Desiccation resistance in pre-diapause, diapause and post-diapause larvae of Choristoneura fumiferana (Lepidoptera: Tortricidae)

Desiccation resistance was examined in pre-diapause, diapause and post-diapause larvae of the spruce budworm, Choristoneura fumiferana (Clemens), in terms of passive water evaporation under three desiccation conditions: freeze-drying, desiccant-drying at 2 degrees C and desiccant-drying at 18 degrees C. Diapausing second instar larvae and post-diapause non-feeding second instar larvae showed strongest desiccation resistance: a significant amount of water was retained after repeated drying under desiccating conditions, while pre-diapause first instar larvae and post-diapause feeding second instar larvae lost almost all their water content after one or two drying cycles. A hibernaculum covering had no effect on water evaporation. While dead larvae tended to lose significantly more water than their living counterparts, particularly among first instar larvae, such an impact was much weaker among diapausing second instar larvae. Desiccation resistance was lost when post-diapause second instar larvae were allowed access to water while the level of desiccation resistance was maintained or enhanced when the larvae did not have access to water. These results are discussed in the context of overwintering ecology of the species and possible mechanisms for the desiccation resistance are also discussed.     

昆虫滞育后的生物学特性

从昆虫滞育后性比、寿命、生殖力和重复滞育等方面对昆虫滞育后的生物学特性进行了概括,并分析了影响昆虫滞育后生物学特性数量表达的因子。这些因子包括:(1)滞育前昆虫的发育速率、取食行为和个体大小;(2)滞育期间的环境条件及昆虫取食行为;(3)滞育持续期;(4)滞育后的取食需求;以及(5)滞育后的温度和光周期。     

Effect of photoperiod and allatectomy on reproduction and (second) diapause induction in post-diapause adult females of the Colorado potato beetle,Leptinotarsa decemlineata Say

Summary

The effect of photoperiod and allatectomy on the rates of oviposition and juvenile hormone (JH) synthesis was studied in female beetles after the break of adult diapause. The results indicate that after diapause, photoperiod has no effect on oviposition rates. However, the photoperiodic effect on diapause induction is restored within a few days after the break of the first diapause. Allatectomy performed within one day after diapause did not induce a second diapause, whereas about 40% of the beetles allatectomized after 3 or more days responded by entering a second diapause. Oogenesis in the operated animals was never completely prevented under long and short day conditions. These preliminary results suggest a change in the sensitivity of post-diapause beetles to photoperiodic and endocrine manipulation as compared with pre-diapause insects.     

麦红吸浆虫滞育发生和解除过程中总脂和甘油三酯含量变化

【目的】脂类与昆虫滞育密切相关。本研究旨在明确麦红吸浆虫Sitodiplosis mosellana (Géhin)滞育过程中脂类物质含量的变化规律,探讨麦红吸浆虫滞育与脂类物质变化的关系。【方法】采用香兰素硫酸显色法测定了2008年5月－2010年1月不同时间陕西杨凌养虫圃内麦红吸浆虫滞育前、滞育期及滞育解除后幼虫和蛹体内总脂和甘油三酯含量。【结果】 滞育前幼虫总脂和甘油三酯含量分别为378.12和291.67 μg/mg,显著高于整个滞育期（P<0.05）。滞育年周期中,冬季总脂和甘油三酯含量最高,翌年1月结茧幼虫二者含量分别为335.29 和275.72 μg/mg,显著高于其他季节（P<0.05）;整个滞育期间,裸露幼虫和结茧幼虫总脂和甘油三酯随季节变化趋势相同,但同期结茧幼虫含量高于裸露幼虫;滞育当年与第2年同期幼虫总脂和甘油三酯含量差异不显著（P>0.05）。滞育解除后,总脂和甘油三酯含量随着幼虫的发育和变态逐渐降低,其中中蛹和后蛹显著低于活动幼虫（P<0.05）。【结论】麦红吸浆虫滞育不同时期幼虫及蛹总脂和甘油三酯含量存在明显差异,其滞育与脂类物质的含量密切相关。     

麦红吸浆虫3-羟基-3-甲基戊二酰辅酶A还原酶基因SmHMGR的克隆及在滞育与发育变态过程中的表达动态

【目的】3-羟基-3-甲基戊二酰辅酶A还原酶(HMGR)是保幼激素(JH)合成途径的限速酶。麦红吸浆虫Sitodiplosis mosellana是一种典型的专性幼虫滞育昆虫。本研究旨在探讨HMGR基因在麦红吸浆虫滞育和发育变态过程中的作用。【方法】通过RT-PCR和RACE技术克隆麦红吸浆虫滞育前幼虫HMGR基因全长cDNA序列;利用生物信息学软件分析HMGR基因核苷酸和其编码的蛋白氨基酸序列特性;采用qPCR技术测定其在麦红吸浆虫滞育不同时期3龄幼虫及不同发育阶段(1-2龄幼虫、预蛹、初蛹、中蛹和后蛹以及雌雄成虫)中的mRNA表达水平。【结果】克隆获得一条麦红吸浆虫HMGR基因全长cDNA序列,命名为SmHMGR(GenBank登录号: MG876766)。该基因全长2 548 bp,其中开放阅读框长2 328 bp,编码775个氨基酸,预测的蛋白分子量为84.16 kD,理论等电点为8.29。序列分析发现该基因编码的蛋白具有HMGR蛋白家族典型的HMG-CoA-reductase-classⅠ催化功能域及其他保守功能基序;序列比对和系统发育分析表明,SmHMGR与达氏按蚊Anopheles darling等长角亚目(Nematocera)昆虫HMGR的相似性最高、亲缘关系最近。SmHMGR在麦红吸浆虫滞育前的3龄早期幼虫中表达量显著升高,进入滞育后一直维持较高水平,并在滞育后静息阶段的当年12月至翌年1月达到最高。SmHMGR在蛹期表达量低于幼虫期,预蛹期表达量最低;在雌成虫中表达量显著高于在蛹和雄成虫中的表达量。【结论】SmHMGR的表达与麦红吸浆虫发育密切相关,可能在滞育诱导、维持及滞育后静息状态的维持及生殖中发挥作用,其表达量的降低可能参与了幼虫到蛹的变态。     

小麦吸浆虫保幼激素酯酶和保幼激素环氧水解酶基因的克隆及在滞育和变态发育过程中的表达动态

【目的】保幼激素(juvenile hormone, JH)在小麦吸浆虫Sitodiplosis mosellana滞育诱导及滞育后静息状态的维持中发挥着重要作用。保幼激素酯酶(hormone esterase, JHE)和保幼激素环氧水解酶(juvenile hormone epoxide hydrolase, JHEH)是调控JH滴度的重要降解酶。本研究旨在探讨JHE和JHEH在小麦吸浆虫滞育和变态发育中潜在功能。【方法】通过RT-PCR和RACE技术从小麦吸浆虫滞育前幼虫克隆JHE和JHEH全长cDNA序列;利用生物信息学软件分析其核苷酸及编码蛋白特性;采用qPCR技术分析其在小麦吸浆虫滞育不同时期(滞育前、滞育期、滞育后静息期和滞育后发育)3龄幼虫及1龄幼虫到成虫不同发育阶段(1-2龄幼虫、预蛹、初蛹、中蛹、后蛹、雌成虫和雄成虫)中的表达水平。【结果】克隆获得了cDNA全长分别为3 102和1 980 bp的小麦吸浆虫SmJHE和SmJHEH基因(GenBank登录号分别为MG876768和MG876769),其开放阅读框分别长1 740和1 371 bp,分别编码579和456个氨基酸,预测蛋白分子量分别为65.67和51.65 kD。SmJHE蛋白含有5个JHE家族特有的保守模块,SmJHEH含有催化三联体Asp228, Asp404和His431及组成阴氧离子洞的两个Tyr(Tyr299和Tyr374)和HGWP花样结构。序列比对和进化分析表明,SmJHE和SmJHEH均与双翅目(Diptera)长角亚目(Nematocera)昆虫同源蛋白氨基酸序列一致性较高,亲缘关系最近。不同滞育时期的表达模式表明,SmJHE和SmJHEH在滞育前期(1龄到滞育前的3龄幼虫早期)表达量变化不明显,进入滞育后表达量基本维持恒定,但均在滞育后静息阶段的当年12月至翌年1月最低。发育表达模式表明,幼虫恢复发育后SmJHE表达量逐渐升高,预蛹期达到最高,在雌成虫中的表达量显著低于雄成虫中的;SmJHEH表达量则在预蛹期最低,在雌成虫中最高。【结论】SmJHE和SmJHEH参与小麦吸浆虫滞育调控,其表达量的降低与滞育后静息阶段JH的累积有关;SmJHE在发育过程中表达量的升高可能参与幼虫到蛹的变态,表达量的降低可能与生殖发育有关。     

Effect of temperature on embryonic development and diapause in Chesias legatella (Lepidoptera: Geometridae)

The changing temperature requirements of Chesias legatella Schiffermüller during embryonic development are investigated. The eggs are laid in the autumn and embryogenesis is interrupted by a short diapause at the germ band stage, after which development proceeds slowly throughout the winter. The effects of temperature during the pre-diapause, diapause and post-diapause phases of development are assessed by examination of the rate and variability of morphogenesis through all the embryonic stages. An examination is presented of the effect of temperature during the late autumn on the rate and uniformity of diapause termination, and consequently on the length of the hatching period the following spring.     

昆虫滞育研究进展

首先对昆虫滞育的若干名词概念作出解释,按照滞育过程的阶段特点划分为3个时期:(1)滞育前期(包括滞育诱导和滞育准备);(2)滞育期(包括滞育进入、滞育维持和滞育解除);(3)滞育后期。然后重点介绍这3个阶段近期分子和细胞水平上的重要工作,对一些科学问题进行讨论。     

Respiration in bumblebee queens: effect of life phase on the discontinuous ventilation cycle

Evidence for discontinuous ventilation cycles (DVC) has been obtained for many insects under various experimental conditions. Here, we show that DVC's exist for queens of the bumblebee Bombus terrestris in different phases of their life cycle: before entering diapause (pre-diapause queens) and when a colony has been established (post-diapause queens). Pre- and post-diapause queens experience distinct environmental conditions (ambient O₂ and CO₂ concentrations) which is reflected in their DVC: both ventilation frequency and amount of carbon dioxide emitted change with differences in ambient CO₂ concentration.     

Two calcium‐binding chaperones from the fat body of the Colorado potato beetle,Leptinotarsa decemlineata (Coleoptera: Chrysomelidae) involved in diapause

Molecular chaperones are crucial for the correct folding of newly synthesized polypeptides, in particular, under stress conditions. Various studies have revealed the involvement of molecular chaperones, such as heat shock proteins, in diapause maintenance and starvation; however, the role of other chaperones in diapause and starvation relatively is unknown. In the current study, we identified two lectin‐type chaperones with calcium affinity, a calreticulin (LdCrT) and a calnexin (LdCnX), that were present in the fat body of the Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae) during diapause. Both proteins possessed an N‐globular domain, a P‐arm domain, and a highly charged C‐terminal domain, while an additional transmembrane domain was present in LdCnX. Phylogenetic analysis revealed distinction at the order level. Both genes were expressed in multiple tissues in larval and adult stages, and constitutively throughout development, though a starvation response was detected only for LdCrT. In females, diapause‐related expression analysis in the whole body revealed an upregulation of both genes by post‐diapause, but a downregulation by diapause only for LdCrT. By contrast, males revealed no alteration in their diapause‐related expression pattern in the entire body for both genes. Fat body‐specific expression analysis of both genes in relation to diapause revealed the same expression pattern with no alteration in females and downregulation in males by post‐diapause. This study suggests that calcium‐binding chaperones play similar and possibly gender‐specific roles during diapause.     

A novel cytochrome P450 gene (CYP4G25) of the silkmoth Antheraea yamamai: cloning and expression pattern in pharate first instar larvae in relation to diapause

A new cytochrome P450 gene, CYP4G25, was identified as a differentially expressed gene between the diapausing and post-diapausing pharate first instar larvae of the wild silkmoth Antheraea yamamai, using subtractive cDNA hybridization. The cDNA sequence of CYP4G25 has an open reading frame of 1674 nucleotides encoding 557 amino acid residues. Sequence analysis of the putative CYP4G25 protein disclosed the motif FXXGXRXCXG that is essential for heme binding in P450 cytochromes. Hybridization in situ demonstrated predominant expression of CYP4G25 in the integument of pharate first instar larvae. Northern blotting analysis showed an intensive signal after the initiation of diapause and no or weak expression throughout the periods of pre-diapause and post-diapause, including larval development. These results indicate that CYP4G25 is strongly associated with diapause in pharate first instar larvae.     

Effect of 20-hydroxyecdysone agonist, tebufenozide, on pre-and post-diapause larvae of Dendrolimus pini (L.)(Lep., Lasiocampidae)

Studies of the effects of a novel insect growth regulator (tebufenozide, RH 5992) in pest lepidopterans are extended to its influence on pre- and post-diapause larvae of the pine-tree lappet, Dendrolimus pini (L.) (Lep., Lasiocampidae). Larval diapause was induced under experimental conditions. The relation between the width of head capsule and each larval instar of the experimental populations is presented. The toxicity of tebufenozide applied topically and per os (orally), and its effects on feeding activity were examined. High doses and concentrations (≥100 ng/larva and ≥0.001%) of tebufenozide were toxic and reduced the feeding intensity in larvae of D. pini , however, post-diapause larvae were less sensitive to oral treatment with tebufenozide then pre-diapause insects. In pre-diapausing insects of the fourth instar a greater than four-fold fall in mortality was observed among the insects treated with low doses/ concentrations (0.1–10 ng/larva and 0.00001%–0.001%, respectively). A remarkable stimulatory effect on feeding behaviour was observed in pre-diapausing IV instar larvae when treated with tebufenozide topically, in a dose of 10 ng/larva. A possible ecdysteroid mimicking effect of tebufenozide onto feeding and mortality among diapause-destined D. pini is discussed.