor potential (ψ) of watered plants increased, whereas the solute potential and the water content per unit dry weight decreased. The large positive psgrdap of acclimated plants indicates that the decreased water content per unit dry weight is related to an increased proportion of tissue dry matter rather than to tissue dehydration.     

Factors affecting the cold hardiness of the peach-potato aphid Myzus persicae

Supercooling point studies were used to investigate the factors influencing the cold hardiness of the peach-potato aphid Myzus persicae, a freezing-susceptible insect. Overwintering adults lost cold hardiness as winter progressed, with a variable proportion showing a marked reduction in supercooling ability. Cold hardiness increased in spring so that all individuals demonstrated extensive supercooling ability typical of aphids reared in the laboratory at 20°C with a long photoperiod; these levels of cold hardiness were maintained in the field during summer and early autumn. First instar nymphs demonstrated considerable cold hardiness all year. Surface moisture caused inoculative freezing in some first instar nymphs and adults when supercooled, but the majority were unaffected. In the laboratory, adults starved for 7 days at 5°C showed distinct losses of supercooling potential equivalent to those observed in the field during mid to late winter. No loss of cold hardiness was found in first instar nymphs starved under the same conditions. The results demonstrate that the cold hardiness characteristics of M. persicae are atypical of those observed in other freezing-susceptible insects and it is suggested that continued feeding during mild winter conditions allows maintenance of cold hardiness particularly in adult aphids, and provides a possible explanation for the successful anholocyclic overwintering of M. persicae during such winters.     

Geographic variation in cold hardiness of eggs and neonate larvae of the yellow-spotted longicorn beetle Psacothea hilaris

Cold hardiness of eggs and neonate larvae of the yellow-spotted longicorn beetle, Psacothea hilaris (Pascoe) was examined using six geographical populations in Japan. Particular attention was paid to cold hardiness of eggs and neonate larvae of the subtropical population (Ishigaki), because the east Japan populations are considered to have been introduced from a subtropical area, and the overwintering stage in the east Japan populations is incidentally shifted from the original mature larval stage to the egg or neonate larval stages. When the eggs were exposed to low temperatures for 1 h, the decrease in hatchability became significant at –12°C in the southernmost two populations (Ishigaki and Naze), and at –16°C in the northern populations. After 1 h exposure to –20°C, few eggs could hatch in the Ishigaki population, whereas 27–55% of the eggs survived in the northern populations. Pre-chilling of the eggs at 10°C for 10 days enhanced the cold hardiness in all populations. This effect was particularly distinct in the subtropical population; the eggs of the Ishigaki population became as cold hardy as those of the northern populations after acclimation. These results suggest that the subtropical population is capable of establishing itself in east Japan, where the winter is cold.     

Tolerance to multiple climate stressors: a case study of Douglas‐fir drought and cold hardiness

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Abscisic acid and mefluidide in the regulation of cold hardiness development in Solanum tuberosum L. potato

Plants of Solanum tuberosum L. potato do not cold acclimate when exposed to low temperature such as 5°C, day/night. When ABA (45 M) was added to the culture medium, stem-cultured plantlets of S. tuberosum, cv. Red Pontiac, either grown at 20°C/15°C, day/night, or at 5°C, increased in cold hardiness from –2°C (killing temperature) to –4.5°C. The increase in cold hardiness could be inhibited in both temperature regimes if cycloheximide (70 M) was added to the culture medium at the inception of ABA treatment. Cycloheximide did not inhibit cold hardiness development, however, when it was added to the culture medium 3 days after ABA treatment.When pot-grown plants were foliar sprayed with mefluidide (50 M), ABA content increased from 10 nmol to 30 nmol g^–1 dry weight and plants increased in cold hardiness from –2°C to about –3.5°C. The increases in free ABA and cold hardiness occurred only in plants grown at 20°C/15°C; neither ABA nor cold hardiness increased in plants grown at 5°C.The results suggest that an increase in ABA and a subsequent de novo synthesis of proteins are required for the development of cold hardiness in S. tuberosum regardless of temperature regime, and that the inability to synthesize ABA at low temperature, rather than protein synthesis, appears to be the reason why S. tuberosum does not cold acclimate.     

A common strategy of cold hardiness in ants of the genus <Emphasis Type="Italic">Myrmica</Emphasis> (Formicidae,Hymenoptera) in Northeast Asia

The biotopic distribution, nest structure, wintering conditions, and cold hardiness of four ecologically contrasting ant species (Myrmica angulinodis, M. kamtschatica, M. bicolor, and M. transsibirica) are considered. The cold hardiness of these species is typical of the genus: the supercooling points vary from −28 to −31°C; cold hardiness levels (LT50%) are higher by 5°C. At this level of cold hardiness, ants can be practically ubiquitous across the whole Hypoarctic (Berman et al., 2007). However, the above Myrmica species are strictly segregated (M. kamtschatica occurs in moss bogs, M. angulinodis and M. transsibirica, on dry and warm south slopes, and M. bicolor, in sandy-gravel floodplains), probably due to different requirements for weather conditions in summer and depth of ground thawing. At present, the excess cold hardiness common to the species in question (exceeding the nest temperature by 5–10°C in different years) is not adaptive and may be considered as preadaptive. It could have been acquired during ancient cold epochs or inherited by the genus as a concomitant result of adaptation not to low temperatures but, for instance, to aridity. These Myrmica species do not undergo selection for resistance to negative temperatures since their current level of cold hardiness is excessive, considering the possible wintering temperatures.     

Overwintering strategy of two weevils infesting three gorse species: When cold hardiness meets plant-insect interactions

The cold hardiness of two closely related weevil species, Exapion ulicis and E. lemovicinum was studied in relation to their life cycles. These two seed-eating weevils reproduce on Ulex plant species with different fruiting phenologies. E. ulicis lays eggs in spring and overwinters as an adult while E. lemovicinum lays eggs in autumn and overwinters as a larva. Adult weevils were collected in natural populations of Brittany (Western France) and characterized with morphological and molecular tools before experiments. We showed that both weevil species exhibited low supercooling points (SCPs) with mean seasonal values below −17 °C. Fresh mass, moisture content and sex were not correlated to supercooling ability. Weevils died upon freezing and the lower lethal temperatures (LLT) were within the range of SCP, indicating that both species are freezing intolerant. Comparison between species for SCP, LLT and survival to exposure at −8 °C in winter showed a higher cold resistance for E. ulicis than for E. lemovicinum. In addition, the seasonal evolution of cold hardiness differed depending on the species. These features suggest that response to cold of weevils is linked to their life cycles, and thus to the life history of their host plants.     

Supercooling ability and cold hardiness of the pollen beetle Meligethes aeneus

Supercooling point (SCP) and cold‐hardiness of the pollen beetle Meligethes aeneus (Fabricius) (Coleoptera: Nitidulidae) were investigated. Mature eggs from the oviduct were supercooled on average to ?28.0 °C and from oilseed rape buds to ?24.4 °C; first instars were supercooled to ?21.0 °C and second instars to ?16.8 °C. Despite their high supercooling ability, none of the eggs survived 24 h exposure to ?2.5 °C. The supercooling ability of adults varied significantly among feeding and non‐feeding beetles: high SCPs prevailed during the whole warm period, being about ?12 °C; low values of SCP of ?20 °C dominated in non‐feeding beetles. In spring and autumn, beetles displayed the same acclimation efficiency: after 1 week of exposure at 2.0 °C with no access to food their SCPs were depressed equally by about 3 °C. Meligethes aeneus beetles have a different response to low temperatures depending on the season. The lowest tolerance was found in reproductively active beetles after emergence from overwintering sites; the time needed to kill 50% of individuals (Ltime₅₀) was 56.2 h at ?7 °C and the lower lethal temperature needed to kill 50% (Ltemp₅₀) after 24 h exposure was ?8.6 °C. Cold hardiness increased from midsummer to midwinter; Ltime₅₀ was 80 h in August, 182.8 h in September, and 418.1 h in January. Lethal temperature after 24 h exposure was ?9.1 °C in August and ?9.8 °C in September. In February, after diapause, the beetles started to loose their cold tolerance, and Ltemp₅₀ was slightly increased to ?9.5 °C. Hibernating beetles tolerated long exposure at ?7 °C well, but mortality was high after short exposure if the temperature dropped below ?9 °C for 24 h. Despite the season, the beetles died at temperatures well above their mean SCP; consequently, SCP is not a suitable index for cold hardiness of M. aeneus.     

Changes in chemical components in the freshwater apple snail, Pomacea canaliculata (Gastropoda: Ampullariidae), in relation to the development of its cold hardiness

The apple snail, Pomacea canaliculata, is an invasive freshwater snail. It increases its cold hardiness before winter. However, the physiological mechanism of cold hardiness in molluscs is poorly understood, especially in freshwater molluscs. In this study, we examined the changes in low molecular weight compounds, glycogen and lipids, in the body of P. canaliculata in association with the development of cold hardiness. When snails without cold hardiness were experimentally cold-acclimated, the amount of glycerol, glutamine, and carnosine increased, while glycogen and phenylalanine decreased. Overwintering cold-tolerant snails collected from a drained paddy field in November also showed increased glycerol in their bodies with decreasing glycogen concentration, compared to summer snails collected from a submerged field. Water content also decreased during the cold acclimation, although the water loss was minimal. These results indicate that the freshwater snail, P. canaliculata enhances cold hardiness by accumulation of some kinds of low molecular weight compounds in its body as some insects do. However, the actual function of each low molecular compound is still unknown.     

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.     

Impact of climate change on cold hardiness of Douglas‐fir (Pseudotsuga menziesii): environmental and genetic considerations

The success of conifers over much of the world's terrestrial surface is largely attributable to their tolerance to cold stress (i.e., cold hardiness). Due to an increase in climate variability, climate change may reduce conifer cold hardiness, which in turn could impact ecosystem functioning and productivity in conifer‐dominated forests. The expression of cold hardiness is a product of environmental cues (E), genetic differentiation (G), and their interaction (G × E), although few studies have considered all components together. To better understand and manage for the impacts of climate change on conifer cold hardiness, we conducted a common garden experiment replicated in three test environments (cool, moderate, and warm) using 35 populations of coast Douglas‐fir (Pseudotsuga menziesii var. menziesii) to test the hypotheses: (i) cool‐temperature cues in fall are necessary to trigger cold hardening, (ii) there is large genetic variation among populations in cold hardiness that can be predicted from seed‐source climate variables, (iii) observed differences among populations in cold hardiness in situ are dependent on effective environmental cues, and (iv) movement of seed sources from warmer to cooler climates will increase risk to cold injury. During fall 2012, we visually assessed cold damage of bud, needle, and stem tissues following artificial freeze tests. Cool‐temperature cues (e.g., degree hours below 2 °C) at the test sites were associated with cold hardening, which were minimal at the moderate test site owing to mild fall temperatures. Populations differed 3‐fold in cold hardiness, with winter minimum temperatures and fall frost dates as strong seed‐source climate predictors of cold hardiness, and with summer temperatures and aridity as secondary predictors. Seed‐source movement resulted in only modest increases in cold damage. Our findings indicate that increased fall temperatures delay cold hardening, warmer/drier summers confer a degree of cold hardiness, and seed‐source movement from warmer to cooler climates may be a viable option for adapting coniferous forest to future climate.     

Cold hardiness in immature stages and adults in the adult‐diapausing spider mite Stigmaeopsis longus

The cold hardiness of arthropods is an important characteristic associated with overwintering success. Cold‐tolerant stages affect overwintering strategy, especially in arthropods that continuously feed on evergreen host plants in temperate regions. However, cold hardiness to mildly low temperature is rarely investigated. In the present study, we estimate the stage‐specific cold hardiness of a population of the spider mite Stigmaeopsis longus (Saito) (Acari: Tetranychidae) occurring on evergreen Sasa bamboo in a temperate area (Kochi City, southwestern Japan). Individuals of each stage developing under diapause‐inducing conditions (LD 10 : 14 h at 20 °C) are maintained at 4 °C (approximating the mean daily minimum temperature for December in Kochi City) for 30 days (eggs are cooled immediately, without short‐day development first). They are then returned to the initial conditions to assess survival rates. The survival rate of adult females (i.e. the diapausing stage) is high (99.5%), as is that of adult males and deutonymphs of both sexes (84.2–98.7% and 89.7–89.8%, respectively). The survival rate of mobile immature stages tends to increase toward adulthood in both sexes, whereas the survival rate of the eggs and quiescent stages is extremely low (16–23% and 0–20%, respectively). The poor cold hardiness of the eggs and quiescent stages presumably prevents the normal development of immature individuals during winter.     

Ultrastructural features and formation of the micropylar apparatus in the cherry fly Rhagoletis cerasi

The micropylar apparatus (MA) in Rhagoletis cerasi (Diptera, Tephritidae) is located at the anterior pole of the egg and consists of two parts: an outer chorion and an inner vitelline membrane. Sperm entry takes place through the micropylar canal, 2.0–2.5 μm in diameter, which penetrates the micropylar endochorion and terminates in the thick vitelline membrane, thus forming the “pocket.” The pore of the micropylar canal, i.e., the micropyle, is covered by the exochorionic tuft. The formation of the MA is accomplished by 40 micropylar cells during oogenesis. These cells secrete the successive eggshell layers: the vitelline membrane, the wax layer, the innermost chorionic layer, the micropylar endochorion, and the exochorion. Two among 40 micropylar cells differentiate and form two tightly connected projections. The latter contain a bundle of parallel microtubules and participate in the formation of the micropylar canal and the pocket. At the tip of the projections there are two thin extensions full of microfilaments. In late developmental stages the two projections and the extensions degenerate and leave the canal and the pocket behind. We also discuss the structural features of the MA in relation to its physiology among Diptera.     

Dynamic thermal time model of cold hardiness for dormant grapevine buds

Background and Aims

Grapevine (Vitis spp.) cold hardiness varies dynamically throughout the dormant season, primarily in response to changes in temperature. The development and possible uses of a discrete-dynamic model of bud cold hardiness for three Vitis genotypes are described.

Methods

Iterative methods were used to optimize and evaluate model parameters by minimizing the root mean square error between observed and predicted bud hardiness, using up to 22 years of low-temperature exotherm data. Three grape cultivars were studied: Cabernet Sauvignon, Chardonnay (both V. vinifera) and Concord (V. labruscana). The model uses time steps of 1 d along with the measured daily mean air temperature to calculate the change in bud hardiness, which is then added to the hardiness from the previous day. Cultivar-dependent thermal time thresholds determine whether buds acclimate (gain hardiness) or deacclimate (lose hardiness).

Key Results

The parameterized model predicted bud hardiness for Cabernet Sauvignon and Chardonnay with an r² = 0·89 and for Concord with an r² = 0·82. Thermal time thresholds and (de-)acclimation rates changed between the early and late dormant season and were cultivar dependent but independent of each other. The timing of these changes was also unique for each cultivar. Concord achieved the greatest mid-winter hardiness but had the highest deacclimation rate, which resulted in rapid loss of hardiness in spring. Cabernet Sauvignon was least hardy, yet maintained its hardiness latest as a result of late transition to eco-dormancy, a high threshold temperature required to induce deacclimation and a low deacclimation rate.

Conclusions

A robust model of grapevine bud cold hardiness was developed that will aid in the anticipation of and response to potential injury from fluctuations in winter temperature and from extreme cold events. The model parameters that produce the best fit also permit insight into dynamic differences in hardiness among genotypes.     

Seasonal changes in cold hardiness of Ophraella communa

Insects can prepare themselves to tolerate subzero temperatures through various physiological changes, such as the alteration in body water or glycerol content. Indeed, it has been hypothesized that increasing glycerol body content has the benefit of decreasing the temperature necessary to freeze their body water and therefore increasing the supercooling point (SCP) and the cold hardiness. We here studied physiological plasticity in cold tolerance in Ophraella communa LeSage (Coleoptera: Chrysomelidae), a potential biological control agent of an invasive plant, the common ragweed, Ambrosia artemisiifolia L. (Asteraceae). Pupae of O. communa were collected from June to October, and the water and glycerol contents and the SCP of emerging adults were assessed. We found that SCP, water, and glycerol contents of beetles fluctuated significantly with season. Glycerol content of males and females increased with decreasing temperature between July and October, and glycerol content reached a maximum in October in the field. The lowest SCP was observed in adults in October prior to overwintering, and the highest SCP was evident in the summer population in July. Thus, cold hardiness of the beetles in the autumn population was significantly higher than in the summer population. We therefore conclude that cold tolerance, via changes in the relative composition of their body fluids and fats, is a plastic trait that can be influenced by fluctuations in abiotic factors (e.g., temperature) throughout the breeding season of the insect.     

Influence of soil moisture on egg cold hardiness in the migratory locust Locusta migratoria (Orthoptera: Acridiidae)

Abstract.  The present study investigates the influence of environmental moisture on cold hardiness of the migratory locust, Locusta migratoria . The water content of locust eggs kept in soil at 30 °C varies according to the moisture content of the substrate. In turn, it can significantly affect the supercooling point of locust eggs (range from −26 to −14.8 °C) and the mortality when exposed to subzero temperatures. Environmental moisture influences the supercooling capacity of eggs and their survival at low temperature. When locust eggs of the same water content are exposed to subzero temperatures under different soil moistures, their mortality varies between short-time exposure and long-time exposure at subzero temperatures. Given a short-time exposure, mortality in wet soil is lower than in dry soil due to the buffering effect of soil water against temperature change. The pattern of egg mortality is reversed after long-time exposure at low temperature, suggesting that inoculative freezing may be an important mortality factor. It is suggested that interactions between soil moisture and low temperature can influence the cold hardiness of locust eggs, and partial dehydration is beneficial to over-wintering eggs of the migratory locust.     

Antiviral effect of the egg wax of Amblyomma cajennense (Acari: Ixodidae)

The control of viral infections, especially those caused by influenza viruses, is of great interest in Public Health. Bio prospection has shown the presence of active principles in the hemolymph of arthropods, and in the salivary gland of ticks, and some of these are of interest for the development of new pharmacological drugs. Ticks lay their eggs in the environment, and to protect them from desiccation and microbial attack they involve the eggs in a waxy layer produced by an organ known as Gené’s Organ. In this study, the eggs wax from tick Amblyomma cajennense (Fabricius) was extracted using ice cold phosphate buffer. The antiviral activity was evaluated with picornavirus and influenza virus. In both cases egg wax was able to inhibit virus replication. For influenza virus, an amount as small as 12 μg/mL of crude egg wax suspension neutralized 128 UHA (hemaglutinant unit) of H₁N₁ influenza virus. With picornavirus, egg wax led to a 256-fold reduction in virus production by L929 cells. Egg wax was not cytotoxic to VERO, MDCK and L929 cell, being observed that the cell morphology was preserved with concentration as high as 2 mg/mL. In addition no genotoxic effect was observed for Vero cells, suggesting a very interesting potential antiviral activity.     

Geographical variation in egg cold hardiness: a study on the adaptation strategies of the migratory locust Locusta migratoria L.

Abstract. 1. For many species of insect, cold hardiness is an important trait that enables a population to develop in the next season and to extend its range. To elucidate the role of cold hardiness of the migratory locust Locusta migratoria L. in its outbreak and distribution areas, egg cold hardiness was examined in locusts derived from four locations from latitude 18°23'N to latitude 41°10'N in eastern China.
2. The supercooling points of eggs from different geographic populations did not differ significantly for the first development stage, with an average ± SE of −24.5 ± 0.51 °C, or for the second stage, −22.06 ± 0.68 °C, however there was a significant difference for the embryonic development phase among the four geographical populations. The egg supercooling point increased gradually from neonatal egg to old egg; eggs prior to hatching always had a much higher supercooling point.
3. Comparisons of the cold hardiness of four populations were carried out by validating the close correlation between latitude and the effects of cold on hatching, low lethal temperature (Ltemp₅₀), and low lethal time (Ltime₅₀). There were significant differences among the four populations; the northern population was more cold hardy than the southern population, and the two mid-latitude populations were intermediately cold hardy.
4. The cold hardiness of all populations was enhanced to various degrees by short-term cold acclimation at 0 °C and 5 °C. For most populations, a 2-day acclimation period seemed to be optimal.     

Development and ultrastructure of the thickened serosa and serosal cuticle formed beneath the embryo in the stonefly Scopura montana Maruyama, 1987 (Insecta,Plecoptera, Scopuridae)

We aimed to describe the development and ultrastructure of the thickened serosa and serosal cuticle formed beneath the embryo of Plecoptera, using Scopura montana of Scopuridae as a euholognathan representative. Using transmission electron microscopy, we found that the egg membranes were composed of a thick exochorion, a thicker endochorion consisting of two sublayers, and an extremely thin vitelline membrane. The egg membrane construction represents a groundplan feature of the euholognathan egg membranes. The serosa converges beneath the embryo to form a thickened serosa, comprising cells in a radial arrangement, in association with the formation of the amnioserosal fold. The thickened serosa then deposits the thickened serosal cuticle, consisting of four layers differing in fine structure and electron density. After achieving its secretory function, the thickened serosa then disintegrates, and the liberated serosal cells float for a short period in the peripheral region of the egg inside. Collectively, our findings should provide the basis for further characterization of the serosal structures concerned, but we were unable to corroborate previous studies assigning the thickened serosa and serosal cuticle in Plecoptera to the water absorption function.     

Age‐dependent changes in tolerance to cold and accumulation of cryoprotectants in overwintering and non‐overwintering larvae of European corn borer Ostrinia nubilalis

Abstract The age‐dependent cold hardiness profile of Ostrinia nubilalis is compared between nondiapausing and diapausing larvae, as well as with field‐collected larvae. The results suggest that both cold tolerance and accumulation of cryoprotectants depends upon the age of O. nubilalis larva. Late fifth‐instar nondiapausing larvae are more cold tolerant than younger fifth‐instars because they show enhanced ability to withstand sub‐zero temperatures. No appreciable difference is observed between the experimental groups of diapausing larvae as far as their supercooling ability and tolerance at sub‐zero temperatures above the supercooling point. In general, both field‐collected and diapausing larvae are more cold tolerant than nondiapausing larvae, indicating a direct link between diapause and cold hardiness. The age of diapausing larvae affects the ability to accumulate glycerol. Glycerol levels of 45‐day‐old diapausing larvae are significantly higher (2.7‐fold) compared with 90‐day‐old diapausing larvae. Moreover, diapausing larvae display a five‐ to 13‐fold higher glycerol content compared with nondiapausing larvae. There is a trend for an age‐dependent cold hardiness profile in O. nubilalis and further tests that could demonstrate a causal relationship between age and cold tolerance are needed.