Plant growth and development influenced by transgenic insertion of bacterial chitinolytic enzymes

The role of the chitinolytic enzymes in plants is not necessarilyrestricted to plant defense. Tomato plants transformed with an endochitinaseand a chitobiosidase gene from Streptomyces albidoflavus andgrowth under greenhouse conditions showed a significant reduction in plantheight, and reduced time to flowering compared with the control(non-transformed) plants. The levels of chitobiosidase and endochitinaseactivity in the transgenic tomato plants were positively correlated with earlyflowering, and negatively correlated with plant height. We have not determinedwhether these effects are exclusively due to the expression of the transgenesof endochitinase and chitobiosidase from S. albidoflavus orthe additive effect of these 2 enzymes combined with the endogenouschitinolytic enzymes produced by the plants. However, when control plants were trimmed,early flowering was observed compared with the controls that were not trimmed, whichindicates that wound induced proteins such as chitinolytic enzymes affect thetime of flowering. In addition, the expression of the endochitinase andchitobiosidase genes significantly increased the number of flowers and fruit onthe plants, resulting in an increase in yield of fruit. One of the primarygoals of crop breeding programs is to increase the productivity of plants. These twogenes were directly associated with plant productivity, and should be studied further.     

Cuticular and non-cuticular substrate influence on expression of cuticle-degrading enzymes from conidia of an entomopathogenic fungus,Nomuraea rileyi

Larval cuticle fromTrichoplusia ni, Helicoverpa (=Heliothis)zea, andHeliothis virescens and a cellulose substrate were used to quantify release of proteolytic, chitinolytic, and lipolytic enzymes by germinating conidia of the entomopathogenic fungus,Nomuraea rileyi. There was no significant difference in conidial viability incubated withT. ni, H. zea or cellulose substrates. Conidial viability onH. virescens cuticle, however, was significantly lower (ca. 19–25%) than the other three substrates. The presence of cuticle substrates, especially cuticle ofT. ni, stimulated germination. The nature of the substrate influenced both the time and quantity of the enzymes expressed. Specific proteases (aminopeptidase, chymoelastase, trypsin) generally were expressed earlier and/or in greater quantities on cuticular than on the cellulose substrate. Although both chitinolytic enzymes (endochitinase, N-acetylglucosaminidase) were detected on all three cuticular substrates, their activity was substantially lower than that of the proteolytic enzymes. Lipase activity was only minimally present. Early concurrent release of both proteases and chitinases suggested that both may be important in the penetration of the larval integument by germinating conidia ofN. rileyi. Expression of proteases and chitinases, especially aminopeptidase and endochitinase was probably a specific response to cuticle, because little or no activity was expressed on the non-host, cellulose substrate.This article reports the results of research only. Mention of a proprietary product in this paper does not constitute a recommendation for use by the US Department of Agriculture.     

Defense-related gene expression and enzyme activities in transgenic cotton plants expressing an endochitinase gene from <Emphasis Type="Italic">Trichoderma virens</Emphasis> in response to interaction with <Emphasis Type="Italic">Rhizoctonia solani</Emphasis>

There are many reports on obtaining disease-resistance trait in plants by overexpressing genes from diverse organisms that encode chitinolytic enzymes. Current study represents an attempt to dissect the mechanism underlying the resistance to Rhizoctonia solani in cotton plants expressing an endochitinase gene from Trichoderma virens. Several assays were developed that provided a powerful demonstration of the disease protection obtained in the transgenic cotton plants. Transgene-dependent endochitinase activity was confirmed in various tissues and in the medium surrounding the roots of transformants. Biochemical and molecular analyses conducted on the transgenic plants showed rapid/greater induction of ROS, expression of several defense-related genes, and activation of some PR enzymes and the terpenoid pathway. Interestingly, even in the absence of a challenge from the pathogen, the basal activities of some of the defense-related genes and enzymes were higher in the endochitinase-expressing cotton plants. This elevated defensive state of the transformants may act synergistically with the potent, transgene-encoded endochitinase activity to confer a strong resistance to R. solani infection. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Partial characterization of chitinolytic enzymes from Streptomyces albidoflavus

Streptomyces albidoflavus NRRL B-16746 secreted three types of chitinolytic enzymes: N -acetyl-glucosaminidase, chitobiosidase and endochitinase. Optimal activity for all three types of enzymes occurred at pH 4–6; however 55–74% of the chitobiosidase and endochitinase activity was detectable at pH 8–10. Chitobiosidase activity originated from two strongly acidic (pI < 3.0) proteins with molecular mass of 27 kDa and 34 kDa, while endochitinase activity originated from five major acidic proteins (pI 5.1, 5.3, 5.75, 5.8–5.9 and 6.4) with molecular mass of 59, 45, 38.5, 27 and 25.5 kDa. Purified chitobiosidases significantly reduced spore germination and germ tube elongation of Botrytis cinerea and Fusarium oxysporum. Chitinolytic enzymes with significant activity at pH 4–10 may be used, transgenically, to reduce the growth and/or development of a broad spectrum of insects and fungi that are major economic pests.     

Midgut cysteine protease-inhibiting activity in Trichoplusia ni protects the peritrophic membrane from degradation by plant cysteine proteases

The action of plant cysteine proteases on the midgut peritrophic membrane (PM) of a polyphagous herbivorous lepidopteran, Trichoplusia ni, was studied. Proteins in PMs isolated from T. ni larvae were confirmed to be highly resistant to the serine proteinases trypsin and chymotrypsin, but were susceptible to degradation by plant cysteine proteases, which is consistent with the known molecular and biochemical characteristics of the T. ni PM proteins. However, the PM proteins were not degraded by plant cysteine proteases in larvae or in the presence of larval midgut fluid in vitro. With further biochemical analysis, cysteine protease-inhibiting activity was identified in the midgut fluid of T. ni larvae. The cysteine protease-inhibiting activity was heat resistant and active in the tested pH range from 6.0 to 10.0, but could be suppressed by thiol reducing reagents or reduced by treatment with catalase. In addition to T. ni, cysteine protease-inhibiting activity was also identified from two other polyphagous Lepidoptera species, Helicoverpa zea and Heliothis virescens. In conclusion, results from this study uncovered that herbivorous insects may counteract the attack of plant cysteine proteases on the PM by inhibiting the potentially insecticidal cysteine proteases from plants in the digestive tract. However, the biochemical identity of the cysteine protease-inhibiting activity in midgut fluid has yet to be identified.     

Effects of wax bloom variation in Brassica oleracea on foraging by a vespid wasp

Foraging by individual Polistes dominulus (Christ) (Hymenoptera: Vespidae) was observed and compared on Brassica oleracea L. plants that differed in surface wax bloom. Twenty-six wasps, previously trained to forage on plants for 4th instar Trichoplusia ni (Walker), were presented in the greenhouse with mixtures of plants with normal wax bloom or genetically reduced wax bloom, and on which T. ni caterpillars had been placed. During foraging, the wasps were observed to slip significantly more frequently from the leaf surfaces of normal-wax bloom plants than from reduced-wax bloom plants (129 vs 63 occurrences) and to retrieve significantly fewer pieces of caterpillar after attack on normal-wax bloom plants (151 vs 223 pieces). Altogether the wasps retrieved 333 caterpillar pieces from reduced-wax bloom plants and 248 pieces from normal-wax bloom plants. Despite these differences, the number of caterpillars attacked and killed did not differ between the two wax bloom types (116 vs 121), nor did handling time for individual attacks (time from contact with prey until prey piece was carried to the nest) (170±12.5 s vs 180±10.5 s). Thus, in contrast with previous reports for smaller predators, wax bloom variation in B. oleracea did not influence the effectiveness of P. dominulus as a predator of T. ni.     

Chitinolytic Enterobacter agglomerans Antagonistic to Fungal Plant Pathogens

Three Enterobacter agglomerans strains which produce and excrete proteins with chitinolytic activity were found while screening soil-borne bacteria antagonistic to fungal plant pathogens. The chitinolytic activity was induced when the strains were grown in the presence of colloidal chitin as the sole carbon source. It was quantitated by using assays with chromogenic p-nitrophenyl analogs of disaccharide, trisaccharide, and tetrasaccharide derivatives of N-acetylglucosamine. A set of three fluorescent substrates with a 4-methylumbelliferyl group linked by (beta)-1,4 linkage to N-acetylglucosamine mono- or oligosaccharides were used to identify the chitinolytic activities of proteins which had been renatured following their separation by electrophoresis. This study provides the most complete evidence for the presence of a complex of chitinolytic enzymes in Enterobacter strains. Four enzymes were detected: two N-acetyl-(beta)-d-glucosaminidases of 89 and 67 kDa, an endochitinase with an apparent molecular mass of 59 kDa, and a chitobiosidase of 50 kDa. The biocontrol ability of the chitinolytic strains was demonstrated under greenhouse conditions. The bacteria decreased the incidence of disease caused by Rhizoctonia solani in cotton by 64 to 86%. Two Tn5 mutants of one of the isolates, which were deficient in chitinolytic activity, were unable to protect plants against the disease.     

Excretome of the chitinolytic bacterium <Emphasis Type="Italic">Clostridium paraputrificum</Emphasis> J4

A strictly anaerobic mesophilic chitinolytic bacterial strain identified as Clostridium paraputrificum J4 was isolated from human feces. In response to various types of growth substrates, the bacterium produced an array of chitinolytic enzymes representing significant components of the J4 strain secretome. The excreted active proteins were characterized by estimating the enzymatic activities of endochitinase, exochitinase, and N-acetylglucosaminidase induced by cultivation in medium M-10 with colloidal chitin. The enzyme activities produced by J4 strain cultivated in medium M-10 with glucose were significantly lower. The spectrum of extracellularly excreted proteins was separated by SDS-PAGE. The chitinase variability was confirmed on zymograms of renatured SDS-PAGE. The enzymes were visualized under ultraviolet light by using 4-methylumbelliferyl derivatives of N-acetyl-β-d-glucosaminide, N,N′-diacetyl-β-d-chitobiose, or N,N′,N˝-triacetyl-β-d-chitotriose for β-N-acetylglucosaminidase, chitobiosidase, or endochitinase activities, respectively. Protein components of the secretome were separated by 2D-PAGE analysis. The distinct protein bands were excised, isolated, and subsequently characterized by using MALDI-TOF/TOF tandem mass spectrometry. The final identification was performed according to sequence homology by database searching.     

Development of the polyembryonic parasitoid Copidosoma floridanum in Trichoplusia ni

Trichoplusia ni (Hübner) (Lepidoptera: Noctuidae) parasitized by the polyembryonic egg-larval parasitoid Copidosoma floridanum (Ashmead) (Hymenoptera: Encyrtidae) attained significantly larger final weights and head capsule widths than unparasitized controls. The difference in weight between parasitized and unparasitized hosts was not entirely accounted for by the weight of the C. floridanum brood. The head capsule widths of all parasitized and unparasitized fifth instars used in the study exceeded the critical threshold of 1.66 mm previously established for T. ni metamorphosis. The critical ratios associated with each T. ni instar of: 1) maximum weight within the instar:head capsule width and 2) maximum weight within the instar:weight at the beginning of the instar differed between parasitized and unparasitized larvae. Development of C. floridanum was synchronized with that of its host. Germ band formation and gastrulation of morulae destined to produce reproductive larvae invariably coincided with the host molt to the ultimate, fifth instar. Reproductive larvae had two instars. Eclosion from the egg to the first instar occurred during day 2 of the host's fifth instar, and ecdysis from the first to the second instar was synchronized with host cocoon spinning. Conversely, embryogenesis of morulae destined to produce precocious larvae began during the host first instar, continued through the second and third instar and ceased during the penultimate, fourth instar. Precocious larvae never molted and died when the host was consumed by the reproductive larvae.

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Résumé T. ni Hübner parasité par le parasitoïde ovo-larvaire C. floridanum Ashmead à développement polyembryonnaire atteint un poids final signficativement plus élevé avec une capsule céphalique plus grosse que les témoins non parasités, sans subir de mues surnuméraires. La différence de poids entre noctuelles parasitées ou non ne correspondait pas entièrement au poids des C. floridanum. Les largeurs des capsules céphaliques de tous les T. ni du cinquième stade dépassaient toutes le seuil critique de 1,66 mm lié à la métamorphose, mais les seuils critiques de taille du corps:largeur de la capsule céphalique et/ou taille et corps, taille initiale du corps au début du stade associé à la mue, différaient chez T. ni parasités ou non. Les développements de T. ni et de C. floridanum étaient synchrones. La formation de la bande germinative et la gastrulation de la morula produisant la multiplication des larves ont coïncidé invariablement avec la mue de l'hôte donnant le dernier stade. Les larves polyembryonnaires ont présenté deux stades. L'éclosion des oeufs s'est produite le deuxième jour du cinquième stade de T. ni, et le passage du premier au second était synchrone de la formation du cocon de l'hôte. Réciproquement, l'embryogenèse de la morula qui donnait des larves précoces commençait pendant le premier stade de l'hôte et se poursuivait à travers les second et troisième, pour cesser pendant le quatrième et pénultième stade.

     

Characterization of Chitinolytic and Antifungal Activities in Marine-Derived Trichoderma bissettii Strains

Trichoderma fungi have been intensively studied for mycoparasitism, and the latter is closely related to their cell-wall degrading enzymes including chitinase. Here, we studied marine-derived Trichoderma spp., isolated from distinct sources and locations, for chitinolytic and antifungal activity. Based on morphological and phylogenetic analyses, two strains designated GJ-Sp1 and TOP-Co8 (isolated from a marine sponge and a marine alga, respectively) were identified as Trichoderma bissettii. This species has recently been identified as a closely related species to Trichoderma longibrachiatum. The extracellular crude enzymes of GJ-Sp1 and TOP-Co8 showed activities of chitobiosidase and β-N-acetylglucosaminidase (exochitinase) and chitotriosidase (endochitinase). The optimum chitinolytic activity of the crude enzymes was observed at 50 °C, pH 5.0, 0–0.5% NaCl concentrations, and the activities were stable at temperatures ranging from 10 to 40 °C for 2 h. Moreover, the crude enzymes showed inhibitory activity against hyphal growth of two filamentous fungi Aspergillus flavus and Aspergillus niger. To the best of our knowledge, this is the first report of the chitinolytic and antifungal activity of T. bissettii.     

Comparative bioactivity of selected extracts from Meliaceae and some commercial botanical insecticides against two noctuid caterpillars, <Emphasis Type="Italic">Trichoplusia ni</Emphasis> and <Emphasis Type="Italic">Pseudaletia unipuncta</Emphasis>

Plant-derived extracts and phytochemicals have long been a subject of research in an effort to develop alternatives to conventional insecticides but with reduced health and environmental impacts. In this review we compare the bioactivities of some plant extracts with those of commercially available botanical insecticides against two important agricultural pests, the cabbage looper, Trichoplusia ni and the armyworm, Pseudaletia unipuncta. Test materials included extracts of Azadirachta indica (neem), A. excelsa (sentang), Melia volkensii, M. azedarach (Chinaberry) and Trichilia americana, (all belonging to the family Meliaceae) along with commercial botanical insecticides ryania, pyrethrum, rotenone and essential oils of rosemary and clove leaf. Most of the extracts and botanicals tested proved to be strong growth inhibitors, contact toxins and significant feeding deterrents to both lepidopteran species. However, there were interspecific differences with T. ni generally more susceptible to the botanicals than the armyworm, P. unipuncta. All botanicals were more inhibitory to growth and toxic (through feeding) to T. ni than to P. unipuncta, except for M. azedarach which was more toxic to P. unipuncta than to T. ni. Athough, pyrethrum was the most toxic botanical to both noctuids, A. indica, A. excelsa, and M. volkensii were more toxic than ryania, rotenone, clove oil and rosemary oil for T. ni. As feeding deterrents, pyrethrum was the most potent against T. ni, whereas A. indica was the most potent against the armyworm. Based upon growth inhibition, chronic toxicity, and antifeedant activity, some of these plant extracts have levels of activity that compare favorably to botanical products currently in commercial use and have potential for development as commercial insecticides.     

Resistance to herbicide and susceptibility to herbivores: environmental variation in the magnitude of an ecological trade-off

Trade-offs can maintain genetic diversity and constrain adaptation; however, their magnitude may depend on ecological factors. I considered whether resistance to the herbicide triazine in Amaranthus hybridus (Amaranthaceae) imposed the trade-off of increasing susceptibility to herbivorous insects. I grew triazine-resistant and triazine-susceptible plants under contrasting levels of light and fertilization, and quantified susceptibility to herbivores using the specialist Disonycha glabrata (Coleoptera: Chrysomelidae) and the generalist Trichoplusia ni (Lepidoptera: Noctuidae). Resistance to triazine increased susceptibility to both species of herbivorous insects, as manifested by greater feeding preference, growth, and survival of herbivores. However, these effects were more pronounced with T. ni and for plants grown under high light. My results demonstrate the presence of a trade-off between resistance to triazine and susceptibility to herbivorous insects that may in turn impose an ecologically based fitness cost, and illustrate the potential for this cost to vary across environments.     

Decreased Response to Feeding Deterrents Following Prolonged Exposure in the Larvae of a Generalist Herbivore, Trichoplusia ni (Lepidoptera: Noctuidae)

We investigated the role of experience with several antifeedants on the feeding behavior of a generalist herbivore, Trichoplusia ni. Second-, third-, and fifth-instar larvae of T. ni were examined for their feeding responses to plant extracts (Melia volkensii, Origanum vulgare) and individual plant allelochemicals (cymarin, digitoxin, xanthotoxin, toosendanin, and thymol), after being exposed to them continually beginning as neonates. All tested instars of T. ni were capable of showing a decreased antifeedant response following prolonged exposure to most of the antifeedants tested compared with their naive conspecifics. Cardenolides (digitoxin and cymarin) were the exceptions. The response to oregano was affected as a result of previous exposure to different concentrations of oregano, unlike M. volkensii, leading us to conclude that T. ni sensitivity varies between stimuli and cannot be generalized. To demonstrate that decreased deterrence following prolonged exposure to M. volkensii was the result of learned habituation, three aversive stimuli were used. A high concentration of the particular antifeedant, xanthotoxin, acted as a noxious stimulus and dishabituated (reversed) the decrement in the antifeedant response to M. volkensii. Cold shock or CO₂ was marginally effective in dishabituating the response. The fact that the decrease in antifeedant response can be dishabituated has implications for pest management.     

Resistance to Alternaria brassicicola in transgenic broccoli expressing a Trichoderma harzianum endochitinase gene

Transgenic broccoli plants expressing a Trichoderma harzianum endochitinase gene were obtained by Agrobacterium tumefaciens-mediated transformation. PCR and Southern blot analysis confirmed the presence of the gene in plants initially selected via resistance to kanamycin. Primary transformants (T₀) and selfed progeny (T₁) were examined for expression of the endochitinase gene using a fluorometric assay and for their resistance to the fungal pathogens Alternaria brassicicola and Sclerotinia sclerotiorum. All transgenic plants with elevated endochitinase activity had the expected 42 kDa endochitinase band in western blot analysis, whereas no such band was detected in the non-transgenic control. Leaves of most mature T₀ plants had 14–37 times higher endochitinase activity than controls; mature T₁ plants had higher endochitinase activity (100–200 times that in controls), in part because of lower control values. T₀ plantlets in vitro or young plants in soil had higher absolute and relative endochitinase activity. When detached leaves of T₀ plants were inoculated with A. brassicicola, lesion size showed a significant negative correlation with endochitinase levels. After inoculation of two-month old T₀ plants with A. brassicicola, all 15 transgenic lines tested showed significantly less severe disease symptoms than controls. In contrast, lesion size on petioles of T₀ and T₁ plants inoculated with S. sclerotiorum was not statistically different from controls.     

Sulfhydryl-reagent alteration of soybean resistance to the cabbage looper, Trichoplusia ni

The hypothesis that the sulfhydryl reagent, N-ethylmaleimide, would function as an elicitor of alterable resistance in soybean (Glycine max) plants to Trichoplusia ni herbivory was tested experimentally under greenhouse conditions. This elicitory chemical, which allows receptor thiols to add across its carbon-carbon double bond, altered the resistance in one or more leaves of plants at one or more intervals after treatment; and thus yielded results supporting the hypothesis. Leaf dipping and soil application were both effective methods of treatment. Results support the interpretation that an elicitor may function in intact plants by altering the integrity of sulfhydryl groups in receptor macromolecules which are also involved in signaling a change in the plant's biosynthesis of characteristic defensive compounds such as phenylpropanoids including antifeedant and antibiotic flavonoids. Induced feeding non-preference by T. ni was highly correlated positively with the amount of glyceollins in the leaf.     

Leaf shape variation and herbivore consumption and performance: a case study with Ipomoea hederacea and three generalists

The effect of leaf shape variation on plant-herbivore interactions has primarily been studied from the perspective of host seeking behavior. Yet for leaf shape to affect plant-herbivore coevolution, there must be reciprocal effects of leaf shape variation on herbivore consumption and performance. We investigated whether alternative leaf morphs affected the performance of three generalist insect herbivores by taking advantage of a genetic polymorphism and developmental plasticity in leaf shape in the Ivyleaf morning glory, Ipomoea hederacea. Across four experiments, we found variable support for an effect of leaf shape genotype on insects. For cabbage loopers (Trichoplusia ni) and corn earworms (Helicoverpa zea) we found opposing, non-significant trends: T. ni gained more biomass on lobed genotypes, while H. zea gained more biomass on heart-shaped genotypes. For army beetworms (Spodoptera exigua), the effects of leaf shape genotype differed depending on the age of the plants and photoperiod of growing conditions. Caterpillars feeding on tissue from older plants (95 days) grown under long day photoperiods had significantly greater consumption, dry biomass, and digestive efficiency on lobed genotypes. In contrast, there were no significant differences between heart-shaped and lobed genotypes for caterpillars feeding on tissue from younger plants (50 days) grown under short day photoperiods. For plants grown under short days, we found that S. exigua consumed significantly less leaf area when feeding on mature leaves than juvenile leaves, regardless of leaf shape genotype. Taken together, our results suggest that the effects of leaf shape variation on insect performance are likely to vary between insect species, growth conditions of the plant, and the developmental stage and age of leaves sampled. Handling editor: May Berenbaum.     

Fitness of Bt‐resistant cabbage loopers on Bt cotton plants

Development of resistance to the insecticidal toxins from Bacillus thuringiensis (Bt) in insects is the major threat to the continued success of transgenic Bt crops in agriculture. The fitness of Bt‐resistant insects on Bt and non‐Bt plants is a key parameter that determines the development of Bt resistance in insect populations. In this study, a comprehensive analysis of the fitness of Bt‐resistant Trichoplusia ni strains on Bt cotton leaves was conducted. The Bt‐resistant T. ni strains carried two genetically independent mechanisms of resistance to Bt toxins Cry1Ac and Cry2Ab. The effects of the two resistance mechanisms, individually and in combination, on the fitness of the T. ni strains on conventional non‐Bt cotton and on transgenic Bt cotton leaves expressing a single‐toxin Cry1Ac (Bollgard I) or two Bt toxins Cry1Ac and Cry2Ab (Bollgard II) were examined. The presence of Bt toxins in plants reduced the fitness of resistant insects, indicated by decreased net reproductive rate (R₀) and intrinsic rate of increase (r). The reduction in fitness in resistant T. ni on Bollgard II leaves was greater than that on Bollgard I leaves. A 12.4‐day asynchrony of adult emergence between the susceptible T. ni grown on non‐Bt cotton leaves and the dual‐toxin‐resistant T. ni on Bollgard II leaves was observed. Therefore, multitoxin Bt plants not only reduce the probability for T. ni to develop resistance but also strongly reduce the fitness of resistant insects feeding on the plants.     

Oviposition preference,larval performance and adaptation of Trichoplusia ni on cabbage and cotton

Most female herbivores ensure to lay eggs where their offspring can develop successfully. The oviposition preferences of females affect strategies in pest management. In this study, the performance of two cohorts of Trichoplusia ni larvae on cabbage and cotton (after they had been transferred from their original host plants) were investigated. The preferences of female moth ovipositing and larval feeding on these two host plants were observed. The results indicated that plants significantly affected oviposition preference of the female adults and development and survival of larvae of T. ni. All females preferred to lay eggs on cabbage than cotton regardless from which host they originated. The detrimental effects of cotton on the development and survival of T. ni larvae originated from cabbage (CaTn) increased with the increase of the larval age when they were transferred. In addition, the host plant change did not significantly affect the development and survival of larvae of T. ni originating from cotton (CoTn). Larvae of CaTn preferred cabbage plants as compared to cotton plants, whereas larvae of CoTn did not show a significant choice. Although the adult females preferred laying eggs on cabbage, they did not show preferences between cotton and cabbage in a Y‐tube olfactometer test. The hypothesis of oviposition preference and performance of larvae was supported by the results of CaTn, whereas they not supported by those from CoTn. Based on these results, the strategy to manage this serious pest was discussed.     

Comparative growth inhibitory and antifeedant effects of plant extracts and pure allelochemicals on four phytophagous insect species

Antifeedant and growth inhibitory effects of crude plant extracts (Melia volkensii and Origanum vulgare) and pure allelochemicals (digitoxin, cymarin, xanthotoxin, toosendanin, thymol and trans‐anethole) were investigated in the cabbage looper (Trichoplusia ni), and in the armyworm (Pseudaletia unipuncta) using different bioassays. Antifeedant effects of M. volkensii, O. vulgare and thymol were investigated in larvae of the diamondback moth (Plutella xylostella), and of O. vulgare and thymol in the Mexican bean beetle (Epilachna varivestis), using leaf disc choice bioassays. M. volkensii was the most potent growth inhibitor for T. ni and P. unipuncta (dietary EC₅₀ = 7.6 and 12.5 p.p.m., respectively) of all the test substances. Cymarin was the second most potent growth inhibitor (EC₅₀ = 132.0 p.p.m.) for T. ni. The most effective feeding deterrents for third instar T. ni larvae were xanthotoxin andM. volkensii (DC₅₀ = 0.9 and 8.3 μg/cm², respectively). M. volkensii was also the most potent feeding deterrent for third instar P. unipuncta, P. xylostella and adult E. varivestis (DC₅₀ = 10.5, 20.7 and 2.3 μg/cm², respectively). Because of interspecific differences in response to feeding deterrents and the lack of a strong relationship between EC₅₀ and DC₅₀ values, we recommend testing a battery of bioassay species with candidate compounds and the use of more than one bioassay. Based on their growth inhibitory and feeding deterrent properties, some of these plant extracts and pure allelochemicals have potential for use as alternative crop protectants against a number of pest species.     

Effect of dietary selenium supplementation on resistance to baculovirus infection

We have examined the effects of dietary selenium (Se) supplementation on larval growth and immunocompetence of the lepidopteran pest, the cabbage looper, Trichoplusia ni. Supplementation of the diet of T. ni larvae with 10–20 ppm Se resulted in a 1 day delay in pupation. The effects of the addition and/or removal of dietary Se on total Se bioaccumulation and sequestration were determined by neutron activation analysis of pupae. Early penultimate instar larvae moved from selenium containing diet to basal diet lost total pupal Se content down to the level of those fed basal diet. Conversely, larvae moved from basal diet to diet containing additional Se rapidly attained pupal Se levels comparable to larvae fed Se throughout larval development. Therefore, dietary Se is rapidly accumulated or lost during larval development, but significant amounts are sequestered from diet into pupae. Larvae were reared on diet supplemented with 5 or 10 ppm Se until the onset of the penultimate instar then infected per os with increasing concentrations of the fatal baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV). Larvae fed Se in the penultimate and ultimate instars were more resistant to viral infection than larvae not fed Se in the final instars. This study indicates that dietary Se levels rapidly impact Se assimilation and sequestration and that tissue Se levels are an important factor in resistance to AcMNPV infection in larval T. ni.