Characterization of antixenosis to Dichelops melacanthus (Hemiptera: Pentatomidae) in soybean genotypes

Soybeans are of great importance in the world agricultural landscape, and their productive potential is significantly reduced by attacks from insect pests. Factors such as the expansion of national agricultural regions, together with no-tillage management and “off-season” maize cultivation, have favored the increase of secondary species such as Dichelops melacanthus (Dallas) (Hemiptera: Pentatomidae), intensifying the damage caused by the soybean stink bug complex. The use of resistant genotypes may be a valuable strategy as an alternative to the excessive use of chemical control in crops. This study evaluated the attractiveness and feeding preferences of the green-belly stink bug in 17 soybean genotypes in different maturity groups (early, semiearly, and late) to characterize the expression of antixenosis resistance. To this end, free-choice tests of attractiveness and food preference were performed under laboratory conditions. The early genotypes PI 171451 and D 75-10169, the semiearly genotypes IAC 78-2318, “IAC 100”, IAC 74-2832, PI 227687, and “IAC 24” and the late genotypes PI 274454, PI 274453, and L 1-1-01 expressed significant levels of antixenosis against adult D. melacanthus. These results will be useful for soybean breeding programs focusing on the stink bug resistance complex.     

The effects of diet on the detection of resistance to Cry1Ac toxin in Australian Helicoverpa armigera Hübner (Lepidoptera: Noctuidae)

The effects of raw or heat-denatured soybean flour in an artificial diet on the detection of Cry1Ac resistance in Helicoverpa armigera were examined. Resistant neonate larvae reared on denatured soybean flour diet showed resistance factors of 7980 and 16,901 at the LC₅₀ and LC_99.9 levels, respectively. By comparison, resistance could not be detected in neonate larvae reared on raw flour diet. Third instar larvae reared on denatured flour diet showed resistance factors of 322 and 21,190 at the LC₅₀ and LC_99.9 levels. Resistance was not detected in third instar larvae reared on raw flour diet. There was 68% survival of resistant neonate larvae on Bollgard II cotton leaf feeding assays, compared to 100% mortality in a susceptible strain. We conclude that detection of CRY1Ac resistance in H. armigera from Australia can be masked, if an artificial diet gives chronic exposure to potent, protease inhibitors present in raw soy flour.     

Effect of dihydrodillapiole on pyrethroid resistance associated esterase inhibition in an Indian population of Spodoptera litura (Fabricius)

Resistance in Spodoptera litura (Fabricius) has been attributed to enhanced detoxification of insecticides by increased levels of esterases, oxidases and/or glutathione S-transferases. Enzyme inhibiting insecticide synergists can be employed to counter increased levels of such enzymes in S. litura. Dihydrodillapiole induced synergism of pyrethroid toxicity was examined in the laboratory-reared third instar larval population of S. litura collected in Delhi (susceptible), and Guntur (resistant) region of Andhra Pradesh, India. The Guntur population was found to be 7.04 and 10.19 times resistant to cypermethrin and lambdacyhalothrin, respectively. The activity of cypermethrin, lambdacyhalothrin and profenophos against susceptible and resistance populations of S. litura, was gradually increased when used along with a plant-derived insecticide synergist dihydrodillapiole. The α-naphthyl acetate hydrolysable esterase activity in Delhi population was less as compared to the Guntur population. Resistance associated esterases in Delhi population were inhibited by pre-treatment with dihydrodillapiole. The esterase level in insect was instantly reduced initially, sustained for about 3 h and equilibrated at 4 h post treatment. The esterase activity of Guntur population was increased to 1.28 μmoles/mg/min at 2 h post treatment and subsequently reduced to lower than 0.70 μmoles at 4-12 h post treatment. The variation in esterase activity is suggestive of its homeostatic regulation in test populations. Dihydrodillapiole thus caused significant reduction of resistance in S. litura to cypermethrin, lambda cyhalothrin and profenophos.     

转基因棉和常规棉叶片对抗Cry1Ac近等基因系棉铃虫生长发育的影响

利用室内饲喂法,以抗Cry1Ac近等基因系棉铃虫为材料,比较转基因棉花33B和SGK321及其对照亲本DP5415和石远321对抗、感棉铃虫生长发育的影响。结果表明,抗性棉铃虫在取食常规棉叶后表现出一定的适合度代价。取食DP5415和石远321两种常规棉花后,抗性品系棉铃虫的幼虫存活率显著低于敏感品系,取食33B和SGK321两种转基因棉花的抗性棉铃虫,不仅其幼虫存活率显著高于敏感品系,而且致死中时间也比敏感品系延长。取食9天后,抗性品系在常规棉花石远321和DP5415上发育到3龄和4龄幼虫的比例显著低于敏感品系,取食33B和SGK321转基因棉花的抗性品系发育到3龄幼虫的比例均显著高于敏感品系。抗性品系在常规棉上的蛹重均显著低于敏感品系,部分取食转基因棉花的抗性品系棉铃虫可以化蛹,而敏感品系不能在转基因棉花上化蛹。     

Seasonality of the common cutworm Spodoptera litura in a soybean ecosystem

Seasonal occurrence of the common cutworm Spodoptera litura (Fab.) (Lepidoptera: Noctuidae) moths captured in synthetic sex pheromone traps and associated field population of eggs and larvae in soybean were examined in India from 2009 to 2011. Male moths of S. litura first appeared in late July or early August and continued through October. Peak male trap catches occurred during the second fortnight of September, which was within soybean reproductive stages. Similarly, the first appearance of S. litura egg masses and larval populations were observed after the first appearance of male moths in early to mid-August, and were present in the growing season up to late September to mid-October. The peak appearance of egg masses and larval populations always corresponded with the peak activity of male moths recorded during mid-September in all years. Correlation studies showed that weekly mean trap catches were linearly and positively correlated with egg masses and larval populations during the entire growing season of soybean. Seasonal means of male moth catches in pheromone traps during the 2010 and 2011 seasons were significantly lower than the catches during the 2009 season. However, seasonal means of the egg masses and larval populations were not significantly different between years. Pheromone traps may be useful indicators of the onset of numbers of S. litura eggs and larvae in soybean fields.     

Pyrethroid resistance and esterase activity in three strains of the cotton bollworm, Helicoverpa armigera (Hübner)

Microplate assay technique for estimation of esterase activity in a single insect was used in combination with dose mortality bioassays to detect pyrethroid resistance in three strains of Helicoverpa armigera and to study the frequency of pyrethroid resistant individuals within the population of the same strain at two larval stages, third and fifth instar. The third and fifth instar larvae of the field strains i.e., Nagpur strain and Delhi strain that displayed high degree of resistance towards deltamethrin, had higher esterase activity compared to a susceptible laboratory strain. The frequency distribution of individuals with elevated esterase activity above 1.00 absorbance unit was correlated with the resistance level of the strains. The frequency of resistant individuals in the third instar larvae of Nagpur strain and Delhi strain were 29% and 23%, respectively compared to 4% in the susceptible strain. The resistant individuals in the resistant strains have markedly increased in the fifth instar larvae with a frequency distribution of 63% and 90% in Delhi strain and Nagpur strain, respectively, while only 14% of individuals was found to have elevated esterase activity in the susceptible strain. The results demonstrated the role of esterase in pyrethroid resistance in H. armigera. Microplate assay proved to be a rapid and reliable biochemical technique for monitoring of pyrethroid resistance in H. armigera.     

Studies on the acetylcholinesterase of Anopheles albimanus resistant and susceptible to organophosphate and carbamate insecticides

Acetylcholinesterase from fourth instar Anopheles albimanus larvae was studied in vitro. The acetylcholinesterase from both the resistant and susceptible strains behaved as a single enzyme “type,” with straight pseudo first-order insecticide inhibition lines which intersected the Y axis at 100%. The enzyme from resistant larvae was more slowly inhibited than the susceptible enzyme; bimolecular rate constants (k_i) differed by approximately 1.2- to 6-fold for a range of organophosphorous compounds and 17- to 1570-fold for the carbamates. There was a good correlation between the levels of resistance and the acetylcholinesterase inhibition rates.     

荧光增白剂对Cry1Ac抗感棉铃虫围食膜的影响及增效作用

为了提高Bt的杀虫活性,运用扫描电镜、SDS-PAGE电泳和生物测定技术,研究了荧光增白剂FB28对棉铃虫Cry1Ac抗性和敏感品系围食膜的影响及对Cry1Ac杀虫蛋白的增效作用。结果表明,正常棉铃虫围食膜表面光滑致密,没有空洞和缝隙。用含1%FB28的人工饲料饲喂刚蜕皮的5龄棉铃虫幼虫2 h后,抗感棉铃虫围食膜表面均出现许多空洞,且敏感棉铃虫的空洞明显大于抗性棉铃虫;随着处理时间的延长,围食膜上的空洞逐渐变小,13 h后恢复正常。用1%FB28离体或活体处理抗、感品系棉铃虫,均可造成围食膜蛋白被降解。用1%FB28与Cry1Ac杀虫蛋白混合饲喂抗、感棉铃虫初孵幼虫,可明显提高棉铃虫的死亡率,且对抗性品系的增效作用明显大于敏感品系,对敏感和抗性品系棉铃虫的增效比分别为2.23和9.34;1%FB28还可以明显增加对抗、感品系3龄棉铃虫幼虫的生长抑制作用。     

甜菜夜蛾对氯虫苯甲酰胺抗性种群选育及鱼尼丁受体基因表达特征

为明确甜菜夜蛾Spodoptera exigua对氯虫苯甲酰胺的抗性发展及抗性种群中鱼尼丁受体(ryanodine receptor,RyR)基因的表达量变化,室内采用饲料混毒法进行甜菜夜蛾对氯虫苯甲酰胺的抗性选育,荧光定量PCR技术研究了抗性和敏感种群之间RyR基因mRNA表达量的差异。结果表明,室内选育31代后获得一个抗性倍数为105.60倍的甜菜夜蛾抗性种群,其mRNA表达量在甜菜夜蛾不同发育阶段及抗性种群和敏感种群之间均存在差异,以表达量最低的卵期作为对照,抗性种群中1龄幼虫表达量最高,是卵期的154.58倍;其次是雄性成虫,其表达量是卵期的101.51倍;2~5龄幼虫分别是卵期的59.56、35.35、72.99和19.84倍。抗性种群中1、2和4龄幼虫mRNA表达量分别是敏感种群的5.99、2.79和2.14倍,其余阶段低于敏感种群。表明甜菜夜蛾对氯虫苯甲酰胺的抗药性可能主要表现在幼虫阶段,RyR基因的表达量变化与氯虫苯甲酰胺诱导有关。     

Fitness cost, cross resistance and realized heritability of resistance to imidacloprid in Spodoptera litura (Lepidoptera: Noctuidae)

Imidacloprid has been used as a key insecticide for controlling sucking insect pests of cotton, whereas Spodoptera litura also has been indirectly exposed to this insecticide in Pakistan. To evaluate the risk of resistance evolution and to develop a better resistance management strategy, a field collected population was selected with imidacloprid in the laboratory. Thereafter, fitness cost, realized heritability and cross resistance of imidacloprid resistance in S. litura were investigated. After 14 generations of selection with imidacloprid, S. litura developed a 137.48-fold resistance to the insecticide. Bioassay revealed that this strain showed cross-resistance to acetamiprid (RR 8.52) and a little to lamdacyhalothrin (1.92) but negative cross-resistance was found to methomyl (−0.19). The resistant strain had a relative fitness of 0.38, with substantially lower rates of larval survival, larval duration, male pupal duration, development time, emergence rate of healthy adults, fecundity, hatchability, and prolonged larval and pupal duration. Mean relative growth rate of the larvae, intrinsic rate of population increase, and biotic potential was lower for the selected populations. The estimated realized heritability (h²) of imidacloprid resistance was 0.15 in the resistant strain of S. litura. Development of the resistance may cost significant fitness for the resistant population. This study provided valuable information for further understanding the impact of imidacloprid resistance on physiological parameters of S. litura and for facilitating the development of resistance management strategies.     

斜纹夜蛾核型多角体病毒对宿主实验种群增长的影响

在室内自然变温条件下,应用生命表方法评价了斜纹夜蛾核型多角体病毒对宿主实验种群增长的影响。结果表明,该病毒对斜纹夜蛾实验种群有明显的控制作用:①随着病毒接种浓度的升高,幼虫病死速率加快;②宿主感染病毒后,存活率、化蛹率降低,5、6龄幼虫历期明显延长,残存蛹羽化所得雌虫产卵量下降,当饲毒浓度达1.09×10~5PIBs/ml时,雌虫产卵量和产卵历期与对照差异显著(P<0.05),但病毒组各处理间无显著差异;③病毒能显著降低种群趋势指数(Ⅰ)值。各病毒组与对照的Ⅰ值差异极显著(P<0.01)。     

Mechanisms of resistance to the juvenoid methoprene in the house fly Musca domestica L

The mechanisms of resistance and cross resistance to the juvenoids methoprene and R-20458 in the house fly, Musca domestica, were examined. Radiolabeled methoprene was found to be metabolized faster in resistant and cross-resistant house fly larvae than in susceptible larvae, and methoprene and R-20458 penetrated more slowly into larvae of the resistant strain. In vivo and in vitro metabolism of methoprene was largely by oxidative pathways followed by conjugation in all strains examined, and little or no ester change of methoprene was noted in vitro. In vitro oxidative metabolism of methoprene, R-20458, juvenile hormone I, and several model substrates was higher in resistant and cross-resistant larvae than in susceptible larvae. Juvenoid functionalities susceptible to metabolic attack by resistant strains are indicated.     

Comparative toxicity,absorption, and metabolism of chlorpyrifos and its dimethyl homologue in methyl parathion-resistant and -susceptible tobacco budworms

Chlorpyrifos (Dowco 179) and its dimethyl homologue, chlorpyrifosmethyl (Dowco 214), were used to study the influence of the O,O-dialkyl group of organophosphorus insecticides on toxicity, absorption, and metabolism among larvae of the tobacco budworm [Heliothis virescens (F.)] from strains that were resistant (R) and susceptible (S) to methyl parathion. In toxicity tests, chlorpyrifos and chlorpyrifosmethyl were more toxic than methyl parathion to 3rd-stage R larvae but less toxic to S larvae. Chlorpyrifosmethyl was more toxic (3–4 ×) than chlorpyrifos to both strains of larvae, and the results of absorption studies indicated that the toxicity differential of the homologues may be explained in part by the more rapid absorption of the dimethyl form. Studies of the in vivo metabolism of both Dowco compounds indicated that each was degraded mainly by the cleavage of the pyridylphosphate linkage. In vitro tests demonstrated that the NADPH-dependent microsomal oxidases were of primary importance in detoxification, while glutathione (GSH)-dependent mechanisms (aryl- and alkyltransferases) present in the soluble cell fractions were of lesser importance. O-dealkylation occurred only with chlorpyrifosmethyl. The R larvae demonstrated greater capability in detoxifying both compounds in the comparative in vivo and in vitro studies of metabolism, but the differences were more apparent during the 5th instar than during the 3rd instar.     

Cry1Ac的绿色荧光蛋白GFP标记及其在棉铃虫幼虫中肠的定位分布

为了明确Cry1Ac蛋白在棉铃虫体内与中肠组织的相互作用,采用重叠PCR方法将Bt-cry1Ac基因和绿色荧光蛋白GFP基因融合,构建含Cry1Ac毒蛋白和绿色荧光蛋白GFP原核表达载体,并在大肠杆菌大量表达。利用荧光显微镜观察发现,表达Cry1Ac-GFP融合蛋白的大肠杆菌在蓝光激发下发出绿色荧光。将含有融合蛋白的菌液拌入人工饲料饲喂3龄棉铃虫幼虫96h,取棉铃虫幼虫中肠做冰冻切片并在荧光显微镜下观察。结果显示,取食含有Cry1Ac-GFP融合蛋白饲料的棉铃虫幼虫中肠能够发出强烈荧光。比较Cry1Ac杀虫蛋白敏感和抗性棉铃虫幼虫中肠的发光部位,敏感棉铃虫幼虫的中肠围食膜已经消失,肠壁细胞发出强烈的荧光,而抗性棉铃虫的围食膜较健全并发出荧光。     

大叶桃花心木的花、叶抽提物对斜纹夜蛾的拒食作用研究

测试了大叶桃花心木花和叶的乙醇抽提物对重要杂食性害虫斜纹夜蛾不同龄期幼虫的拒食活性。结果表明,大叶桃花心木花、叶抽提物50μg/mg浓度对斜纹夜蛾3龄、5龄试虫均具有较高的拒食作用,10μg/mg浓度对斜纹夜蛾3龄、5龄幼虫拒食作用较低,25μg/mg浓度对斜纹夜蛾3龄幼虫拒食作用高于5龄幼虫,且呈显著差异。大叶桃花心木花抽提物对斜纹夜蛾的拒食活性高于叶抽提物。     

Insecticidal potential of Bowman-Birk proteinase inhibitors from red gram (Cajanus cajan) and black gram (Vigna mungo) against lepidopteran insect pests

The present study assessed the insecticidal potential of purified Bowman-Birk proteinase inhibitors from red gram (RgPI) and black gram (BgPI) towards lepidopteran insect pests. Both RgPI and BgPI showed remarkable inhibitory activity against midgut trypsin-like proteinases of Achaea janata. While, BgPI showed moderate to low inhibitory activity towards the midgut trypsin-like proteinases of Helicoverpa armigera, Spodoptera litura, Papilio demoleus, Amsacta albistriga, Corcyra cephalonica, Bombyx mori and Daphnis nerii, RgPI showed low to negligible inhibitory activity. The isoinhibitors of RgPI and BgPI were stable in presence of trypsin and chymotrypsin of bovine pancreatic origin, as well as larval midgut proteinases of A. janata, H. armigera and S. litura. The midgut trypsin-like proteinases of A. janata and H. armigera as well as midgut trypsin- and chymotrypsin-like proteinases of S. litura were susceptible to inhibition by RgPI and BgPI. Feeding assays displayed a dose-dependent decrease in body weight and survival rate of these larvae with increasing concentration of inhibitors supplemented in the diet. RgPI displayed significant reduction in body weight and survival rate of A. janata larvae compared with BgPI. Conversely, BgPI showed significant reduction in body weight of S. litura larvae compared with RgPI.     

A new derivative of fipronil: Effect of adding a glycinyl group to the 5-amine of pyrazole on phloem mobility and insecticidal activity

Glycinyl fipronil, a new fipronil derivative with 5-amine acylated by glycine, was synthesized. The phloem mobility of the synthesized derivative was higher that of the parent compound in intact soybean seedling and its insecticidal activity against the 3rd instar larvae of Plutella xylostella was comparable with that of fipronil.     

The biochemistry of insecticide resistance in Anopheles sacharovi: Comparative studies with a range of insecticide susceptible and resistant Anopheles and Culex species

Fourth instar larvae, the progeny from wild-caught Anopheles sacharovi females, were subjected to a number of biochemical tests and the results were compared to those from similar tests on laboratory insecticide resistant and susceptible strains of anopheline and culicine mosquitoes. DDT resistance in An. sacharovi is associated with the ability to rapidly metabolise DDT to DDE. The organophosphorus and carbamate resistance was not associated with quantitative changes in esterases, multifunction oxidases, or glutathione S-transferase. The acetylcholinesterase was less sensitive to malaoxon and propoxur than laboratory susceptible An. albimanus, and plots of inhibition suggest that the population was polymorphic for more than one form of acetylcholinesterase. Metabolism studies on malathion and pirimiphos methyl did not indicate resistance due to increased metabolism. There was no evidence of penetration barriers contributing to resistance to either DDT or malathion, and there was no indication of any resistance to pirimiphos methyl in our tests.     

Synergist induced susceptibility of tobacco caterpillar, <Emphasis Type="Italic">Spodoptera litura</Emphasis> (Fabricius) from Kerala,India exposed to conventional insecticides

The present study was undertaken to assess the insecticide resistance developed in various field collected population of S. litura and to induce susceptibility by using the synergists. Third-instar larvae collected from three different locations of Kerala viz., Thiruvananthapuram (TVM), Pathanamthitta (PTA) and Alappuzha (ALP) were exposed to conventional insecticides like chlorpyriphos, quinalphos, lambda-cyhalothrin and cypermethrin by leaf dip bioassay and resistance ratios were calculated by using the baseline data generated for respective insecticides using susceptible strain. Resistance ratios recorded were 1965, 840 and 320 against chlorpyriphos, 605, 255 and 59 against quinalphos, 926, 250 and 108 against lambda-cyahlothrin and 2566, 534 and 396 against cypermethrin respectively for TVM, PTA and ALP populations. The effect of selected synergists viz., piperonyl butoxide (PBO), diethyl maleate (DEM) and triphenyl phosphate (TPP) was studied in combination with respective test insecticides against the highly resistant population of S.litura collected from TVM of Kerala. The population was tested with insecticide in combination of the above synergists at different ratios. When PBO, TPP and DEM at ratio of 1:4 were used the synergistic ratio was 8.47, 7.26 and 3.98 for chlorpyriphos, 6.09, 5.26 and 3.05 for quinalphos, 13.37, 4.53 and 7.39 for lambda cyhalothrin and 4.77, 3.36 and 3.40 for cypermethrin respectively. PBO showed highest synergistic activity against both the organophosphates tested followed by DEM and TPP. Highest synergistic activity against synthetic pyrethroids also was shown by PBO, followed by TPP and DEM. The results obtained from the present study revealed that PBO at 1:4 ratio showed higher synergism with the test insecticides against the resistant populations of S.litura and proved to be an effective molecule alternate for breaking the resistance against conventional organophosphates and synthetic pyrethroids.