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.     

Plant-natural enemy association in the tritrophic systemCotesia rubecula-Pieris rapae-brassicaceae (cruciferae): II. Preference ofC. rubecula for landing and searching

The responses of the parasitoidCotesia rubecula to differently damaged cabbages were recorded during a series of choice tests. To determine if flyingC. rubecula can discriminate differences in the blend of volatiles emitted by cabbages damaged by different causes and how plant volatiles released from a distant source affect the searching behavior ofC. rubecula once searching on a plant, wasps were presented with a choice of plants located one behind the other and separated by a distance of 15 cm. The sources of damage were: cabbage damaged by the host (Pieris rapae), by a nonhost lepidopteran herbivore (Plutella xylostella), by a nonhost, noninsect herbivore (snail), and by mechanical means. The results showed that the site of first landing and the time spent searching on the leaves was influenced by the type of damage inflicted on plants. Wasps preferred to land on cabbages damaged by host and nonhost species of Lepidoptera over those damaged by snails and mechanical means. No preference was observed for first landing between cabbages damaged by the two species of Lepidoptera or between cabbages damaged by snails and mechanical means. Cabbage damaged byP. rapae was searched most intensively, followed by cabbage damaged byP. xylostella, cabbage damaged by snails, and cabbage damaged by mechanical means.C. rubecula differentiates between the volatile blends emitted by differently damaged cabbages, and it is attracted to volatiles related to recent lepidopteran damage. Wasps searched longer on freshly damaged than on leaves with older damage.     

Host-marking pheromones in lepidoptera,with special reference to twoPieris spp

Adult females of several lepidopteran species avoid oviposition near conspecific eggs. Laboratory and field studies indicate that visual factors as well as chemical host markers may be involved in egg avoidance behavior. The oviposition-deterring pheromones (ODP) of two pierids,P. brassicae andP. rapae have been studied in considerable detail. The ODPs are probably produced in the female accessory glands and contain volatile and nonvolatile components. They are perceived by nonspecialized antennal or tarsal receptors, respectively. The ODP ofP. brassicae affects oviposition ofP. rapae females and vice versa. ODPs show promise for insect control programs, but more information on oviposition behavior in the presence of ODP under field conditions is required.     

Chemical and experiential basis for rejection ofTropaeolum majus byPieris rapae larvae

Rejection of nasturtium,Tropaeolum majus, by cabbage-reared larvae ofPieris rapae has been explained by the presence of feeding deterrents in the nastrutium foliage. Sensitivity to the deterrents develops as neonate larvae feed on cabbage. The most prominent deterrent compound, which is present in nasturtium at a concentration of 40 mg/100 g fresh leaves, was identified as chlorogenic acid. When neonate larvae were fed on a cabbage leaf treated with high concentrations of deterrent-containing extracts of nasturtium foliage, they remained insensitive to the deterrents, so they accepted nasturtium when transferred as second instars. When neonate larvae were reared on a cabbage leaf treated with 0.1 mg chlorogenic acid, ca. 35% of the second instars accepted nasturtium. Similar dietary exposure of neonates to the subunits of chlorogenic acid, caffeic acid and quinic, acid resulted in much less or no effect on the rejection behavior of second instars. The results suggest that the combined effects of specific chemical constituents of nasturtium can explain the rejection of this plant by larvae ofP. rapae, but if larvae are continuously exposed to these compounds immediately after hatching, they apparently become habituated to the feeding deterrents. The lack of activity of the subunits of chlorogenic acid suggests that specific structural features are necessary for a dietary constituent to cause such habituation or suppression of sensitivity development.     

Quantity Does Matter: How Feces Are Used for Host Stage Selection by Granary Weevil Parasitoid Lariophagus distinguendus

The present paper examines the mechanisms of host stage selection in Lariophagus distinguendus, a larval parasitoid of the granary weevil Sitophilus granarius endophytic in wheat grain. The amount of host feces accumulating in infested grains was on average, 0.2, 0.3, 1.5, and 4 mg for first, second, third, and fourth instars, respectively. A maximum of 9 mg feces was found in grains with fourth instars. In bioassays with female parasitoids, only a little drumming and drilling behavior was observed on grain models treated with feces extract equivalent to 0.2 mg and 0.4 mg feces. Most drumming and drilling occurred on models with extracts equivalent to 4 and 8 mg feces. These results correspond to literature data demonstrating that older host larvae (fourth instars) are preferred for oviposition. Thus, host stage selection in L. distinguendus is apparently achieved by assessing the quantity of feces present in infested grains.     

Plant-natural enemy association in tritrophic system,Cotesia rubecula-Pieris rapae-brassicaceae (Cruciferae). III: Collection and identification of plant and frass volatiles

To elucidate the identity of the volatile compounds that could be involved in the searching behavior of the parasitoidCotesia rubecula Marshall (Hymenoptera: Braconidae), the volatiles released by cabbage and frass of Lepidoptera feeding on cabbage were collected and analyzed using a gas chromatograph-mass spectrometer. The volatiles emitted by intact cabbage were -pinene, -pinene, myrcene, 1,8-cineole,n-hexyl acetate,cis-3-hexen-1-yl acetate, and dimethyl trisulfide. Mechanical damage on an intact plant induced the release of two more compounds,trans-2-hexenal and 1-methoxy-3-methylene-2-pentanone. Current feeding by larvae ofPieris rapae L. (Pieridae) induced the plant to release all the compounds released after mechanical damage and additionally 4-methyl-3-pentenal and allyl isothiocyanate. Current feeding by larvae ofPlutella xylostella L. (Plutellidae) induced the plant to release all the compounds present after mechanical damage and additionally allyl isothiocyanate. The volatiles emitted after feeding by the lepidopterans had ceased were the same as those emitted by cabbage damaged by mechanical means. The blend of volatiles emitted by frass was comprised of plant chemicals, mainly sulfur compounds. Frass ofP. rapae emitted allyl isothiocyanate, methyl isothiocyanate, methyl propyl sulfide, dimethyl trisulfide,S-methyl methane thiosulfinate, 4-methyl-3-pentenal,trans-2-hexenal, and 2,3-dihydro-4-methyl furan. Frass ofP. xylostella emitted only dimethyl trisulfide andS-methyl methane thiosulfinate. The blend of volatiles emitted by frass is herbivore-species specific.     

Relative importance of infochemicals from first and second trophic level in long-range host location by the larval parasitoidCotesia glomerata

Recently parasitoids were hypothesized to encounter a reliability-detectability problem relating to chemical stimuli from the first and second trophic level, when searching for hosts. The relative role of infochemicals originating from the host,Pieris brassicae (second trophic level), and its food plant, cabbage (first trophic level), have been investigated with respect to long-range host location by the larval parasitoidCotesia glomerata. Flight-chamber dual choice tests showed that uninfested cabbage plants are least attractive to female wasps. Host larvae and their feces were more attractive than clean plants but far less attractive than artificially damaged and herbivore-damaged plants. The plant-host complex, with host larvae actively feeding on the plant, was the most attractive odor source for the parasitoids. The data indicate that one of the solutionsC. glomerata uses to solve the reliability-detectability problem is to respond to infochemicals that are emitted from herbivore-damaged plants. Whether these infochemicals are herbivore-induced synomones that are produced by the plant remains to be demonstrated. Infochemicals emitted by the herbivore or its by-products are of little importance in the foraging behavior ofC. glomerata.     

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.     

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.     

Herbivore-Induced Plant Volatiles Mediate In-Flight Host Discrimination by Parasitoids

Herbivore feeding induces plants to emit volatiles that are detectable and reliable cues for foraging parasitoids, which allows them to perform oriented host searching. We investigated whether these plant volatiles play a role in avoiding parasitoid competition by discriminating parasitized from unparasitized hosts in flight. In a wind tunnel set-up, we used mechanically damaged plants treated with regurgitant containing elicitors to simulate and standardize herbivore feeding. The solitary parasitoid Cotesia rubecula discriminated among volatile blends from Brussels sprouts plants treated with regurgitant of unparasitized Pieris rapae or P. brassicae caterpillars over blends emitted by plants treated with regurgitant of parasitized caterpillars. The gregarious Cotesia glomerata discriminated between volatiles induced by regurgitant from parasitized and unparasitized caterpillars of its major host species, P. brassicae. Gas chromatography-mass spectrometry analysis of headspace odors revealed that cabbage plants treated with regurgitant of parasitized P. brassicae caterpillars emitted lower amounts of volatiles than plants treated with unparasitized caterpillars. We demonstrate (1) that parasitoids can detect, in flight, whether their hosts contain competitors, and (2) that plants reduce the production of specific herbivore-induced volatiles after a successful recruitment of their bodyguards. As the induced volatiles bear biosynthetic and ecological costs to plants, downregulation of their production has adaptive value. These findings add a new level of intricacy to plant–parasitoid interactions.     

<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.     

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.     

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.     

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.     

Rejection of host plant by larvae of cabbage butterfly: Diet-dependent sensitivity to an antifeedant

Garden nasturtium,Tropaeolum majus (Tropaeolaceae), is an acceptable host plant for the cabbage butterfly,Pieris rapae. Eggs are readily laid on the plant and hatching larvae feed and develop into normal pupae and adults. However, when second- to fifth-instar larvae were transferred from cabbage to nasturtium, they refused to feed and starved to death. Similar results were obtained when larvae were transferred from other host plants to nasturtium. However, larvae that were reared on nasturtium readily accepted cabbage as a new host plant. We have demonstrated the presence of strong antifeedants in nasturtium foliage and identified the most prominent active compound as chlorogenic acid. However, larvae reared on nasturtium had limited sensitivity, and larvae reared on a wheat germ diet were completely insensitive to the antifeedants. Larvae apparently develop sensitivity to the deterrent as a result of feeding on other host plants, whereas continuous exposure to the deterrent causes habituation or suppression of sensitivity development. The results demonstrate that dietary experience can dramatically affect the response of an insect to a potentially antifeedant compound in a plant.     

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.     

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.     

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.     

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.