In vivo inhibition of Helicoverpa armigera gut pro-proteinase activation by non-host plant protease inhibitors

We evaluated 22 different host and non-host plant protease inhibitors (PIs) for in vivo inhibition of Helicoverpa armigera gut pro- and proteinases, and their biological activity against the pod borer, H. armigera, the most important pest of agriculture and horticultural crops worldwide. In vitro activation of H. armigera gut pro-proteinases (HaGPPs) in larvae fed on non-host plant PIs showed significant in vivo inhibition of HaGPPs activation in solution as well as in gel assays. The larvae fed on diet incorporated with Datura alba ness PIs showed highest inhibition of HaGPPs, followed by Psophocarpus tetragonolobus. Non-host plant PIs from Pongamia pinnata, Mucuna pruriens, Capsicum annuum, and Nigela sativa showed maximum inhibitory potential towards HaGPs in vivo, and also exhibited moderate level of inhibition of pro-proteinases. However, some of non-host plant PIs, such as those from Penganum harmala and Solanum nigrum, and the principal host plant PIs, viz., Cicer arietinum and Cajanus cajan did not inhibit HaGPP activity. Pro-proteinase level increased with the growth of the larvae, and maximum HaGPP activity was observed in the fifth-instars. Larvae fed on diets with D. alba ness PIs showed greater inhibition of HaGPPs as compared to the larvae fed on diets with P. tetragonolobus. Low concentrations of partially purified HaGPs treated with gut extract of larvae fed on D. alba ness showed that out of 10 proteinase isoforms, HaGPs 5 and 9 were activators of pro-proteinases. Larval growth and development were significantly reduced in the larvae fed on the non-host plant PIs, of which D. alba ness resulted in highest stunted growth of H. armigera larvae. The in vivo studies indicated that non-host plant PIs were good candidates as inhibitors of the HaGPs as well as HaGPPs. The PIs from the non-host plants can be expressed in genetically engineered plants to confer resistance to H. armigera.     

Characterization of the Mamestra configurata (Lepidoptera: Noctuidae) larval midgut protease complement and adaptation to feeding on artificial diet,Brassica species,and protease inhibitor

The midgut protease profiles from 5th instar Mamestra configurata larvae fed various diets (standard artificial diet, low protein diet, low protein diet with soybean trypsin inhibitor [SBTI], or Brassica napus) were characterized by one‐dimensional enzymography in gelatin gels. The gut protease profile of larvae fed B. napus possessed protease activities of molecular masses of approximately 33 and 55 kDa, which were not present in the guts of larvae fed artificial diet. Similarly, larvae fed artificial diet had protease activities of molecular masses of approximately 21, 30, and 100 kDa that were absent in larvae fed B. napus. Protease profiles changed within 12 to 24 h after switching larvae from artificial diet to plant diet and vice versa. The gut protease profiles from larvae fed various other brassicaceous species and lines having different secondary metabolite profiles did not differ despite significant differences in larval growth rates on the different host plants. Genes encoding putative digestive proteolytic enzymes, including four carboxypeptidases, five aminopeptidases, and 48 serine proteases, were identified in cDNA libraries from 4th instar M. configurata midgut tissue. Many of the protease‐encoding genes were expressed at similar levels on all diets; however, three chymoptrypsin‐like genes (McSP23, McSP27, and McSP37) were expressed at much higher levels on standard artificial diet and diet containing SBTI as was the trypsin‐like gene McSP34. The expression of the trypsin‐like gene McSP50 was highest on B. napus. The adaptation of M. configurata digestive biochemistry to different diets is discussed in the context of the flexibility of polyphagous insects to changing diet sources. Published 2010 Wiley Periodicals, Inc.     

Compensatory proteolytic responses to dietary proteinase inhibitors from Albizia lebbeck seeds in the Helicoverpa armigera larvae

Plant proteinase inhibitors (PIs) have been shown to reduce the growth rates in larvae of numerous insect species. On the other hand, insects can also regulate their proteinases against plant PIs. In the present study, we report the compensatory activities of Helicoverpa armigera (Hubner) (Lepidoptera: Noctuidae) gut proteinases against the PIs of Albizia lebbeck seeds. Total of ten proteinase inhibitor bands were detected in the seed extract of A. lebbeck. Bioassays were conducted by feeding H. armigera larvae on diet containing partially purified PIs from A. lebbeck seeds. Results show that larval growth and survival was significantly reduced by A. lebbeck PIs. We found that higher activity H. armigera gut proteinase (HGP) isoforms observed in the midgut of control larvae were inhibited in the midgut of larvae fed on test diet. Some HGP isoforms were induced in the larvae fed on PI containing test diet; however, these isoforms showed lower activity in the larvae fed on control diet. Aminopeptidase activities were significantly increased in the midgut of larvae fed on test diet. A population of susceptible and resistant enzymes was observed in the midgut of H. armigera, when fed on diet containing PIs from A. lebbeck seeds. Our initial observations indicate that H. armigera can regulate its digestive proteinase activity against non-host plant PIs, too. It is important to study the exact biochemical and molecular mechanisms underlying this phenomenon in order to develop PI-based insect control strategies.     

Effects of the mixed biocide Bacillus thuringiensis–abamectin on the development of the parasitoid Microplitis mediator and its host Helicoverpa armigera

The effects of the mixed biocide Bacillus thuringiensis Berliner with abamectin (BtA) on the development of the parasitoid Microplitis mediator (Haliday) (Hymenoptera: Braconidae) and its cotton bollworm host, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae), were evaluated in the laboratory. Weight gain in larvae of H. armigera was initially delayed, but larval developmental period increased and pupal weight increased when they were fed on a diet containing BtA. Due to increased longevity of the host larvae, the susceptible period to parasitization of H. armigera by M. mediator increased when the host larvae were reared on diets containing BtA at concentrations of 0.5, 1, 2, and 4 μg g^?1. The longevity of female and male parasitoids significantly decreased when newly emerged wasps were fed a honey solution containing 200 μg ml^?1 BtA in comparison with those fed only a honey solution. Mean longevity was significantly prolonged when parasitoids were fed a honey solution and BtA–honey solution in comparison with those fed BtA–distilled water, distilled water, or nothing. There were no significant differences compared with the control in any biological characteristics for the offspring of female parasitoids fed the honey solutions containing BtA at concentrations of 50, 100, and 200 μg ml^?1; characteristics measured include the egg‐larval period, pupal weight, male and female pupal periods, adult fresh weight, and adult longevity. When female parasitoids parasitized host larvae that had been fed the diet containing BtA, their male and female pupal periods were significantly prolonged compared with the control (without BtA).     

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.     

Effects of class-specific,synthetic, and natural proteinase inhibitors on life-history traits of the cotton bollworm Helicoverpa armigera

Herbivorous insects have more difficulty obtaining proteins from their food than do predators and parasites. The scarcity of proteins in their diet requires herbivores to feed voraciously, thus heavily damaging their host plants. Plants respond to herbivory by producing defense compounds, which reduce insect growth, retard development, and increase mortality. Herbivores use both pre- and postdigestive response mechanisms to detect and avoid plant defense compounds. Proteinase inhibitors (PIs) are one example of plant compounds produced as a direct defense against herbivory. Many insects can adapt to PIs when these are incorporated into artificial diets. However, little is known about the effect of PIs on diet choice and feeding behavior. We monitored the diet choice, life-history traits, and gut proteinase activity of Helicoverpa armigera larvae using diets supplemented with synthetic and natural PIs. In choice experiments, both neonates and fourth-instar larvae preferred the control diet over PI-supplemented diets, to varying degrees. Larvae that fed on PI-supplemented diets weighed less than those that fed on the control diet and produced smaller pupae. Trypsin-specific PIs had a stronger effect on mean larval weight than did other PIs. A reduction of trypsin activity but not of chymotrypsin activity was observed in larvae fed on PI-supplemented diets. Therefore, behavioral avoidance of feeding on plant parts high in PIs could be an adaptation to minimize the impact of this plant's defensive strategy.     

Responses of midgut amylases of Helicoverpa armigera to feeding on various host plants

Midgut digestive amylases and proteinases of Helicoverpa armigera, a polyphagous and devastating insect pest of economic importance have been studied. We also identified the potential of a sorghum amylase inhibitor against H. armigera midgut amylase. Amylase activities were detected in all the larval instars, pupae, moths and eggs; early instars had lower amylase levels which steadily increased up to the sixth larval instar. Qualitative and quantitative differences in midgut amylases of H. armigera upon feeding on natural and artificial diets were evident. Natural diets were categorized as one or more members of legumes, vegetables, flowers and cereals belonging to different plant families. Amylase activity and isoform patterns varied depending on host plant and/or artificial diet. Artificial diet-fed H. armigera larvae had comparatively high amylase activity and several unique amylase isoforms. Correlation of amylase and proteinase activities of H. armigera with the protein and carbohydrate content of various diets suggested that H. armigera regulates the levels of these digestive enzymes in response to macromolecular composition of the diet. These adjustments in the digestive enzymes of H. armigera may be to obtain better nourishment from the diet and avoid toxicity due to nutritional imbalance. H. armigera, a generalist feeder experiences a great degree of nutritional heterogeneity in its diet. An investigation of the differences in enzyme levels in response to macronutrient balance and imbalance highlight their importance in insect nutrition.     

Effects of the Cry1Ac toxin of Bacillus thuringiensis on Microplitis mediator, a parasitoid of the cotton bollworm, Helicoverpa armigera

Interactions between the cotton bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae), its larval parasitoid Microplitis mediator (Haliday) (Hymenoptera: Braconidae), and the Cry1Ac toxin of Bacillus thuringiensis Berliner were evaluated under laboratory conditions. The growth of H. armigera larvae was delayed and its pupal rate and pupal weight decreased when they were fed on a diet containing Cry1Ac toxin. Due to the lowered growth rate of the host larvae, the time available for parasitization of H. armigera by M. mediator increased when the host larvae were reared on a diet containing Cry1Ac toxin at concentrations of 0.5, 1, 2, and 4 µg g^?1. The longevity of female and male parasitoids was not significantly affected when newly emerging wasps fed on honey solutions containing three different concentrations of Cry1Ac toxin (125, 250, and 500 µg ml^?1). When female parasitoids were fed on honey solutions containing Cry1Ac, their offsprings’ egg and larval development period, pupal weight, length of pupation, adult weight, and adult longevity did not change significantly in most of the treatments compared with controls. When the female parasitoids parasitized host larvae that had been fed on a diet containing 0.5, 1, 2, 4, and 8 µg g^?1 Cry1Ac toxin, their offsprings’ eggs and larvae were significantly delayed. Their pupal weight, adult weight, and adult longevity were also significantly less than controls.     

取食含不同植物源成分的饲料对香樟齿喙象幼虫生长发育及体内解毒酶活性的影响

【目的】明确取食含不同植物源成分的饲料对香樟齿喙象Pagiophloeus tsushimanus幼虫生长发育以及体内解毒酶活性的影响,以探究寄主植物对香樟齿喙象食性与抗性的作用。【方法】以3种樟科植物(香樟Cinnamomum camphora和浙江桂Cinnamomum chekiangensis和浙江楠Phoebe chekiangensis)枝条粉末为植物源成分,配制成半人工饲料。以半人工饲料进行室内饲养后,分别测定取食半人工饲料后香樟齿喙象幼虫的发育历期、体重、死亡率和化蛹率,以及饥饿12 h后取食含不同植物源成分的半人工饲料的4龄幼虫体内解毒酶(谷胱甘肽-S-转移酶、羧酸酯酶和乙酰胆碱酯酶)活性时序变化。【结果】香樟齿喙象幼虫的发育历期在取食含不同植物源成分的半人工饲料之间存在显著差异。取食含香樟成分的半人工饲料的幼虫发育历期最短(100.13 d),取食含浙江楠成分的半人工饲料的幼虫发育历期最长 (123.33 d);整个幼虫期的死亡率以取食含浙江桂成分的半人工饲料的幼虫最高(41.67%),以取食含香樟成分的半人工饲料的幼虫最低(10.00%);化蛹率以取食含香樟成分的半人工饲料的最高(87.32%);取食含香樟成分的半人工饲料的幼虫体重增长速率也明显高于取食含浙江桂或浙江楠成分的半人工饲料的幼虫。饥饿12 h后取食含不同植物源成分的半人工饲料的香樟齿喙象4龄幼虫体内GST和 AChE活性并无显著差异(P＞0.05);而取食含香樟成分的半人工饲料的幼虫CarE活性在60 h时大幅度上升。【结论】说明香樟齿喙象对天然寄主香樟具有较强的生理适应性,这可能与其专食性密切相关;同时,也表明香樟齿喙象可以通过调节自身生长发育和解毒代谢功能来适应不同寄主植物。     

Maintenance of narrow diet breadth in the monarch butterfly caterpillar: response to various plant species and chemicals

In order to better understand the maintenance of a fairly narrow diet breadth in monarch butterfly larvae, Danaus plexippus L. (Lepidoptera: Nymphalidae: Danainae), we measured feeding preference and survival on host and non-host plant species, and sensitivity to host and non-host plant chemicals. For the plant species tested, a hierarchy of feeding preferences was observed; only plants from the Asclepiadaceae were more or equally preferred to Asclepias curassavica, the common control. The feeding preferences among plant species within the Asclepiadaceae are similar to published mean cardenolide concentrations. However, since cardenolide data were not collected from individual plants tested, definitive conclusions regarding cardenolide concentrations and plant acceptability cannot be made. Although several non-Asclepiadaceae were eaten in small quantities, all were less preferred to A. curassavica. Additionally, these non-Asclepiadaceae do not support continued feeding, development, and survival of first and fifth-instar larvae. Preference for a host versus a non-host (A. curassavica versus Vinca rosea) increased for A. curassavica reared larvae as compared to diet-reared larvae suggesting plasticity in larval food preferences. Furthermore, host species were significantly preferred over non-host plant species in bioassays using a host plant or sucrose as a common control. Larval responses to pure chemicals were examined in order to determine if host and non-host chemicals stimulate or deter feeding in monarch larvae. We found that larvae were stimulated to feed by some ubiquitous plant chemicals, such as sucrose, inositol, and rutin. In contrast, several non-host plant chemicals deterred feeding: caffeine, apocynin, gossypol, tomatine, atropine, quercitrin, and sinigrin. Additionally the cardenolides digitoxin and ouabain, which are not in milkweed plants, were neutral in their influence on feeding. Another non-milkweed cardenolide, cymarin, significantly deterred feeding. Extracts of A. curassavica leaves were tested in bioassays to determine which components of the leaf stimulate feeding. Both an ethanol extract of whole leaves and a hexane leaf-surface extract are phagostimulatory, suggesting the involvement of both polar and non-polar plant compounds. These data suggest that the host range of D. plexippus larvae is maintained by both feeding stimulatory and deterrent chemicals in host and non-host plants.     

The life parameters of a parasitoid Microplitis mediator (Hymenoptera: Braconidae), reared on cotton bollworm Helicoverpa armigera (Hübner) with Cry1Ac diet

Microplitis mediator (Haliday) is an important endoparasitoid of the cotton bollworm, Helicoverpa armigera (Hübner) in northern China. Interactions among H. armigera, its larval parasitoid M. mediator, and Cry1Ac over two generations were evaluated under laboratory conditions. The results indicated that the developmental period of M. mediator offspring's eggs and larvae were significantly delayed and pupal and adult weight were significantly less compared to the control when the female parasitoids parasitized H. armigera larvae that fed on diet containing 1, 2, 4 and 8 µg/g Cry1Ac. The female parasitoids emerged from the host fed diet containing 8 µg/g Cry1Ac could oviposit in healthy hosts, and their offspring's biological parameters (egg–larval period, pupal weight and adult weight), parasitism rate, abnormal cocoon rate and adult emergence were not significantly affected. Cry1Ac was detected in larvae and hemolymph of H. armigera, but not in the larvae of M. mediator. The results suggest that the observed significant effects on several fitness parameters of the F₁ M. mediator developed from H. armigera fed Cry1Ac diet most likely were host-quality mediated rather than direct effects of Cry1Ac.     

Organellar and Cytosolic Localization of Four Phosphoribosyl Diphosphate Synthase Isozymes in Spinach

We report on the efficacy of proteinase inhibitors (PIs) from three host plants (chickpea [Cicer arietinum], pigeonpea [Cajanus cajan], and cotton [Gossypium arboreum]) and three non-host (groundnut [Arachis hypogea], winged bean [Psophocarpus tetragonolobus], and potato [Solanum tuberosum]) in retarding the growth of Helicoverpa armigera larvae, a devastating pest of important crop plants. Enzyme assays and electrophoretic analysis of interaction of H. armigera gut proteinases (HGPs) with PIs revealed that non-host PIs inhibited HGP activity efficiently whereas host PIs were ineffective. In the electrophoretic assay, trypsin inhibitor activity bands were detected in all of the host and non-host plants, but HGP inhibitor activity bands were present only in non-host plants (except cotton in the host plant group). H. armigera larvae reared on a diet containing non-host PIs showed growth retardation, a reduction in total and trypsin-like proteinase activity, and the production of inhibitor-insensitive proteinases. Electrophoretic analysis of PI-induced HGP showed differential regulation of proteinase isoforms. Interestingly, HGP activity induced in response to dietary potato PI-II was inhibited by winged bean PIs. The optimized combination of potato PI-II and winged bean PIs identified in the present study and their proposed successive use has potential in developing H. armigera-resistant transgenic plants.     

Diet-induced food preference by Manduca sexta larvae: acceptable non-host plants elicit a stronger induction than host plants

Diet-induced changes in food preference by fifth instar larvae of the tobacco hornworm, Manduca sexta (Johan.) (Lepidoptera, Sphingidae), were examined. Two groups of larvae with different diet experience were used: larvae reared on a host or on an acceptable non-host plant species. Each group of larvae was offered a choice between leaf discs from each rearing plant species (2-plant choice test) and food preference was measured as the consumption of one plant species relative to that of the other plant species. Diet-induced changes in preferences were larger with the host versus acceptable non-host plant pairs Solanum pseudocapsicum (L.) versus Raphanus sativus (L.), Lycopersicon esculentum (Mill.) versus Vigna sinensis (Savi), and Datura innoxia (L.) versus V. sinensis than with the host versus host plant pairs L. esculentum versus Capsicum annuum (L.) and L. esculentum versus D. innoxia. To examine how much the food preference had been altered for each test plant species alone, two other groups of larvae were offered a choice between leaf discs from a single plant species and filter paper discs laced with distilled water (1-plant choice test). Larvae preferred the dietary plant species more strongly than the non-dietary plant species in tests using the following plant species: for C. annuum with C. annuum and L. esculentum as diets, for V. sinensis with V. sinensis and L. esculentum or D. innoxia as diets, and for R. sativus with R. sativus and S. pseudocapsicum as diets. The preference for the hosts L. esculentum and D. innoxia did not change significantly after rearing larvae on different hosts or on an acceptable non-host. Thus, diet-induction by M. sexta larvae results in an enhancement of preference for the dietary plant species which is much stronger with acceptable non-hosts than with hosts.     

Desiccation tolerance of the resurrection plant Ramonda serbica is associated with dehydration-dependent changes in levels of proteolytic activities

The unique response of desiccation-tolerant, or resurrection plants, to extreme drought is accompanied by major changes in the protein pool, raising the possibility of the involvement of proteases. We detected and characterized proteases present in their active state in leaf extracts of desiccated Ramonda serbica Pan?., a resurrection plant from the Balkan Peninsula. Plants desiccated under laboratory conditions and maintained in anhydrobiosis for 4 and 14 months revived upon rehydration. Protease activities were determined spectrophotometrically in solution and by zymography on gels. Several endo- and aminopeptidases were detected and characterized by their pH profiles. Their enzyme class was determined using specific inhibitors. Those with higher activities were a serine endopeptidase active against Bz-Arg-pNA with a pH optimum around 9, and aminopeptidases optimally active at pHs from 7 to 9 against Leu-pNA, Met-pNA, Phe-pNA, Pro-pNA and Ala-pNA. The levels of their activities in leaf extracts from desiccated plants were significantly higher than those from rehydrated plants and from regularly watered plants, implying their involvement in the recovery of vegetative tissues from desiccation.     

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.     

Comparison of the growth and development of beet armyworm,Spodoptera exigua (Hübner) (Lepidoptera: Noctuidae) on two different diets

Beet armyworm (Spodoptera exigua (Hübner)) (Lepidoptera: Noctuidae) is the most economically important sugar beet (Beta vulgaris) pest worldwide. In this study, a comparison was made between two different diets: one was based on Merkx diet (Holidic diet) and the other was based on sugar beet leaf (Oligidic diet). Results showed that the whole development time from larvae to adult between two diets (Merkx and leaf) was not significantly different. For example, developmental time from first instar larvae to adult in Merkx diet was 11.33?days, whilst developmental time of larvae to adult when larvae fed with sugar beet leaf was 10.33?days. However, analysis of variance showed that in some cases like development time of the first instar, third instar and fifth instar larvae and pupae was significantly different between two treatments (p?<?0.05). Larval weight showed differences when larvae fed on Merkx diet and sugar beet leaves. For example, significant differences were shown between first, third and fourth instar larvae weight when larvae fed on Merkx diet and sugar beet leaves (p?<?0.05). However, significant differences were not observed between weight of second and fifth instar as well as pupae weight when larvae fed on Merkx diet and sugar beet leaves (p?>?0.05).     

Endogenous and induced monooxygenase activity in gypsy moth larvae feeding on natural and artificial diets

NADPH oxidase activity was measured in third to sixth instar gypsy moth larvae fed oak or pine foliage. Activity levels ranged from 400 to 1,900 pmol NADPH oxidized/min/mg microsomal protein, but enzyme activity was not correlated with host plant ingested. Similarly, activity levels in larvae fed diets containing inducers, such as the terpenoid α-pinene or pentamethylbenzene, ranged from 700 to 1,500 pmol NADPH oxidized/min/mg protein, levels that were comparable to those measured for larvae fed control diets. O-demethylase activity in older instar gypsy moth larvae fed pine averaged 109 pmol p-nitrophenol/min/mg protein, and activity levels in those fed diet containing α-pinene ranged from 22 to 55 pmol/min/mg protein. Although statistically significant, these induced O-demethylase levels are well below those observed for Heliothis zea larvae. Our findings indicate that monooxygenases play a minor, if any, role in the ability of later instar gypsy moth larvae to develop successfully on pine foliage.     

Changes in immune responses of Helicoverpa armigera Hübner followed by feeding on Knotgrass,Polygonum persicaria agglutinin

There is no study implying the effect of plant lectins on insect immune elements in both challenged and non‐challenged conditions with entomopathogenic agents. Lectins may bind to immune receptors on the surface of insect hemocytes, thus inducing or even disabling common immune functions including hemocyte counts, nodulation/encapsulation, phenoloxidase activity, and synthesis of antimicrobial peptides. In the present study, effect of Polygonum persicaria L. agglutinin (PPA) on immune responses of Helicoverpa armigera Hübner was investigated by feeding artificial diet treated to the larvae. Subsequently hemocyte count and expression of some immune‐related genes were considered for analyses. The two groups of larvae including control and PPA‐treated (1%) were divided into four subgroups of intact, Tween‐80 injected, latex‐bead injected and Beauveria bassiana‐injected. Except for intact larvae, the highest numbers of total and differential hemocyte counts were recorded 12 hr postinjection, however, the PPA‐fed larvae showed a significantly lower hemocyte counts compared to control. The number of nodules in PPA‐fed larvae was significantly lower than control, but the injected larvae of both control and PPA showed the highest nodulation 24 hr postinjection. Although the highest activity of phenoloxidase was observed 12 and 24 hr postinjection but its activity significantly decreased in PPA‐fed larvae compared to control. Gene expression of antimicrobial peptides including attacin, cecropin, and peptidoglycan receptor proteins were significantly decreased in artificial diet‐fed larvae containing PPA and then injected by B. bassiana spores and latex bead compared to control. These results clearly indicate adverse effects of PPA on immune responses in H. armigera.     

Partial characterization of the proteolytic enzymes in the gut of the banana weevil, Cosmopolites sordidus, and effects of soybean Kunitz trypsin inhibitor on larval performance

The proteolytic enzymes in the gut of the banana weevil, Cosmopolites sordidus (Germar) (Coleoptera: Curculionidae), have been characterized. Both larvae and adults rely on a complex proteolytic system based on at least cathepsin D‐, cathepsin B‐, trypsin‐, chymotrypsin‐, leucine aminopeptidase‐, carboxypeptidase A‐, and carboxypeptidase B‐like activities. All endoproteolytic activities were higher in the anterior section of the gut, whereas the exopeptidases were evenly distributed in the anterior and middle sections, and almost no activity was detected in the posterior section. Gelatin‐containing gels confirmed the spatial organization of the proteolytic digestive process. According to this proteolytic profile, the STI (soybean Kunitz trypsin inhibitor) was tested in vivo to establish its potential as a resistance factor against C. sordidus. Newly hatched larvae fed on diets containing 0.2% (w/w) STI experience lower survival rates and display significant reductions in larval growth. Biochemical analysis carried out on guts of larvae reared on STI‐treated diet showed a reduction of trypsin‐like activity compared to that from larvae fed on control diet. This decrease was compensated with an induction of cathepsin B, whereas cathepsin D, chymotrypsin, and leucine aminopeptidase were not affected. These results are discussed as a basis for selecting appropriate inhibitors to obtain transgenic banana and plantain plants with enhanced resistance to this pest.     

Host plant species affects the quality of the generalist Trichoplusia ni as a host for the polyembryonic parasitoid Copidosoma floridanum

Diet of herbivorous insects can influence both the herbivores and their natural enemies. We examined the direct and indirect effects of diet on the interactions between the polyphagous herbivore Trichoplusia ni Hübner (Lepidoptera: Noctuidae) and its polyembryonic parasitoid Copidosoma floridanum Ashmead (Hymenoptera: Encyrtidae). To determine how host plant species and host plant iridoid glycoside content affect host caterpillars and their parasitoids, parasitized and unparasitized T. ni were given leaves of either Plantago lanceolata L., which contains the iridoid glycosides aucubin and catalpol, Plantago major L. (Plantaginaceae), which contains only aucubin, or Taraxacum officinale F.H. Wigg (Asteraceae), which contains neither. Survival of unparasitized T. ni was much lower when fed P. major compared with the other two host plants, whereas pupae were smallest when fed T. officinale and developed most slowly when fed P. lanceolata as larvae. Neither aucubin nor catalpol were detected in intact Plantago‐fed T. ni larvae or their hemolymph, and only trace amounts of aucubin were detected in frass, suggesting that these compounds are mostly metabolized in the midgut and are not encountered by the parasitoid. Copidosoma floridanum clutch size was almost doubled when reared from P. lanceolata‐fed T. ni compared with T. officinale‐fed larvae and tripled compared with P. major‐fed larvae, although the percent of parasitoids surviving to adulthood was uniformly high regardless of host diet. The observed variation in C. floridanum fitness among host diets is likely mediated by the effect of the diets on host quality, which in turn may be influenced more by other factors in the host plants than their iridoid glycoside profiles. Interactions between plant metabolites, generalist herbivores like T. ni, and their parasitoids may be predominantly indirect.