RNAi介导的HvIAP1基因沉默对茄二十八星瓢虫存活和发育的影响

RNAi抗虫技术,是一种新型绿色环保的害虫防控方法,具有广阔的应用前景.茄二十八星瓢虫Henosepilachna vigintioctopunctata是茄科植物上的重要害虫,对作物生产造成了严重的经济损失.本研究用饲喂法RNAi探究了沉默凋亡抑制蛋白1(inhibitor of apoptosis protein 1,IAP1)表达对茄二十八星瓢虫生长发育的影响.结果显示,HvIAP1在茄二十八星瓢虫的每个发育阶段均表达,1龄幼虫和3龄幼虫的表达量最高,成虫的表达量最低.取食200 ng/μL dsHvIAP1浸泡处理的叶片2 d后,可导致92％的茄二十八星瓢虫1龄幼虫死亡,同时在取食dsHvIAP124 h后,HvIAP1基因的表达量下降了2.40倍;另外,单头取食200 ng的dsHvIAP1可导致70％的4龄幼虫死亡,同时在取食48 h后,HvIAP1基因的表达量下降了2.58倍.并且RNAi处理后1龄幼虫和4龄幼虫大部分个体在48 h内出现急性取食障碍现象,基本不取食直至死亡.上述结果表明,HvIAP1基因在茄二十八星瓢虫的生长发育过程中起重要作用,当该基因表达受阻,会直接影响茄二十八星瓢虫的取食、抑制其生长发育而死亡.这一研究表明HvIAP1基因可作为潜在的防治茄二十八星瓢虫的RNAi靶标基因.     

马铃薯瓢虫和茄二十八星瓢虫体表细微结构的比较研究

本研究利用扫描电镜对马铃薯瓢虫Henosepilachnavigintioctomaculata (Motschulsky)和茄二十八星瓢虫H .vigintioctopunctata (Fabricius)体表的细微结构进行了比较研究。结果表明 ,马铃薯瓢虫和茄二十八星瓢虫在细微结构上存在明显差异。第一 ,2种瓢虫鞘翅表面凹陷内 ,毛的着生位置不同 ,马铃薯瓢虫的毛着生于凹陷中心 ,茄二十八星瓢虫的毛着生于凹陷边缘 ;第二 ,鞘翅表面凹陷的深浅不同 ,马铃薯瓢虫的凹陷较深于茄二十八星瓢虫。这为 2种瓢虫的形态学研究提供了细微结构方面的依据     

茄子品种对茄二十八星瓢虫及其寄主蜂生长发育的影响

茄子不同品种对茄二十八星瓢虫及瓢虫标腹姬小蜂的影响结果表明,上海紫茄、呼杂34^#、托塔姆3^#、洋红茄4个品种中,取食上海紫茄的瓢虫发育历期明显短于取食其它3个品种的瓢虫。其幼虫－蛹历期比在托塔姆3^#上取食的瓢虫发育历期快5.69d。瓢虫柄腹姬小蜂在取食上海紫茄的瓢虫幼虫内发育出蜂数多、性比高,但寄生率稍低。而在托塔姆3^#上取食的幼虫内发育出蜂数少、羽化率低、蜂体小、但寄生率稍高。该蜂喜在3,4龄幼虫内寄生,在其上寄生蜂出蜂多、性比高、蜂的个体大。     

瓢虫柄腹姬小蜂对茄二十八星瓢虫功能反应的研究

研究了瓢虫柄腹姬小蜂功能反应。结果表明,在供试温度下,瓢虫柄腹姬小蜂对茄二十八星瓢虫的功能反应均符合Holling圆盘方程Ⅱ型,以25℃下的搜寻效率最高,其模型为Nα=1．94N／(1 0．39N),在茄枝叶、龙葵枝叶、酸酱枝叶和西红柿枝叶4种叶片上,以茄枝叶、龙葵枝叶上的搜寻效率最高,分别为1．85和1．94。瓢虫柄腹姬小蜂对不同密度的2、3、4龄幼虫的功能反应,以4龄的搜寻效率最高,其次为3龄,最低为2龄,实验中未发现瓢虫柄腹姬小蜂寄生1龄的幼虫。不同处理方法、不同药剂影响瓢虫柄腹姬小蜂对不同寄主密度的功能反应,其中阿维菌素组的搜寻效率与对照组没有显著性差异,苦皮藤索组的搜寻效率稍有降低,而敌敌畏、氯氰菊酯处理组的搜寻效率显著下降,处理时间相对延长。     

马铃薯瓢虫与茄二十八星瓢虫SCAR标记的建立及应用

为了探索快速鉴定马铃薯瓢虫Henosepilachna vigintioctomaculata(Motschulsky)和茄二十八星瓢虫Henosepilachna vigintioctopunctata(Fabricius)的分子生物学方法,本研究在随机扩增多态性DNA(random amplified polymorphic DNA,RAPD)的基础上,分别设计了可以鉴别两个物种的序列特征扩增区域(sequence characterized amplified regions,SCAR)标记。从随机合成的60条引物中筛选出来2条特异性引物(分别为OPI-6和OPJ-15),引物OPI-6在马铃薯瓢虫中扩增出约750 bp的特异性条带,引物OPJ-15在茄二十八星中扩增出约750 bp的特异性条带,根据测序结果设计了两对SCAR引物对筛选结果进行验证,发现根据OPI-6的测序结果所设计的SCAR引物(OPI-6 test)仅能在马铃薯瓢虫中扩增出645 bp的条带,而根据OPJ-15的测序结果所设计的SCAR引物(OPJ-15 test)仅能在茄二十八星瓢虫中扩增出436 bp的条带。这两对SCAR引物能够准确、稳定且快速地区分马铃薯瓢虫与茄二十八星瓢虫,对这两种害虫的精准防控具有重要意义。     

不同温度下茄二十八星瓢虫的实验种群生命表

近年来,茄二十八星瓢虫Henosepilachna vigintioctopunctata在江汉平原地区危害严重,本研究旨在探明温度对该地区茄二十八星瓢虫生长发育和繁殖的影响,组建茄二十八星瓢虫的实验种群生命表,为该虫防治提供一定的理论依据。本研究采用室内恒温饲养方法,分别设置19℃、22℃、25℃、28℃、31℃和34℃等6个温度,测定各温度下用马铃薯饲养不同虫态的生长发育历期。结果表明,茄二十八星瓢虫在不同恒温条件下各虫态的发育历期随着温度的升高而逐渐缩短,19℃恒温下发育历期最长为3219 d,31℃恒温下仅为1315 d。幼虫期和蛹期的发育起点温度分别为1093和1213℃,日有效积温为18604和5706 d·℃。在19-31℃5个温区下幼虫和蛹的存活率都很高,并依据该温区内的发育起点温度和有效积温建立了发育历期预测式与Logistic模型。在25℃时单雌平均产卵量达到最大值107450粒,世代存活率和种群趋势指数最高,分别为9333%和50147,高温和低温均不利于其种群数量增加。     

茄二十八星瓢虫在茄子叶片上垂直方向的附着力

本文采用电子天平称重法对茄二十八星瓢虫的2龄幼虫、3龄幼虫、4龄幼虫、雌成虫和雄成虫在茄子叶片上垂直方向的附着力进行测定,为进一步研发和利用气流控制技术防治农业害虫提供技术参数。研究结果表明,2龄幼虫、3龄幼虫、4龄幼虫、雌成虫和雄成虫垂直方向的最大附着力分别为7.3±0.3 mN,8.6±0.6 mN,12.0±0.3 mN,15.8±1.3 mN和13.0±1.0 mN。茄二十八星瓢虫垂直方向的附着力从大到小的顺序均为:雌成虫雄成虫;4龄幼虫3龄幼虫2龄幼虫。幼虫的附着力/虫体体重系数由大到小的顺序为:2龄幼虫3龄幼虫4龄幼虫,所有处理的附着力与虫体体重的相关性均不显著。附着力/虫体重量的系数范围为47.8-90.8,身体重量占附着力1.1%-2.1%。结果进一步表明,在设计旋风式气流吸虫机的频率等参数时,要考虑茄二十八星瓢虫在茄子叶片上的附着力远大于其体重的情况。     

茄二十八星瓢虫成虫天花板爬行行为研究

【目的】探明茄二十八星瓢虫Henosepilachna vigintioctopunctata(Fabricius)成虫的天花板爬行行为,为气流控制技术防治农业害虫提供技术参数。【方法】本研究通过采用录像观察法对茄二十八星瓢虫雌雄成虫的天花板爬行行为进行观测,明确其成虫的运动行为参数,并分析了其步速分别与步距、步频和负荷因数的关系。【结果】茄二十八星瓢虫雌雄成虫天花板爬行时的速度为3.77~19.32 mm/s,步距为2.63~4.51 mm,步频范围为1.32~4.74 Hz。步距和步频均与速度之间存在极显著线性关系,速度的增加同时依靠步距和步频的增加,但步距的变化相对步频的变化小;前足、中足和后足的负荷因数均随着速度的提高而降低,且负荷因数均与速度存在显著相关性;一个步态周期内,足的摆动时间均远小于支撑时间。【结论】茄二十八星瓢虫成虫天花板爬行时速度的增加主要是依靠步频的提高来实现。     

蠋蝽对茄二十八星瓢虫的捕食功能反应

【目的】明确蠋蝽对茄二十八星瓢虫的捕食能力。【方法】在实验室条件下测定蠋蝽5龄若虫对茄二十八星瓢虫2和4龄幼虫的捕食功能反应及搜寻效应。【结果】蠋蝽5龄若虫对茄二十八星瓢虫2和4龄幼虫的捕食行为均符合Ⅱ型功能反应;随着茄二十八星瓢虫密度的增加,蠋蝽的搜寻效应逐渐降低,蠋蝽5龄若虫对茄二十八星瓢虫2龄幼虫的搜寻效应明显大于4龄幼虫;蠋蝽5龄若虫对茄二十八星瓢虫2和4龄幼虫的实际最大捕食量分别为15和10头。【结论】蠋蝽5龄若虫对茄二十八星瓢虫幼虫具有较好的捕食作用。     

茄二十八星瓢虫为害对不同基因型番茄叶内三种抗性酶活性的影响

【目的】研究茉莉酸合成相关基因在番茄抗茄二十八星瓢虫Henosepilachna vigintioctopunctata Fabricius中的作用,探讨茉莉酸信号因子在植物防御过程中起到的作用。【方法】利用3种基因型番茄茉莉酸合成缺失突变体spr2、茉莉酸合成过量表达体35s、野生型番茄Wt为材料,通过经2龄茄二十八星瓢虫分别取食6、24、48 h后,利用分光光度计测定3种基因型番茄叶片内苯丙氨酸解氨酶(Phenylalanineammonialyase,PAL)、胰蛋白酶抑制剂(Protease inhibitors,PI)和脂氧合酶(Lipoxygenase,LOX)酶活性的动态变化。【结果】35s型番茄叶片内PI活性显著高于另两种番茄叶片,而spr2型番茄叶片PI活性最低。瓢虫为害后,3种基因型番茄PI酶活性明显提高,在35s和Wt型番茄中,PI和LOX酶活性在胁迫24 h和6 h达到最高值,而spr2型番茄没有显著变化。受到瓢虫侵染的系统叶中,PI和LOX酶活性也受到了诱导,系统叶变化趋势同危害叶相同,但诱导的PI和LOX酶活性明显低于危害叶。在危害处理过程中,均呈现上升-下降的趋势,在系统叶中,则PAL活性没有显著变化。番茄叶片受害后,PI和LOX酶活性上升辐度大于PAL,说明PI和LOX对取食胁迫响应比PAL更敏感。【结论】35s茉莉酸合成过量表达体的番茄对茄二十八星瓢虫有一定的抗虫作用,因此,茉莉酸在植物抗性中起了一定作用。     

Rearing the 28-spotted ladybird beetle,Henosepilachna vigintioctopunctata (Coleoptera: Coccinelidae), with a switchover from host plant leaves to artificial diet

In this study, I tested an artificial diet, Insecta LFS, for rearing the 28-spotted ladybird beetle, Henosepilachna vigintioctopunctata (Fabricius). H. vigintioctopunctata larvae could not be reared through all of the larval stages on the artificial diet. However, initially they could be reared on tomato leaves up to the second or third instar, and thereafter exclusively on the artificial diet. The larval and pupal periods were not significantly different from those reared only on tomato leaves. For females reared by the diet-switching method, the preoviposition period was significantly longer and the number of eggs laid significantly lower than for those on tomato leaves. However, these differences did not become a hindrance to laboratory rearing. This rearing method saves labor involving obtaining fresh plant leaves and produces a reliable supply of the insects.     

Oral delivery of dsHvlwr is a feasible method for managing the pest Henosepilachna vigintioctopunctata (Coleoptera: Coccinellidae)

RNA interference(RNAi)techniques have emerged as powerful tools that facilitate development of novel management strategies for insect pests,such as Henosepilachna vigintioctopunctata(Coleoptera:Coccinellidae),which is a major pest of solanaceous plants in Asia.In this study,the potential of oral delivery of in vvYro-synthesized and bacterially expressed double-stranded H.vigintioctopunctata lesswright(Iwr)gene(dsHvlwr)to manage of H.vigintioctopunctata was investigated.Our results showed that the gene Hvlwr had a 480-bp open reading frame and encoded a 160-amino acid protein.Hvlwr expression levels were greater in the fat body than other tissue types.Hvlwr silencing led to greater H.vigintioctopunctata mortality rates and appeared to be time-and partially dose-dependent,likely as a result of the number of hemocytes increasing with dsRNA concentration,but decreasing with time.Bacterially expressed dsHvlwr that was applied to leaf discs caused 88%,66%,and 36%mortality in 1st instars,3rd instars,and adults after 10,10,and 14 d,respectively;when applied to living plants,there was greater mortality in 1 st and 3rd instars,but there was no effect on adults.Furthermore,dsHvlwr led to improved plant protection against H.vigintioctopunctata.Our study shows an effective dietary RNAi response in H.vigintioctopunctata and that Hvlwr is a promising RNAi target gene for control of this pest species.     

Seasonal Occurrence of Henosepilachna vigintioctopunctata (F.) (Coleoptera: Coccinellidae) and Its Parasitoid on Ashwagandha in India

The outbreak of Henosepilachna vigintioctopunctata (F.) was noticed on a medicinal plant, Withania somnifera Dunal during 2004-05. The population level of the pest was reached its peak in August. The parasitoid Pediobius foveolatus caused 51.94%±12.20% parasitism. The pest completed its life cycle in 20.15±1.50 days on W. somnifera. The longevity of the male and female adults of the pest was 22.07±3.71 and 31.07±4.38 days, respectively. The gravid females of H. vigintioctopunctata laid an average of 287.64±33.38 eggs during their oviposition period of 10.40±2.80 days.     

异色瓢虫成虫耐寒能力的季节性变化

为研究异色瓢虫Harmonia axyridis自然种群耐寒能力的季节性变化,测定了其过冷却点、体内含水量及总脂肪含量和低温存活力。结果表明：异色瓢虫成虫低温存活力呈现出明显的季节性变化,越冬前成虫的耐寒性显著强于夏季成虫和越冬后成虫。冷驯化（5℃, 5 d）可以显著提高夏季成虫的低温存活力。雌雄成虫过冷却点和体内含水量随气温的降低而降低,升高而升高。过冷却点7月份最高,分别为−7.6℃和−8.0℃;越冬中期（2008-01-15）最低,分别为−18.1℃ 和−16.9℃。雌雄成虫体内含水量9月份最高,分别为66.87%和68.01%,10月份显著降低,越冬后期（2008-02-19）最低,分别为52.94%和51.53%。越冬期间过冷却点和体内含水量显著低于其他时期。而雌雄成虫体内总脂肪含量在越冬开始就达到最高,分别为50.07%和47.93%,随后又逐渐降低,越冬期间显著高于其他时期。由此可知异色瓢虫自然种群的耐寒性呈现出明显的季节性变化,文中还就异色瓢虫自然种群耐寒性影响因素及其越冬策略进行了讨论。     

Genome size increase in the phytophagous ladybird beetle Henosepilachna vigintioctomaculata species complex (Coleoptera: Coccinellidae)

This study reports genome size (C‐value) estimates for seven species of ladybird beetles (Coleoptera: Coccinellidae) in Japan using flow cytometry. The results demonstrated genome sizes of 1.0–1.4 Gb in four closely related phytophagous ladybird beetles belonging to the Henosepilachna vigintioctomaculata species complex. These values were approximately two times larger than that of a congeneric phytophagous ladybird beetle H. vigintioctopunctata (0.66 Gb), and of two very distantly related common carnivorous ladybird beetles, Harmonia axyridis (0.46 Gb) and Coccinella septempunctata (0.42 Gb). These lines of evidence suggest that rapid and large genome size increase occurred just after the branching of the common ancestor of the H. vigintioctomaculata species complex from other ladybird species.