Interruptin B对小菜蛾和亚洲玉米螟幼虫的拒食活性及对斜纹夜蛾卵巢细胞的毒力

为了研究Interruptin B的杀虫活性, 初步探讨其杀虫作用机理, 本研究采用叶碟法和叶片浸渍法测定了Interruptin B对小菜蛾Plutella xylostella和亚洲玉米螟Ostrinia furnacalis 3龄幼虫的拒食活性和毒杀活性; 采用注射法和血球计数法研究对斜纹夜蛾Spodoptera litura幼虫血细胞数的影响; 采用四甲基偶氮唑盐［3-(4, 5-dimethylthiazole-2-yl)-2, 5-diphenyltetrazolium bromide, MTT］法测定对斜纹夜蛾卵巢细胞(SL细胞)的增殖抑制作用; 采用倒置显微镜观察对SL细胞形态、密度和贴壁率的影响。结果显示：处理后48 h, Interruptin B对亚洲玉米螟和小菜蛾3龄幼虫的拒食中浓度(AFC₅₀）分别为108.56 和143.88 μg/mL, 致死中浓度(LC₅₀）分别为346.23和194.32 g/mL; 处理后24 h和48 h, 对SL细胞的抑制中浓度(IC₅₀）分别为13.51和4.11 g/mL。Interruptin B处理SL细胞后12-48 h, 细胞密度随时间的延长而下降, 与对照差异显著(P<0.05）; 处理后3, 6, 12, 24和48 h, SL细胞变形率分别为28.18%, 83.90%, 85.18%, 93.93%和100%, 细胞贴壁率分别为96.59%, 89.90%, 83.18%, 5.06%和1.09% , 处理后6-48 h, SL细胞变形率和贴壁率与对照差异显著(P<0.05）。Interruptin B对小菜蛾和亚洲玉米螟3龄幼虫具有良好的拒食和毒杀活性, 能显著降低斜纹蛾幼虫血细胞数量, 对SL细胞具有良好的增殖抑制作用, 可显著降低SL细胞生长密度, 改变细胞形态, 降低贴壁率, 表明该化合物可通过影响昆虫细胞活力毒杀活体昆虫。     

50种热带地区植物对斜纹夜蛾的拒食活性

在室内测定了采自海南儋州两院本部及昌江霸王岭的26科50种热带地区植物56个样品的乙醇提取物对斜纹夜蛾四龄幼虫的拒食活性,结果表明：假药(根)、肉桂(叶)、密脉崖鹿角藤(叶)、牛筋果(叶)、鸦胆子(枝叶)、过江龙(枝叶)、牛心番荔枝(叶)等具有较高的拒食活性,其拒食率分别为92．50％、76．25％,、73．95％、57．50％、56．85％、56．58％、和50．42％。此结果为进一步从这些热带植物中提取、分离和纯化高活性的拒食作用活性成分奠定了基础。     

番荔枝内酯化合物布拉它辛对斜纹夜蛾的杀虫活性及对SL细胞的致凋亡作用

为了明确番荔枝内酯化合物布拉它辛的杀虫活性和探索杀虫作用机理,本研究采用浸叶法测定该化合物对斜纹夜蛾Spodoptera litura幼虫的生物活性,采用MTT检测法和流式细胞术,研究该化合物对斜纹夜蛾离体培养卵巢细胞（SL细胞）的细胞毒力和致细胞凋亡作用,以及对SL细胞线粒体膜电位的影响。结果表明: 布拉它辛不仅对斜纹夜蛾幼虫具有良好的拒食活性,处理后24 h,对2龄和3龄幼虫的AFC₅₀值分别为60.25 μg/mL和86.73 μg/mL,还对幼虫生长有良好的抑制作用。布拉它辛处理SL细胞后24 h和48 h,IC50值分别为22.32 μg/mL和10.03 μg/mL。流式细胞仪检测结果表明,布拉它辛对SL细胞具有明显的致凋亡作用,可导致SL细胞线粒体膜电位显著下降。本研究表明布拉它辛对斜纹夜蛾幼虫具有良好的拒食活性与生长抑制作用, 并且该化合物能明显抑制SL细胞的增殖,诱导细胞凋亡,降低线粒体膜电位。因此,布拉他辛具有广阔的研究应用前景。     

广藿香酮对斜纹夜蛾的拒食作用研究

广藿香酮Pogostone,是一种新型的、被认为具有应用潜力的植物源杀虫活性物质。为探明广藿香酮的杀虫活性,本试验以斜纹夜蛾Spodoptera litura作为试虫,采用选择性和非选择性拒食活性测定法,在室内分别测定了不同浓度的广藿香酮对斜纹夜蛾的拒食活性以及广藿香酮对不同龄期的斜纹夜蛾的拒食作用,研究结果表明,1 mg/mL、2 mg/mL、4 mg/mL、8 mg/mL、10 mg/mL的浓度下,广藿香酮对三龄斜纹夜蛾幼虫的选择性拒食率分别为56.5%、62.24%、80.01%、82.40%、90.03%;非选择性拒食率分别为62.48%、71.78%、84.15%、86.35%、92.71%。浓度为4 mg/mL的广藿香酮对2龄、3龄、4龄、5龄斜纹夜蛾幼虫的选择性拒食率分别为78.56%、80.25%、66.80%、40.35%;非选择性拒食活性分别为78.21%、83.02%、77.36%、40.24%。可见,浓度为4 mg/mL的广藿香酮对三龄斜纹夜蛾的拒食效果较好。     

杀菌剂对斜纹夜蛾SL细胞系和幼虫的生物活性

以MTT法筛选了19种杀菌剂对斜纹夜蛾Spodoptera litura SL细胞的毒杀活性,并以该方法研究了三唑类杀菌剂对SL细胞的毒力。结果表明: 福美双、烯唑醇、己唑醇、氟硅唑、苯霜灵、苯醚甲环唑、戊唑醇和腈菌唑等杀菌剂对SL细胞具有优异的毒杀活性。三唑类杀菌剂腈菌唑、烯唑醇、己唑醇和戊唑醇处理SL细胞48 h后,LC₅₀值分别为21.94 μg/mL、 23.80 μg/mL、 33.16 μg/mL和47.63 μg/mL。以考马斯亮蓝G₂₅₀法研究了腈菌唑对SL细胞中蛋白质含量和乳酸脱氢酶(LDH)漏出率的影响, 20 μg/mL腈菌唑处理12 h、24 h、48 h和72 h后,SL细胞中蛋白质含量分别降低8.55%、25.95%、42.95%和67.05%;处理24 h和48 h后,SL细胞的LDH漏出率分别为30.66%和32.05%。以浸叶喂食法处理斜纹夜蛾3龄幼虫,三唑类杀菌剂可显著抑制试虫体重增长。以0.5 μg/头、 1.0 μg/头和2.0 μg/头剂量的腈菌唑注射处理72 h后,斜纹夜蛾4龄幼虫血细胞数量分别降低12.31%、 25.96%和25.73%;腈菌唑注射处理48 h和72 h后,对斜纹夜蛾幼虫的LD₅₀值分别为1.59 μg/头和1.53 μg/头。结果显示以离体培养细胞为对象,从现有杀菌剂中寻找新的杀虫剂先导化合物具有良好的研究潜力。     

刚竹叶提取物对斜纹夜蛾酚氧化酶抑制作用研究

采用酶标仪法测定了刚竹叶提取物对斜纹夜蛾酚氧化酶(PO)活性的影响。结果表明:6种不同溶剂(极性大小:去离子水乙醇丙酮乙酸乙酯氯仿石油醚)获得的竹叶提取物均对PO表现为抑制作用(三氯甲烷提取物在0-300 mg/L范围内,对PO表现为激活作用),且随提取物浓度的升高,抑制作用有逐渐增强的趋势。丙酮提取物对PO的抑制效果最显著(IC50为550 mg/L),其次为乙酸乙酯提取物和乙醇提取物(IC50分别为575 mg/L和700 mg/L);丙酮提取物对斜纹夜蛾酚氧化酶的抑制效果属于可逆过程,抑制类型为非竞争性抑制,抑制常数Ki为158.2 mg/L。     

斜纹夜蛾NPV对取食不同蔬菜的斜纹夜蛾幼虫的毒力差异

采用生物测定法,比较了斜纹夜蛾ＮＰＶ对取食芋叶等７科８种蔬菜的斜纹夜蛾幼虫的毒力反应差异性。结果表明ＮＰＶ对取食番茄叶的幼虫毒力最高,比取食芋叶的毒力高６．８２倍;但再经菜心饲养一代后,ＮＰＶ的毒力差异消失。     

苦瓜叶提取物对亚洲玉米螟的生物活性及对斜纹夜蛾卵巢细胞的毒力

【目的】为研究苦瓜(Momordica charantia L.)叶正丁醇萃取物对亚洲玉米螟Ostrinina furnacalis(Güenée)生长发育及生殖力的影响,并探讨其作用机理。【方法】采用饲料混药法测定了苦瓜叶正丁醇萃取物对亚洲玉米螟幼虫存活率、发育历期和雌成虫产卵量的影响。【结果】用0.025%、0.1%和0.2%浓度的苦瓜叶正丁醇萃取物混合饲料饲喂亚洲玉米螟2龄幼虫3 d后,幼虫存活率明显降低,校正死亡率分别为27.43%、39.23%和54.93%,且发育历期明显延长,雌成虫产卵量与对照相比分别下降了12.63%、22.74%和37.27%。进一步的研究结果表明,苦瓜叶正丁醇萃取物对斜纹夜蛾卵巢细胞(SL)的形态、增殖和生长密度有明显的抑制作用。处理后的SL细胞皱缩变圆,细胞核偏移甚至消失,胞质外泄。处理24、48和72 h后,对SL细胞增殖的IC50值分别为180.96、158.54和122.87μg/mL;处理6~72 h后,SL细胞相对生长密度随药剂浓度和处理时间的增加而明显下降;用125μg/mL浓度处理SL细胞2、3、4和5 d后,SL细胞对葡萄糖吸收的抑制率分别为7.66%、13.97%、19.45%和27.30%。【结论】苦瓜叶正丁醇萃取物对SL细胞表现出的较强毒杀作用可能是其导致亚洲玉米螟生殖能力降低的主要作用机制之一。     

竹提取物对棉铃虫幼虫及菜青虫的拒食活性

研究结果表明 ,质量浓度为 1 0g L的 1 0种供试竹提取物对棉Helicoverpaarmigera铃虫均具有较强的拒食作用 ,1 2h拒食率最高达 95 . 0 3 % ,最低为 74. 0 7% ,其中毛金竹 (Phyllostachysnigravar.henonis)、白纹短穗竹 (Brachystachyumalbostriatum)提取物对棉铃虫 3龄幼虫的AFC50 (拒食中浓度 )分别为 2 . 0g L、2. 7g L ,但 2 4h、48h拒食率均明显下降。对菜青虫 3龄pierisrapae幼虫的拒食效果以凤凰竹 (Bambusamultiplex)提取物最好 ,2 4h拒食率为 71. 69% ;对 4龄菜青虫的拒食效果以毛金竹提取物最好 ,拒食率为71 . 45 % ,其中毛金竹、白纹短穗竹 2种竹提取物对菜青虫 3龄幼虫的AFC50 分别为 2 .68g L和 3 . 3 7g L。研究结果对于开发环境友好农药以及充分利用竹类资源均具有重要意义。     

苦瓜叶乙酸乙酯提取物对斜纹夜蛾实验种群的抑制作用

应用生命表方法评价了苦瓜Momordica charantia叶乙酸乙酯提取物与人工饲料混合饲喂斜纹夜蛾Spodoptera litura 3龄幼虫后对其实验种群增长的影响,旨在为探明苦瓜叶提取物的作用方式和作用机理以及田间应用提供科学依据。结果表明,苦瓜叶乙酸乙酯提取物对斜纹夜蛾幼虫的生长发育有显著的抑制作用。随着处理浓度的增加,幼虫体重的增长也随着减慢,发育历期明显延长,死亡率也随着提高。用提取物浓度为0.032%、0.04%、0.08%和0.16%的人工饲料饲喂3龄幼虫,2d后的体重增长抑制率分别为76.3%、79.9%、97.6%和111.2%。0.16%浓度处理的幼虫化蛹率明显降低,成虫的羽化率和产卵量也明显下降。苦瓜叶乙酸乙酯提取物能显著降低斜纹夜蛾的种群趋势指数值(I),与对照相比,0.032%、0.04%、0.08%和0.16%浓度处理的种群控制指数(IIPC)分别是0.59、0.56、0.29和0.20。说明苦瓜叶乙酸乙酯提取物不仅对斜纹夜蛾的生长发育有明显的抑制作用,而且对斜纹夜蛾的繁殖及其种群增长也有明显的控制作用。     

杀菌剂丙环唑对斜纹夜蛾的毒性

在研究比较了11种农药对斜纹夜蛾Spodoptera litura细胞（简称SL细胞）的毒杀活性基础上,选择毒杀活性最高的杀菌剂丙环唑,对其毒理学机理进行进一步研究。结果表明,丙环唑的细胞毒力最高,在100 μg/mL浓度下处理后48 h,SL细胞的死亡率为98.08％。处理后36 h,丙环唑对SL细胞的LC₅₀值为20.31 μg/mL。丙环唑能明显降低SL细胞的蛋白质含量。以0.5 μg/头的丙环唑注射斜纹夜蛾4龄幼虫,处理后72 h,试虫血淋巴总含量及血细胞数分别下降了26.80％和25.26％;在1.0 μg/头的剂量下,则分别下降了37.67%和36.32%。以0.5 μg/头和1.0 μg/头的丙环唑注射处理后,斜纹夜蛾幼虫体重显著降低。此外,丙环唑能降低斜纹夜蛾幼虫血淋巴含糖量及血淋巴蛋白质含量。在注射处理后96 h和120 h,丙环唑对斜纹夜蛾4龄幼虫的LD₅₀值分别为0.59 μg/头和0.45 μg/头。丙环唑对SL细胞和斜纹夜蛾幼虫均具有较好的毒杀活性,显示出丙环唑类似物控制害虫的可能性。     

Insecticidal activities of Ginkgo biloba seed coat extracts against Spodoptera exigua (Lepidoptera: Noctuidae)

The present study relates to a methanol extract of the seed coat of Ginkgo biloba, and tested particularly on the third instar larvae of Spodoptera exigua. The extract was found to have an inhibitory effect on the growth of the larvae besides bringing a change in the nutrient reserves in the body of the insect. Topical application of five different doses of the methanol extract resulted in a mortal effect to third instar larvae of S. exigua that is very much dependent on the dose as well as duration of exposure. Lower doses revealed lower mortality after 24 h of application. At doses of 1.00, 2.00, 4.00, 8.00 and 16.00 ng/larva, mortalities were 9.25, 26.07, 50.32, 56.28 and 92.44%, respectively. The dose for 50% mortality (LD₅₀) of methanol extracts by applied by a topical method with 1 µL of acetone solution was 1.92 ng/larva. Nutrient reserves like protein, glycogen and lipid are known to regulate pupation and adult emergence. These reserves have been found to be lower in treated larvae, indicating the insecticidal role of methanol extracts from G. biloba against third instar larvae of S. exigua.     

Laboratory evaluation of flight activity of the common cutworm, Spodoptera litura (Lepidoptera: Noctuidae)

Abstract  The flight activity of Spodoptera litura in tethered conditions is evaluated using a computer-mediated flight-mill in the laboratory. The 3–4-day-old moths fly longer and farther than those of other ages. Male and female moths exhibit similar flight activity. Mating status does not influence the flight duration and distance of 2-day-old females. However, these two flight parameters with a 6-day-old mated female is significantly lower than that of unmated ones. The optimum temperature for flight ranged from 16–24°C, whereas the optimum RH ranged from 60%–100%. During 72-h period, the total flight duration and distance of 1-day-old male and female moths were 19.6 h (± 5.8) and 83.3 km (± 28.4), and 24.0 h (± 7.0) and 105.4 km (± 37.4), respectively. These results indicate that S. litura has a great potential to undertake long-distance migratory flights.     

Bioefficacy of Ceasalpinea bonduc (L.) Roxb. against Spodoptera litura Fab. (Lepidoptera: Noctuidae)

Bioefficacy of hexane, chloroform and ethyl acetate extracts of the seeds of Ceasalpinea bonduc (L.) Roxb. was studied against third instar larvae of Spodoptera litura at 0.25, 0.5, 1, 2.5 and 5.0% concentrations. Significant antifeedant, larvicidal and pupicidal activities and least LC₅₀ values were observed in chloroform extract. The chloroform extract was subjected to fractionation using silica gel column chromatography. Six fractions were obtained; among these, the third fraction showed high antifeedant, larvicidal and pupicidal activities at 1000 ppm concentration. Abnormalities in adults were also observed. All the activities were concentration dependent. C. bonduc could be useful in integrated pest management programme.     

Growth inhibitory activity of Millettia pachycarpa (Bentham) extracts against Tobacco Caterpillar,Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae)

Abstract

Millettia pachycarpa (Bentham), a perennial climbing tree, found endemic in north-east India used by tribes for treating various ailments. To ascertain the potential of M. pachycarpa for pest management, the present investigation was formulated against a polyphagous pest, Spodoptera litura (Fabricius). Growth inhibitory activity of various extracts and extract fractions was evaluated against third instar larvae of S. litura through topical as well as diet incorporation method. It was observed that dichloromethane extract showed very good growth inhibition in both topical (GI₅₀ 7.77?ppm) and diet incorporation methods (GI₅₀< 5?ppm). The growth inhibitory potential of this plant is also demonstrated by the presence of various abnormalities and reduction in larval and pupal weight.     

Exposure to herbicides reduces larval sensitivity to insecticides in Spodoptera litura (Lepidoptera: Noctuidae)

Herbicides and insecticides are widely used in modern agriculture. It has been reported in various studies that application of insecticides can increase tolerance of herbivorous insects to insecticides. However, limited information exists on susceptibility to insecticides when insects are exposed to herbicides. This study was conducted to investigate the potential impact of the herbicides trifluralin and 2-methyl-4-chlorophenoxyacetic acid sodium salt (MCPA-Na) on the susceptibility of the nocturnal moth Spodoptera litura to the insecticides X-cyhalothrin, phoxim and bifenthrin. We found that larvae exposed to trifluralin or MCPA-Na became significantly less susceptible to both insecticides than nonexposed control larvae. Herbicide-treated larvae did not show altered growth under the used test conditions. However, heads of herbicide-treated larvae showed increased expression of the acetylcholinesterase genes SI Ace I and SI Ace 2. Moreover, the fat body and midgut of herbicide-treated larvae displayed elevated expression of detoxification genes (the carboxylesterase gene SI CarE;the glutathione S-transferase genes SlGSTe2 and SlGSTe3\ the cytochrome P450 monooxygenase genes CYP6B48, CYP9A40 and CYP321B1). The CYP6B48 gene exhibited highest inducibility. In conclusion, the data of this study suggest that exposure of S. litura larvae to herbicides may stimulate detoxification mechanisms that compromise the efficacy of insecticides.     

氟铃脲对斜纹夜蛾酚氧化酶活性的影响

【目的】探讨氟铃脲对斜纹夜蛾Spodoptera litura酚氧化酶活性的影响.【方法】应用酶动力学法,在确定斜纹夜蛾酚氧化酶最适反应条件基础上,测定氟铃脲与酚氧化酶直接作用及氟铃脲处理幼虫后酚氧化酶的活性.【结果】以邻苯二酚为底物,斜纹夜蛾酚氧化酶的最适pH 6.5,最适温度30℃.氟铃脲没有和酚氧化酶结构中的铜离子结合,而是直接与酶蛋白发生作用.氟铃脲浓度低于1 153 mg/L对离体酚氧化酶有激活作用,在138 mg/L激活作用最大,浓度高于1 153 mg/L则产生抑制作用.用46 mg/L和92 mg/L亚致死剂量氟铃脲连续处理5龄幼虫不同时间后,酚氧化酶活性均显著提高,且高浓度的激活作用大于低浓度的激活作用.进一步测定药剂处理幼虫24,48和72 h的血淋巴、表皮和头部酚氧化酶活性,随着氟铃脲处理时间延长,两个剂量的氟铃脲对血淋巴、表皮及头部酚氧化酶的激活作用逐渐增大,氟铃脲对血淋巴酚氧化酶的激活作用最大,表皮次之,头部最低.氟铃脲处理幼虫后,其预蛹和蛹酚氧化酶的活性也显著提高,表现出一定的后效应.【结论】一定浓度范围内,氟铃脲对斜纹夜蛾酚氧化酶具有激活作用.     

斜纹夜蛾铁硫亚基蛋白的表达及功能鉴定

铁硫亚基蛋白(rieske iron-sulfur protein, RISP) 是线粒体复合物Ⅲ的关键蛋白亚基之一, 在呼吸链电子传递过程中起到重要作用。本研究通过RT-PCR克隆得到斜纹夜蛾Spodoptera litura RISP基因SlitRISP的ORF, 构建了pET32a-SlitRISP原核表达载体, SDS-PAGE和Western blot检测结果显示, SlitRISP原核表达蛋白以包涵体的形式存在于菌体沉淀中, 且RISP抗体可成功用于该蛋白的免疫印迹检测。为了进一步鉴定SlitRISP在斜纹夜蛾离体细胞系SL-1中的功能, 通过向细胞内转染siRNA, 利用RNAi技术沉默SL-1中的SlitRISP。qRT-PCR结果表明, 分别经50 nmol/L和100 nmol/L siRNA处理48 h后, SL-1中SlitRISP的表达均几乎完全被抑制; Western blot结果显示, SL-1中SlitRISP含量显著低于CK。当SL-1 SlitRISP被成功沉默后, 通过检测SL-1线粒体膜电位、 细胞ATP含量和细胞增殖抑制率鉴定RISP在线粒体电子传递过程中的重要作用。流式细胞仪测定结果表明, 经50 nmol/L和100 nmol/L siRNA处理24 h后, SL-1线粒体膜电位相对于CK分别降低23.52%和11.32%, 而处理48 h后, SL-1线粒体膜电位则分别升高5.58%和27.66%; siRNA处理24 h和48 h后, SL-1 ATP含量相对于CK分别降低82.71%和84.50%, 最终导致SL-1细胞增殖抑制率分别为53.64%和67.94%。这些结果表明SlitRISP在SL-1中参与线粒体膜电位的形成和细胞ATP的合成。介于RISP在线粒体电子传递链中的重要作用, 其可能成为新型杀虫作用靶标, 这可为研制新型呼吸抑制剂提供参考。     

Effect of plant secondary metabolites on common cutworm,Spodoptera litura (Lepidoptera: Noctuidae)

The effect of various flavonoids, lectins and phenyl β‐_D‐glucoside on larval survival, weights and the activities of digestive (total serine protease and trypsin) and detoxifying (esterase and glutathione‐S‐transferase) enzymes of Spodoptera litura larvae at 7 days after treatment was studied through diet incorporation assay. Flavonoids (rutin, chlorogenic acid, quinic acid, caffeic acid, naringenin, quercitin, kaempferol, myricetin, catechin, and ferulic acid) were incorporated in artificial diet at 100, 500 and 1000 ppm, lectins: groundnut leaf lectin (GLL), concavalin A (ConA) and phenyl β‐_D‐glucoside at 1, 2 and 5 μg/mL. Flavonoids such as rutin, quercitin and kaempferol at 1000 ppm were more toxic to S. litura larvae than quinic acid, caffeic acid, naringenin, myricetin, catechin, and ferulic acid. Larval growth and development were significantly reduced in S. litura larvae fed on a diet with GLL and ConA at 5 μg/mL compared to the larvae fed at 2 and 1 μg/mL concentrations. The larvae fed on flavonoid‐treated diets showed significant reduction in serine protease, trypsin and esterase activities. The flavonoids such as rutin, chlorogenic acid, quinic acid, naringenin, quercitin, kaempferol and myricetin, and lectins, GLL and ConA can be utilized in insect control programs.