Antifeedant Effects of Some Novel Terpenoids on Chrysomelidae Beetles: Comparisons with Alkaloids on an Alkaloid-Adapted and Nonadapted Species

Structure–dose–feeding deterrency relationships were compared between the Colorado potato beetle, Leptinotarsa decemlineata (Say), and the western corn rootworm, Diabrotica virgifera virgifera LeConte, using 15 alkaloids, terpenoids, and phenolic derivatives. The former species, a specialist herbivore on selected alkaloid-rich Solanaceae species, was on average 100-times less sensitive to the antifeedant effects of alkaloids, but more similarly sensitive to the terpenoids and phenolics than the latter species, a generalist flower herbivore predominantly on Graminae, Cucurbitaceae, and Compositae species. Antifeedant ED₅₀ values for the potato beetle and corn rootworm, each from closely related subfamilies of Chrysomelidae, ranged over four orders of dose magnitude among the 15 compounds with major species differences in stereosensitivity to -hydrastines and analog sensitivity with the silphinenes. Extremes in sensitivity ranged from silphinene, a rare tricyclic sesquiterpene that is 53 times more active on the potato beetle to aconitine, which is 430 times more antifeedant to the corn rootworm. Among silphinene and its two hydrolysis derivatives, there was not a strong correlation between antifeedant potency and injected toxicity for the two beetle species, but there was correlation between behavioral activity and galeal taste cell electrophysiological threshold and frequency responses. That all of the established GABA- and glycinergic compounds tested were antifeedant for both species suggests a shared molecular mechanism for antifeedant taste chemoreception in these divergent Chrysomelidae species. Moreover, the wide differences in antifeedant sensitivities among these and other chrysomelids to a suite of ligand-gated ion channel antagonists implicate a common protein neuroreceptor type with extraordinary heterogeneity in beetle taste.     

Effects ofneo-clerodane diterpenes fromTeucrium on feeding behavior of colorado potato beetle larvae

Treatment of potato leaf disks with fourneo-clerodane diterpenes fromTeucrium (eriocephalin, teucrin-A, teucvin, and teuscorolide) significantly reduced feeding by larvae ofLeptinotarsa decemlineata (Colorado potato beetle). Choice and no-choice tests suggest that teuscorolide acts as a feeding deterrent, whereas the antifeedant activity of teucrin-A, teucvin, and eriocephalin is likely associated with a toxic mode of action. Nutritional tests, antifeedant simulation assays, and posttreatment studies confirmed that teucrin-A can be categorized by its mode of action as a toxin, rather than as a feeding deterrent.     

Antifeedant Compounds from Three Species of Apiaceae Active Against the Field Slug, Deroceras reticulatum (Muller)

Extracts of volatiles from foliage of three plants in the Apiaceae, Conium maculatum L. (hemlock), Coriandrum sativum L. (coriander), and Petroselinum crispum Mill. (Nym.) (parsley), previously shown to exhibit antifeedant activity in assays with the field slug, Deroceras reticulatum (Muller) (Limacidae: Pulmonata), were studied further to identify the active components. Coupled gas chromatography-mass spectrometry (GC-MS) and neurophysiological assays using tentacle nerve preparations resulted in the identification of 11 active compounds from the three extracts. Wheat flour feeding bioassays were used to determine which of these compounds had the highest antifeedant activity. One of the most active compounds was the alkaloid gamma-coniceine, from C. maculatum. The role of potentially toxic alkaloids as semiochemicals and the potential for using such compounds as crop protection agents to prevent slug feeding damage is discussed.     

Systemic induction of feeding deterrents in cotton plants by feeding ofSpodoptera SPP. Larvae

Cotton,Gossypium hirsutum L., has been shown to exhibit systemic induced resistance to arthropods under certain conditions. We conducted experiments to determine the effects of previous feeding ofSpodoptera exigua Hübner andSpodoptera littoralis (Boisd.) larvae on feeding behavior, growth, and survival of larvae subsequently feeding on cotton. In one feeding choice test,S. exigua larvae preferred young leaves from undamaged control plants to undamaged young leaves from a previously damaged plant. Feeding deterrence was noticeable after only 6 hr of initial feeding damage by larvae, and there was almost complete deterrence after 30 and 54 hr of continuous feeding. In a second feeding choice test,S. littoralis larvae fed more on mature leaves from undamaged control plants than on undamaged mature leaves from previously damaged plants. In no-choice tests, third instars ofS. littoralis fed undamaged young leaves from damaged plants did not gain weight and died by the seventh day, whereas larvae fed young leaves from undamaged control plants gained weight and pupated within 11 days. Sixth instars ofS. littoralis fed either old damaged leaves, old undamaged leaves, or young undamaged leaves all from previously damaged plants gained weight slowly and took more than 12 days to pupate, whereas larvae fed young leaves from undamaged plants gained weight rapidly and pupated within five days of the beginning of the experiment.     

Feeding and oviposition preferences of sweet potato weevil,Cylas formicarius elegantulus (Summers), on storage roots of sweet potato cultivars with differing surface chemistries

Cores from sweet potato [Ipomoea batatas (L.) Lam.] storage roots (Centennial, Jewel, Resisto, and Regal cultivars) were presented to sweet potato weevils [Cylas formicarius elegantulus (Summers) (Coleoptera; Curculionidae)] in multiple-choice, limited-choice, and no-choice bioassays. Centennial, a susceptible cultivar in field-plot experiments, was preferred for feeding and oviposition by female weevils in choice bioassays, and for ovi-position in no-choice bioassays, compared to three other cultivars. Analysis of root surface chemistry showed a tentatively identified triterpenol acetate in Centennial, which was not found in the more resistant cultivars; another root surface component was found in higher concentrations in the more resistant cultivars.     

Differential toxicity of juglone (5-hydroxy-1,4-naphthoquinone) and related naphthoquinones to saturniid moths

The preferred hosts of the saturniid mothActias luna include members of the Juglandaceae, whose foliage contain the toxin juglone (5-hydroxy-1,4-naphthoquinone). The performance ofActias luna andCallosamia promethea was compared when fourth-instar larvae of each were fed birch foliage, a mutually acceptable food plant, or birth supplemented with 0.05% (w/w) juglone.A. luna fed juglone exhibited no changes in developmental time or mortality compared to a diet without juglone. In contrast, juglone-supplemented diets, when fed toC. promethea, caused negative growth rate, and a 3.6-fold decrease in consumption rate. The performance ofA. luna also was compared on birch and walnut; larvae developed and grew more rapidly on an all-walnut vs. an all-birch diet. To examine the effect of 1,4-naphthoquinone structure onA. luna survival, first instars were fed on birch supplemented with varying concentrations of juglone (J), menadione (M), plumbagin (P), or lawsone (L). In diets supplemented at 0.05% (w/w), none of the compounds produced effects significantly different from controls. In diets supplemented at 0.5% (w/w), the treatments produced significant toxic effects in the order P>M=L>J for mortality, and P>L>M=J for increased developmental time. Late-instarA. luna are clearly resistant to juglone compared toC. promethea, and early-instarA. luna are resistant to several related 1,4-naphthoquinones. These results suggest a chemical basis for host choice among saturniids. In addition, the luna-walnut system may be a valuable model for studying quinone detoxication.     

Phenolic Metabolites in Leaves of the Invasive Shrub, Lonicera maackii, and Their Potential Phytotoxic and Anti-Herbivore Effects

Lonicera maackii is an invasive shrub in North America for which allelopathic effects toward other plants or herbivores have been suspected. We characterized the major phenolic metabolites present in methanol extracts of L. maackii leaves. In addition, we examined the effects of methanol–water extracts of L. maackii leaves on seed germination of a target plant species and on feeding preference and growth rate of a generalist insect herbivore. A total of 13 individual major and minor compounds were detected in crude leaf extracts by high-performance liquid chromatography coupled to electronspray ionization-tandem mass spectrometry (ESI-MS/MS). Extracts were dominated by two major flavones, apigenin and luteolin, and their glucoside derivatives, apigenin-7-glucoside and luteolin-7-glucoside. Quantities of these compounds, along with chlorogenic acid, varied between two sampling points. Leaf extracts that contained these compounds were inhibitory to seed germination of Arabidopsis thaliana. In addition, treatment of artificial diet with leaf extracts deterred feeding of the generalist herbivore, Spodoptera exigua, in choice experiments but had no effect on growth rate in short-term no-choice bioassays. Purified apigenin tended to deter feeding by S. exigua and inhibited seed germination of A. thaliana. We conclude that leaves of L. maackii contain phenolic compounds, including apigenin and chlorogenic acid, capable of having biological effects on other plants and insects.     

Antifeedant activity of extracts from neem,Azadirachta indica,to strawberry aphid,Chaetosiphon fragaefolii

Leaf disk choice test bioassays demonstrated that formulated neem seed oil (NSO) was equally deterrent to first- and third-instar nymphs and adult strawberry aphids,Chaetosiphon fragaefolii (Cockerell). Concentrations of NSO resulting in 50% feeding deterrence were approximately 1.1% for this species. The rapid disruption of aphid feeding (<1 hr) was not related to the presence of the limonoid azadirachtin, and deterrence likely results from the combined activity of several compounds. Activity toC. fragaefolii disappeared within 12–24 hr following application to strawberry in the greenhouse. NSO was deterrent to only half of the six aphid species tested. The antifeedant properties of neem do not appear to contribute significantly to the control of aphids and the viruses they transmit.     

Interactions amongHeliothis virescens larvae,cotton condensed tannin and the CryIA(c) δ-endotoxin ofBacillus thuringiensis

The potential interactions among a plant-produced allelochemical, a phytophagous insect, and an endotoxin produced byBacillus thuringiensis were investigated using purified cotton condensed tannins, the CryIA(c)-endotoxin fromB. thuringiensis subsp.kurstaki strain HD-73, and larvae ofHeliothis virescens. Purified condensed tannin from cotton fed to neonateH. virescens reduced feeding and mortality caused by insecticidal crystals ofB. thuringiensis. In fifth instars, tannin reduced relative growth rate (RGR), relative consumption rate (RCR), but antagonized the effects of the crystal-endotoxin. Tannin did not deter feeding of fifth instars in choice tests with cellulose-ester disks. Masking tannin from interacting with the dietary ingredients of artificial diets and crystal protein by encapsulation in alginate gel suggested that tannin adversely affected feeding after ingestion.These results suggest that insect control tactics that employ-endotoxins in microbial insecticides and transgenic cotton plants may not be compatible when used in conjunction with plants containing high tannin concentrations.     

Identification of Semiochemicals Released During Aphid Feeding That Attract Parasitoid Aphidius ervi

Herbivore induced release of plant volatiles mediating the foraging behavior of the aphid parasitoid Aphidius ervi was investigated using the pea aphid, Acyrthosiphon pisum, feeding on broad bean, Vicia faba. Behavioral responses were studied using an olfactometer and a wind tunnel. Volatiles obtained by air entrainment of aphid infested plants were more attractive to A. ervi than those from uninfested plants, in both behavioral bioassays. GC-EAG of both extracts showed a number of peaks associated with responses by A. ervi, but with some differences between extracts. Compounds giving these peaks were tentatively identified by GC-MS and confirmed by comparison with authentic samples on GC, using two columns of different polarity. The activity of pure compounds was further investigated by EAG and wind tunnel assays. Results showed that, of the compounds tested, 6-methyl-5-hepten-2-one was the most attractive for A. ervi females, with linalool, (Z)-3-hexen-1-yl acetate, (E)--ocimene, (Z)-3-hexen-1-ol, and (E)--farnesene all eliciting significantly more oriented flight behavior than a solvent control. Foraging experience significantly increased parasitoid responses to these compounds, with the exception of (E)--farnesene. Time-course GC analysis showed that feeding of A. pisum on V. faba induced or increased the release of several compounds. Release of two of these compounds (6-methyl-5-hepten-2-one and geranic acid) was not induced by the nonhost black bean aphid, Aphis fabae. During the analysis period, production of (E)--ocimene remained constant, but 6-methyl-5-hepten-2-one, linalool, geranic acid, and (E)--farnesene appeared during the first day after A. pisum infestation and increased in concentration with increasing time of aphid feeding.     

Aphid deterrence by glucose esters in glandular trichome exudate of the wild tomato,Lycopersicon pennellii

Settling of the potato aphid,Macrosiphum euphorbiae, on feeding membranes was deterred by methanolic leaf rinses ofLycopersicon pennellii, or of its F₁ with tomato,L. esculentum. The active compounds in theL. pennellii rinsates were identified as 2,3,4-tri-O-acylglucoses bearing short to medium chain length fatty acids. These compounds are localized in the glandular exudate of the type IV trichomes and may accumulate to levels in excess of 400 g/cm². In choice assays, purified glucose esters fromL. pennellii reduced aphid settling at concentrations as low as 25 g/cm²; at concentrations of 150 g/cm² or more, all aphids avoided treated areas. Glucose esters were also active in deterring aphid settling in no-choice assays. At 100 g/ cm², these esters resulted in increased levels of mortality after 48 hr.     

Effects of juglone on growth,photosynthesis, and respiration

The impacts of juglone on plant growth and several other physiological functions were evaluated in this study. Juglone inhibitedLemna minor growth, chlorophyll content, and net photosynthesis at treatments between 10 and 40M. Soybean leaf disks vacuum infiltrated with as little as 10M juglone had reduced photosynthesis. Oxygen evolution by chloroplasts isolated fromPisum sativum was inhibited by juglone with an I₅₀ of 2M. Micromolar treatments of juglone stimulated oxygen uptake in mitochondria isolated fromGlycine max. These data suggest perturbations of chloroplast and mitochondrial functions may contribute to plant growth reductions observed in juglone-mediated allelopathy.This study was supported by National Science Foundation grant STI-8902066.     

Larval exposure to oviposition deterrents alters subsequent oviposition behavior in generalist,Trichoplusia ni and specialist,Plutella xylostella moths

The present study was undertaken to determine the effects of larval feeding experience on subsequent oviposition behavior of the resulting moths. Larvae of the cabbage looper (Trichoplusia ni, Noctuidae) and the diamondback moth (Plutella xylostella, Plutellidae) were exposed to the phenylpropanoid allelochemical trans-anethole (at 100 ppm fw in artificial diet) or the limonoid allelochemical toosendanin (10 ppm sprayed on cabbage leaves). Both compounds had been shown to deter oviposition in naïve moths in previous choice tests. Moths developing from experienced larvae (both sexes) showed a decrease in oviposition deterrence response when given a choice between control and treated leaves, unlike naïve moths. This phenomenon, analogous to habituation to feeding deterrents in lepidopteran larva, occurred irrespective of duration of feeding on the deterrent compound. We also observed that F₁larvae resulting from experienced moths (previously exposed to toosendanin as larvae) grew as well on toosendanin-treated foliage as on control foliage. In contrast, growth of F₁larvae from naïve moths was significantly impaired by toosendanin. These results demonstrate that host-selection behavior in cabbage looper (a generalist) and diamondback moth (a specialist) may be shaped by feeding experience according to Hopkins' Host Selection Principle in addition to chemical legacy.     

Insect antifeedant activity associated with compounds isolated from species ofLonchocarpus andTephrosia

The antifeedant activity of a series of 21 chalcones, flavanes, and flavanones isolated from the generaLonchocarpus andTephrosia (Leguminosae) was assessed by behavioral and electrophysiological bioassays against larvae ofSpodoptera littoralis andS. exempta. The antifeedant activity is related to the molecular structure of the compounds, and possible modes of interaction with the insect taste receptors are discussed.     

Systemic Induction of Allelochemicals in Glanded and Glandless Isogenic Cotton by Spodoptera exigua Feeding

Induction of systemic resistance to feeding of beet armyworm, Spodoptera exigua, was investigated in two isogenic lines of Stoneville 213 cotton, Gossypium hirsutum, that differed in the presence of pigment glands. In laboratory bioassays, larvae strongly preferred to feed on glandless cotton plants when presented a choice between undamaged terminal leaves of undamaged glanded and glandless plants. Feeding damage inflicted by S. exigua larvae on the two oldest leaves of glanded plants seven days prior to feeding bioassays caused larvae to prefer by 33-fold the undamaged terminal foliage from undamaged plants compared to that from damaged plants. Feeding damage on glandless plants caused only a 2.6-fold greater preference for terminal foliage from undamaged plants over foliage from previously damaged plants. Extracts of terminal foliage from glanded cotton damaged seven days earlier had significantly greater quantities of terpenoid aldehydes (hemigossypolone, gossypol, and heliocides) than did foliage from undamaged glanded plants. Terpenoid aldehydes were undetectable in extracts of both undamaged and previously damaged glandless plants. The profile of volatile compounds collected from the headspace of mechanically damaged terminal leaves of undamaged glanded and glandless plants differed. Both cotton isolines released large quantities of lipoxygenase products (hexenyl alcohols, acetates, and butyrates), but glandless plants released only small amounts of mono- and sesquiterpenes compared to glanded plants. Glandless plants damaged seven days prior to volatile collection released significantly greater quantities of lipoxygenase products, -ocimene, and - and -farnesene than did undamaged glandless plants. Previously damaged glanded plants released significantly greater quantities of all mono- and sesquiterpenes and hexenyl acetates and butyrates, but not alcohols. The relative importance of volatile compounds versus terpenoid aldehydes in induced feeding deterrence in cotton to S. exigua larvae is still unclear.     

A new method for fast isolation of insect antifeedant compounds from complex mixtures

Bioassay-guided isolation of bioactive natural substances requires monitoring of all fractionation and purification steps using a bioassay system. This is often a long and tedious process, especially with insect feeding bioassays. We report here a new method, based on the principle of bioautography, for a quick isolation of insect antifeedant compounds. TLC plates, after development, are coated with a thin layer of artificial diet and fed toSpodoptera litura larvae. The location of uneaten areas is then compared with theR _fvalues of the TLC spots, in order to determine rapidly the active fractions. This methods allows for a very fast determination of the most active antifeedant compounds in a complex mixture and considerably speeds up the isolation process. This new method was successfully applied in the study of antifeedant activity of several plant samples, and results are presented here for a model plant,Skimmia japonica (Rutaceae). Using this new method, the compounds responsible for the feeding-deterrent activity, three furanocoumarins (bergapten, xanthotoxin, and oxypeucedanin), were quickly and efficiently identified.     

Olfactory Reception of Conspecific Aggregation Pheromone and Plant Odors by Nymphs of the Predator, Podisus maculiventris

Olfactory reception of 23 odorants, including plant volatiles and male-produced aggregation pheromone, by third and fifth instars of the spined soldier bug (SSB) Podisus maculiventris was investigated by using electroantennograms (EAGs). Both nymphal stages were sensitive to male-produced aggregation pheromone components (E)-2-hexenal, benzyl alcohol, and -terpineol. The plant volatile, (E)-2-hexen-1-ol (a chemical known to be released by plants in response to prey feeding over the short-term), elicited the largest EAGs of all volatiles tested. While third instars were sensitive to nonanal, only fifth instars responded to both nonanal and (±)-linalool, both compounds released systemically by plants in response to feeding by potential prey. Antennal extirpation experiments showed that sensilla responsive to hexan-1-ol, (E)-2-hexenal, and -terpineol are situated mainly on the terminal antennal segment. The results support the hypothesis that P. maculiventris nymphs use both plant volatiles and pheromone components in locating potential prey and other behaviors.     

Chemical Ecology of the Defense of Two Nymphalid Butterfly Larvae Against Ants

We analyzed the behavioral responses of the ants Camponotus rufipes and Solenopsis geminata towards all instars of Dione junio and Abananote hylonome. We also analyzed ant behavior towards hexane extracts of larvae and extracts of the spines and neck glands of the fifth instars of both species and identified the chemical compounds present. Larvae of both species were repellent to ants from the first instar onward. Later instars survived ant attacks better than earlier instars. The spines and neck glands of the larvae influenced the behavior of C. rufipes. The chemical compounds contained in the hexane extracts of whole first and fifth instars and in the spines and neck glands of fifth instars were principally carboxylic acids and terpenes. Further bioassays confirmed the repellent effect of some of these acids toward ants.     

Feeding Response to Host and Nonhost Compounds by Males and Females of the Spruce Bark Beetle <Emphasis Type="Italic">Ips typographus</Emphasis> in a Tunneling Microassay

Research on host selection by bark and wood boring insects has concentrated on flight orientation behavior. Less is known of the factors that govern the steps successive to host landing. Here, we discuss chemical factors involved in host acceptance by bark beetles and a new microassay. Adult males and females of Ips typographus were offered an artificial diet treated with various concentrations of different plant-derived compounds (host terpenes and nonhost compounds) in a no-choice mode. Beetles were tested individually in a glass tube for 4 hr, and the length of feeding was measured and compared to a control (diet with only solvent). The first effect was diet rejection, especially when nonhost compounds were tested at high concentrations. Most compounds reduced feeding, in proportion to concentration. Females were fed more readily than males after addition of both host and nonhost compounds. Diet removal was significantly affected by all the tested factors (sex, compound, dose) as well as by their interactions. With increased concentrations, males were more responsive than females to antifeedants, as all compounds (except juglone) showed clear sex differences of diet consumption. 3-Octanol, 1-hexanol, and a Green Leaf Volatile (GLV)-blend (three C6 alcohols) showed the strongest antifeedant effects, which started at a low dose (0.1%) and had a low Effective Dose 50 (ED50, 0.3–1%). In contrast, host monoterpenes, limonene and -pinene, inhibited feeding at high doses (10–30%) only with ED50 > 10%. The highest Antifeedant Indexes (AFIs) were shown by verbenone, carvone, and 1-hexanol (0.90–1.00). Both host and nonhost compounds inhibited feeding at some concentration. No significant stimulation of feeding by any host compound at concentrations reported in the literature as optimal were found, with the possible exception of -pinene at low concentrations in females.     

Effect of Trichilia americana Extract on Feeding Behavior of Asian Armyworm, Spodoptera litura

We investigated the desensitization of Asian armyworms, Spodoptera litura, to a crude methanolic extract of wood Trichilia americana. Following repeated exposures to the extract over four days, desensitization was observed under no-choice conditions at low extract concentration (0.5 g/cm²). No desensitization was observed under choice conditions or at a higher concentration (5.0 g/cm²). Repeated exposures of larvae to the extract over 7.5 hr showed that larvae increasingly tolerated the extract due to hunger, rather than showing true desensitization. When larvae were exposed to the extract in test arenas of different sizes, larvae showed reduced feeding and desensitization as arena size increased. Finally, treatment of whole cabbage plants with extract resulted in larvae abandoning the plant. The crude extract was effective at protecting the plants, although desensitization did not occur, because the larvae moved away from the plant.