Effect of protein type and quantity on growth and development of larval Heliothis zea and Spodoptera exigua and the endoparasitoid Hyposoter exiguae

Based on the use of casein, soy protein, and corn gluten in artificial diets, we show that larval growth of Heliothis zea (Boddie) and Spodoptera exigua (Hubner) (Lepidoptera: Noctuidae) is significantly reduced both by a deficit and surfeit of dietary protein. The optimal level of protein varies with protein type. Similar protein treatments were also used to examine the influence of host diet on growth and development of the endoparasitoid Hyposoter exiguae (Viereck) (Hymenoptera: Ichneumonidae). Dietary protein regimens reducing host larval growth to the greatest extent are most stressful to the parasitoid, irrespective of protein type. Parasitoid growth appeared to be regulated by the nutritional status of the host. The parasitoid required the host to reach ca 20 mg before it could successfully complete its larval development and pupate. This suggests that host hormones may also be involved in parasitoid development. The implications of these findings for theory of tritrophic interactions are discussed.

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Zusammenfassung Mit Hilfe des Gebrauchs von Kasein, Sojaprotein und Maisgluten in künstlicher Diät zeigen wir, dass das Wachstum der Larven von Heliothis zea (Boddie) und Spodoptera exigua (Hubner) (Lepidoptera: Noctuidae) signifikant reduziert ist durch Defizit oder Überschuss von Protein in der Diät. Der optimale Proteingehalt hängt von der Proteinart ab. Ähnliche Proteindiäten wurden auch benutzt, um den Einfluss von Wirtsdiät auf Wachstum und Entwicklung des Endoparasiten Hyposoter exiguae (Viereck) (Hymenoptera: Icheneumonidae) zu untersuchen. Diejenigen Proteingehaltsstufen, die das Larvenwachstum des Wirts zum grössten Ausmass reduzieren, produzieren den grössten Stress für den Parasiten, unabhängig von der Proteinart. Das Wachstum des Parasiten scheint durch den Ernährungszustand des Wirtes reguliert zu werden. Für die erfolgreiche Larvenentwicklung und Verpuppung des Parasiten war ein Gewicht des Wirtes von ca. 20 mg Voraussetzung. Das deutet auf die Möglichkeit hin, dass Wirtshormone für die Entwicklung des Parasiten eine Rolle spielen können. Schlussfolgerungen aus diesen Ergebnissen für die Theorie tritrophischer Wechselbeziehungen werden diskutiert.

     

Preparation and molecular characterization of a polyclonal antibody as an efficient cutworm reference protein

Antibodies, which are wildly used in molecular researches, are proteins secreted by plasma cells that can specifically recognize specific antigens. To obtain a stable reference antibodies are important for protein analysis. In this study, a capable glyceraldehyde phosphate dehydrogenase (GAPDH) antiserum against most noctuid larval protein samples was prepared. Firstly, the full-length gapdh was amplified from Spodoptera exigua larvae to construct the prokaryotic expression vector. The purified His-tag fused GAPDH protein was used to immunize rabbits for the antiserum preparation. Protein samples extracted from 11 insect species distributed across seven families and three orders were used to perform the Western bolt analysis. The bright specific bands were detected in S. exigua, S. litura, Helicoverpa armigera, and Mythimna seperata protein samples, indicating that this antiserum may be capable of detecting noctuid larval encoded GAPDH. Further immunofluorescence identification of H. armigera and S. exigua paraffin section slides with GAPDH antiserum exhibited an identical cyto-stained image. The GAPDH antiserum prepared in this study is useful and efficient as reference antibodies in studies involving noctuid larval protein samples in other related fields.     

Apis iridescent virus and "clustering disease" of Apis cerana

NPV of Spodoptera littoralis was completely inactivated in vitro following 10 min of exposure to a temperature higher than 90°C, but survived 3 weeks at ?20°C. At pH 12, some 75% of the infectivity was lost. Measurable proteolysis in vitro of the polyhedral protein by a larval midgut extract could be obtained only when the pH of the reaction mixture was raised to an unnatural level of 10.5, the natural pH of the midgut content being 8.5 or 9.5 according to different authors. The plant growth retardant Phosfon synergized mortality caused by the NPV. The virus could be cross-transmitted to two congeneric species of Spodoptera (S. exigua and S. litura), but could not infect any of four tested species belonging to other genera of the Moctuid family.     

Responses of representative midgut detoxifying enzymes from Heliothis zea and Spodoptera frugiperda to trichothecenes

The relative levels of representative midgut detoxifying enzymes were measured in last instar larvae of Heliothis zea and Spodoptera frugiperda after the insects had been fed for 48 h on diets containing representative trichothecenes, which are protein synthesis inhibitors. 4-Nitroanisole O-demethylation was induced (as indicated by increased enzyme activity) by 1.6-fold in H. zea and by 6.1-fold in S. frugiperda when fed 250 ppm of deoxynivalenol, a dose that severely retarded growth of the insects. Induction of this activity was also noted with lower levels of deoxynivalenol in both species, and with diacetoxyscirpenol and T-2 toxin in H. zea. 1-Chloro-2,4-dinitrobenzene glutathione transferase conjugation was induced by c. 1.3-fold with deoxynivalenol at 250 ppm and 25 ppm, and with T-2 toxin at 25 ppm in H. zea; but was slightly inhibited (c. 10–20%) in S. frugiperda in some cases.1-Naphthyl acetate hydrolysis was generally unaffected by all trichothecenes tested at all doses in H. zea, but was inhibited by c. 30% in S. frugiperda by lower doses of all three trichothecenes. However, a new 1-naphthyl acetate esterase band was noted after exposure to 250 ppm of deoxynivalenol, and to some extent lower doses of deoxynivalenol and the other trichothecenes in both insect species. The hydrolysis of a radiolabeled model trichothecene, 4-monoacetoxyscripenol, was induced by 3.1-fold in H. zea and 2.7-fold in S. frugiperda with 250 ppm of deoxynivalenol (the only compound and dose tested). This activity could be inhibited by nearly 100% with 10⁻⁴ M paraoxon, suggesting a serine-hydroxyl esterase was involed. Although this activity could not be recovered from gels, indirect competitive inhibition assays with 1-naphthyl acetate suggested the induced band could be responsible for some of the hydrolysis of the 4-monoacetoxyscirpenol.     

Vitamin requirements of larvae of Sitophilus oryzae

Improvements were made in the meridic larval diet for Sitophilus oryzae by replacing the minerals and vitamins supplied by dietary brewer's yeast and wheat germ with mineral and vitamin mixtures. The effects of different concentrations of individual vitamins were studied with the improved diet containing 20% casein. In later tests the dietary casein was replaced with a mixture of 15 amino acids. The results indicated that these larvae, which contain an associated bacteria-like micro-organism, required thiamine, nicotinic acid, pyridoxine, folic acid, and biotin in the diet. No requirement for riboflavin or pantothenic acid could be demonstrated on either the 20% casein diet or the amino acid diet possibly due to contamination of the cornstarch with these two vitamins. In addition, the larvae did not require choline or inositol for the growth of one larval generation.The asymbiotic larvae of S. granarius failed to develop on the improved casein diet indirectly implicating the symbiotes in a nutritional rôle; however, the symbiotes present in S. oryzae apparently do not provide B vitamins. Larvae of S. oryzae failed to develop when the concentration of casein was reduced to 10% while growth was maintained with a 10% concentration of the amino acid mixture. Casein is not an optimal source of amino acids for this species.     

A host-plant specialist, Helicoverpa assulta, is more tolerant to capsaicin from Capsicum annuum than other noctuid species

Plant secondary compounds not only play an important role in plant defense, but have been a driving force for host adaptation by herbivores. Capsaicin (8-methyl-N-vanillyl-6-nonenamide), an alkaloid found in the fruit of Capsicum spp. (Solanaceae), is responsible for the pungency of hot pepper fruits and is unique to the genus. The oriental tobacco budworm, Helicoverpa assulta (Lepidoptera: Noctuidae), is a specialist herbivore feeding on solanaceous plants including Capsicum annuum, and is one of a very few insect herbivores worldwide capable of feeding on hot pepper fruits. To determine whether this is due in part to an increased physiological tolerance of capsaicin, we compared H. assulta with another specialist on Solanaceae, Heliothis subflexa, and four generalist species, Spodoptera frugiperda, Heliothis virescens, Helicoverpa armigera, and Helicoverpa zea, all belonging to the family Noctuidae. When larvae were fed capsaicin-spiked artificial diet for the entire larval period, larval mortality increased in H. subflexa and H. zea but decreased in H. assulta. Larval growth decreased on the capsaicin-spiked diet in four of the species, was unaffected in H. armigera and increased in H. assulta. Food consumption and utilization experiments showed that capsaicin decreased relative consumption rate (RCR), relative growth rate (RGR) and approximate digestibility (AD) in H. zea, and increased AD and the efficiency of conversion of ingested food (ECI) in H. armigera; whereas it did not significantly change any of these nutritional indices in H. assulta. The acute toxicity of capsaicin measured by injection into early fifth instar larvae was less in H. assulta than in H. armigera and H. zea. Injection of high concentrations produced abdominal paralysis and self-cannibalism. Injection of sub-lethal doses of capsaicin resulted in reduced pupal weights in H. armigera and H. zea, but not in H. assulta. The results indicate that H. assulta is more tolerant to capsaicin than the other insects tested, suggesting that this has facilitated expansion of its host range within Solanaceae to Capsicum after introduction of the latter to the Old World about 500 years ago. The increased larval survival and growth due to chronic dietary exposure to capsaicin suggests further adaptation of H. assulta to that compound, the mechanisms of which remain to be investigated.     

A Comparison of Growth and Development of Three Major Agricultural Insect Pests Infected with Heliothis virescens ascovirus 3h (HvAV-3h)

Ascoviruses are double-stranded DNA viruses that are pathogenic to lepidopteran hosts, particularly noctuid larvae. Infection of a larva is characterized by retarded growth, reduced feeding and yellowish body color. In this paper, we reported the growth and development of three major agricultural noctuid insect pests, Helicoverpa armigera (Hübner), Spodoptera exigua (Hübner) and Spodoptera litura (Fabricius), infected with Heliothis virescens ascovirus 3h (HvAV-3h). Using 10-fold serial dilutions (0 to 7) of HvAV-3h-containing hemolymph to infect S. litura larvae, we found no significant difference in larval mortalities from 0 to 10³-fold dilutions; however, significant differences were observed at 10⁴-fold dilution and above. Using a 10-fold dilution of HvAV-3h-containing hemolymph to infect H. armigera, S. exigua and S. litura larvae, we found that the growth and development were significantly affected. All infected larvae could not pupate; the survival times of treated H. armigera, S. litura and S. exigua larvae were significantly longer than untreated control larvae. Body weight showed significant difference between treated and untreated control group from day 1 after inoculation in H. armigera and S. exigua, but day 2 in S. litura. Additionally, food intake also showed significant difference between treated and untreated control group from day 2 after inoculation in H. armigera and S. litura, but day 3 in S. exigua.     

Role of the exoprotease InA in the pathogenicity of Bacillus thuringiensis in pupae of Hyalophora cecropia

Two geographical biotypes of Nomuraea rileyi (from Ecuador and the United States) were topically bioassayed against seven lepidopteran species, i.e., Anticarsia gemmatalis, Heliothis zea, Heliothis virescens, Heliothis subflexa, Pseudoplusia includens, Spodoptera exigua, and Trichoplusia ni. There was an average difference of 1.7-fold in mortality in how cultures of the same insect species from different sources responded to topical applications of either biotype of N. rileyi. Regression equations and LC₅₀ values were obtained for each insect species and fungal biotype combination. Larvae of S. exigua were equally susceptible to both biotypes of N. rileyi. Although larvae of A. gemmatalis were moderately susceptible to the Ecuadoran biotype, they were relatively nonsusceptible to the Mississippian biotype. Species of Heliothis (H. zea, H. virescens, and H. subflexa) were about equally susceptible to the Mississippian biotype. Larvae of H. subflexa and H. virescens, however, were significantly less susceptible than H. zea to the Ecuadoran biotype. When the integumental barrier was breached via intrahemocoelic injections, larvae of H. virescens were as susceptible as H. zea larvae to blastospores of either biotype of N. rileyi.     

Growth and development of two polyphagous lepidopterans fed high- and low-tannin sericea lespedeza

The biological impact of consumption of sericea lespedeza (Lespedeza cuneata (Dumont) G. Don) genotypes varying in tannin content was examined for two generalist insect herbivores, Heliothis zea Boddie and Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae). Foliage of high- and lowtannin genotypes did not substantially affect the growth and development of either species when incorporated into meridic diet except at large concentrations where a diet containing high tanin genotypes reduced larval weight and delayed pupation of both species. Fresh foliage of sericea lespedeza with varying levels of tannin did not adversely affect larval growth and development of S. frugiperda. All genotypes were a poor host for H. zea in that most larvae died before pupation. Initial larval weight of H. zea was not consistently different between high- and low-tannin genotypes. Except for one low-tannin genotype having a greater efficiency of conversion of digested diet than the other genotypes, foliage tannin content had little effect on diet assimilation and utilization and larval developmental and consumption rates of stages 6 and 7 H. zea larvae. H. zea neonates also did not show a significant preference for any genotypes. Therefore, tannin content of sericea lespedeza had relatively little effect on the growth and development of these generalist insect defoliators which suggests that low-tannin genotypes of sericea lespedeza will not be substantially more susceptible to defoliation by these species. The poor performance of H. zea on all L. cuneata genotypes suggests that the plant may contain factors other than tannin that inhibit the growth and development of this species or that sericea lespedeza lacks essential nutrients for proper development of H. zea.

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Résumé Les effets de la consommation de phénotypes de la légumineuse S. lespedeza (Lespedeza cuneata) (Dumont) (G. Don) dont les teneurs en tanin diffèrent, ont été examinés sur 2 insectes généralistes; Heliothis zea Boddie et Spodoptera frugiperda (J. E. Smith) (Lep.: Noctuidae). Le feuillage de phénotypes à teneur élevée ou basse en tanin n'a pas affecté considérablement la croissance et le développement des 2 espèces, quand il a été incorporé dans un régime méridique, sauf aux fortes concentrations où un régime contenant le phénotype riche en tanin a réduit le poids larvaire et retardé la nymphose des 2 espèces. Le feuillage frais de S. lespedeza avec différentes teneurs en tanin n'a pas nui au développement et à la croissance larvaire de S. frugiperda. Tous les phénotypes ont constitué un hôte désavantageux pour H. zea dont toutes les chenilles sont mortes avant la nymphose. Les poids initiaux des chenilles de H. zea ne différaient pas significativement suivant la pauvreté ou la richesse en tanin des phénotypes. A l'exception d'un phénotype pauvre en tanin ayant une plus grande efficacité de transformation lors de la digestion, la teneur en tanin du feuillage a eu peu d'effet sur l'assimilation du repas, son utilisation, les taux de consommation et le développement larvaire des stades 6 et 7 des chenilles d'H. zea. Les chenilles néonates de H. zea n'ont présenté aucune préférence significative pour l'un des phénotypes. Par conséquent, la teneur en tanin de S. lespedeza a eu relativement peu d'effet sur la croissance et le développement de ces deux généralistes défoliateurs, ce qui suggère que les phénotypes de S. lespedeza pauvres en tanin ne sont pas nettement plus susceptibles de défoliation par ces espèces. Les faibles performances de H. zea sur tous les phénotypes de S. lespedeza suggèrent que cette plante peut contenir des facteurs autres que les tanins qui inhibent la croissance et le développement de cette espèce ou que S. lespedeza manque de certains éléments nécessaires au développement de H. zea.

     

A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity

Crystal toxin Cry1Ca from Bacillus thuringiensis has an insecticidal spectrum encompassing lepidopteran insects that are tolerant to current commercially used B. thuringiensis crops (Bt crops) expressing Cry1A toxins and may be useful as a potential bioinsecticide. The mode of action of Cry1A is fairly well understood. However, whether Cry1Ca interacts with the same receptor proteins as Cry1A remains unproven. In the present paper, we first cloned a cadherin-like gene, SeCad1b, from Spodoptera exigua (relatively susceptible to Cry1Ca). SeCad1b was highly expressed in the larval gut but scarcely detected in fat body, Malpighian tubules, and remaining carcass. Second, we bacterially expressed truncated cadherin rSeCad1bp and its interspecific homologue rHaBtRp from Helicoverpa armigera (more sensitive to Cry1Ac) containing the putative toxin-binding regions. Competitive binding assays showed that both Cry1Ca and Cry1Ac could bind to rSeCad1bp and rHaBtRp, and they did not compete with each other. Third, Cry1Ca ingestion killed larvae and decreased the weight of surviving larvae. Dietary introduction of SeCad1b double-stranded RNA (dsRNA) reduced approximately 80% of the target mRNA and partially alleviated the negative effect of Cry1Ca on larval survival and growth. Lastly, rSeCad1bp and rHaBtRp differentially enhanced the negative effects of Cry1Ca and Cry1Ac on the larval mortalities and growth of S. exigua and H. armigera. Thus, we provide the first lines of evidence to suggest that SeCad1b from S. exigua is a functional receptor of Cry1Ca.     

A kidney bean trypsin inhibitor with an insecticidal potential against Helicoverpa armigera and Spodoptera litura

In the present study, trypsin inhibitor extracts of ten kidney bean seed (Phaseolus vulgaris) varieties exhibiting trypsin and gut trypsin-like protease inhibitor activity were tested on Helicoverpa armigera and Spodoptera litura. Trypsin inhibitor protein was isolated and purified using multi-step strategy with a recovery of ~15 % and purification fold by ~39.4. SDS-PAGE revealed a single band corresponding to molecular mass of ~15 kDa and inhibitory activity was confirmed by reverse zymogram analyses. The inhibitor retained its inhibitory activity over a broad range of pH (3–11), temperature (40–60 °C) and thermostability was promoted by casein, CaCl₂, BSA and sucrose. The purified inhibitor inhibited bovine trypsin in 1:1 molar ratio. Kinetic studies showed that the protein is a competitive inhibitor with an equilibrium dissociation constant of 1.85 μM. The purified trypsin inhibitor protein was further incorporated in the artificial diet and fed to second instar larvae. A maximum of 91.7 % inhibition was obtained in H. armigera, while it was moderate in S. litura (29 %) with slight varietal differences. The insect bioassay showed 40 and 22 % decrease in larval growth followed by 3 and 2 days delay in pupation of H. armigera and S. litura, respectively. Some of the adults emerged were deformed and not fully formed. Trypsin inhibitor protein was more effective against H. armigera as it showed 46.7 % mortality during larval growth period compared to S. litura (13.3 %).     

Role of Bacillus thuringiensis Cry1 δ Endotoxin Binding in Determining Potency during Lepidopteran Larval Development

Five economically important crop pests, Manduca sexta, Pieris brassicae, Mamestra brassicae, Spodoptera exigua, and Agrotis ipsilon, were tested at two stages of larval development for susceptibility to Bacillus thuringiensis toxins Cry1Ac, Cry1Ca, Cry1J, and Cry1Ba. Bioassay results for M. sexta showed that resistance to all four Cry toxins increased from the neonate stage to the third-instar stage; the increase in resistance was most dramatic for Cry1Ac, the potency of which decreased 37-fold. More subtle increases in resistance during larval development were seen in M. brassicae for Cry1Ca and in P. brassicae for Cry1Ac and Cry1J. By contrast, the sensitivity of S. exigua did not change during development. At both larval stages, A. ipsilon was resistant to all four toxins. Because aminopeptidase N (APN) is a putative Cry1 toxin binding protein, APN activity was measured in neonate and third-instar brush border membrane vesicles (BBMV). With the exception of S. exigua, APN activity was found to be significantly lower in neonates than in third-instar larvae and thus inversely correlated with increased resistance during larval development. The binding characteristics of iodinated Cry1 toxins were determined for neonate and third-instar BBMV. In M. sexta, the increased resistance to Cry1Ac and Cry1Ba during larval development was positively correlated with fewer binding sites in third-instar BBMV than in neonate BBMV. The other species-instar-toxin combinations did not reveal positive correlations between potency and binding characteristics. The correlation between binding and potency was inconsistent for the species-instar-toxin combinations used in this study, reaffirming the complex mode of action of Cry1 toxins.     

Specificity of induced resistance in the tomato, Lycopersicon esculentum

Specificity in the induced responses of tomato foliage to arthropod herbivores was investigated. We distinguished between two aspects of specificity: specificity of effect (the range of organisms affected by a given induced response), and specificity of elicitation (ability of the plant to generate distinct chemical responses to different damage types). Specificity of effect was investigated by examining the effect of restricted feeding by Helicoverpa zea on the resistance of tomato plants to an aphid species (Macrosiphum euphorbiae), a mite species (Tetranychus urticae), a noctuid species (Spodoptera exigua), and to a phytopathogen, Pseudomonas syringae pv. tomato. Prior H. zea feeding was found to increase the resistance of tomato plants to all four organisms. Specificity in elicitation was investigated by examining the effect of aphid feeding on the activities of four defense-related proteins and on the suitability of foliage for S. exigua. Aphid feeding was found to induce peroxidase and lipoxygenase activities but not polyphenol oxidase and proteinase inhibitor activities; this response is distinct from the response to H. zea feeding, which induces polyphenol oxidase and proteinase inhibitors but not peroxidase. Leaflets which had been fed upon by aphids were better sources of food for S. exigua than were leaflets which had not been fed upon by aphids. Studies of both these aspects of specificity are needed to understand the way in which plants coordinate and integrate induced responses against insects with other physiological processes. Received: 20 December 1996 / Accepted: 2 July 1997     

Bioinsecticidal activity of Talisia esculenta reserve protein on growth and serine digestive enzymes during larval development of Anticarsia gemmatalis

Plants synthesize a variety of molecules to defend themselves against an attack by insects. Talisin is a reserve protein from Talisia esculenta seeds, the first to be characterized from the family Sapindaceae. In this study, the insecticidal activity of Talisin was tested by incorporating the reserve protein into an artificial diet fed to the velvetbean caterpillar Anticarsia gemmatalis, the major pest of soybean crops in Brazil. At 1.5% (w/w) of the dietary protein, Talisin affected larval growth, pupal weight, development and mortality, adult fertility and longevity, and produced malformations in pupae and adult insects. Talisin inhibited the trypsin-like activity of larval midgut homogenates. The trypsin activity in Talisin-fed larvae was sensitive to Talisin, indicating that no novel protease-resistant to Talisin was induced in Talisin-fed larvae. Affinity chromatography showed that Talisin bound to midgut proteinases of the insect A. gemmatalis, but was resistant to enzymatic digestion by these larval proteinases. The transformation of genes coding for this reserve protein could be useful for developing insect resistant crops.     

Joint toxicity of methoxyfenozide and lufenuron on larvae of Spodoptera exigua Hübner (Lepidoptera: Noctuidae)

The joint action and sublethal effects of methoxyfenozide and lufenuron were measured against Spodoptera exigua. Methoxyfenozide and lufenuron exhibited optimum synergistic toxicity on S. exigua at a mass ratio of 4:6, and the co-toxicity coefficient (CTC) was 165.705. Third instars larvae of S. exigua were treated with methoxyfenozide (LC₁₅ = 21.004 ng/cm²), lufenuron (LC₁₅ = 27.134 ng/cm²), or a mixture of methoxyfenozide and lufenuron (MML, LC₁₅ = 16.503 ng/cm²) through feeding for 72 h. Ingestion of MML by larvae significantly inhibited larval and pupal weights and pupation rate, and prolonged the larval and pupal development of S. exigua compared to individual treatment ofmethoxyfenozide or lufenuron. Both methoxyfenozide and MML treatments significantly decreased the fertility of female S. exigua. No significant changes were observed in case of adult emergence and egg hatching for different treatments. The MML-treated S. exigua exhibited significantly lower activities of polyphenol oxidase (PPO) and cytochrome P450 (CYP450) than those in S. exigua treated separately with methoxyfenozide or lufenuron. Finally, methoxyfenozide, lufenuron, and MML treatments decreased chitinase, acetylcholinesterase (AChE), carboxylesterase (CarE), and glutathione-s-transferase (GST) activities in S. exigua.     

Induced plant responses to multiple damagers: differential effects on an herbivore and its parasitoid

Herbivore-induced plants responses can affect the preference and performance of herbivores and their natural enemies. These responses may vary depending on the identity and number of herbivore species feeding on the plant so that when herbivores from different guilds feed on plants, the interactions between plants, herbivores, and natural enemies may be disrupted. Tomato plants were damaged either by the caterpillar Spodoptera exigua, or the aphid Macrosiphum euphorbiae, or damaged by both herbivores, or undamaged controls. We measured the preference and performance of S. exigua and its parasitoid Cotesia marginiventris, and activity of proteinase inhibitors (PI) as an indicator of induced resistance. Compared to undamaged plants, caterpillar damage reduced the number of eggs laid by S. exigua adults, reduced growth, consumption, and survival of larval S. exigua and C. marginiventris, and increased activity of PIs 43%; but did not increase attraction of C. marginiventris. While pupal mass of S. exigua was not affected, the pupal mass of C. marginiventris decreased on caterpillar-damaged plants compared to controls. In contrast, plants damaged by aphids were preferred for oviposition by S. exigua, and had increased larval consumption and survival, compared to controls. Aphid feeding did not affect the preference or performance of C. marginiventris, or PI activity, compared to controls. While oviposition was deterred on caterpillar-damaged plants, plants damaged by both herbivores received the same amount of oviposition as controls. The attraction of C. marginiventris to plants damaged by caterpillars and aphids was increased compared to controls. However, plants damaged by both herbivores had similar PI activity, larval growth and survival of S. exigua and C. marginiventris, as plants singly damaged by caterpillars. Overall, the preference component for both the herbivore and parasitoid was more strongly affected by damage due to multiple herbivores than the performance component.     

The endophytic fungal entomopathogens Beauveria bassiana and Purpureocillium lilacinum enhance the growth of cultivated cotton (Gossypium hirsutum) and negatively affect survival of the cotton bollworm (Helicoverpa zea)

The effects of two entomopathogenic fungal endophytes, Beauveria bassiana and Purpureocillium lilacinum, were assessed on the growth of cultivated cotton (Gossypium hirsutum) and development of the cotton bollworm (Helicoverpa zea). In two replicate greenhouse trials, cotton plants were inoculated as seed treatments with two concentrations of B. bassiana or P. lilacinum conidia and evaluated for effects on both plant dry biomass, number of nodes and number of developing flowers (squares). We similarly treated cotton plants and evaluated H. zea performance using no-choice in planta assays starting at the 2nd larval instar. Treatment with both fungal endophytes resulted in a significant increases in plant dry biomass (ANOVA, P = 0.024). Plant developmental stage and number of squares were also significantly enhanced in the endophyte treated plants (ANOVA, P = 0.005 and P = 0.027, respectively). The survivorship of H. zea was significantly different among the endophyte treatment groups (Kaplan–Meier, P = 0.02), where insects feeding on control plants exhibited higher survival than insects on the endophyte treated plants. There were no significant endophyte treatment effects on larval or pupal weights of H. zea individuals. There was no endophyte effect on days to pupation among treatments, but there was a marginal effect on days to eclosion (Kaplan–Meier, P = 0.07). Overall, our results demonstrate (i) the positive plant growth enhancing effects of the target endophyes on cultivated cotton under greenhouse conditions and (ii) the negative effects of endophytic P. lilacinum and B. bassiana on H. zea survivorship and development using whole plant assays.     

Identification of Bacillus thuringiensis Delta-Endotoxin Cry1C Domain III Amino Acid Residues Involved in Insect Specificity

Cry1C domain III amino acid residues involved in specificity for beet armyworm (Spodoptera exigua) were identified. For this purpose, intradomain III hybrids between Cry1E (nontoxic) and Cry1E-Cry1C hybrid G27 (toxic) were made. Crossover points of these hybrids defined six sequence blocks containing between 1 and 19 of the amino acid differences between Cry1E and G27. Blocks B, C, D, and E of G27 were shown to be required for optimal activity against S. exigua. Block E was also required for optimal activity against the tobacco hornworm (Manduca sexta), whereas block D had a negative effect on toxicity for this insect. The mutagenesis of individual amino acids in block B identified Trp-476 as the only amino acid in this block essential, although not sufficient by itself, for full S. exigua activity. In block D, we identified a seven-amino-acid insertion in G27 that was not in Cry1E. The deletion of either one of two groups of four consecutive amino acids in this insertion completely abolished activity against S. exigua but resulted in higher activity against M. sexta. Alanine substitutions of the first group had little effect on toxicity, whereas alanine substitutions of the second group had the same effect as its deletion. These results identify groups of amino acids as well as some individual residues in Cry1C domain III, which are strongly involved in S. exigua-specific activity as well as sometimes involved in M. sexta-specific activity.