Arrestant responses of southwestern corn borer larvae to free amino acids: Structure-activity relationships

The leaf-feeding resistance of corn or maizeZea mays L. to the southwestern corn borer, SWCB,Diatraea grandiosella Dyar has been attributed at least in part to decreased protein, increased crude fiber, and increased hemicellulose in the whorls of resistant genotypes. In this study, individual amino acids and sugars were evaluated as arrestants, with the objective of identifying those that gave weak or negative responses. Several structure-activity relationships were identified. Larvae responded to three-carbonn alkyl alpha amino acids more than to two-, four-, five-, and six-carbon compounds. Amino acids with terminal isopropyl functions gave decreased responses relative to theirn-alkyl counterparts. Dicarboxylic acids and their amides gave the lowest responses of all classes of amino acids. The normally occurring basic amino acids were all good arrestants. The guanido [HN:C(NH₂)NH-] function was somewhat important to an arrestant response, as was the number of methylenes between the alpha and omega amino functions of diaminon-alkyl amino acids. Hydroxy amino acids were generally good arrestants unless the hydroxyl was located on a ring system. The two sugars present in expressed corn whorl juice, glucose and fructose, gave poor responses. However, two other sugars, mannose and arabinose, whose C-2 hydroxyls are conformationally in the axial position, were strongly arrestant. Formulated amino acid mixtures based on their content in whorl juice were as strong arrestants as whorl juice. However, the relative contributions of amino acids and sugars that are weak arrestants to the resistance of corn to SWCB larvae is uncertain because amino acid analyses did not reveal significantly higher contents of these amino acids in the whorl juices of resistant lines.     

Hemicellulose is an important leaf-feeding resistance factor in corn to the fall armyworm

The fall armyworm,Spodoptera frugiperda (J. E. Smith), (FAW) is a major pest of corn,Zea mays L., in the southeastern United States. The damage to pretassel corn is caused by larvae feeding primarily on immature inner whorls. In this study, resistant lines were found to contain more crude fiber in whorls, mostly hemicellulose and cellulose. While hemicellulose, chiefly an arabinoxylan, was higher in resistant (R) lines than in susceptible (S) lines, the distribution of constituent neutral sugars was very similar in the lines. Both lines also containedp-coumaric and ferulic acids. These phenolic acids are known to occur both in the free state and in the cell wall as complexes bound by ester linkages to the arabinose moiety of the arabinoxylan.¹³C NMR data showed that the intensity of the carbonyl carbon (184 ppm) in resistant hemicellulose was stronger, indicating a greater degree of cross-linking. Thus, resistant hemicellulose is both structurally different from susceptible hemicellulose and present in greater quantities. In two of three laboratory dietary tests, FAW larval weight gains were significantly higher on diets with (S) hemicellulose incorporated at the same level as (R) hemicellulose. Therefore, resistance to the FAW appears to be correlated with both a greater amount and a higher degree of cross-linking of the hemicellulose of (R) lines.Mention of a trademark, proprietary product, or vendor does not constitute a guarantee of warranty of the product by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products or vendors that may also be suitable.     

Physiological, Nutritional, and Biochemical Bases of Corn Resistance to Foliage-Feeding Fall Armyworm

Three corn (Zea mays) germplasm lines [i.e., Ab24E (susceptible control), Mp708 (resistant control), and a locally selected partial inbred line FAW7050 (resistant)] were examined for Spodoptera frugiperda (J.E. Smith; Lepidoptera: Noctuidae) resistance. Nutritional [i.e., total protein content, amino acids, glucose, total nonstructural carbohydrates (TNC), protein to TNC (P/C) ratios] and biochemical (i.e., peroxidase and lipoxygenase 3) properties in the seedlings of these corn lines were examined to categorize resistance mechanisms to S. frugiperda. Physiological changes in photosynthetic rates also were examined in an attempt to explain nutritional and biochemical dynamics among corn germplasm lines and between insect-infested and noninfested corn plants within a germplasm line. Results indicated that S. frugiperda larvae survived better and developed faster in susceptible Ab24E than in resistant FAW7050 or Mp708. The three germplasm lines differed in resistance mechanisms to S. frugiperda, and the observed patterns of resistance were probably collective results of the P/C ratio and defensive proteins. That is, the susceptibility of Ab24E to S. frugiperda was due to a high P/C ratio and a low level of induced defensive compounds in response to insect herbivory, while the resistance of FAW7050 resulted from elevated defensive proteins following insect herbivory, low P/C ratio, and elevated defensive proteins in Mp708 contributed to its resistance to S. frugiperda. The elevated protein amounts in resistant Mp708 and FAW7050 following S. frugiperda injury were likely due to greater conversion of photosynthates to defensive proteins following the greater photosynthetic rates in these entries. Greater photosynthetic capacity in Mp708 and FAW7050 also led to higher amino acid and glucose contents in these two lines. Neither amino acid nor lipoxygenase 3 played a critical role in corn resistance to S. frugiperda. However, high inducibility of peroxidase may be an indicator of S. frugiperda susceptibility as observed elsewhere.     

Sources of fall armyworm,Spodoptera frugiperda (Lepidoptera: Noctuidae), kairomones eliciting host-finding behavior inCotesia (=Apanteles) marginivenitris (Hymenoptera: Braconidae)

Bioassay responses inCotesia marginiventris (Cresson) females to materials derived from fall armyworm (FAW) larvae,Spodoptera frugiperda (J. E. Smith), were most intense for frass and somewhat less intense for larval and pupal cutical materials, scales, exuviae, silk, and oral secretion, with FAW larval hemolymph eliciting only a slight response. The highest percentage of ovipositor probing was caused by frass (100%) and moth scales (90%). Various types of corn-leaf damage when assayed alone did not produce responses as intense as when assayed in combination with frass, cuticle material, and oral secretion. Parasitoid response was somewhat better to frass derived from FAW larvae feeding on corn and peanut leaves than from larvae feeding on the foliage of soybeans, Bermuda grass, cowpeas, or laboratory diet. Hexane and chloroform were better than methanol and water for extracting active material from FAW frass, and chloroform was the best of these solvents for extracting corn leaves. Serial dilutions of frass extracts resulted in a reduction in parasitoid response.This paper was submitted in partial fulfillment of the requirements for Doctor of Philosophy, Department of Entomology and Nematology, University of Florida, Gainesville.     

Effects of Peruvian Maize Extracts on Growth, Development, and Fecundity of the European Corn Borer

Twelve Peruvian maize, Zea mays, accessions were selected because of their relatively high level of field resistance to first-generation European corn borer (ECB), Ostrinia nubilalis, larval leaf-feeding. Water extracts of freeze-dried, powdered, leaf tissue were incorporated into a standard ECB diet, fed to larvae, and the effects on larval growth, development, and fecundity were measured. Larval and pupal weights were monitored as were the time elapsed in the larval, pupal, and adult stages. Adult fecundity and egg fertility were recorded. The experiment was a randomized block design (larvae and pupae) or a completely randomized design (adults) and analyzed with ANOVA ( = 0.05). Pairwise comparisons were made between groups of insects grown on diets containing extracts from the Peruvian lines, a standard diet, or diets containing extracts of a known susceptible inbred, and a known resistant inbred line. Survival was analyzed with a chi-squared test ( = 0.05). Two Peruvian accessions significantly reduced female larval and pupal weights, extended pupal and adult development time, and decreased survival of pupae and adults. Water extracts also had a pronounced impact on males; two accessions significantly reduced pupal weight and extended the time required to pupate, and one reduced male survival to adults. The results indicate that water-soluble factors from resistant Peruvian accessions inhibit the growth, developmental time, and survival of ECB. These resistance factors could be useful in the development of maize germplasm with insect-resistant traits.     

Effects of cotton plant allelochemicals and nutrients on behavior and development of tobacco budworm

Female moths of the tobacco budworm,Heliothis virescens (F.), oviposit in the terminals of the cotton plant,Gossypium hirsutum (L.). The hatched larvae migrate to the terminal area and then to small squares (buds), on which they feed, finally burrowing into the anthers where they grow and develop. They attempt to avoid gossypol glands as they feed. Chemically related evidence explains, in part, these observations. The calyx crown of resistant lines (which is avoided) is high in the terpenoid aldehydes (TAs) including gossypol. HPLC data showed that the gossypol content of both susceptible and resistant glanded lines is equal, while the hemigossypolone and heliocides H₁ and H₂ are greatly increased in resistant lines and presumably are more closely associated with resistance. Analysis for total amino acids in cotton square tissues showed that there was a gradation from the calyx and calyx crown, which were lowest, to the anthers, the site of final insect development, which were highest. Synthetic diets mimicking amino acid distribution in anthers were found to be successful for larval growth and development.     

2-hydroxy-4,7-dimethoxy-1,4-benzoxazin-3-one (N-O-Me-DIMBOA), a possible toxic factor in corn to the southwestern corn borer

The southwestern corn borer (SWCB),Diatraea grandiosella Dyar, is a major pest of corn,Zea mays L., in the southern United States. The damage to corn is caused primarily by larval feeding on leaf, ear, and stem tissues. In this study, 2-hydroxy-4,7-dimethoxy-1,4-benzoxazin-3-one (N-O-Me-DIMBOA) was identified by MS and NMR as present in corn whorl surface waxes. This compound has evidently not been isolated previously, but its glucoside has been reported in corn, wheat, andCoix lachryma. It is present in the waxes in a higher concentration than DIMBOA (2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one) and 6-MBOA (6-methoxybenzoxazolinone). It was toxic to the SWCB in a stress diet, but it was less toxic to this insect than 6-MBOA when incorporated in the standard rearing diet. Nevertheless, it may have some role in the resistance of corn to the SWCB because the total surface wax content is higher in resistant lines than in susceptible lines.Mention of a trademark, proprietary product, or vendor does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products or vendors that may also be suitable.     

Activation of plant foliar oxidases by insect feeding reduces nutritive quality of foliage for noctuid herbivores

The foliage and fruit of the tomato plantLycopersicon esculentum contains polyphenol oxidases (PPO) and peroxidases (POD) that are compartmentally separated from orthodihydroxyphenolic substrates in situ. However, when leaf tissue is damaged by insect feeding, the enzyme and phenolic substrates come in contact, resulting in the rapid oxidation of phenolics to orthoquinones. When the tomato fruitwormHeliothis zea or the beet army-wormSpodoptera exigua feed on tomato foliage, a substantial amount of the ingested chlorogenic acid is oxidized to chlorogenoquinone by PPO in the insect gut. Additionally, the digestive enzymes of the fruitworm have the potential to further activate foliar oxidase activity in the gut. Chlorogenoquinone is a highly reactive electrophilic molecule that readily binds cova-lently to nucleophilic groups of amino acids and proteins. In particular, the —SH and —NH₂ groups of amino acids are susceptible to binding or alkylation. In experiments with tomato foliage, the relative growth rate of the fruitworm was negatively correlated with PPO activity. As the tomato plant matures, foliar PPO activity may increase nearly 10-fold while the growth rate of the fruitworm is severely depressed. In tomato fruit, the levels of PPO are highest in small immature fruit but are essentially negligible in mature fruit. The growth rate of larvae on fruit was also negatively correlated with PPO activity, with the fastest larval growth rate occurring when larvae fed on mature fruit. The reduction in larval growth is proposed to result from the alkylation of amino acids/protein byo-quinones, and the subsequent reduction in the nutritive quality of foliage. This alkylation reduces the digestibility of dietary protein and the bioavailability of amino acids. We believe that this mechanism of digestibility reduction may be extrapolatable to other plant-insect systems because of the ubiquitous cooccurrence of PPO and phenolic substrates among vascular plant species.     

Toxicity of allelochemicals from wild insect-resistant tomatoLycopersicon hirsutum f.glabratum toCampoletis sonorensis,a parasitoid ofHeliothis zea

Greenhouse-grown plants of five tomato lines varying in their level of 2-tridecanone-mediated resistance toManduca sexta (L.) andLeptinotarsa decemlineata (Say) were found to adversely affect larvae ofCampoletis sonorensis (Cameron), a larval endoparasitoid ofHeliothis zea (Boddie), in a manner directly related to their level of resistance. The parasitoid larvae, which emerge as fifth instars from their host and construct a cocoon on the foliage of their hosts' host plant, suffered extensive mortality during cocoon spinning on highly resistant foliage. Mortality was greatest (82%) on the highly resistant plants ofLycopersicon hirsutum f.glabratum (accession PI 134417) and an F₁ backcross [(L. esculentum × PI 134417) × PI 134417] selection. Mortality was intermediate (40 and 28%, respectively) on backcross selections with moderate and low levels of resistance and least (8%) on susceptibleL. esculentum. Removal of the glandular trichomes, which contain 2-tridecanone in their tips, from the foliage eliminated differences in parasitoid mortality among plant lines.Bioassays of 2-tridecanone indicated that it is acutely toxic to fifth instarC. sonorensis larvae at the quantities associated with highly resistant foliage and produces symptoms identical to those observed on resistant foliage. 2-Undecanone, a second methyl ketone present in the glandular trichomes of resistant foliage, was also toxic toC. sonorensis larvae, but significantly less so than 2-tridecanone. The results support the hypothesis that 2-tridecanone is responsible for the observed mortality ofC. sonorensis larvae during cocoon construction on resistant foliage.     

Leaf profile of maize resistance factors to European corn borer,Ostrinia nubilalis

The feeding preference of European corn borer larvae for immature whorl tissue of maize was examined by conducting leaf bioassays and quantifying resistance factors along the length of mid-whorl leaves from the maize synthetic BS9(C4) developed by recurrent selection for resistance. Potential resistance factors that were quantified included percent foliar nitrogen, gravimetric determination of soluble metabolites and fiber, soluble phenolics and hydroxamic acids, cell-wall-bound phenolics, leaf toughness, and UV absorbance of the epidermal cell wall determined by microspectrophotometry. Larvae consumed immature tissue at a higher rate than more mature tissue outside of the whorl, despite higher levels of DIMBOA in immature tissue. Consumption rate was highly negatively correlated with epidermal cell wall absorbance and leaf toughness. Fiber content and phenolic fortification of cell walls are proposed as the major resistance components that influence European corn borer feeding preference within the resistant synthetic BS9(C4).PRC contribution no. 1562.     

Protective action of midgut catalase in lepidopteran larvae against oxidative plant defenses

Catalase activity was detected in the midgut tissues and regurgitate of several lepidopteran pests of the tomato plant. Greatest activity in the midgut was detected in larvalHelicoverpa zea, followed bySpodoptera exigua, Manduca sexta, andHeliothis virescens. We present evidence that catalase, in addition to removing toxic hydrogen peroxide, may inhibit the oxidation of plant phenolics mediated by plant peroxidases. Small amounts of larval regurgitate significantly inhibited foliar peroxidase activity via removal of hydrogen peroxide. Treatment of foliage with purified catalase nearly eliminated peroxidase activity and was superior as a larval food source compared to untreated foliage. Tomato foliar peroxidases oxidize an array of endogenous compounds including caffeic acid, chlorogenic acid, rutin, coumaric acid, cinnamic acid, and guaiacol. The oxidized forms of these compounds are potent alkylators of dietary and/or cellular nucleophiles (e.g., thiol and amino functions of proteins, peptides, and amines). When tomato foliar protein was pretreated with peroxidase and chlorogenic acid and incorporated in artificial diet, larval growth was reduced compared to larvae fed untreated protein. Thus, the diminution of peroxidase activity and removal of hydrogen peroxide by catalase may represent an important adaptation to leaf-feeding. The secretion of catalase in salivary fluid during insect feeding is also suggested to be a potential mechanism for reducing hydrogen peroxide formation as an elicitor of inducible plant defenses.     

Variation among and within mountain birch trees in foliage phenols,carbohydrates, and amino acids,and in growth ofEpirrita autumnata larvae

Leaf quality of the mountain birch (Betula pubescens ssp.tortuosa) for herbivores was studied at several hierarchical levels: among trees, among ramets within trees, among branches within ramets, and among short shoots within branches. The experimental units at each level were chosen randomly. The indices of leaf quality were the growth rate of the larvae of a geometrid,Epirrita autumnata, and certain biochemical traits of the leaves (total phenolics and individual phenolic compounds, total carbohydrates and individual sugars, free and protein-bound amino acids). We also discuss relationships between larval growth rate and biochemical foliage traits. Larval growth rates during two successive years correlated positively at the level of tree, the ramet, and the branch, indicating that the relationships in leaf quality remained constant between seasons both among and within trees. The distribution of variation at different hierarchical levels depended on the trait in question. In the case of larval growth rate, ramets and short shoots accounted for most of the explained variation. In the case of biochemical compounds, trees accounted for most of the variance in the content of total phenolics and individual low-molecular-weight phenolics. In the content of carbohydrates (total carbohydrates, starch, fructose, glucose, and sucrose) and amino acids, variation among branches was generally larger than variation among trees. Variation among ramets was low for most compounds. No single leaf trait played a paramount role in larval growth. Secondary compounds, represented by phenolic compounds, or primary metabolites, particularly sugars, may both be important in determining the suitability of birch leaves for larvae. If phenols are causally more important, genet-specific analyses of foliage chemistry are needed. If sugars are of primary importance, within-genet sampling and analysis of foliage chemistry are necessary.     

Behavioral and biological responses ofCotesia marginiventris to kairomones of the fall armyworm,Spodoptera frugiperda

Potential kairomone sources of the fall armyworm (FAW),Spodoptera frugiperda (J.E. Smith), were bioassayed with females ofCotesia marginiventris (Cresson) in Petri dishes (10 cm diam). MatedC. marginiventris females, ranging in age from 1 to 3 days exhibited the most intense bioassay responses to potential sources of kairomone. Contacting a FAW-damaged corn leaf modified the pattern of movement inC. marginiventris from random to one exhibiting a significant increase in klinokinesis. No significant differences were present in kairomone responses of nonconditioned and conditioned parasitoids and parasitoid response to kairomones did not change throughout the photophase (0800–1800 hr). Removal of one, both, or the first eight antennal segments reduced or eliminated the response of the parasitoid to kairomones. Female parasitoids did not exhibit a preference for corn leaves damaged by a particular fall armyworm instar and parasitization rates were highest in larvae 48 hr old.Submitted in partial fulfillment of the requirements for Doctor of Philosophy, Department of Entomology and Nematology, University of Florida, Gainesville.     

How contact foraging experiences affect preferences for host-related odors in the larval parasitoidCotesia marginiventris (Cresson) (Hymenoptera: Braconidae)

Responses of individual females of the parasitoidCotesia marginiventris to the odors of four different complexes of host larvae feeding on leaves were observed in a four-arm olfactometer. The plant-host complexes were composed of fall armyworm (FAW) larvae or cabbage looper (CL) larvae feeding on either corn or cotton seedlings. Prior to testing, each female was given a brief foraging experience on a plant-host complex and was then exposed to the odors of the same complex in the olfactometer. The experienced females responded to familiar odors in a dose-related manner, and these responses were virtually identical to all four complexes. Preferences for the odors of one of two plant-host complexes were tested in dual choice situations. Generally, FAW odors were preferred over CL odors and corn odors over cotton odors. A short foraging experience significantly affected the females' odor preferences in favor of the odors released by the experienced complex. Additional experiments revealed that neither longer bouts of experience nor bouts that included ovipositions resulted in a stronger change in preference. Experience affected preference in combinations where only the host species was varied as well as in combinations where only the plant species was varied. The results, therefore, strongly indicate that both the plants and the hosts somehow are involved in the production and/or release of the semiochemicals that attractC. marginiventris.Mention of a proprietary product does not constitute an endorsement or the recommendation for its use by the USDA.     

Reassessment of the role of gut alkalinity and detergency in insect herbivory

Previously it was reported that significant amounts of the tomato phenolic, chlorogenic acid, were oxidized in the digestive system of generalist feedersSpodoplera exigua andHelicoverpa zea. The covalent binding of the oxidized phenolic (i.e., quinone) to dietary protein exerts a strong antinutritive effect against larvae. In this study, we examined the fate of ingested chlorogenic acid in larvalManduca sexta, a leaf-feeding specialist of solanaceous plants. Significant amounts of chlorogenic acid were bound to excreted protein byM. sexta when larvae fed on tomato foliage. However, in the case ofM. sexta we suggest that the strong alkalinity and detergency of the midgut may minimize the antinutritive effects of oxidized phenolics. The solubility of tomato leaf protein is significantly greater at pH 9.7, representative of the midgut ofM. sexta, than at pH 8.0, representative of the midguts ofH. zea and S. exigua. We suggest that this increase in solubility would compensate for any loss in bioavailability of essential amino acids caused by the covalent binding of chlorogenic acid to amino acids. Furthermore, lysolecithin, a surfactant likely to contribute to the detergent properties of the midgut fluid, was shown to enhance protein solubility as well as inhibit polyphenol oxidase activity. The adaptive significance of gut alkalinity and detergency is discussed.Approved by the Director of the Arkansas Agricultural Experiment Station.     

Plant defenses: Chlorogenic acid and polyphenol oxidase enhance toxicity ofBacillus thuringiensis subsp.kurstaki toHeliothis zea

Two chemicals implicated in resistance of the tomato plant, chlorogenic acid and polyphenol oxidase, are known to form orthoquinones in damaged plant tissue. Orthoquinones have been reported to alkylate –NH₂ and –SH groups of proteins and amino acids, altering solubility, digestibility, and, for some pathogenic viruses, infectivity. Here we explore effects of quinone alkylation on toxicity of an important microbial insecticide,Bacillus thuringiensis subsp.kurstaki (BTk), to larvalHeliothis zea. BTk incubated with these phytochemicals and fed to larvae was more toxic than untreated BTk. Similar but less dramatic results arose when BTk was incubated with polyphenol oxidase alone. Digestibility experiments suggest that alkylation enhanced the solubilization and/or proteolysis of crystal protein in vivo. Implications of our results for compatibility of BTk with host-plant resistance and biological control are discussed.     

Inhibition ofCampoletis sonorensis parasitism ofHeliothis zea and of parasitoid development by 2-tridecanone-mediated insect resistance of wild tomato

Field populations ofHeliothis spp. were sampled for levels of naturally occurring larval parasitism on six tomato lines varying in levels of 2-tridecanone-mediated resistance toManduca sexta (L.) andLeptinotarsa decemlineata (Say). Second and third instars were parasitized byCampoletis sonorensis (Cameron) (Hymenoptera: Ichneumonidae) andCotesia (=Apantales)marginiventris (Cresson) (Hymenoptera: Braconidae) in 1984 through 1986 and byMicropletis croceipes (Cresson) (Hymenoptera: Braconidae) in 1986. Differences in parasitism by individual and multiple species among host plants were not demonstrated. However, levels of parasitism were low and variable among replicates. Total larval parasitism averaged across all plant lines was less than 6% in 1984 and 1986 and approximately 11% in 1985. In laboratory cage studies,C. sonorensis parasitized fewerH. zea larvae on tomato foliage with high levels of 2-tridecanone than on foliage with low levels. RearingH. zea on diet containing 2-tridecanone and 2-undecanone did not alter incidence of parasitism byC, sonorensis; nor did rearing parasitizedH. zea larvae on chemically treated host diets precondition the parasitoid to higher or lower mortality when transferred to foliage as a substrate for cocoon spinning, regardless of the foliage genotype. However, parasitoid survival during cocoon spinning on foliage varied significantly among plant lines in a manner corresponding to the level of 2-tridecanone-mediated resistance of the foliage. Parasitoid mortality was greatest on highly resistant foliage and lowest on susceptible foliage.     

Evaluation of flavonoids inGossypium arboreum (L.) cottons as potential source of resistance to tobacco budworm

Asiatic cottons [Gossypium arboreum (L.)] have been investigated as a source of resistance to the tobacco budworm [Heliothis virescens (Fab.)] because their diversely colored petals have been presumed to contain various allelochemicals. However, we found that larvae fedG. arboreum squares (buds) grew about equally compared with those fed squares from commercialG. hirsutum lines. The best source of resistance was found in severalG. hirsutum double-haploid (DH) lines. In our investigation of allelochemicals, G.arboreum lines were found to contain much less gossypol in leaves, squares (buds), and petals thanG. hirsutum L. lines. Flavonoids were significantly higher inG. arboreum lines only in petals. Of 22G. arboreum lines from which squares were gathered and fed to tobacco budworm (TBW) larvae in the laboratory, larval growth was not significantly decreased on any, but larval survival was decreased on six. When the square flavonoids were isolated and incorporated in laboratory diets for the TBW, moderate toxicity was observed. However, the estimated toxicities were not greater than those of the same flavonoid isolates fromG. hirsutum lines. The most prevalent flavonoids, all previously found in G.arboreum plant tissues, were gossypetin 8-0-glucoside and gossypetin 8-0-rhamnoside, neither of which were present inG. hirsutum tissue. Quercetin 3-0-glucoside, quercetin-3-0-glucoside, and quercetin 7-0-glucoside were also present in significant amounts in both species. Gossypetin 8-0-rhamnoside and gossypetin 8-0-glucoside were the most toxic flavonoids tested (the ED₅₀% was estimated to be 0.007 and 0.024) and therefore may prove to be contributing factors of resistance to TBW feeding.Mention of a trademark, proprietary product, or vendor does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products or vendors that may also be suitable.     

Avoidance of antinutritive plant defense: Role of midgut pH in Colorado potato beetle

The fate of the tomato foliar phenolic, chlorogenic acid, in the digestive systems of Colorado potato beetleLeptinotarsa decemlineata (Coleoptera: Chrysomelidae) andHelicoverpa tea (Lepidoptera: Noctuidae) is compared. In larvalH. zea and other lepidopteran species previously examined, approximately 35–50% of the ingested chlorogenic acid was oxidized in the digestive system by foliar phenolic oxidases (i.e., polyphenol oxidase and peroxidase) from the tomato plant. The oxidized form of chlorogenic acid, chlorogenoquinone, is a potent alkylator of dietary protein and can exert a strong antinutritive effect upon larvae through chemical degradation of essential amino acids. In contrast, inL. decemlineata less than 4% of the ingested dose of chlorogenic acid was bound to protein. In vitro experiments to determine the influence of pH on covalent binding of chlorogenic acid to protein showed that 30–45% less chlorogenic acid bound to protein at pHs representative of the beetle midgut (pH 5.5–6.5) than at a pH representing the lepidopteran midgut (pH 8.5). At an acidic pH, considerably more of the alkylatable functional groups of amino acids (–NH₂, –SH) are in the nonreactive, protonated state. Hence, polyphenol oxidases are unlikely to have significant antinutritive effects against the Colorado potato beetle and may not be a useful biochemical source of resistance against this insect. The influence of feeding by larval Colorado potato beetle on foliar polyphenol oxidase activity in tomato foliage and its possible significance to interspecific competition is also considered.Approved by the Director of the Arkansas Agricultural Experiment Station.     

Sequestration ofVeratrum alkaloids by specialistRhadinoceraea nodicornis konow (Hymenoptera,Tenthredinidae) and its ecoethological implications

The larvae of the specialist sawflyRhadinoceraea nodicornis Konow (Hymenoptera, Tenthredinidae) store in their hemolymph ceveratrum alkaloids originating from the host plantVeratrum album L. (Liliales, Melanthiaceae). The major alkaloid found in the hemolymph is 3-acetyl-zygadenine. Qualitative and quantitative data showed that the plant alkaloid 3-angeloylzygadenine is most probably metabolized in the larval gut to zygadenine and then acetylated. A still unidentified alkaloid with a molecular weight of 591 Da was detected in plant leaves as well as in the gut, hemolymph, and excrement of larvae. Protoveratrine A and B, on the other hand, seem to be degraded by the larvae. These findings indicate that the pathway of ceveratrum alkaloids inR. nodicornis larvae is fourfold: direct sequestration, metabolism followed by sequestration, excretion of intact alkaloids, and degradation. In contrast, no ceveratrum alkaloids were detected in the hemolymph and excrement of larvae of the generalist sawflyAglaostigma sp. fed withV. album leaves. Bioassays with the antMyrmica rubra L. proved that the hemolymph ofR. nodicornis larvae is highly deterrent and toxic. In bioassays evaluating defensive efficiency against predators (ants, spiders, and bushcrickets), no larvae were eaten. Ceveratrum alkaloids were also detected in the hibernating prepupae ofR. nodicornis. In feeding bioassays, the shrewCrocidura russula Hermann rarely fed upon prepupae, suggesting that this stage is also protected from predation to some degree. In field surveys, the only parasitoids recorded were two ichneumonid species that are believed to be specialized onR. nodicornis. Bioassays and field observations enable us to suppose thatR. nodicornis and its enemies produce a food web of ion connectance.