Temperature dependence-independence of antifreeze turnover in Eurosta solidaginis (Fitch)

Temperature effects on antifreeze metabolism were investigated in two populations (northern and southern) of the golden rod gallfly, Eurosta solidaginis. Sorbitol production was temperature dependent and was triggered by short-term exposure to < +10°C. The maximal rate sorbitol synthesis occurred at 0°C. For both populations, sorbitol was rapidly catabolized during warm acclimation at +20°C. During the first 12 h of warm acclimation, sorbitol levels decreased by 36% (19.7 ± 0.6) to 12.6 ± 1.2 μg/mg) and by 83% (to 3.3 ± 1.7 μg/mg) after 48 h in the northern population. The southern population decreased sorbitol levels 64% (11.8 ± 0.69 to 4.2 ± 0.62) after 48 h. The southern population resynthesized more sorbitol than did the northern population upon re-exposure to 0°C. Glycerol levels increased linearly during the experimental period independent of temperature.     

Divergent mechanisms of frost-hardiness in two populations of the gall fly, Eurosta solidaginsis

Two populations of the gall fly Eurosta solidaginsis utilize different strategies to endure seasonal exposure to temperatures below freezing. Both populations are freezing tolerant. In north temperate populations, supercooling points rise from ?10.2°C to ?6.2°C following exposures to temperatures below freezing. This level is maintained throughout winter and ensures frequent and prolonged periods of tissue freezing. South temperate populations depress the supercooling point to ?14.2°C during autumn and early winter, and this depression precludes extracellular ice formation during periods of supra-optimal temperature fluctuations. During mid-winter, supercooling points rise to the same level as in northern groups.Both populations accumulate three principal cryoprotective agents following first frost exposures (glycerol, sorbitol and trehalose). Cryoprotectants levels do not peak in northern populations until 4–6 weeks after first frost. In southern populations the accumulation profile is characterized by a high initial rate of synthesis, a protective overshoot and pronounced seasonal fluctuations. The relative survival advantages of each strategy are discussed.     

中华通草蛉成虫抗寒能力季节性变化

中华通草蛉（Chrysoperla sinica）成虫在自然条件下抗寒能力的季节性变化研究表明,雌、雄成虫的抗寒能力均呈现出季节性的变化趋势,即随着冬季低温的到来,其抗寒能力逐渐增强,冬季过后又随气温的回升,其抗寒力逐渐减弱.雌、雄成虫的体内含水量、过冷却点（SCP）和结冰点（FP）均随气温的降低而降低,升高而升高,但体内总脂肪的含量却随气温的降低而升高,升高而降低.越冬代在越冬的前期和中期,雌、雄成虫体内的含水量和SCP均显著低于生长季节的其它各代,越冬代后期的含水量和SCP与生长季节其它各代没有显著差异.经回归分析发现,雌、雄成虫体内含水量与SCP之间均存在极显著负相关关系（p＜0.01）;越冬代成虫（特别是越冬前期）的体内总脂肪含量显著高于其它时期,并且雌成虫体内的总脂肪含量与SCP之间呈显著正相关（p＜0.05）,雄成虫体内总脂肪含量与SCP之间没有显著相关性（p＞0.05）.在一年的不同世代中,雌、雄成虫体内含水量只是在越冬前期有显著差异,SCP和体内总脂肪含量却在越冬后期有显著差异.     

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

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

Simulated winter to summer transition in diapausing adults of the lady beetle (Hippodamia convergens): supercooling point is not indicative of cold-hardiness

ABSTRACT. Adults of Hippodamia convergens Guerin-Meneville (Coleoptera: Coccinellidae) form massive overwintering aggregations in the Sierra Nevada Mountains of California. These diapausing pre-reproductive adults may remain in the aggregations for up to 10 months. Beetles were collected from overwintering sites in Nevada County, California, and held at 4C for 6 weeks. In order to simulate the winter to summer transition, the beetles were transferred to 20C and 68% r.h. under a LD 12:12h cycle. Supercooling points increased slightly from - 16C on day 0 to - 12.9CC on day 21. In contrast, low temperature tolerance decreased markedly within 2 weeks. On day 0 nearly all beetles survive 2 h of exposure to - 5C, whereas only 50% survived this treatment on day 4. These data demonstrate the general lack of correlation between the supercooling point and the lower lethal temperature in warm-acclimated beetles. Within 1 day of transfer to 20C, oxygen consumption decreased by 26%. This decrease continued through day 14 when the rate of oxygen consumption had decreased to 40% of initial values. Beetles acclimated to 20C selected temperatures significantly higher than cold-acclimated beetles. The parametric shifts observed in this study are consistent with a physiological transition from cold-hardiness and energy conservation during winter to one of dispersal, feeding and reproduction in summer beetles.     

赤松毛虫越冬幼虫生化物质变化与抗寒性的关系

昆虫的抗寒性与其体内生化物质的种类、含量密切相关。测定了赤松毛虫(Dendrolimusspectabilis)越冬幼虫体内和血淋巴内抗寒物质含量的变化,结果表明,血淋巴内小分子碳水化合物总量1月份为10月份的3.8倍,其中以葡萄糖、海藻糖和山梨醇增加幅度较大,依次为10月份的10.08倍、2.84倍、7.44倍。3月份有较大幅度的下降,虫体内糖原含量下降了56.5%。越冬期苏氨酸、丝氨酸、谷氨酸、丙氨酸、胱氨酸增加幅度较大,分别比越冬前增加了493.8%、433.7%、474.2%、21.5%和47.1%,甘氨酸、天门冬氨酸、蛋氨酸、亮氨酸、酪氨酸、苯丙氨酸、组氨酸、精氨酸、脯氨酸含量则有较大幅度的降低。越冬幼虫体内脂肪含量下降,蛋白质含量上升。应用蛋白质垂直平板电泳法对糖蛋白进行电泳分析发现糖蛋白含量增加,为越冬前的300%。综合分析认为赤松毛虫越冬幼虫体内抗寒物质系统为小分子碳水化合物类(山梨醇、海藻糖、葡萄糖)-糖蛋白-氨基酸类(丙氨酸、苏氨酸、谷氨酸等)。该系统可随生态条件的改变而变化。     

Neuropeptides associated with the central nervous system of the cabbage root fly, Delia radicum (L)

The peptidome of the central nervous system of adult cabbage root fly, Delia radicum (L) was investigated using matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS). Over twenty neuropeptides were identified from three different tissue sources, the combined brain/suboesophageal ganglion (SOG), the retrocerebral complex, and the thoracic-abdominal ganglion (TAG). A number of peptides were identified in all three tissues, including allatostatins, short neuropeptide F-like peptides, corazonin, a pyrokinin, and a myosuppressin. Adipokinetic hormone was restricted to the retrocerebral complex. Other peptides, including FMRFamides and sulfakinins were detected only in the brain/SOG and TAG. Some peptides, notably myoinhibitory peptides and tachykinins, which have been identified in other fly species, were not detected in any tissue sample. This study has structurally characterized for the first time, the neuropeptides from adult D. radicum.     

The ontogeny of cold tolerance in the gall fly, Eurosta solidagensis

The lower lethal temperature of many insects indicates an overwintering flexibility as a result of either extensive supercooling or production of cryoprotectants. Ontogenetically, the gall fly (Eurosta solidagensis) utilizes both means of seasonal cryoprotection. All stages except third instar larvae demonstrate supercooling points well below the lowest temperature normally experienced by that particular stage. The third instar larvae exhibit a high supercooling point but are well protected by a cryoprotectant system consisting of glycerol, sorbitol, and trehalose. Glycerol is accumulated, possibly from triglyceride sources, during early autumn and reaches plateau levels (0·6 M) by early winter. Sorbitol synthesis is delayed until freezing exposures and reaches a plateau with glycerol at 0·3 M. It is not until mid-winter that peak trehalose levels are reached (300 mg %). All cryoprotectant levels are a reflection of haemolymph concentrations.Laboratory acclimation experiments further quantify these results. Trehalose synthesis is time and temperature dependent and appears to be affected by developmental processes.     

桔小实蝇不同发育阶段过冷却点的测定

对桔小实蝇Bactrocera dorsalis (Hendel)不同发育阶段的过冷却点进行了测定。结果表明：同一虫期个体间的过冷却点出现不同程度的变异,但均服从正态分布。不同虫期的过冷却点差异显著,其中蛹的过冷却点最低（-12.2℃～-15.0℃）。老熟幼虫的过冷却点为-8.1℃。成虫的过冷却点最低值为7日龄雄虫（-10.5℃）和雌虫（-10.1℃）,最高值为60日龄雄虫（-5.9℃）和雌虫（-6.4℃）,但同一发育时期的雌、雄成虫之间的过冷却点没有差异。测定结果提示蛹期最有可能是该虫在温带地区越冬的虫态。     

Examining the relationship between hemolymph phenoloxidase and resistance to a DNA virus, Plodia interpunctella granulosis virus (PiGV)

We have a detailed understanding of invertebrate immune responses to bacteria and fungal pathogens, but we know less about how insects respond to virus challenge. Phenoloxidase (PO) functions as an important immune response against many parasites and pathogens and is routinely used as a measure of immune competance. We examine the role of haemolymph PO activity in Plodia interpuncetella's response to its natural granulosis virus (PiGV). Larvae were challenged with virus by both oral inoculation of occluded virus (the natural infection route) and direct intrahaemocoelic injection of budded virus. Haemolymph was collected at time points post-viral challenge using a novel method that allows the volume of haemolymph to be quanitified. The haemolmyph was collected without killing the larvae so that haemolymph samples from individuals that developed viral disease could be distinguished from samples collected from those that fought off infection. The level of haemolymph PO activity in resistant larvae did not differ from control larvae. Therefore we have no evidence that PO is involved in resistance to virus in the haemocoel whether larvae are challenged naturally by oral innoculation or directly by intraheamocoelic injection. Phenoloxidase may therefore not be a relevant metric of immunocompetence for viral infection.     

The replication of a cytoplasmic polyhedrosis virus from Chrysodeixis eriosoma (Lepidoptera: Noctuidae) in Spodoptera frugiperda cells

A cytoplasmic polyhedrosis virus (CPV) from Chrysodeixis eriosoma (Lepidoptera: Noctuidae) replicated in Spodoptera frugiperda cells. Low rates of infection were achieved, even at high multiplicities of infection and TCID₅₀ assays showed that there was negligible release of virus particles from infected cells. In an infected focus assay, based on formation of PIB, the dose-response data demonstrated that a single particle could initiate infection. No loss of infectivity occurred in virus preparations stored at 4°, ?20°, or ?90°C, but infectivity of virus stored at 20°C declined sharply. A small isometric virus contaminant was present in some CPV preparations and its interaction with the CPV is discussed. Limited CPV infection was achieved in Trichoplusia ni cells, but attempts to infect Aedes aegypti cells were unsuccessful.     

Attalea phalerata and Bactris glaucescens (Arecaceae, Arecoideae): Phenology and pollination ecology in the Pantanal, Brazil

We compared the reproductive and vegetative phenology and pollination ecology of sympatric Attalea phalerata and Bactris glaucescens palms (Arecaceae) in the Pantanal, Brazil, in a riparian forest area subject to periodic flooding. Attalea phalerata has a solitary stem and produces staminate, pistillate and rarely bisexuals inflorescences that open during the day. Bactris glaucescens has multiple stems and has bisexual flowers with crepuscular/nocturnal anthesis. Both species present bud break and leaf-budding during the year. Attalea phalerata shows continual annual flowering with ripening of fruits during the dry season. For B. glaucescens flowering occurs simultaneously with fruiting for about seven months, and fruit production can be influenced by temperature and level of flooding. The difference in the timing of anthesis of the flowers ensures production of floral resources during both day and night when both species of palms are flowering. The floral structures of both species have morphological characteristics associated with pollination by insects (mainly beetles), such as the presence of odor, stigmatic secretion, heat production, and adhesive substances (“pollenkitt”) in the pollen. In A. phalerata, the main pollinators were Mystrops sp. (Nitidulidae) and Madarini (Curculionidae). Derelomus sp. (Curculionidae) and Paratenetus sp. (Tenebrionidae) beetles visited B. glaucescens during the day and could have pollinated the flowers at these occasions.     

Immunocompetence of the red turpentine beetle, Dendroctonus valens LeConte (Coleoptera: Curculionidae, Scolytinae): Variation between developmental stages and sexes in populations in China

Immune defense imposes fitness costs as well as benefits, so organisms should optimize, not maximize, their immune function through their life cycle. We investigated this issue in the red turpentine beetle, Dendroctonus valens LeConte (Coleoptera: Curculionidae, Scolytinae), which is a pine-killing invasive beetle in China, though it is usually considered as a secondary pest in its native range of North America. We hypothesized that pathogen pressure may affect these beetles differently throughout their life history. We measured the insect's immunocompetence throughout life, determining encapsulation ability and phenoloxidase activity in larval stages, pupae and adults. Pupae had the highest encapsulation ability, but encapsulation was not different between final instar larvae and adults. Phenoloxidase (PO) activity was highest in final instar larvae and pupae, followed by the second instar larvae and adults. Total phenoloxidase activity increased significantly from the second instar larval stage to pupae, and then decreased in adults. Although the second instar larvae had the lowest phenoloxidase activity, more than 90% of total PO existed in the hemolymph in the form of the active enzyme, as compared with pupae, in which over 60% of PO occurred as a proenzyme. Both active PO and total PO were much higher in females than in males, though no significant differences were detected between the encapsulation ability of male and female adults. This result suggests the existence of a sexual dimorphism of immunocompetence in D. valens adults. Variations in immunocompetence across developmental stages suggest that D. valens adopts diverse investment strategies in immunocompetence during different stages. Potential reasons for variation in immunocompetence among developmental stages and between the sexes of D. valens are discussed.     

Regulation of greenhouse sciarid fly populations using Tetradonema plicans (Nematoda: Mermithoidea)

Tetradonema plicans (Tetradonematidae) satisfied many of the requirements for an efficient biological control agent. It was highly pathogenic and specific to the target hosts, sciarid flies, particularly Bradysia paupera (Lycoriidae). It could be mass-reared, stored as eggs for up to one year at 10°C, and applied to the soil surface as eggs in water. It had high reproductive capacity, each female producing up to 12,000 viable eggs. Since T. plicans was a naturally occurring parasite B. paupera both life cycles could be synchronized.     

Enzyme activity profiles in an overwintering population of freeze-tolerant larvae of the gall fly,Eurosta solidaginis

The activity of some enzymes of intermediary metabolism, including enzymes of glycolysis, the hexose monophosphate shunt, and polyol cryoprotectant synthesis, were measured in freeze-tolerant Eurosta solidaginis larvae over a winter season and upon entry into pupation. Flexible metabolic rearrangement was observed concurrently with acclimatization and development. Profiles of enzyme activities related to the metabolism of the cryoprotectant glycerol indicated that fall biosynthesis may occur from two possible pathways: 1. glyceraldehyde-phosphate glyceraldehyde glycerol, using glyceraldehyde phosphatase and NADPH-linked polyol dehydrogenase, or 2. dihydroxyacetonephosphate glycerol-3-phosphate glycerol, using glycerol-3-phosphate dehydrogenase and glycerol-3-phosphatase. Clearance of glycerol in the spring appeared to occur by a novel route through the action of polyol dehydrogenase and glyceraldehyde kinase. Profiles of enzyme activities associated with sorbitol metabolism suggested that this polyol cryoprotectant was synthesized from glucose-6-phosphate through the action of glucose-6-phosphatase and NADPH-linked polyol dehydrogenase. Removal of sorbitol in the spring appeared to occur through the action of sorbitol dehydrogenase and hexokinase. Glycogen phosphorylase activation ensured the required flow of carbon into the synthesis of both glycerol and sorbitol. Little change was seen in the activity of glycolytic or hexose monophosphate shunt enzymes over the winter. Increased activity of the -glycerophosphate shuttle in the spring, indicated by greatly increased glycerol-3-phosphate dehydrogenase activity, may be key to removal and oxidation of reducing equivalents generated from polyol cryoprotectan catabolism.Abbreviations 6PGDH 6-Phosphogluconate dehydrogenase - DHAP dihydroxy acetone phosphate - F6P fructose-6-phosphate - F6Pase fructose-6-phospha-tase - FBPase fructose-bisphosphatase - G3P glycerol-3-phosphate - G3Pase glycerol-3-phosphate phophatase - G3PDH glycerol-3-phosphate dehydrogenase - G6P glucose-6-phosphate - G6Pase glucose-6-phosphatase - G6PDH glucose-6-phosphate dehydrogenase - GAK glyceraldehyde kinase - GAP glyceraldehyde-3-phosphate - GAPase glyceraldehyde-3-phosphatase - GAPDH glyceraldehyde-3-phosphate dehydrogenase - GDH glycerol dehydrogenase - GPase glycogen phosphorylase - HMS hexose monophosphate shunt - LDH lactate dehydrogenase - NADP-IDH NADP⁺-dependent isocitrate dehydrogenase - PDHald polyol dehydrogenase, glyceraldehyde activity - PDHgluc polyol dehydrogenase, glucose activity - PFK phosphofructokinase - PGI phosphoglucoisomerase - PGK phosphoglycerate kinase - PGM phosphoglucomutase - PK pyruvate kinase - PMSF phenylmethylsulfonylfluoride - SoDH sorbitol dehydrogenase - V _max maximal enzyme activity - ww wet weight