Kairomones for the egg parasiteTrichogramma evanescens Westwood

In observation-cage experiments some new contact kairomones for the egg parasiteTrichogramma evanescens Westwood are demonstrated.T. evanescens females search significantly longer on cabbage leaves treated with the wing scales of two hosts,Pieris brassicae L. andP. rapae L. Further, egg washes ofP. brassicae containing an oviposition deterrent pheromone for the butterflies, were found to have a contact-kairomonal effect on the parasite.T. evanescens females search significantly longer on cabbage leaves sprayed with a methanol or water wash ofP. brassicae eggs than on leaves treated with the solvent only.Hymenoptera: Trichogrammatidae.Lepidoptera: Pieridae.     

Role of volatile inforchemicals emitted by feces of larvae in host-searching behavior of parasitoidCotesia rubecula (Hymenoptera: Braconidae): A behavioral and chemical study

The role of volatile infochemicals emitted by feces of larvae in the host-searching behavior of the parasitoidCotesia rubecula was evaluated during single- and dual-choice tests inside a wind tunnel. The following treatments were tested: feces produced by second and fourth instars ofPieris rapae (preferred host), second instars ofP. brassicae (inferior host), second instars ofP. napi (nonhost), and wet feces of second instars ofP. rapae. During a single-choice situation females ofC. rubecula oriented to all types of feces tested. When a preference was to be made,C. rubecula preferred feces of second instars ofP. rapae over that of fourth, feces ofP. rapae over that ofP. brassicae, feces ofP. napi over that ofP. brassicae, and wet over normal host feces. No preference was exhibited between feces of second instars ofP. napi and that of second instars ofP. rapae. The relative importance of infochemicals from host feces versus plant damage caused by host larvae to the searching behavior ofC. rubecula was also evaluated. Plant damage was more important to the searching females than host feces when feces were present in specific concentrations in relation to damage. The volatiles released by normal and wet feces of second instars ofP. rapae, wet feces of fourth instars ofP. rapae, and normal and wet feces ofP. brassicae were collected and identified. Overall, 85 chemical compounds were recorded belonging to the following chemical groups: alcohols, ketones, aldehydes, esters, isothiocyanates, sulfides, nitriles, furanoids, terpenoids and pyridines. The blend of chemicals emitted by feces of different instars ofP. rapae and different species ofPieris exhibited an instar and species specificity in both quantity and quality. Wetting of normal feces increased the amount of volatile chemicals released, and it was also responsible for the appearance of new compounds. The role of feces of larvae in the host-seeking behavior ofC. rubecula is discussed.     

Jasmonic Acid-Induced Changes in <Emphasis Type="BoldItalic">Brassica oleracea</Emphasis> Affect Oviposition Preference of Two Specialist Herbivores

Jasmonic acid (JA) is a key hormone involved in plant defense responses. The effect of JA treatment of cabbage plants on their acceptability for oviposition by two species of cabbage white butterflies, Pieris rapae and P. brassicae, was investigated. Both butterfly species laid fewer eggs on leaves of JA-treated plants compared to control plants. We show that this is due to processes in the plant after JA treatment rather than an effect of JA itself. The oviposition preference for control plants is adaptive, as development time from larval hatch until pupation of P. rapae caterpillars was longer on JA-treated plants. Total glucosinolate content in leaf surface extracts was similar for control and treated plants; however, two of the five glucosinolates were present in lower amounts in leaf surface extracts of JA-treated plants. When the butterflies were offered a choice between the purified glucosinolate fraction isolated from leaf surface extracts of JA-treated plants and that from control plants, they did not discriminate. Changes in leaf surface glucosinolate profile, therefore, do not seem to explain the change in oviposition preference of the butterflies after JA treatment, suggesting that as yet unknown infochemicals are involved.     

Biological evidence of an oviposition-deterring pheromone inLobesia botrana Den. et Schiff. (Lepidoptera,Tortricidae)

Females of the European grapevine moth (Lobesia botrana Den. et Schiff.) usually deposit isolated eggs on flowers and berries of the grapevine (Vitis vinifera L.). We have investigated whether an epideictic pheromone could be present on the egg surface to explain this spacing behavior. About 21,000 eggs ofL. botrana were washed in cold methanol, and the biological activity was tested in a two-choice bioassay offering treated and nontreated areas. Different dilutions of the extract were tested in methanol, which was inactive alone. At the dose of four egg equivalents per microliter of extract, the number of eggs laid by theL. botrana females was reduced by as much as 57% on the treated areas compared to nontreated areas. A longer-term suppression of oviposition (at least 24 hr) following an exposure to the extract occurred for two doses (0.6 and 4.0 eggs/l) of egg extract. Our results strongly suggest the occurrence of an oviposition-deterring pheromone (ODP) on the eggs ofL. botrana. The ecological value of these results is discussed.     

A chemical basis for differential acceptance ofErysimum cheiranthoides by twoPieris species

Wormseed mustard,Erysimum cheiranthoides, is unacceptable as a host for the cabbage butterfly,Pieris rapae. However, it is preferred for oviposition byPieris napi oleracea in the greenhouse. Isolation and identification of the oviposition stimulants toP. napi oleracea were accomplished by C₁₈ open-column chromatography, TLC, ion-exchange chromatography, HPLC, UV, and NMR spectroscopy. Glucoiberin and glucocheirolin were identified as the most active stimulants. The extracted glucoiberin was as stimulatory as glucocheirolin, although its concentration in theErysimum plants was about 10 times lower than that of glucocheirolin. These glucosinolates were only weak stimulants toP. rapae. Furthermore,P. rapae was strongly deterred by the cardenolides, erysimoside and erychroside, fromE. cheiranthoides, andP. napi oleracea was less sensitive to these compounds. No other deterrent toP. napi oleracea was detected in this plant species. The results explain the differential acceptance ofE. cheiranthoides by these twoPieris species.     

Relative activities of glucosinolates as oviposition stimulants forPieris rapae andP. napi oleracea

The relative activities of 10 glucosinolates in stimulating oviposition byP. rapae andP. napi oleracea were compared under the same conditions. When tested at the same concentration, the structurally different glucosinolates stimulated both butterfly species to widely varying degrees. In most cases,P. rapae was more sensitive to aromatic and indole glucosinolates than to aliphatic representatives. This species responded even less to alkyl thio and sulfinyl glucosinolates. However,P. napi oleracea responded strongly to these aliphatic and sulfur-containing members of the group, and the relative activities of aromatic and aliphatic glucosinolates did not show a clear pattern for this species.P. napi oleracea was much more sensitive to low concentrations of sinigrin than wasP. rapae. The threshold concentration for response ofP. napi oleracea to sinigrin was 10^–8 M, which was 100 times lower than forP. rapae, butP. rapae was more sensitive thanP. napi oleracea to changes in glucosinolate concentrations. For bothPieris species, an optimal concentration was reached, above which the response remained constant or tended to decrease.     

<Emphasis Type="Italic">Barbarea vulgaris</Emphasis> Glucosinolate Phenotypes Differentially Affect Performance and Preference of Two Different Species of Lepidopteran Herbivores

The composition of secondary metabolites and the nutritional value of a plant both determine herbivore preference and performance. The genetically determined glucosinolate pattern of Barbarea vulgaris can be dominated by either glucobarbarin (BAR-type) or by gluconasturtiin (NAS-type). Because of the structural differences, these glucosinolates may have different effects on herbivores. We compared the two Barbarea chemotypes with regards to the preference and performance of two lepidopteran herbivores, using Mamestra brassicae as a generalist and Pieris rapae as a specialist. The generalist and specialist herbivores did not prefer either chemotype for oviposition. However, larvae of the generalist M. brassicae preferred to feed and performed best on NAS-type plants. On NAS-type plants, 100% of the M. brassicae larvae survived while growing exponentially, whereas on BAR-type plants, M. brassicae larvae showed little growth and a mortality of 37.5%. In contrast to M. brassicae, the larval preference and performance of the specialist P. rapae was unaffected by plant chemotype. Total levels of glucosinolates, water soluble sugars, and amino acids of B. vulgaris could not explain the poor preference and performance of M. brassicae on BAR-type plants. Our results suggest that difference in glucosinolate chemical structure is responsible for the differential effects of the B. vulgaris chemotypes on the generalist herbivore. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Does the imported cabbageworm,Pieris rapae,use an oviposition deterring pheromone?

Eggs ofPieris rapae (L.) contain a water- and methanol-soluble oviposition-deterring pheromone (ODP) that is avoided by ovipositing females offered treated and untreated cabbage leaves in a choice situation. Water extracts of female accessory glands also deter oviposition. Egg washes and gland extracts stimulate a contact chemoreceptor in sensilla on the fore tarsi. Electroantennogram (EAG) recordings show that antennal receptors respond to volatiles from conspecific eggs, but not to volatiles from eggs ofMamestra brassicae. No differences were detected between North American and European insects with regard to production and perception of ODP. These results differ from conclusions in the literature based upon field observations. It is concluded that this discrepancy in due to methodological differences.     

Cardenolides as oviposition deterrents to twoPieris species: Structure-activity relationships

Oviposition responses ofPieris rapae andP. napi oleracea to 18 cardenolides were compared under the same conditions. Effects of different concentrations of selected cardenolides were also tested. Most of the compounds were deterrent to oviposition by both insects, but to significantly different degrees.P. rapae were strongly deterred by K-strophanthoside, K-strophanthin-, cymarin, convallatoxin, oleandrin, erysimoside, erychroside, and gitoxigenin. The most deterrent compounds forP. napi oleracea were erychroside, cymarin, erysimoside, convallatoxin, and K-strophanthoside. Strophanthidin-based glycosides were more deterrent than digitoxigenin-based ones, and the number and type of sugar substitutions can have profound effects on activity. Both similarities and contrasts were found in responses ofP. rapae andP. napi oleracea to these cardenolides. Cymarin was equally deterrent to bothPieris species at all concentrations tested. However, when compared withP. rapae, P. napi oleracea was less sensitive to most of the cardenolides.P. napi oleracea was insensitive to K-strophanthin- and oleandrin at 0.5 × 10^–4 M, which were highly deterrent toP. rapae.     

Antiaphrodisiacs in pierid butterflies: a theme with variation!

Male Pieris napi butterflies previously have been shown to synthesize and transfer an antiaphrodisiac, methyl salicylate (MeS), to females at mating. This substance curtails courtship and decreases the likelihood of female remating. Here, we show that similar systems occur in Pieris rapae and Pieris brassicae. In P. rapae, ¹³C-labeling studies showed that males utilize the amino acids phenylalanine and tryptophan as precursors to MeS and indole, respectively. These volatiles are transferred to females at mating and function as antiaphrodisiacs, as demonstrated by field tests entailing painting MeS, indole, or a mixture on the abdomens of virgin females and assessing their attractiveness to wild males. With P. brassicae, ¹³C-labeling studies showed that males use phenylalanine as a precursor to synthesize benzyl cyanide, which was demonstrated to function as an antiaphrodisiac by field tests similar to those for P. rapae. This communication system exhibits both similarities and differences among the three species; in P. napi and P. rapae, males are fragrant but transfer a volatile antiaphrodisiac to females that is completely different from the male odor, whereas in P. brassicae the antiaphrodisiac transferred by male to female is identical with male odor.     

Chemical constituents of an unacceptable crucifer,Erysimum cheiranthoides,deter feeding byPieris rapae

The wild cruciferErysimum cheiranthoides was found to contain extractable constituents that deterred feeding by larvae of the crucifer specialistPieris rapae when applied to cabbage leaf disks in both choice and nochoice bioassays. High-performance liquid chromatography was used to separate the extract into several fractions, two of which retained the feeding deterrent activity of the extract. UV-absorption spectra of the fractions suggested that one contained cardenolides similar or identical to those reported to deter oviposition byP. rapae onE. cheiranthoides. The other active fraction evidently contains a compound that deters larval feeding but not adult oviposition. The results suggest that the chemical defense ofE. cheiranthoides depends on two types of compounds acting on separate developmental stages of the insect.     

Plant-natural enemy association in the tritrophic system,Cotesia rubecula-Pieris rapae-brassiceae (cruciferae): I. Sources of infochemicals

The role of airborne infochemicals in host selection by the parasitoidCotesia rubecula (Marshal) (Hymenoptera: Braconidae) was examined in a wind tunnel. To elucidate the role of volatile chemicals in attractingC. rubecula to cabbage infested by the host [Pieris rapae L. (Lepidoptera: Pieridae)], the potential sources of volatiles related toP. rapae infestation on cabbage were tested individually. The responses of females to nonhost plant species, bean and geranium, as well as to frass of a nonhost lepidopteran were also examined.C. rubecula was attracted to cabbage previously infested byP. rapae and to frass and regurgitate ofP. rapae. No attraction was observed to larvae ofP. rapae alone. Females were also attracted to mechanically damaged cabbage, cabbage previously infested byPlutella xylostella L. (Lepidoptera: Plutellidae) (a nonhost lepidopteran herbivore), and cabbage previously infested by snails (a nonhost, noninsect herbivore). Intact cabbage, bean, and geranium plants elicited no attraction. A low frequency of attraction was observed to mechanically damaged bean and geranium. Attraction was also observed to frass ofP. xylostella. Volatiles from cabbage related to damage, and volatiles from frass and regurgitate of the host seem to play an important role in guidingC. rubecula to plants infested by its host.     

Oviposition stimulants inBarbarea vulgaris forPieris rapae andP. napi oleracea: isolation,identification and differential activity

The closely related butterflies,Pieris rapae andP. napi oleracea, readily laid eggs onBarbarea vulgaris in greenhouse cages. When offered a choice between cabbage andB. vulgaris, P. rapae showed no preference, butP. napi oleracea preferredB. vulgaris. Bioassays of extracts ofB. vulgaris foliage revealed the presence of oviposition deterrent(s) in l-butanol extracts as well as stimulants in the postbutanol water extracts. However, the deterrent effect was apparently outweighed by the strong stimulatory effect in the whole plants. The postbutanol water extract was preferred over an equivalent cabbage extract by both species, but more significantly in the case ofP. napi oleracea. The stimulants were isolated by open column chromatography and HPLC, and the activity was associated with three glucosinolates.P. napi oleracea was more sensitive thanP. rapae to the natural concentration of compounds1 and3, whereas both species were strongly stimulated to oviposit by natural concentrations of compound2. Compounds1 and2 were identified as (2R)-glucobarbarin and (2S)-glucobarbarin, respectively, and3 was identified as glucobrassicin, on the basis of their UV, mass, and NMR spectra. When the pure compounds were tested at the same concentrations applied to bean plants, the (2R)-glucobarbarin at 0.2 mg/plant was preferred over a standard cabbage extract by both butterfly species. However, at a dose of 0.02 mg/plant,P. rapae preferred the cabbage extract whereasP. napi oleracea still preferred the (2R)-glucobarbarin. No such difference in response of the two species to the same two concentrations of (2S)-glucobarbarin was obtained. The results indicate a distinct difference in sensitivity of these butterflies to the epimers of glucobarbarin, and the differences in behavioral responses of the two butterfly species depend to a large extent on the concentration of stimulant present.     

Oviposition stimulants and deterrents regulating differential acceptance ofIberis amara byPieris rapae andP. napi oleracea

Iberis amara (Cruciferae) contains both stimulants and deterrents that are involved in regulating oviposition byPieris rapae andP. napi oleracea. The most active deterrents toP. rapae isolated from butanol extracts of the plant were found to be 2-O--d-glucosyl cucurbitacin I and 2-O--d-glucosyl cucurbitacin E. However,P. napi oleracea was behaviorally insensitive to these compounds and was only weakly deterred by other individual fractions of the butanol extract. Stimulant activity of the postbutanol water extract ofI. amara was associated with glucosinolates. The most abundant of these was identified as sinigrin, and a relatively minor component was shown to be glucoiberin. The isolated sinigrin was more stimulatory toP. rapae than was the glucoiberin-containing fraction, butP. napi oleracea was stimulated as strongly by the glucoiberin fraction, even though the concentration of this compound was much lower. The contrasting responses of the twoPieris species to the deterrents and stimulants inI. amara can explain the differential acceptance of the plant by these butterflies.     

Perception of Oviposition-Deterring Pheromone by Cabbage Seed Weevil (Ceutorhynchus assimilis)

Following oviposition into a pod of oilseed rape (Brassica napus), the female cabbage seed weevil (Ceutorhynchus assimilis) marks the pod with oviposition-deterring pheromone (ODP) by brushing it with her eighth abdominal tergite. On an unmarked pod, oviposition site selection was always accompanied by intensive antennation of the pod. Females approaching a freshly ODP-marked pod brought their antennae within 1 mm of the pod but usually did not antennate it before rejecting it for oviposition. Females with the clubs of their antennae amputated continued to discriminate pods from stems or petioles as oviposition sites but showed no behavioral response to ODP. Extracts of volatiles air-entrained from ovipositing weevils failed to inhibit oviposition. Air passed over a behaviorally active extract of ODP did not elicit a detectable electroantennogram response. By contrast, when presented as a gustatory stimulus to the sensilla chaetica of the antennal club, a behaviorally active extract of ODP from postdiapause, gravid females elicited a strong electrophysiological response. This response usually involved more than one cell and displayed a phasic–tonic time course over the recording period of 10 sec. Extract from prediapause (and hence sexually immature) females elicited neither behavioral nor electrophysiological (contact) responses. Thus the ODP of the cabbage seed weevil is sensed primarily by contact chemoreception at the sensilla chaetica of the antennae, and the electrophysiological responses recorded from these gustatory sensilla are of value as the basis of a bioassay to assist identification of the active constituent(s) of the pheromone.     

Chemical constituents ofErysimum cheiranthoides deterring oviposition by the cabbage butterfly,Pieris rapae

Avoidance ofErysimum cheiranthoides for oviposition byPieris rapae has been attributed to the presence of water-soluble deterrents. The active material was extracted inton-butanol and isolated by a series of HPLC separations. TLC of the active fraction and visualization of individual constituents with Kedde's reagent indicated that cardenolides are responsible for deterring oviposition. UV spectra were also characteristic of cardenolides. Bioassays of selected known cardenolides revealed a general lack of activity, except for cymarin, which was as strongly deterrent as the most prominent cardenolide isolated in pure form fromE. cheiranthoides. The results suggest that cardenolides in this plant can explain its escape from cabbage butterflies, but specific structural features of the glycosides are necessary for oviposition-deterring activity.     

Olfactory responses of the parasitoidDiaeretiella rapae (Hymenoptera: Aphidiidae) to odor of plants,aphids, and plant-aphid complexes

Diaeretiella rapae (M'Intosh) (Hymenoptera: Aphidiidae) is a parasitoid of several aphid species, including the Russian wheat aphid (RWA),Diuraphis noxia (Mordvilko), and the cabbage aphid (CA).Brevicoryne brassicae (L.). The response of matedD. rapae females to odors from wheat, cabbage, and plant-host complexes was investigated using a four-choice olfactometer. Experienced parasitoids, but not inexperienced females, responded positively to odors of the wheat-RWA complex in a no-choice test. In choice tests, experienced parasitoids did not respond to odors of uninfested cabbage and wheat leaves, but did respond positively to aphid-infested plants and to aphids alone. The response ofD. rapae to the cabbage-CA complex and to CA alone was significantly greater than to the wheat-RWA complex and RWA alone, suggesting an innate odor preference for crucifer-feeding aphids.     

Chemical stimulants and deterrents regulating acceptance or rejection of crucifers by cabbage butterflies

GravidPieris rapae butterflies oviposit on many, but not all, crucifers. Rejection ofErysimum cheiranthoides andCapsella bursa-pastoris was initially explained by the presence of chemical deterrents in the plants. Analyses and bioassays of plant extracts indicated the absence of oviposition stimulants inC. bursa-pastoris, but similar chemical separation ofE. cheiranthoides extracts revealed the presence of stimulants as well as deterrents. Choice tests illustrate how acceptance or rejection of a plant by an insect may depend on the balance of positive and negative chemical stimuli within the plant.     

Seven-week persistence of an oviposition-deterrent pheromone

Cabbage leaves sprayed with a water solution of the oviposition-deterrent pheromone ofPieris brassicae remain deterrent to ovipositing females for at least 14 days. Under laboratory conditions, the pheromone, when dried on a glass surface, retains activity for a period of at least 7 weeks. After 7 days under vacuum conditions, some pheromone is still present, indicating a low volatility and / or high stability of (an active fraction of) the pheromone. After 125 eggs are slowly rinsed with 300 ml water, they still release detectable quantities of the pheromone.     

Isolation and identification of oviposition deterrents to cabbage butterfly,Pieris rapae,fromErysimum cheiranthoides

Avoidance of some crucifer species by the crucifer specialist,Pieris rapae, has been attributed to the presence of oviposition deterrents in these plants. Studies on one such unacceptable plant,Erysimum cheiranthoides, have resulted in the isolation ofn-butanol-soluble deterrents from the alcoholic extract of foliage. The active fraction contained three cardiac glycosides, which were isolated by reversed-phase HPLC and by open column chromatography on silica gel. Chemical and spectral evidence (UV, [¹H]NMR, and FAB-MS) led to the characterization of these compounds as erysimoside (1), erychroside (2), and erycordin (3). Erysimoside and erychroside were strongly deterrent toPieris rapae, but erycordin was inactive. Both active compounds have the same aglycone, strophanthidin (5) and the inner sugar in both cases is a 2,6-dideoxy hexose to which the outer sugar is attached at position C-4. These structural features, which are absent in the inactive compound (3), may represent specific requirements for oviposition deterrent activity.