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Abstract: Overwintering adults of the spruce bark beetle, Ips typographus (L.) showed an unusually complex sugar/polyol cryoprotectant system. The major components of the multiple system were: glucose (177.6mmolL−1, March); trehalose (175.0mmolL−1, December); sorbitol (147.9mmolL−1, January); mannitol (81.2mmolL−1, March); and erythritol (40.7mmolL−1, March) (in the parentheses, the maximum concentrations are shown and the month when they were reached). Other minor components were glycerol, fructose, threitol, myo-inositol, arabinitol and ribitol. Distinct seasonal patterns of accumulation/depletion in various components were found. Glycerol, trehalose and glucose started to accumulate first, during early autumn, when the air temperatures fluctuated between 20 and 0°C, and diapause beetles continued in feeding. Glycerol was depleted, glucose remained stable and trehalose continued in accumulation during late autumn when the temperatures oscillated around 0°C. During early winter severe frosts reaching −20°C came, the beetles terminated their diapause and trehalose was partially depleted, while mannitol, sorbitol, fructose, threitol and erythritol started to accumulate. Cold weather continued also during late winter when the beetles remained quiescent. During this period, trehalose was re-accumulated, threitol and erythritol continued to increase, mannitol remained stable and sorbitol, fructose decreased. All cryoprotectans were finally cleared in the beetles which were spontaneously leaving bark during early spring. The seasonal maximum of total concentration of all cryoprotectants (578.2mOsmolL−1) was reached in March. Such a concentration results in colligative depression of melting point of body fluids down by 1.08°C only. It suggests that the potential cryoprotective effect of accumulated sugars and polyols was related rather to their non-colligative functions. [Copyright &y& Elsevier]

Adults of the insect Pyrrhocoris apterus acquire chill tolerance through the process of autumnal acclimatization. Field and laboratory experiments were conducted to separate the triggering effects of low temperatures, desiccation and diapause progression on the physiological characteristics related to chill tolerance with emphasis on the restructuring of glycerophospholipid (GPL) composition. Changes in relative proportions of major molecular species of glycerophosphoethanolamines (GPEtns) and glycerophosphocholines (GPChols) in thoracic muscle and fat body tissues were followed using HPLC coupled to electrospray ionisation mass spectrometry. The increase in relative proportion of 1-palmitoyl-2-linoleyl-sn-GPEtn at the expense of 1,2-dilinoleyl-sn-GPChoI was the most prominent feature of the complex change observed in both tissues during autumnal acclimatization in the field. The relative proportion of total GPEtns increased, while the proportion of total GPChoIs decreased. The relative proportion of unsaturated fatty acyls slightly decreased. A similar restructuring response was seen during acclimatization in the field and cold acclimation in the laboratory. By contrast, the GPL changes related to desiccation and diapause progression were relatively small, differed qualitatively from the cold-acclimation response, and were accompanied with no increase of chill tolerance. Other features of autumnal acclimatization, i.e. depression of supercooling capacity and accumulation of polyhydric alcohols, were also triggered solely by low temperatures. [ABSTRACT FROM AUTHOR]

The capacity to accumulate winter polyols (mainly ribitol and sorbitol) during cold-acclimation in Pyrrhocoris apterus is restricted only to the adults that have previously entered diapause. The enzymatic complement involved in polyol biosynthesis was found to differ in a complex manner between diapause and non-diapause adults. Nearly 100% of glycogen phosphorylase (GPase) was present in its active form in non-diapause adults irrespective of their acclimation status. In contrast, less than 40% of GPase was present in its active form in diapause adults prior to cold-acclimation and the inactive form was rapidly activated upon transition from 5 to 0 °C, concomitantly with the start of rapid polyol accumulation. The flow of carbon released by activation of glycogen degradation might be routed to the pentose cycle because the activity of glucose-6-P dehydrogenase (G6P-DH) was significantly higher and it increased with cold-acclimation in diapause adults while it was relatively low and it decreased with cold-acclimation in non-diapause adults. Reducing equivalents in the form of NADPH, which were generated in the pentose cycle, might require re-oxidation. Such re-oxidation might be achieved during reduction of sugars to polyols. The activity of NADP(H)-dependent aldose reductase (AR) was about 20-fold higher in diapause than in non-diapause adults. Similarly, the activity of NAD(H)-dependent polyol dehydrogenase (PDH) was higher in diapause adults. In addition, we found a very high activity of an unusual enzyme, NADP(H)-dependent ketose reductase (KR), exclusively in diapause adults. KR might be involved in reduction of fructose to sorbitol. Although its affinity for fructose as a substrate was low (KM=0.64 M), its activity was about 10-fold higher than that of PDH with fructose. Moreover, the activity of KR significantly increased with cold-acclimation while that of PDH remained unchanged. Different electrophoretic mobilities in PAGE gel suggested that KR and PDH are two different enzymes with specific requirement for NADP(H) or NAD(H), respectively, as co-factors. [Copyright &y& Elsevier]

Three acclimation groups [i.e. non-diapause (LD), diapause (SD) and diapause, cold-acclimated (SDA)] of the adult bugs Pyrrhocoris apterus differed markedly in their levels of chill tolerance. Survival time at a sub-zero, but non-freezing, temperature of-5°C (Lt50) extended from 7.6 days, through 35.6 days, to >60 days in the LD, SD and SDA insects, respectively. The time necessary for recovery after chill-coma increased linearly with the increasing time of exposure to -5°C, and the steepness of the slope of linear regression decreased in the order LD>SD>SDA. The capacity to prevent/counteract leakage of Na+ down the electrochemical gradient (from haemolymph to tissues) during the exposure to -5°C increased in the order LD