麦红吸浆虫滞育发生和解除过程中保护酶活力动态

采用保护酶活性测试盒分别测定了麦红吸浆虫滞育前、滞育期及滞育解除后过氧化物酶（POD）、超氧化物歧化酶（SOD）和过氧化氢酶（CAT）等3种保护酶的活力.结果表明：幼虫从老熟到进入滞育的初期,3种保护酶的活力均呈下降趋势.滞育年周期中,SOD和CAT活力对环境温度的反应相同,即低温促进其活力升高,高温导致其活力下降;POD活力与环境温度和滞育发育有关;整个滞育期间,裸露幼虫和结茧幼虫3种保护酶的活力随季节变化趋势相同,但同期的裸露幼虫活力略高于结茧幼虫;不同滞育年限幼虫3种保护酶的活力差异不显著.滞育解除后,3种保护酶的活力均随生长发育进程逐渐升高.     

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。     

麦红吸浆虫幼虫不同滞育时期体内山梨醇含量及山梨醇脱氢酶基因表达水平变化

【目的】本研究旨在明确麦红吸浆虫Sitodiplosis mosellana(Géhin)滞育进程中山梨醇含量及山梨醇脱氢酶基因(SmSDH)mRNA表达水平的变化规律,探讨麦红吸浆虫滞育与山梨醇含量及SmSDH表达水平的关系。【方法】分别采用比色法和实时荧光定量PCR方法,对2013年5月-2014年3月不同时间陕西杨凌养虫圃内麦红吸浆虫滞育前后及滞育期幼虫体内山梨醇含量及SmSDH2a和SmSDH2b mRNA表达水平进行分析。【结果】麦红吸浆虫幼虫滞育前山梨醇含量最高(为9.41μg/mg);滞育期间,12月份含量最高(为8.50μg/mg);滞育解除后山梨醇含量为3.25~3.49μg/mg,显著低于滞育前和整个滞育期(P0.05)。山梨醇脱氢酶基因SmSDH2a及SmSDH2b在滞育进程中mRNA表达水平变化趋势相同,且与滞育强度递减趋势一致,即进入滞育后表达水平显著降低(P0.05)并一直稳定在较低水平,滞育解除后逐渐升高并达到最高。滞育期间,同期不同滞育状态幼虫之间山梨醇含量和SmSDH2a表达水平差异不明显,但裸露幼虫SmSDH2b表达水平始终高于结茧幼虫。【结论】山梨醇和SmSDH参与麦红吸浆虫滞育调控,与滞育维持和解除密切相关。     

麦红吸浆虫幼虫滞育期间糖类物质变化

分析研究了麦红吸浆虫Sitodiplosis mosellana (Gehin) 幼虫滞育期间总糖、海藻糖和糖原含量的变化。结果表明： 不同滞育年限幼虫的总糖含量无显著差异。在滞育年周期中,不同时期的滞育幼虫总糖含量明显不同。幼虫老熟入土滞育以前的总糖含量明显高于入土滞育期间。入土滞育过程中,越冬期和春季的总糖含量明显高于夏季和秋季。同期的裸露幼虫与结茧幼虫的总糖含量仅在秋季有一定的差异,其他时间基本相似。结茧幼虫滞育期间的海藻糖含量一直维持较高的水平,说明其滞育期间的糖醇积累型属于海藻糖积累型;秋季与春季糖原含量的增加可能与麦红吸浆虫的滞育解除或滞育强度的变化有一定的关系。     

低温对麦红吸浆虫滞育解除及蜕皮激素受体基因EcR和热激蛋白基因Hsp70和Hsp90表达水平的影响

【目的】本研究旨在明确破茧率和破茧所需时间作为典型的专性幼虫期滞育昆虫麦红吸浆虫Sitodiplosis mosellana滞育解除指标的可行性,探讨蜕皮激素受体基因EcR和热激蛋白基因Hsp70和Hsp90在低温解除滞育中的作用。【方法】9月上旬采自田间的麦红吸浆虫滞育幼虫在低温(4℃)和自然变温处理不同时间(0~90 d)后转移至24℃化蛹,调查幼虫的破茧率、化蛹率及破茧和化蛹所需时间;采用qPCR技术分析4℃低温终止滞育并在24℃下恢复发育的幼虫EcR, Hsp70和Hsp90的mRNA水平。【结果】不同时间低温(4℃)和自然变温处理的麦红吸浆虫滞育幼虫破茧率和化蛹率以及破茧和化蛹所需时间均存在显著差异。未经低温(4℃)处理的幼虫在水分满足的条件下破茧率为71.3%,但均不能化蛹;低温(4℃)处理60 d内,随着处理时间的延长破茧率和化蛹率逐渐升高,破茧和化蛹所需时间逐渐缩短;与4℃低温处理30~60 d相比,自然变温处理30~60 d的幼虫化蛹率显著较低;4℃低温处理60 d和自然变温处理90 d后幼虫的化蛹率均超过91%。低温(4℃)处理显著提高了麦红吸浆虫幼虫EcR, Hsp70和Hsp90的表达量,其中处理30 d时其表达量最高,随着低温(4℃)处理时间的延长,表达量逐渐降低,大部分幼虫滞育解除后表达量趋于恒定。【结论】低温能显著促进麦红吸浆虫的滞育解除,效果优于9-10月自然变温;破茧率及破茧所需时间可作为评判麦红吸浆虫幼虫滞育强度的参考,但不能独立作为滞育解除的指标;EcR,Hsp70和Hsp90的表达水平与滞育强度密切相关,在麦红吸浆虫滞育解除中发挥潜在作用。     

麦红吸浆虫热激蛋白基因SmHsp40的克隆及其在滞育进程中和极端温度胁迫下的表达

【目的】麦红吸浆虫Sitodiplosis mosellana是一种典型的专性滞育昆虫,滞育过程中经历酷暑和严寒。本研究旨在探讨麦红吸浆虫热激蛋白基因Hsp40在滞育中的作用。【方法】以麦红吸浆虫滞育前幼虫为材料,利用RT-PCR和RACE技术克隆Hsp40全长cDNA序列;利用生物信息学软件分析该基因及其编码蛋白的特性;采用实时荧光定量PCR技术分析其在滞育不同阶段幼虫及越夏滞育幼虫在短期(≤120 min)极端高温(35～50℃)和越冬滞育幼虫在短期极端低温(0～-15℃)胁迫下的表达模式。【结果】克隆获得麦红吸浆虫Hsp40基因,命名为SmHsp40(GenBank登录号:MH001167),其cDNA全长1 553 bp,开放阅读框长为1 074 bp,编码357个氨基酸,相对分子量为40.87 kD,理论等电点为5.31。推导的氨基酸序列中含有Hsp40蛋白家族典型的J结构域和HPD基序。蛋白序列相似性和系统进化关系分析表明,SmHsp40蛋白与冈比亚按蚊Anopheles gambiae等长角亚目(Nematocera)昆虫Hsp40的相似性最高、亲缘关系最近。麦红吸浆虫滞育不同时期幼虫SmHsp40表达量存在显著差异,进入滞育后表达量上调;滞育进程中,夏季7-8月和冬季12月至翌年1月表达量显著高于其他季节。与未处理的对照相比,40℃处理越夏滞育幼虫和-10℃处理越冬滞育幼虫1 h明显诱导SmHsp40的表达,但超过45℃或低于-15℃极端温度处理诱导效果均不明显。【结论】SmHsp40参与了麦红吸浆虫滞育进程,可能在滞育状态的维持及滞育期间耐热和耐寒能力的调控方面起重要作用。     

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。     

麦红吸浆虫滞育习性研究

1986～1991年研究了麦红吸浆虫的滞育习性,提出了该由主动延长滞育和被动延长滞育的两种形式,探讨了土壤湿度、温度对延长滞育的影响,从体重、羽化率、卵胚数、死亡率等方面测定了延长滞育幼虫的生殖势能,文章最后讨论了麦红吸浆虫多峰羽化的特点和延长滞育的生物学意义。     

用重复序列引物PCR分析不同滞育状态麦红吸浆虫DNA多态性

应用 5 个重复序列引物,通过 PCR 扩增试验,研究了不同滞育状态麦红吸浆虫 DNA 的多态性。结果表明:1) 5 个重复序列引物共扩增了 104 条核酸带,分子量介于 202~1 163bp 之间;2) 不同滞育状态的麦红吸浆虫的平均相似性指数范围在 0.573~0.936 之间,在种群 3 和种群 4 之间,最大的平均相似性指数是 0.936,在种群 1、3、4、5 和种群 6、7、8 之间平均相似性指数较高(均超过 0.81);3)不同滞育状态麦红吸浆虫群体的遗传变异大部分来自群体间,产生于群体间的变异为62.41%,来自群体内的变异为37.59%;4) 根据对不同滞育状态间遗传距离的聚类分析,麦红吸浆虫的滞育类型可分为三大类。首次报道了不同滞育状态麦红吸浆虫量化的DNA多态性。     

麦红吸浆虫蜕皮激素受体（EcR）基因的克隆与表达分析

为了研究蜕皮激素受体（EcR）在麦红吸浆虫Sitodiplosis mosellana (Géhin)滞育活动中的作用, 利用RT PCR和RACE技术克隆得到了麦红吸浆虫蜕皮激素受体基因cDNA全长, 并通过Real-time quantitative PCR研究了其表达情况。该cDNA全长序列被命名为SmEcR （GenBank登录号： KC491135）, 其开放阅读框长1 386 bp, 编码461个氨基酸残基。其蛋白预测分子量52.90 kD, 理论等电点6.24。该蛋白与其他已报道的昆虫EcR蛋白具有很高的同源性, 其中与迟眼蕈蚊Bradysia coprophila中相应蛋白的氨基酸序列一致性高达92%。SmEcR在麦红吸浆虫不同滞育时期和不同虫态中均有表达, 且在不同滞育时期、 不同虫态中的表达量差异很大。在滞育不同时期以11月表达量最高, 12月表达量最低; 在不同虫态以麦穗幼虫中的表达量较低, 而成虫中的表达水平很高。本研究为进一步明确SmEcR在麦红吸浆虫滞育调控中的作用奠定了基础。     

越冬过程中桃小食心虫结茧和裸露幼虫体内耐寒性物质动态变化

为比较桃小食心虫Carposina niponensis Walsingham越冬过程中种群耐寒性的差异, 本文研究了其结茧和裸露幼虫自然种群的耐寒性,分别测定了不同时期桃小食心虫越冬幼虫过冷却点、体内含水量、总脂肪、总蛋白和总糖含量。结果表明：从10月至翌年2月份结茧和裸露越冬幼虫的过冷却点、结冰点、总蛋白含量、总糖和总脂肪含量及结茧幼虫的含水量均呈下降趋势。裸露幼虫含水量持续增加,在2月达到最高61.42%,在其存活的整个时期的含水量高于同时期结茧幼虫含水量。裸露幼虫的总糖含量持续降低,在2月份含量达到最低（0.65 μg/mg）,存活的整个时期显著低于同时期的结茧幼虫总糖含量。在裸露幼虫存活的整个时期总蛋白含量和总脂肪含量都和结茧幼虫没有显著差异(P>0.05)。在越冬过程中, 裸露幼虫体内高含水量和低含糖量可能是其不能正常越冬的原因。     

CHANGES OF GLYCEROL CONTENT IN DIAPAUSE LARVAE OF THE ORANGE WHEAT BLOSSOM MIDGE, SITODIPLOSIS MOSELLANA (GEHIN) IN VARIOUS SEASONS

Abstract  The glycerol contents in diapause larvae of the orange wheat blossom midge, Sitodiplosis mosellana (Gehin), collected from various seasons, were measured. The results showed that there was less glycerol content in larvae during living on the wheat head. Content of glycerol began to increase significantly when the larvae left the wheat head and entered the soil. A change trend of upper- lower- upper- lower in larvae glycerol contents during diapause in soil was observed from June to April of next year. More glycerol could be examined in larvae collected in summer and winter than in spring and autumn. There was not more glycerol in cocooned larvae than that in non-cocooned larvae during various seasons from the point of statistics. Comparing the glycerol content of larvae being diapause in the first year with that of larvae in the second year, there was yet no obvious difference when larvae were collected in the same season belonged to different years. Therefore, it is shown that the content of glycerol in larvae of the wheat midge in diapause is affected mainly by the seasons or diapause intensity.     

小麦吸浆虫滞育幼虫甘油含量的季节变化(英文)

采用比色分析方法测定了小麦吸浆虫Sitodiplosis mosellana滞育期间幼虫甘油含量的季节变化。结果表明,幼虫离穗入土以前,甘油含量很低,仅为2.96±1.19μg/mg。幼虫离穗入土后,甘油含量急剧升高。入土后5d,甘油含量即上升为6.58±1.11μg/mg;至入土后17d,甘油含量达到高峰,12.21±2.21μg/mg。以后随时间的短期变化其含量变化不大,但随季节变化表现了一定的规律性,呈现了上升——下降——再上升——再下降的变化趋势,即夏季和冬季含量较高,秋季和春季含量较低。结茧幼虫和裸露幼虫比较,结茧幼虫的甘油含量在盛夏和冬季略高于裸露幼虫,秋季和春季时二者含量相当,两者在统计学上并无显著的差异。同一季节的当年滞育幼虫和进入第2年滞育幼虫的甘油含量无明显差异。由此可见,小麦吸浆虫幼虫滞育期间,甘油含量受季节或滞育深度的影响较大。     

The influence of humidity on the effect of Steinernema feltiae against diapausing codling moth larvae (Cydia pomonella L.) (Lepidoptera: Tortricidae)

Codling moth (CM) is a serious and global pest of pome fruit. It overwinters in cryptic habitats as cocooned diapausing larvae. Field trials with the entomopathogenic nematode Steinernema feltiae (Rhabditida: Steinernematidae) report control of diapausing CM of up to 70%, but results are variable. The objective of this study was to define environmental conditions favouring the performance of the nematodes. Cocooned larvae were more susceptible than non-cocooned larvae. S. feltiae was unable to infect CM at a water activity (aw-values) < or = 0.9. Mortality of cocooned larvae was reported at lower aw-values than of non-cocooned larvae. Exposure time and impact of external relative humidity (RH) was studied. Mortality of cocooned larvae did not further increase after half an hour of exposure, whereas the mortality increased with increasing exposure time in non-cocooned larvae. LC50 and LC90 considerably decreased with increasing RH. The influence of the relative humidity was less pronounced when surpassing 80% than the effect of the volume of applied water. When S. feltiae was formulated in a surfactant-polymer-formulation (SPF), mortality significantly increased when compared to application in water.     

Rat brain glycolysis regulation by estradiol-17 beta.

The effect of estradiol-17 beta on the activities of glycolytic enzymes from female rat brain was studied. The following enzymes were examined: hexokinase (HK, EC 2.7.1.1), phosphofructokinase (PFK, EC 2.7.1.11), aldolase (EC 4.1.2.13), glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12), phosphoglycerate kinase (EC 2.7.2.3), phosphoglycerate mutase (EC 2.7.5.3), enolase (EC 4.2.1.11) and pyruvate kinase (PK, EC 2.7.1.40). The activities of HK (soluble and membrane-bound), PFK and PK were increased after 4 h of hormone treatment, while the others remained constant. The changes in activity were not seen in the presence of actinomycin D. The significant rise of the activities of the key glycolytic enzymes was also observed in the cell culture of mouse neuroblastoma C1300 treated with hormone. Only three of the studied isozymes, namely, HKII, B4 and K4 were found to be estradiol-sensitive for HK, PFK and PK, respectively. The results obtained suggest that rat brain glycolysis regulation by estradiol is carried out in neurons due to definite isozymes induction.     

Influence of humidity and a surfactant-polymer-formulation on the control potential of the entomopathogenic nematode <Emphasis Type="Italic">Steinernema feltiae</Emphasis> against diapausing codling moth larvae (<Emphasis Type="Italic">Cydia pomonella</Emphasis> L.) (Lepidoptera: Tortricidae)

The codling moth (Cydia pomonella L.) is a serious pest of pome fruit. Diapausing cocooned larvae overwinter in cryptic habitats in the soil or in the bark of infested trees. The entomopathogenic nematode Steinernema feltiae (Filipjev) (Rhabditida: Steinernematidae) is used to control diapausing codling moth larvae. The objective of this study was to define environmental conditions favouring the performance of the nematodes. Cocooned larvae were more susceptible than non-cocooned larvae. Susceptibility of pupae was low. To determine the influence of decreasing water activity (a_w-value) on the activity of the nematodes, mortality of codling moth larvae and Galleria mellonella L. were tested in sand-sodium-polyacrylate mixtures of variable water activity. S. feltiae was able to infect both insects at a_w-values >0.9. Cocooned larvae of both insects died at lower a_w-values than non-cocooned larvae. Mortality of cocooned larvae did not further increase after half an hour of exposure to nematodes, whereas the mortality of non-cocooned larvae increased with increasing exposure time. LC₅₀ and LC₉₀ considerably decrease with increasing RH. The negative influence of the relative humidity (macro environment) was less important than the effect of the water activity in the bark substrate (micro environment). The micro environment can be manipulated by applying S. feltiae with higher volumes of water. A surfactant-polymer-formulation significantly increased nematode efficacy and can buffer detrimental environmental effects.     

Fuel use in hawkmoth (Amphion floridensis) flight muscle: enzyme activities and flux rates

The fuels used by the hawkmoth Amphion floridensis to power flight are determined by nectar-feeding, with fed moths using primarily carbohydrate and unfed moths using primarily fat. To investigate the metabolic pathways underlying fuel-use flexibility in this species, we measured the maximal activities of several key metabolic enzymes in the flight muscle of fed and unfed individuals, for which metabolic rates and fuel utilization had been previously determined. Hexokinase (HK) and phosphofructokinase (PFK) occur at high activities and, during carbohydrate-fueled flight, are estimated to operate at fractional velocities comparable to those of exclusively carbohydrate-utilizing insects. Females exhibited higher glycolytic enzyme activities than did males, and males regulated PFK activity according to nectar feeding. Although beta-hydroxyacyl-CoA dehydrogenase (HOAD) was found at high activities, carnitine palmitoyl transferase (CPT) was not detectable, suggesting that fatty acids may be utilized via a carnitine-independent pathway during flight. Principal component analysis revealed a tendency for the activities of citrate synthase, HK, PFK, and HOAD to be positively correlated among individuals, as well as a lesser tendency for the activities of glycolytic vs. mitochondrial enzymes to be negatively correlated with each other. However, the principal components did not correlate with variation in either oxygen consumption rate or fuel use in vivo, suggesting that variation in enzyme concentration did not determine differences among individuals in metabolic performance during flight. J. Exp. Zool. 290:108-114, 2001.