EAG-Active Herbivore-Induced Plant Volatiles Modify Behavioral Responses and Host Attack by An Egg Parasitoid

Volatiles emitted by plants in response to feeding by Lygus species were tested in neurophysiological, behavioral, and parasitism trials with Anaphes iole, an egg parasitoid of Lygus. Electroantennogram analyses indicated that A. iole antennae responded to most herbivore-induced plant volatiles (HIPVs) tested and that females were usually more responsive than males. Antennal responses to (Z)-3-hexenyl acetate and methyl salicylate were among the strongest. Behavioral assays in a four-arm olfactometer demonstrated that response of female wasps to (Z)-3-hexenyl acetate varied greatly depending on preconditioning regime. Preconditioning wasps to complex host-plant odors led to stronger preference than did a single preconditioning stimulus, i.e., (Z)-3-hexenyl acetate. In a horizontal wind tunnel, female wasps were attracted by methyl salicylate and alpha-farnesene. Parasitism of Lygus lineolaris eggs by A. iole in a cotton field was greater when the eggs were associated with (Z)-3-hexenyl acetate or alpha-farnesene than with controls. Overall, the results of this study show that A. iole can perceive a variety of plant volatiles released after its host damages plants, that the degree of associative learning in A. iole can be manipulated based on preconditioning regime, and that single synthetic HIPVs are attractive to A. iole and can be used to increase attack rates on host eggs. Therefore, it appears that HIPVs have potential for use in suppression of Lygus population densities.     

Response of Egg Parasitoid Telenomus busseolae to Sex Pheromone of Sesamia nonagrioides

The olfactory cues involved in the host-finding behavior of Telenomus busseolae Gahan, a solitary egg parasitoid of the corn stalk borer, Sesamia nonagrioides (Lefebvre), were studied. The female wasp was attracted to odors from calling virgin females of S. nonagrioides in a Y-tube olfactometer. No response was elicited by noncalling virgin female moths and/or mated female moths. A four-component commercially available sex pheromone of the corn borer was also tested, as were the individual components of the mixture. T. busseolae respond to the whole blend [(Z)-11-hexadecenyl acetate, (Z)-11-hexadecenol, (Z)-11-hexadecenal, and dodecyl acetate in the ratio of 8.5:1:1:2]. Z11–16:OH was the only compound that elicited no significant activity. These results suggest that the sex pheromone released by the host female S. nonagrioides is used by T. busseolae as an indirect cue to locate egg masses of this host.     

Herbivore-Induced Plant Volatiles Can Serve as Host Location Cues for a Generalist and a Specialist Egg Parasitoid

Herbivore-induced plant volatiles are important host finding cues for larval parasitoids, and similarly, insect oviposition might elicit the release of plant volatiles functioning as host finding cues for egg parasitoids. We hypothesized that egg parasitoids also might utilize HIPVs of emerging larvae to locate plants with host eggs. We, therefore, assessed the olfactory response of two egg parasitoids, a generalist, Trichogramma pretiosum (Tricogrammatidae), and a specialist, Telenomus remus (Scelionidae) to HIPVs. We used a Y-tube olfactometer to tests the wasps’ responses to volatiles released by young maize plants that were treated with regurgitant from caterpillars of the moth Spodoptera frugiperda (Noctuidae) or were directly attacked by the caterpillars. The results show that the generalist egg parasitoid Tr. pretiosum is innately attracted by volatiles from freshly-damaged plants 0–1 and 2–3 h after regurgitant treatment. During this interval, the volatile blend consisted of green leaf volatiles (GLVs) and a blend of aromatic compounds, mono- and homoterpenes, respectively. Behavioral assays with synthetic GLVs confirmed their attractiveness to Tr. pretiosum. The generalist learned the more complex volatile blends released 6–7 h after induction, which consisted mainly of sesquiterpenes. The specialist T. remus on the other hand was attracted only to volatiles emitted from fresh and old damage after associating these volatiles with oviposition. Taken together, these results strengthen the emerging pattern that egg and larval parasitoids behave in a similar way in that generalists can respond innately to HIPVs, while specialists seems to rely more on associative learning.     

Chemicals Used for Host Recognition by the Granary Weevil Parasitoid Lariophagus distinguendus

In order to develop methods for mass rearing of the parasitic wasp Lariophagus distinguendus, we characterized and identified chemicals used by this parasitoid to recognize wheat grains infested by its host, larvae of the granary weevil, Sitophilus granarius, a major pest of stored grain worldwide. Bioassays revealed that drumming and drilling behavior of the parasitoids on grain models of filter paper treated with host feces was as intense as on host-infested grains. Thus, chemicals from feces are highly important to recognize infested grains. Extracts of the feces with hexane or dichloromethane applied on grain models were able to provoke drumming and drilling activity. Hexane extract of feces was as active as infested grain itself. Subsequent fractionation of the hexane extract by adsorption chromatography revealed that highest activity was recovered in the dichloromethane fraction. This fraction was characterized by the presence of -tocopherol, -tocopherol, -tocotrienol, cholesterol, ergostenol, and -sitosterol. Synthetic chemicals in proportions found in the active dichloromethane fraction stimulated drumming behavior in bioassays. The significance of these results with respect to the mass propagation of L. distinguendus as natural enemies for the control of the granary weevil is discussed.     

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.     

Cattle and Amblyomma variegatum Odors Used in Host Habitat and Host Finding by the Tick Parasitoid, Ixodiphagus hookeri

The response of mated naive Ixodiphagus hookeri females to cattle and Amblyomma variegatum nymphal odors was tested in a Y-tube olfactometer. I. hookeri females were attracted to cattle urine, dung, and odors from tick-free feeding sites of A. variegatum nymphs on cattle, e.g., dewlaps, front heels, and hind heels. Tick-free scrotal odors did not attract the parasitoids. Furthermore, odors from off-host unfed and fed A. variegatum nymphs did not attract the parasitoids, despite an increase in the number of the nymphs to amplify any odor signal. A blend of odors from feeding on-host nymphs and cattle scrota attracted the parasitoids. In T-tube bioassays, I. hookeri females were attracted to hexane washes and fecal extracts of A. variegatum nymphs.     

Kairomonal Effects of Sawfly Sex Pheromones on Egg Parasitoids

The response of the two eulophid egg parasitoid species Chrysonotomyia ruforum and Dipriocampe diprioni to sex pheromones of their sawfly hosts Diprion pini and Neodiprion sertifer was studied in olfactometer bioassays. Females of C. ruforum were arrested when exposed to odor of the tested major sex pheromone components of Diprion pini [acetate and propionate of (2S,3R,7R)-3,7–dimethyl-2–tridecanol] or Neodiprion sertifer [(2S,3S,7S)-3,7–dimethyl-2–pentadecyl acetate]. This behavioral response of C. ruforum was observed whether (1) parasitoid females had oviposition experience with D. pini eggs or not, (2) parasitoid females emerged from D. pini eggs of a French population or from N. sertifer eggs of a Finnish population, and (3) the tested sex pheromone concentration was low (1 ng/l hexane) or high (100 ng/l hexane). However, C. ruforum did not behaviorally respond to a stereoisomer of the major sex pheromone component of N. sertifer, which is known to act as a sex pheromone antagonist [antagonist = (2S,3R,7R)-3,7–dimethyl-2–pentadecyl acetate]. Thus, C. ruforum responded stereospecifically to the S,S,S configured pheromone of N. sertifer, but not to the S,R,R configuration. Females of the parasitoid D. diprioni were also arrested by the tested major sex pheromone components of the host D. pini. The kairomonal effects of diprionid host sex pheromones on these egg parasitoids are compared to known responses of egg parasitoids to lepidopteran sex pheromones.     

Anti-aphrodisiac Compounds of Male Butterflies Increase the Risk of Egg Parasitoid Attack by Inducing Plant Synomone Production

During mating in many butterfly species, males transfer spermatophores that contain anti-aphrodisiacs to females that repel conspecific males. For example, males of the large cabbage white, Pieris brassicae (Lepidoptera: Pieridae), transfer the anti-aphrodisiac, benzyl cyanide (BC) to females. Accessory reproductive gland (ARG) secretion of a mated female P. brassicae that is deposited with an egg clutch contains traces of BC, inducing Brussels sprouts plants (Brassica oleracea var. gemmifera) to arrest certain Trichogramma egg parasitoids. Here, we assessed whether deposition of one egg at a time by the closely related small cabbage white, Pieris rapae, induced B. oleracea var. gemmifera to arrest Trichogramma wasps, and whether this plant synomone is triggered by substances originating from male P. rapae seminal fluid. We showed that plants induced by singly laid eggs of P. rapae arrest T. brassicae wasps three days after butterfly egg deposition. Elicitor activity was present in ARG secretion of mated female butterflies, whereas the secretion of virgin females was inactive. Pieris rapae used a mixture of methyl salicylate (MeSA) and indole as an anti-aphrodisiac. We detected traces of both anti-aphrodisiacal compounds in the ARG secretion of mated female P. rapae, whereas indole was lacking in the secretion of virgin female P. rapae. When applied onto the leaf, indole induced changes in the foliar chemistry that arrested T. brassicae wasps. This study shows that compounds of male seminal fluid incur possible fitness costs for Pieris butterflies by indirectly promoting egg parasitoid attack.     

Effect of Female Age on Daily Cycle of Sexual Pheromone Emission in Gregarious Egg Parasitoid Anaphes listronoti

The temporal pattern of pheromone emission by Anaphes listronoti females was established in a four-arm olfactometer by observing, at 2-hr intervals, the response of <2-hr-old males to odor produced by individual virgin females of <1, 1, and 2 days of age. This is the first evidence of a sexual pheromone in a Mymaridae. Under a 16L:8D photoperiod and 24°C constant temperature, the responsiveness of males to females of different ages varied significantly during the photophase. When <1-day-old females were used, males made significantly more final choices in the pheromone field than odorless fields at 4 and 6 hr after the onset of the photophase, and their walking speed was significantly higher from 4 to 12 hr, suggesting that females began to release a long-range pheromone during this period. Final choices and high walking speed were observed earlier with 1- and 2-day-old females than with <1-day-old females, and there was a significant decrease in male responses at 6 hr after the onset of the photophase, suggesting a bimodal temporal pattern of sexual pheromone emission.     

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.     

Identification and Synthesis of a Kairomone Mediating Host Location by Two Parasitoid Species of the Cassava Mealybug Phenacoccus herreni

Two encyrtid species, Acerophagus coccois and Aenasius vexans, parasitoids of the cassava mealybug Phenacoccus herreni use a contact kairomone from the body surface of their host as a host-location stimulant. The kairomone was synthesized and identified as O-caffeoylserine based on a combination of chromatographic methods. The synthetic compound was determined to be active.     

Identification of Volatile Synomones, Induced by Nezara viridula Feeding and Oviposition on Bean spp., That Attract the Egg Parasitoid Trissolcus basalis

Bean plants (Vicia faba L. and Phaseolus vulgaris L.) damaged by feeding activity of Nezara viridula (L.) (Heteroptera: Pentatomidae), and onto which an egg mass had been laid, produced volatiles that attracted the egg parasitoid Trissolcus basalis (Wollaston) (Hymenoptera: Scelionidae). Extracts of volatiles of broad bean and French bean plants induced by adults of N. viridula as a result of their feeding activity, oviposition activity, and feeding and oviposition activity combined were analyzed by gas chromatography-mass spectrometry (GC-MS), and tested in Y-tube olfactometer bioassays as attractants for T. basalis females. In extracts from undamaged leguminous plants, green-leaf volatiles were absent or scarcely detected, and monoterpenes and sesquiterpenes were present at trace levels. No significant differences were detected in the profiles of volatiles of undamaged plants, and undamaged plants on which bugs were allowed only to lay eggs. In contrast, feeding and oviposition by adults of N. viridula induced in both leguminous plants a significant increase in terpenoids such as linalool, (E)-beta-caryophyllene, (E,E)-4,8,12-trimethyl-1,3,7,11-tridecatetraene, and (3E)-4,8-dimethyl-1,3,7-nonatriene, which was induced only in French bean plants. Quantitative comparisons revealed increased levels of (E)-beta-caryophyllene in extracts from feeding-damaged plants with N. viridula egg masses compared to feeding-damaged plants without egg masses. In Y-tube olfactometer bioassays, T. basalis females were attracted by extracts of both leguminous plants only when N. viridula adults were allowed to feed and oviposit upon them. Fractionation of extracts of volatiles from broad bean plants with N. viridula feeding damage and egg masses yielded two fractions. but only the fraction containing (E)-beta-caryophyllene was attractive to the egg parasitoid. These findings indicate that N. viridula feeding and oviposition induce leguminous plants to produce blends of volatiles that are characterized by increased amounts of (E)-beta-caryophyllene, and these blends attract female T. basalis. The role of (E)-beta-caryophyllene as a potential synomone for T. basalis is discussed.     

Two Regional Strains of a Phoretic Egg Parasitoid, Telenomus euproctidis (Hymenoptera: Scelionidae), That Use Different Sex Pheromones of Two Allopatric Tussock Moth Species as Kairomones

The egg parasitoid, Telenomus euproctidis Wilcox (Hymenoptera: Scelionidae), is phoretic on females of two allopatrically distributed tussock moths, Euproctis pseudoconspersa (Strand) and Euproctis taiwana (Shiraki) (Lepidoptera: Lymantriidae). Crossing experiments between the two regional parasitoid strains indicated no evidence for their reproductive isolation. More wasps were found on the locally occurring host, E. pseudoconspersa, than on E. taiwana, when virgin females of the two moth species were exposed concurrently in the field for 24 hr in Ibaraki Japan. In Ibaraki, many wasps were caught in traps baited with the synthetic sex pheromone of E. pseudoconspersa, 10,14-dimethylpentadecyl isobutyrate (10Me14Me-15:iBu), but none with that of E. taiwana, (Z)-16-methyl-9-heptadecenyl isobutyrate (16Me-Z9-17:iBu) or blank traps. In Okinawa, Japan, more wasps were found on E. taiwana than on E. pseudoconspersa, and many wasps were caught in traps baited with 16Me-Z9-17:iBu, but only a few with 10Me14Me-15:iBu, and none with blank traps. These results suggest that local wasp strains discriminate between the two sex pheromones, and they strongly prefer the sex pheromone of the moth occurring at their location.     

The Gastropod Menace: Slugs on Brassica Plants Affect Caterpillar Survival through Consumption and Interference with Parasitoid Attraction

Terrestrial molluscs and insect herbivores play a major role as plant consumers in a number of ecosystems, but their direct and indirect interactions have hardly been explored. The omnivorous nature of slugs makes them potential disrupters of predator-prey relationships, as a direct threat to small insects and through indirect, plant-mediated effects. Here, we examined the effects of the presence of two species of slugs, Arion rufus (native) and A. vulgaris (invasive) on the survivorship of young Pieris brassicae caterpillars when feeding on Brassica rapa plants, and on plant attractiveness to the main natural enemy of P. brassicae, the parasitoid Cotesia glomerata. In two separate predation experiments, caterpillar mortality was significantly higher on plants co-infested with A. rufus or A. vulgaris. Moreover, caterpillar mortality correlated positively with slug mass and leaf consumption by A. vulgaris. At the third trophic level, plants infested with slugs and plants co-infested with slugs and caterpillars were far less attractive to parasitoids than plants damaged by caterpillars only, independently of slug species. Chemical analyses confirmed that volatile emissions, which provide foraging cues for parasitoids, were strongly reduced in co-infested plants. Our study shows that the presence of slugs has the potential to affect insect populations, directly via consumptive effects, and indirectly via changes in plant volatiles that result in a reduced attraction of natural enemies. The fitness cost for P. brassicae imposed by increased mortality in presence of slugs may be counterbalanced by the benefit of escaping its parasitoids.     

Pheromonal Discrimination of Sex by Male and Female Leopard Geckos (Eublepharis macularius)

The ability of male and female Eublepharis macularius to discriminate among pheromones of males and females and a blank control was investigated. Stimuli were presented on ceramic tiles in the animal's home cages. Males tongue-flicked at significantly lower rates in response to male stimuli than to female and control stimuli. Males also performed aggressive behaviors toward male, but not female or control, stimuli, and tail vibrations toward female, but not male or control, stimuli. Mean tongue-flick rates by females did not differ significantly among conditions. Discrimination of male pheromones by females was demonstrated by greater performance of labial-licking and chin-rubbing in response to male stimuli than to either female or control stimuli.     

Enhancement of Attraction to Sex Pheromones of Spodoptera exigua by Volatile Compounds Produced by Host Plants

We measured the effects of exposure to volatile compounds produced by host plants on the rate of capture of male Spodoptera exigua using synthetic sex pheromones. Exposure to volatile compounds stimulated strong electroantennographic responses of male S. exigua. The behavioral responses of male moths to combinations of sex pheromone and volatile compounds were tested in wind tunnel experiments. When lures were baited with synthetic sex pheromone plus benzaldehyde, phenylacetaldehyde, (Z)-3-hexenyl acetate, or linalool, respectively, the landing rate of S. exigua males was increased by 101.4%, 79.6%, 60.6%, and 34.3%, respectively, compared to sex pheromone alone. In field tests, traps baited with either pheromone + (E)-2-hexenal, pheromone + phenylacetaldehyde, pheromone + (Z)-3-hexenyl acetate, or pheromone + (Z)-3-hexenol enhanced moth catches by 38.8%, 34.6%, 24.6%, and 20.8%, respectively compared to traps baited with pheromone alone. In a second field experiment, more S. exigua males were trapped with a combination of a synthetic sex pheromone blend and several individual host plant volatiles compared to synthetic sex pheromone alone. These results suggest that some host plant volatiles enhance the orientation response of S. exigua male moths to sex pheromone sources.     

Field Discrimination of Oilseed Rape, Brassica napus Volatiles by Cabbage Seed Weevil, Ceutorhynchus assimilis

The response of the cabbage seed weevil, Ceutorhynchus assimilis to yellow water traps baited with some components of oilseed rape (Brassica napus) odor was studied in a series of field experiments. Four isothiocyanates (NCS), five other amino acid derivatives (aromatic compounds), and two fatty acid derivatives are detected by peripheral olfactory receptors of C. assimilis. 3-Butenyl, 4-pentenyl, and 2-phenylethyl NCS were attractive when released individually during the period when weevils were immigrating to the host crop, but allyl NCS was not. A mixture of these four isothiocyanates was attractive during the periods of weevil immigration and dispersal. However, during the host colonization phase, 2-phenylethyl NCS, like the mixture of the four isothiocyanates, elicited no response or was repellent. Phenylacetonitrile and benzyl alcohol were attractive during the colonization phase. The fatty acid derivative (Z)-3-hexen-1-ol did not affect weevil catch in an experiment conducted when the insects were already established in the crop, but it depressed the catch in another conducted when weevils were dispersing from the crop. These results suggest that yellow traps baited with 2-phenylethyl NCS can be used to monitor immigration into crops in the spring and that phenylacetonitrile may be useful for assessing numbers of weevils colonizing the crop throughout the summer.     

Semiochemical-Mediated Location of Host Habitat by Apanteles carpatus (Say) (Hymenoptera: Braconidae), a Parasitoid of Clothes Moth Larvae

In Y-tube olfactometer bioassays, adult Apanteles carpatus (Say), were attracted to beaver or rabbit pelts infested with larvae of the casemaking clothes moth (CCM)Tinea pellionella L. Porapak Q-captured volatiles from a CCM-infested beaver pelt were also very attractive, whereas isolated CCM larvae or larval feces were not. Coupled gas chromatographic–electroantennographic detection (GC-EAD) analysis of the Porapak Q volatile extract revealed two compounds that elicited responses by A. carpatus antennae. Coupled GC–mass spectrometry (MS) in electron impact and chemical ionization modes of these compounds indicated, and GC-MS and GC-EAD of authentic standards confirmed, that they were nonanal and geranylacetone. While each compound singly did not attract A. carpatusa 1:1 blend of both compounds was as attractive as the volatile extract. Because these compounds are host habitat-derivedA. carpatus must be a habitat rather than host specialist, responding to kairomonal indicators of localized and specific habitats such as animal hair or feather. The tritrophic interaction between A. carpatusits clothes moth hosts and their animal-derived habitats is similar to the well-studied relationship between parasitoids of insect herbivores and their host plant habitats.     

Mineralization of Calcium Carbonate Induced by Egg Substrate and an Electric Field

Egg substrates, including eggshell, egg membranes, and egg white, exert a significant influence on CaCO₃ biomineralization. CaCO₃ crystallization at different temperatures, concentrations, and with various egg substrates was systematically investigated via rapidly mixing solution method. The crystals were determined by scanning electron microscopy, Fourier transform infrared spectroscopy, and X‐ray diffraction. As the CaCl₂ concentration increased, the CaCO₃ crystal form gradually changed from calcite to vaterite. Under rising temperature, the single‐hole vaterite and besom‐like aragonite were formed on the raw and boiled egg white substrate, respectively. The effects of electric field on CaCO₃ crystallization were also explored. The findings may offer a novel approach to CaCO₃ synthesis.     

Discrimination Between Self-Produced Pheromones and Those Produced by Individuals of the Same Sex in the Lizard Cordylus cordylus

Male and female Cordylus cordylus can discriminate between tiles labeled by their own pheromones and tiles labeled by individuals of the same sex, as shown by elevated tongue-flick rates in which the tongue contacts only air above the tiles labeled by other individuals and tongue-flicks in which the tongue contacts the tiles themselves. Potential pheromone sources for these discriminations are the femoral glands, cloacal glands, generation glands, ventral skin, and excretory products. Although studied in few species, pheromonal discriminations between self and other individuals, familiar and unfamiliar individuals, and kin and unrelated individuals appear to be broadly distributed in lizards, occurring in Iguania and in both Gekkonoidea and Scincomorpha within Scleroglossa. Both sexes of C. cordylus defend territories against both sexes. An ability to distinguish pheromones of other individuals of the same sex from self-produced pheromones would allow detection of intruders, but pheromonal discriminations among individuals would be more useful. Adaptive functions of pheromonal discriminations for residents and nonresidents and discriminatory abilities required are discussed. Tongue-flicks touching labeled tiles differed between experimental conditions, suggesting vomerolfactory discrimination, but the interpretation of a similar difference for tongue-flicks that contacted no substrate is problematical. Such air tongue-flicks might indicate sampling of volatile molecules for delivery to the vomeronasal organs. Experiments are needed to conclusively determine the sensory bases of the discriminations and the role of air tongue-flicks.