Identification of a fluorescent compound in the cuticle of the train millipede Parafontaria laminata armigera

The train millipede (Parafontaria laminata armigera) emits a blue fluorescence (λ(max)=455 nm) under black light (350 nm). The isolated fluorescent compound from the cuticle of P. laminata armigera was identified as pterin-6-carboxylic acid. The structure of this compound was identified by fluorescent, HPLC, and mass spectrometric (ESI-ion trap MS) analyses, and then compared with an authentic sample.     

Characterization of dihydrolipoamide dehydrogenase from the mitochondria of Helicoverpa armigera,a pest resistant to insecticides

Dihydrolipoamide dehydrogenase (DHLDH) was isolated from the mitochondria of Helicoverpa armigera, a destructive pest which has developed resistance to commonly used insecticides. The flavoenzyme was purified 17.98‐fold to homogeneity with an overall yield of 10.53% by employing ammonium sulfate precipitation, hydroxylapatite chromatography and CM‐Sephadex chromatography. The purified enzyme exhibited the specific activity of 18.7 U/mg and was characterized as a dimer with a subunit mass of 66 kDa. The enzyme showed specificity for nicotinamide adenine dinucleotide – hydrogen (NADH) and lipoamide, as substrates, with Michaelis‐Menten constants (K_m) of 0.083 mmol/L and 0.4 mmol/L, respectively. The reduction reaction of lipoamide by the enzyme could be explained by ping‐pong mechanism. The spectra of DHLDH showed the maximum absorbance at 420 nm, 455 nm and 475 nm. The enzyme activity was strongly inhibited by mercurial and arsenical compounds. The N‐terminal sequence of Ha‐DHLDH showed homology with those of mammalian and arthropod DHLDH. Since H. armigera has developed high levels of resistance to commonly used insecticides, biochemical properties of the metabolic enzymes such as DHLDH, could be helpful to develop insecticidal molecules for the control of H. armigera, with a different mode of action.     

Systematic differences in the population density of green spruce aphid, Elatobium abietinum in a provenance trial of Sitka spruce, Picea sitchensis

Quantitative bioassay techniques were used to measure the susceptibility of Heliothis armigera to three nuclear polyhedrosis viruses (NPVs): H. armigera singly-enveloped NPV (HaSNPV), H. zea SNPV (HzSNPV) and H. armigera multiply-enveloped NPV (HaMNPV). Viruses were identified by EcoRI restriction endonuclease analysis. Electrophoretic profiles of DNA fragments revealed that the HaSNPV isolate was a previously undescribed genotypic variant. Bioassays with neonate and 6-day-old larvae measured small but significant differences in virulence between the three viruses. HzSNPV was the most virulent for neonate larvae with a median lethal dose (LD₅₀) of five polyhedra. HaMNPV was least virulent for 6-day-old larvae, with a LD₅₀ of 1400 polyhedra compared with 640–670 polyhedra for HaSNPV and HzSNPV. In addition, the median lethal time (LT₅₀) for infection with HaMNPV in neonate larvae was approximately 1·7 days longer than for the other viruses. Although they varied in virulence, each of the three viruses was sufficiently virulent to have considerable potential as a microbial control agent of H. armigera.     

Behavioral and electrophysiological responses of Helicoverpa assulta, H. armigera (Lepidoptera: Noctuidae), their F1 hybrids and backcross progenies to sex pheromone component blends

Two sibling species, Helicoverpa assulta and Helicoverpa armigera both use (Z)-9-hexadecenal and (Z)-11-hexadecenal as their sex pheromone components but in almost reversed ratios, 93:7 and 3:97, respectively. H. assulta and H. armigera males performed upwind flight in response to the H. assulta sex pheromone blend (93:7). H. armigera responded strongly to the H. armigera blend (3:97), whereas H. assulta males remained inactive upon exposure to this blend. Both species gave clear dose-dependent electrophysiological responses to (Z)-11-hexadecenal. However, (Z)-9-hexadecenal evoked strong dose-dependent electrophysiological responses in H. assulta males but not in H. armigera. The two male F₁ hybrids exhibited similar behavioral responses to two sex pheromone blends and electrophysiological responses to two pheromone components as H. armigera males. This indicated that H. armigera genes appear dominant in determining the behavioral response and electrophysiological responses. Behavioral and electrophysiological responses of backcrosses of male F₁ hybrids (H. armigera female × H. assulta male) with female H. assulta and H. armigera were close to that of H. assulta and H. armigera, respectively. However, backcrosses of female F₁ hybrids (H. assulta female × H. armigera male) with male H. assulta and H. armigera showed reduced behavioral responses but normal electrophysiological responses compared to males of the respective parental line.     

In silico exploration of inhibition potency of maslinic acid,a plant based triterpene,against Helicoverpa armigera serine protease

Gut proteases are accountable for survival of Helicoverpa armigera on protein rich parts of plant devastating many important agricultural crops. The aim of present study was to identify potential natural compounds having inhibitory potency against Helicoverpa armigera gut proteases. We have modeled structure of H. armigera serine protease (UniProt ID: O18447) and analyzed its interactions with maslinic acid (Zinc ID: ZINC38140521). A 3D model was generated using bovine trypsin in complex with analogues of sunflower inhibitor 1 as template with the help of Chimera Modeler 1.11. The PROCHECK and Modfold analysis have revealed 81.8% of residue in favored region. The POOL and COACH analysis have revealed 18 amino acids in the active site. In the 10 ns MD simulations of modeled structure, the RMSD of the protein backbone increased slightly and later stabilized from 7 ns to 10 ns. The modeled structure was stabilized at gyration distance of about 1.65 nm at 7 ns. Potential hit compounds from the ZINC database identified in this study showed good inhibitory bindings with modeled structure. Among these compounds maslinic acid, a plant based pentacyclic triterpenes was found to be potent lead compound with good binding affinity (−9.5 kcal/mol). RMSD profile was <0.45 nm for complex with stabilization at about 18,000 ps (18 nm) suggesting stable interaction. This work demonstrates reasonable in silico inhibitory action of maslinic acid against H. armigera serine protease and depicts utility of in silico methodologies for designing competent strategies against dreaded insect pests like H. armigera.     

不同饲喂下棉铃虫肠道内生菌的分离鉴定及脱毒效果研究

[目的]本研究对比饲喂正常饲料和棉粕饲料下棉铃虫(Helicoverpa armigera)肠道中可培养微生物的异同,并对筛选菌株进行棉酚耐受及降解作用的研究,为棉铃虫肠道微生物在棉源饲料中的脱毒作用提供理论和实验支持.[方法]通过饲喂不同饲料、醋酸棉酚单一碳源微生物选择性培养,分离饲喂正常饲料和棉粕饲料下棉铃虫肠道可...     

Impact of some insecticides and their mixtures on the population of tomato borers,Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae) and Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) in tomato crop at Upper Egypt

Impact of some insecticides and their mixtures (Emamectin benzoate, Imidaclopride, Chlorfenapyr, Indoxacarb, Profenofos, Pyridalyl, Methomyl and Teflubenzuron) were evaluated against the tomato borers, Tuta absoluta (Meyrick) and Helicoverpa armigera (Hübner) population in tomato crop at Upper Egypt. Results of LC₅₀ values indicated that Emamectin benzoate was the most toxic compound (LC₅₀ 0.461%) against T. absoluta, larvae. The carbamate insecticide, Methomyl, followed this descending order, showing almost similar toxicity, while Teflubenzuron and Chlorfenapyr were less effective (LC₅₀ 1.054 and 3.165%, respectively). Pyridalyl was found to be the most effective insecticide against larval stage of H. armigera. (LC₅₀ 0.513%). The corresponding toxicities of the other tested insecticides, arranged according to their LC₅₀ values in descending order were as follow: Methomyl, Emamectin benzoate, Profenofos, Imidaclopride, Teflubenzuron, Indoxacarb and Chlorfenapyr. Their LC₅₀ values were ranged between (0.513 and 0.872%). While, their toxicity indexes were ranged between (92.432 and 58.830%, respectively). On the other hand, there were no differences occurred among the treatments. Mean of percent infestation was decreased six weeks after six sprayings, and the per cent reduction in infestation was 79.73, 80.22, 78.41, 80.88, 80.50, 78.30, 79.64 and 78.25% using Emamectin benzoate, Methomyl, Imidaclopride, Pyridalyl, Profenofos, Indoxacarb, Teflubenzuron and Chlorfenapyr, respectively. The efficiency of the tested insecticides was increased with increasing the number of sprays from two to six causing reduction in insect borer infestation ranged between 68.02 and 80.88%, respectively. Generally, the tested insecticides and their mixtures achieved a considerable reduction in T. absoluta and H. armigera population.     

Binding of Cry δ-endotoxins to brush border membrane vesicles of Helicoverpa armigera and Helicoverpa punctigera （Lepidoptera： Noctuidae）

The relatively low susceptibility ofHelicoverpa armigera to CrylAc, its history of resistance to chemical insecticides and the seasonal decline in expression of CrylAc in transgenic cotton necessitated the development of cotton expressing two insecticidal proteins to provide sustainable control of this multinational pest. To manage the resistance issue, it was essential that the second insecticidal protein have a significantly different mode of action to CrylAc. A common feature of resistance to CrylA proteins in several species as well as H. armigera has been a change in the binding site. A study of binding sites for some Cry proteins in the brush border membrane vesicles （BBMV） ofH. armigera and Helicoverpa punctigera was undertaken. The binding affinity for CrylAc was higher than for CrylAb, matching their relative toxicities, and CrylAc and CrylAb were found to share at least one binding site in both I-1. armigera and I-1. punctigera. However Cry2Aa did not compete with CrylAc for binding and so could be used in transgenic cotton in combination with CrylAc to control H. armigera and manage resistance. Variation in the susceptibilities of three different H. armigera strains to CrylAc correlated with the parameter Bmax/Kcom.     

Bioefficacy and mode of action of rocaglamide from Aglaia elaeagnoidea (syn. A. roxburghiana) against gram pod borer,Helicoverpa armigera (Hübner)

Abstract: Rocaglamide, a highly substituted benzofuran, was isolated and identified as the main biologically active component in Aglaia elaeagnoidea (syn. A. roxburghiana) for gram pod borer Helicoverpa armigera (Hübner). Addition of rocaglamide to an artificial diet retarded the growth of neonate larvae in a dose‐dependent manner with EC₅₀ values of 0.76 p.p.m. These values compared favourably with azadirachtin (EC₅₀ = 0.23 p.p.m.). However, azadirachtin was apparently more potent than rocaglamide in inducing growth inhibition via oral administration to these first stadium larvae. The candidate compound was found to have LD₅₀ and LD₉₅ values of 0.40 and 1.02 μg per larva, respectively, in topical application against third instar larvae 96 h post‐treatment. However, these values for azadirachtin were 8.16 and 25.8 μg per larva for the same period. This shows that azadirachtin was less effective against third instar H. armigera larvae in inducing acute toxicity via topical treatment in comparison with rocaglamide. However, severe morphological larval deformities were observed in such azadirachtin‐treated larvae during the process of ecdysis. The cytotoxic nature of rocaglamide was established by evaluating dietary utilization and the results did not implicate any antifeedant effect but the toxicity‐mediated effect due to reduced efficiency of conversion of ingested food. It was obvious that feeding deterrence is not the primary mode of action but a centrally mediated effect, which could be due to the induced cytotoxicity at non‐specific cellular levels.     

Increased food utilization indices and decreased proteolytic activity in Helicoverpa armigera larvae fed sublethal Bacillus thuringiensis‐treated diet

The cotton bollworm Helicoverpa armigera is one of the most devastating insect pests. A set of protease enzymes allows this species to feed on different host plant species. Control measures in agriculture often involve the application of the pathogenic bacterium Bacillus thuringiensis subsp. kurstaki (Btk). In the present study, the effects of sublethal Btk doses are evaluated with respect to the food utilization indices and proteolytic activities of Helicoverpa armigera. Accordingly, the H. armigera larvae are fed with artificial diet containing sublethal Btk doses (LC₅, LC₁₀, LC₁₅, LC₂₀ and LC₂₅) and a Btk‐free diet as control. All but one of the food utilization indices we measured is observed to increase significantly with increasing Btk doses. By contrast, the specific activity of total protease, chymotrypsin and elastase enzymes decrease significantly with an increasing Btk concentration. We conclude that Btk was not toxic to H. armigera larvae and any damage that it causes can be compensated for by H. armigera larvae via various mechanisms. In conclusion, increased nutritional indices in the larvae fed with Btk diet represent an important issue that needs to be considered to avoid the pest establishing Bt resistant populations. Meanwhile, the lack of effect of Btk sublethal concentrations on trypsin enzyme specific activity can bolster this challenge.     

A UV Tolerant Mutant of Bacillus thuringiensis subsp. kurstaki Producing Melanin

A UV-resistant mutant (Bt-m) of Bacillus thuringiensis subsp. kurstaki, producing a dark brown pigment, identified as melanin, was studied. Bt-m had higher larvicidity against Heliothis armigera than its parent. Survival of Bt-m spores and their insecticidal activity to irradiation at 254 nm and 366 nm were higher than those of the parent. The only toxic polypeptide produced by Bt-m was Cry1Ac (130 kDa); it lost cry1Aa, cry2Aa, and cry2Ab. Received: 2 April 2001 / Accepted: 14 May 2001     

Effect of repeated in vitro sub-culturing on the virulence of Metarhizium anisopliae against Helicoverpa armigera (Lepidoptera: Noctuidae)

The effect of repeated conidial sub-culturing of Metarhizium anisopliae on its virulence against Helicoverpa armigera (Hübner) was studied. The LT₅₀ observed against third instar larvae of H. armigera for the first sub-culture was 3.4 days; it increased to 4.5 and 5.6 days for the 20th and the 40th sub-cultures, respectively. The LT₅₀ values after passage of the 40th sub-culture on H. armigera decreased to 4.4 and 3.7 days for the 40th (first in vivo) and the 40th (fifth in vivo) passages, respectively. Similarly, the LC₅₀ of M. anisopliae towards third instar larvae of H. armigera increased from the first sub-culture (0.17×10⁴) to (3.0×10⁴) for the 40th conidial transfers on potato dextrose agar and again decreased to 0.74×10⁴ and 0.23×10⁴ in the 40th (first in vivo) and the 40th (fifth in vivo) passage, respectively. Similar trends for LC₅₀ and LT₅₀ values were seen when sugarcane woolly aphid, Ceratovacuna lanigera Zehntner was used as a host. Significant variation in appressorium formation and cuticle-degrading enzyme production such as chitinase, chitin deacetylase, chitosanase and protease during subsequent sub-culturing and passage through H. armigera was observed. Though there was no effect on internal transcribed spacer (ITS) sequence pattern, interestingly, in randomly amplified polymorphic DNA (RAPD), significant differences in the band intensities and in the banding pattern for different sub-cultures of M. anisopliae were observed. As stable virulence towards the insect pest is desirable for commercialisation of a mycoinsecticide, such changes in virulence due to repeated in vitro transfer need to be monitored and minimised.     

P-glycoprotein ATPase from the resistant pest, Helicoverpa armigera: Purification, characterization and effect of various insecticides on its transport function

Helicoverpa armigera is a major pest of agricultural crops and has developed resistance to various insecticides. A P-glycoprotein (Pgp) with ATPase activity likely to be involved in insecticide resistance was purified and characterized from insecticide-resistant H. armigera. The purification was 18-fold with 3% yield. The optimum pH and temperature were found to be 7.4 and 30-40 °C, respectively. Kinetic studies indicated that this enzyme had a K_m value of 1.2 mM for ATP. Pgp from H. armigera was partially sequenced and found to be homologous to conserved sequences of mammalian Pgps. Pesticides stimulated H. armigera Pgp ATPase activity with a maximum stimulation of up to 40%. Quenching of the intrinsic tryptophan fluorescence of purified Pgp was used to quantitate insecticide binding. Using the high-affinity fluorescent substrate, tetramethylrosamine, transport was monitored in real time in proteoliposomes containing H. armigera Pgp. The presence of Pgp could be one of the reasons for insecticide resistance in this pest.     

Monitoring of four major lepidopteran pests in Korean cornfields and management of Helicoverpa armigera

Lepidopterans such as Helicoverpa armigera are emerging pests of corn in Korea, causing huge yield losses and deteriorating the quality of the corn crop. We monitored four major lepidopteran pests in major corn growing areas of Korea by employing sex pheromone traps from 2012 to 2015 to understand population dynamics. H. armigera, Ostrinia furnacalis, Spodoptera litura, Spodoptera exigua and Mythimna separata were collected from April to October. Among these species, H. armigera was found to be the main pest based on crop damage intensity (80–90% of the total damage). H. armigera was generally observed during the earing stage of corn and would migrate to other fields during September to October for overwintering. We also tested select insecticides against larvae of H. armigera in the laboratory. The most effective insecticide was indoxacarb, which was applied at specific times to manage H. armigera in cornfields. Optimal timing of spraying was estimated at about 70 days after planting (earing season) to control for H. armigera. Additionally, two species of parasitoid (Therion circumflexum and Ophioninae sp.) were identified from H. armigera. However, more extensive surveys are needed to organize a control program based on natural enemies.     

Host crop influence on the susceptibility of the American bollworm, Helicoverpa armigera, to Bacillus thuringiensis ssp. kurstaki HD-73

Studies on the susceptibility of F₁ neonates of Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) collected from chickpea in Delhi and cotton in Punjab, Haryana and Rajasthan in northern India, to Bacillus thuringiensis ssp. kurstaki HD‐73, and the impact of host crop diets on insect susceptibility, were carried out by diet incorporation bioassays. The susceptibility of F₁ neonates of H. armigera to Bacillus thuringiensis ssp. kurstaki HD‐73 ranged from twofold (LC₅₀ 96 h, 84.5–164.2 µg (ai) l^?1) for chickpea to about fivefold (LC₅₀ 96 h, 51.1–247.7 µg (ai) l^?1) for cotton. The F₁ neonates of insects collected from pearl millet were twice as tolerant as those collected from cotton and sunflower at Sirsa to B. thuringiensis ssp. kurstaki HD‐73, suggesting that there was an influence of host crops on insect susceptibility. Insects originally collected from cotton fields at Bhatinda and reared for four generations on a chickpea‐based meridic diet were used to initiate host‐specific colonies of H. armigera. These host‐specific colonies were allowed to complete one generation on meridic diets prepared with different hosts, viz., cabbage, cauliflower, chickpea, green pea, pearl millet, and pigeon pea. Larvae of H. armigera were heaviest on the 15th day, and had a higher growth rate on a pigeon pea‐based diet than all other host diets. The larval period was shorter on chickpea and pigeon pea, with higher percentage pupation than all other host‐diets. The pupal weight of H. armigera was greater on chickpea and pigeon pea diets than on other host diets. The growth and development of larvae was significantly poorer on pearl millet diet than on other host diets. The F₁ neonates of H. armigera belonging to cabbage, cauliflower, and pearl millet host‐specific colonies were more susceptible than those belonging to chickpea, green pea, and pigeon pea host‐specific colonies to B. thuringiensis ssp. kurstaki HD‐73, suggesting the importance of proteinaceous nutrients in tolerance. The F₁ neonates of the pearl millet colony of H. armigera grown on a chickpea‐diet for 4 days were significantly more tolerant to B. thuringiensis ssp. kurstaki HD‐73 than those reared on the pearl millet‐based diet. These studies show the impact of the host diet of H. armigera on tolerance to B. thuringiensis.     

Acute toxicity and sub-lethal effects of a pyrethroid (cypermethrin) on survival,development and fitness of Helicoverpa armigera

The chickpea pod borer, Helicoverpa armigera (Hübner) is one of the most destructive pests of India and most of the countries. In this experiment, sub-lethal effects of cypermethrin were evaluated against sixth instar larvae of this pest that had ingested insecticide-treated chickpea pods. The LC₅₀, LC₃₀ and LC₁₀ values of cypermethrin were 80.38, 26.65 and 5.41?mg a.i.?L^?1, respectively, against sixth instar. The sub-lethal effects on fitness, eggs, larvae, pupae, adult longevity and reproduction were observed in H. armigera population that survived exposure to the sub-lethal doses of LC₅₀, LC₃₀ and LC₁₀ of cypermethrin. Survivorship was reduced to 36?days if H. armigera exposed to the sub-lethal dose of LC_50, whereas it was 42?days in unexposed groups. Larvae treated with LC₅₀ dose exhibit lowest intrinsic rate of increase (r_m) (0.0413 females/female/day) and highest (0.0517) with the unexposed group. Time taken by H. armigera to complete one generation (T_c) was 40.09?days, while this time significantly lowered to 32.85?days when exposed to the sub-lethal dose of LC₃₀. Hatching eggs were greatly reduced in the larvae exposed to cypermethrin at all the sub-lethal doses. About 430 eggs/female/generation were recorded after treatment with 80.38?mg/L as compared with untreated females (1390). Larvae that were exposed to sub-lethal concentrations of cypermethrin exhibited lower pupal weight and prolonged pupal developmental times compared control larvae. The overall fitness performance was better in the unexposed population as compared to the population exposed to sub-lethal doses.     

Preparation of cyanobacterial peptide toxin as a biopesticide against cotton pests

Glycine-rich peptide toxin of cyanobacterium Scytonema MKU 106 was purified. UV spectral analysis showed an absorption maximum at 228 nm and the molecular mass was less than 12 kDa. The mortality rate of American boll worms (Helicoverpa armigera) was about 80% and 40% 84 h after treatment with 0.001% crude and purified peptide toxins respectively; 100% mortality was observed after 108 h treatment with both purified and crude peptide toxins. The LC₅₀ (lethal concentration to 50% of the population) for Heliothis larvae after 96 h was 8.3 μg/ml purified peptide toxin and 6.2 μg/ml crude peptide toxin. Observations also show that the peptide toxin at 0.01% concentration acts as a biopesticide and at high (0.1%) concentrations it will act as an anti-feeding compound for Stylepta derogata (leaf-roller) larvae of the cotton crop. Received: 22 May 1996 / Accepted: 8 July 1996     

Suspension concentrate formulation of Bacillus thuringiensis var. kurstaki for effective management of Helicoverpa armigera on sunflower (Helianthus annuus)

A local strain DOR Bt-1 belonging to Bacillus thuringiensis var. kurstaki (Bt.k) was multiplied through solid state fermentation and the resulting technical powder was milled and sieved to obtain particles of various sizes. Efficacy of Bt.k. against larvae of Helicoverpa armigera was found to increase with the decrease in particle size. Boric acid was found to be synergistic to DOR Bt-1 technical powder. LC₅₀ of the Bt and boric acid mixture (75:25) was lower at 89.63 mg/100 mL in comparison to Bt alone at 116.75 mg/100 mL. A suspension concentrate formulation with DOR Bt-1 technical was developed using boric acid as an adjuvant. The formulation was found to be highly effective against H. armigera in laboratory bioassays with an LC₅₀ of 185 µL (containing 53.36 mg Bt). The formulation gave effective control of H. armigera on sunflower within 3 days after spray even at the lowest dose of 1.0 mL/L under field conditions.     

The inhibition of molting fluid enzymes of Pod borer,Helicoverpa armigera (Hubner) by an aspartic protease inhibitor: Its effects on insect development

In this study, the inhibition of molting fluid enzymes from Helicoverpa armigera by an aspartic protease inhibitor, Alkalophilic Thermophilic Bacillus Inhibitor (ATBI) purified from Bacillus sp. is reported. The in vitro experiments showed 80% inhibition (IC₅₀= 48 µM) of hemoglobin hydrolyzing and 95% inhibition (IC₅₀= 35 µM) of chitin hydrolyzing activity from molting fluid by ATBI (IC₅₀ value is the ATBI concentration for 50% inhibition of total enzymatic activity). The treatment of H. armigera larvae with 400 µM ATBI recorded 20% larval mortality, 27.77% deformed pupae and 12.22% deformed adults. The LC₅₀ value (Concentration of ATBI calculated to give 50% mortality) calculated for insect population was found to be 330.06 µM. Similarly, significant variations in mean larval and pupal weight, no. of eggs laid per female and percent hatching of eggs were observed at higher concentrations of ATBI. The results may provide the basis for the selection of non-host inhibitors to develop a H. armigera insecticide formulation.