Evidence for a Host-Marking Pheromone in White Spruce Cone Fly, Strobilomyia neanthracina

Cone flies (Strobilomyia spp.) lay eggs on coniferous cones, and larvae must complete development within the cone on which the eggs are laid. Previous field surveys showed that egg distributions of several species are uniform, suggesting that females avoid ovipositing on cones with conspecific eggs or larvae. In both the field and laboratory, S. neanthracina females walked around cones following oviposition, touching their mouthparts to the cone as they walked. In laboratory bioassays, where females were presented with a cone on which they or another female had oviposited and touched with mouthparts, or one without an egg, females laid preferentially on cones without eggs. However, females laid randomly when presented with a cone on which a female had oviposited but been prevented from touching with her mouthparts following oviposition or another cone without an egg. This indicates that females deposit a host-marking pheromone with their mouthparts following oviposition and that this deters further oviposition on marked hosts. Laboratory and field bioassays indicated that the host-marking pheromone is located in the head and thorax of female flies.     

Factors Accounting for the Variability in the Behavioral Response of the Onion Fly (Delia antiqua) to n-Dipropyl Disulfide

The onion fly Delia antiqua is a specialist herbivore attacking only onions and closely related Allium species. n-Dipropyl disulfide (Pr₂S₂) has long been known to be attractive to onion flies in the laboratory and in the field. However, the insect's response is highly variable. Using behavioral bioassays we found that Pr₂S₂ was highly attractive to gravid, mated female onion flies, but did not stimulate oviposition. The response of female onion flies was concentration dependent. The physiological state of the flies (i.e., age, mating status, egg load/oviposition experience) also affected their responsiveness. The response of both sexes of onion flies varied with age, but females were always more strongly attracted than males. Responsiveness of females increased during the first 10 days after emergence. It stayed at a high level until 21 days after which the experiment was terminated. The responsiveness of males reached a maximum at 6–7 days after which it declined. Mated, gravid females responded more strongly to Pr₂S₂ than unmated, gravid females. Females deprived of the opportunity to oviposit were more attracted compared to females that had oviposited on cut onions prior to the experiment. Electroantennograms (EAG) of females revealed a higher response to stimulation compared to males. The EAG-response of females was not affected by mating status.     

Host selection byHylemya antiqua laboratory bioassay and methods of obtaining host volatiles

An oviposition bioassay for the onion maggot,Hylemya antiqua (Meigen), is described in which females, in response to volatile stimulants, oviposit through small apertures onto moistened filter paper. Onion volatiles that act as attractants and oviposition stimulants were captured in Porapak Q from air passed over chopped onions in glass chambers. Pentane extracts from odor-impregnated Porapak Q induced 30–50% of the oviposition that occurred in response to 15-g onion-slice stimuli. Extracts presented in pentane on waxed dental cotton wicks induced more constant oviposition over a 3-day period than extracts on unwaxed wicks. Extract of the Porapak Q-captured volatiles from bulbs of fresh, actively growing onions elicited a much stronger response than did stem and leaf extracts from the same onions. The bioassay techniques and chemical procedures developed in this study could be used in chemical isolation programs for host attractants or oviposition stimulants forH. antiqua or similar species.Diptera:Anthomyiidae.Research supported by National Research Council of Canada, Grant Nos. A3881 and A3706.     

Influence of different habitats and mating on olfactory behavior of onion flies seeking ovipositional hosts

Using traps baited with natural and synthetic onion volatiles, we examined the effects of different habitats and mating on the olfactory behavior of laboratory-reared and wild onion flies. Rankings of olfactory treatments as host-finding stimuli for females were dependent on their mating status and the habitat in which they were foraging. In habitats devoid of hosts, traps baited with individual alkyl sulfides were as effective as 4-day-old chopped onions and more effective than 1-day-old onions in eliciting host-finding behavior in laboratory-reared unmated females (LUF) and laboratory-reared mated females (LMF). However, upwind dispersal and percent recapture were always significantly greater in LUF. In one experiment, Pr₂S₂ was 19 times more attractive to LMF in a fallow field, as than it was in an onion field. Reduced effectiveness of alkyl sulfides as host-finding stimuli in onion fields probably results in part because they are less findable, but more importantly because of a change in searching behavior after females have mated. Evidence to support the latter contention is that traps baited with alkyl sulfides and onions were equally findable by unmated females in both habitats. The behavior of LMF was identical to that of wild females, whereas the behavior of LUF was identical to wild males. The hypothesis that olfactory host-finding behavior in onion flies is modified by the resource level was upheld. Alkyl sulfides appear to be the primary, and possibly the only, chemical effectors of host-finding at the patch level of resource distribution, whereas the complex blend emitted by aged, chopped, or damaged onions appears to be acting at the final level of host-finding, while egg-laying females are moving between adjacent hosts in search of an optimal oviposition site.     

Evidence for the accessory glands as the site of production of the oviposition attractant and/or stimulant ofLutzomyia longipalpis (Diptera: Psychodidae)

The phlebotomine sandflyLutzomyia longipalpis Lutz and Neiva, the vector of visceral leishmaniasis in South America, has recently been shown to produce an oviposition semiochemical. In the present study it was found that a nonpolar extract of eggs was attractive and/or stimulatory to ovipositing females. A Chromatographic investigation indicated the presence of similar compounds in accessory glands and egg extracts. Extract of accessory gland was also found to elicit a positive oviposition response. It is concluded that the pheromone is produced in the accessory glands and is secreted onto the eggs during oviposition.     

Plant response to eggs vs. Host marking pheromone as factors inhibiting oviposition byPieris brassicae

Pieris brassicae L. butterflies secrete miriamides onto their eggs. These avenanthramide alkaloids are strong oviposition deterrents when sprayed onto a cabbage leaf. However, these compounds could not be detected in cabbage leaves from which egg batches had been removed two days after deposition and that still showed oviposition deterrency. It was concluded that the miriamides were not directly responsible for the avoidance by females of occupied leaves while searching for an oviposition site. Evidence was obtained that cabbage leaves themselves produce oviposition deterrents in response to egg batches. Fractions containing potent oviposition deterrents could be isolated from surface extracts of leaves from which previously laid egg batches had been removed. The term host marking pheromone that was used previously is not applicable in this case.     

Earthworm mucus stimulates oviposition in a predatory fly (Diptera: Anthomyiidae)

Coenosia tigrina larvae feed on earthworms. We hypothesized that earthworm mucus contains a kairomone that stimulates oviposition behavior in adultC. tigrina females, thus minimizing the search area in the soil required for newly eclosed larvae to find earthworms. In bioassays, adult females responded with extension of the ovipositor 25–43% of the time to earthworm-mucus-soaked filter paper disks compared to 6–7% in response to water-soaked disks. Ovipositor extension on mucus-soaked disks was followed by egg-laying 29% of the time and 0% of the time on water-soaked disks. Egg-laying byC. tigrina followed a diurnal periodicity, with most eggs laid in the latter half of the photophase even in the absence of earthworm mucus. More eggs were deposited from 1600 to 1800 hr by females given access to earthworm mucus during that period than were deposited by females not given access. There was no difference in the number of eggs deposited from 0600 to 0800 hr, by females given access to earthworm mucus or not. This is a time of day when few eggs are normally laid. This paper is the first report of an earthworm-produced kairomone in an insect-earthworm interaction. The kairomone may have potential for enhancing biological control of the onion maggot,Delia antiqua, which is a prey of adultC. tigrina.     

Oviposition Stimulants for Sulfur ButterflyColias erate poliographys: Cyanoglucosides as Synergistis Involved in Host Preference

Host-plant chemicals stimulating oviposition by a Leguminosae-feeding pierid butterflyColias erate poliographyswere isolated and identified from one of its primary host plants, white clover (Trifolium repens). Females readily deposited eggs in response to methanolic extracts of the plant, and subsequent partition of the extracts with organic solvents revealed that chemical constituents critical for host recognition reside in the water-soluble fraction. Further fractionation of the hydrosoluble fraction by column chromatography led to the separation of an active fraction and two cyanoglucosides, linamarin and lotaustralin. Conspicuous oviposition response was evoked by unidentified polar compound(s), while these cyanoglucosides exerted no stimulatory activity by themselves. However, ovipositing females preferred samples containing either of the two cyanoglucosides. In dual-choice bioassays, significantly more eggs were laid on samples admixed with the cyanoglucosides, suggesting that the cyanoglucosides serve as synergistic oviposition stimulants and could play an important role in host selection.     

Influence of Synthetic Oviposition Pheromone and Volatiles from Soakage Pits and Grass Infusions Upon Oviposition Site-Selection of Culex Mosquitoes in Tanzania

The response of Culex mosquitoes to (5R,6S)-6-acetoxy-5-hexadecanolide (the synthetic oviposition pheromone, SOP), emanations from soakage pit water (SPW), and grass infusions (GI) was studied in pit latrines in Muheza, Tanzania. Water treated with the synthetic oviposition pheromone received more egg rafts of Culex quinquefasciatus Say and Cx. cinereus Theobald than did the untreated water (P < 0.001). The residual activity of SOP did not decrease over a nine-day period, with the geometric mean number of egg rafts laid in the SOP-treated water varying daily between 2.5 and 8.9 as compared to 0.1–1.6 egg rafts laid in tap water. However, SOP did not attract ovipositing mosquitoes to nonbreeding sites, and both treatment and control received few eggs. SPW and GI attracted ovipositing Cx. quinquefasciatus, Cx. cinereus, and Cx. tigripes Grandpré and de Charmoy with the number of eggs rafts deposited in bowls containing SPW or GI being larger than that deposited in bowls with tap water (P < 0.05). More egg rafts of Cx. quinquefasciatus were deposited in SOP-treated water than in SPW (P < 0.05). When SOP was compared with SOP + SPW, more egg rafts of Cx. quinquefasciatus were deposited in bowls with the latter combination than in bowls containing SOP or SPW only (P < 0.05), indicating a synergistic effect between SOP and SPW. Similarly, when SOP-treated water and GI were compared with SOP + GI, more egg rafts of Cx. quinquefasciatus were laid in the latter bowl, indicating a synergistic effect between SOP and GI. The result suggest that the combined use of SOP and organically enriched water can be employed in monitoring of Cx. quinquefasciatus for control programs. This is the first record of the attraction of Cx. cinereus and Cx. tigripes to oviposition stimuli.     

Search for potent attractants of onion flies

Of various chopped vegetables tested,Allium spp. high in propyl-containing alkyl sulfides (e.g.,cepa group) caught the most onion flies in trapping tests in the field. Fly catches to chopped onion increased with bait quantity. Attractancy of chopped onion changed dramatically during aging in the field; catch increased over the first few days, peaked at ca. fivefold over fresh material by 3–5 days, and then declined sharply. This age-dependent increase in attraction was not seen for garlic (known to have antimicrobial properties) nor with chopped onion mixed with chopped garlic. These data suggested that attraction of onion flies to onions was strongly influenced by microbial activity associated with decomposing onions. The bacteriumKlebsiella pneumoniae was identified as a major colonizer of onions maximally attractive to onion flies. This increased attraction is not due to the previously reported microbially produced volatiles ethyl acetate and tetramethyl pyrazine.Diptera: Anthomyiidae.Paper No. 11047 of the Michigan State University Agricultural Experiment Station.     

Production of onion fly attractants and ovipositional stimulants by bacterial isolates cultured on onion

Decomposing onions at certain microbial successional stages produce potent volatile attractants and ovipositional stimulants of the onion fly,Delia antiqua (Diptera: Anthomyiidae). A reproducible source of these compounds was obtained by culturingErwinia carotovora var.carotovora (EC) on sterile onion tissue. In laboratory choice tests, EC-inoculated onion was more attractive thanKlebsiella pneumoniae (KP) cultured on onion, EC cultured on potato (a nonhost of onion fly), or the chemical synthetic baits dipropyl disulfide and an aqueous solution of 2-phenylethanol and pentanoic acid. Onion flies were mildly attracted to potato after inoculation with EC, but females did not accept EC-inoculated potato for oviposition. This work emphasizes that sources of semiochemicals may need to be defined microbiologically as well as physically and chemically.Diptera: Anthomyiidae.Paper No. 12106 of the Michigan State University Agricultural Experiment Station.     

Oviposition in Delia platura (Diptera, Anthomyiidae): The Role of Volatile and Contact Cues of Bean

The choice of a suitable oviposition site by female insects is essential for survival of their progeny. Both olfactory and contact cues of the oviposition site may mediate this choice. The polyphagous Delia platura (Diptera: Anthomyiidae), a severe agricultural pest of numerous crops, lays eggs in the soil close to germinating seeds. Maggots feed upon the cotyledons. Only little is known about the cues guiding oviposition behavior. In this study, the effects of both olfactory and contact cues of beans (Phaseolus vulgaris) on oviposition of D. platura females were tested. Egg deposition on germinated beans was preferred to egg deposition on ungerminated beans or on beans in different postgerminating developmental stages. Olfactory cues of germinating beans alone stimulated female flies to lay eggs. Additional contact cues of germinating beans seemed to enhance the response, but the difference was not significant. Surface extracts of germinating beans sprayed on surrogate beans showed that both polar and nonpolar substances stimulated oviposition of D. platura flies. Gas chromatography-electroantennographic detection recordings of head space samples of germinating beans showed positive response of females to different compounds. We conclude that olfaction plays a major role when D. platura females are searching for oviposition sites. Volatile compounds released from germinating beans such as 4-hydroxy-4-methyl-2-pentanone, 1-hepten-3-one, 1-octen-3-ol, and 3-octanone should be considered as key compounds that mediate oviposition behavior. The use of different sensory modalities by closely related species of Delia is discussed.     

Oviposition response ofLobesia botrana females to long-chain free fatty acids and esters from its eggs

Avoidance of occupied ovisposition sites supposes that females perceive information related to their own progency. Fatty acids identified from egg extracts have been reevaluated using a different extraction method, and we have investigated the dose-dependent oviposition response of European grape vine moths (Lobesia botrana) to myristic acid, palmitic acid, palmitoleic acid, stearic acid, oleic acid, linoleic acid, methyl palmitate, methyl oleate, and ethyl palmitate; all except ethyl palmitate have been identified from eggs ofL. botrana. A methylene dichloride extract of eggs fromL. botrana revealed the presence of saturated free fatty acids (myristic, palmitic, and stearic) and unsaturated acids (palmitoleic, oleic, linoleic, and linolenic) in amounts ranging from 3.9 ng/egg equivalent for myristic acid to 30 ng/egg equivalent for palmitic and oleic acids. The extract also contained traces of methyl palmitate and methyl stearate. The greatest avoidance indexes were observed in response to palmitic, palmitoleic, and oleic acids. All the other compounds tested caused weaker responses. A reduction in the number of eggs laid was observed when moths were exposed to each of the esters applied at 0.3 µg per application spot. Reduction in eggs laid was also observed at a 10-fold higher dose of oleic acid. The present results confirm that general and simple molecules can be involved in the regulation of oviposition site selection and that they may participate in chemical marking of the eggs.     

Onion fly and little house fly host finding selectively mediated by decomposing onion and microbial volatiles

Responses of onion flies,Hylemya antiqua (Meigen), to various synthetic onion and microbial volatiles as well as volatiles from microbial cultures and decomposing onions were tested to characterize the most effective host-finding stimuli. Of nine onion and microbial volatiles tested individually, only the known attractant,n-dipropyl disulfide, caught significant numbers of flies. However, a blend of these volatiles attracted more flies than any single chemical, includingn-dipropyl disulfide. In another experiment, agar plates inoculated with microorganisms from decomposing onions did not attract onion flies. However, cut onions inoculated with microorganisms and conditioned 4 days caught more onion flies than freshly cut onions andn-dipropyl disulfide. These results suggest that a blend of chemicals, rather than a single key chemical, is the more effective host-finding stimulus, and that microbial activity enhances the attractancy of a blend of onion volatiles. Large numbers ofFannia canicularis (L.), the little house fly, responded to the microbial cultures, demonstrating the existence of a potent attractant for this muscid.Diptera: Anthomyiidae.Diptera: Muscidae.Paper No. 9470 of the Michigan State University Agricultural Experiment Station. Received for publication May 23, 1980.     

Experience-Induced Habituation and Preference Towards Non-Host Plant Odors in Ovipositing Females of a Moth

In phytophagous insects, experience can increase positive responses towards non-host plant extracts or induce oviposition on non-host plants, but the underlying chemical and behavioral mechanisms are poorly understood. By using the diamondback moth, Plutella xylostella, its host plant Chinese cabbage, and a non-host plant Chrysanthemum morifolium, as a model system, we observed the experience-altered olfactory responses of ovipositing females towards volatiles of the non-host plant, volatiles of pure chemicals (p-cymene and α-terpinene) found in the non-host plant, and volatiles of host plants treated with these chemicals. We assessed the experience-altered oviposition preference towards host plants treated with p-cymene. Naive females showed aversion to the odors of the non-host plant, the pure chemicals, and the pure chemical-treated host plants. In contrast, experienced females either became attracted by these non-host odors or were no longer repelled by these odors. Similarly, naive females laid a significantly lower proportion of eggs on pure chemical-treated host plants than on untreated host plants, but experienced females laid a similar or higher proportion of eggs on pure chemical-treated host plants compared to untreated host plants. Chemical analysis indicated that application of the non-host pure chemicals on Chinese cabbage induced emissions of volatiles by this host plant. We conclude that induced preference for previously repellent compounds is a major mechanism that leads to behavioral changes of this moth towards non-host plants or their extracts.     

Oviposition Responses by Hessian Fly, Mayetiola destructor, to Wheats Varying in Surfaces Waxes

Leaf waxes are known to contain oviposition stimulants for Hessian fly. In dual-choice tests comparing seedlings of each of three nonglaucous lines of wheat with its corresponding glaucous line, Hessian flies laid similar numbers of eggs on each genotype. However, when plants from these genotypes were tested at the flag-leaf stage, Hessian flies deposited 25–100% more eggs on the nonglaucous genotype compared to normal wax genotype in each pair. Leaf waxes extracted from the genotypes in one of these pairs (Avalon and nonglaucous Avalon) at the seedling and the flag-leaf stage and tested on paper leaf models elicited oviposition responses similar to those observed on the intact plants. Scanning electron microscopy showed differences in crystal density on nonglaucous and glaucous genotypes only at the flag-leaf stage. Gas chromatography–mass spectrometry of the leaf wax extracts showed that fly oviposition responses were associated with differences in the chemical composition of the waxes. This is the first report of Hessian fly oviposition responses varying among genotypes of wheat with different surface wax composition.     

Influence of Stimuli from Camellia japonica on Ovipositional Behavior of Generalist Herbivore Epiphyas postvittana

Stimuli from the plant Camellia japonica that influence the host-finding and acceptance behaviors of adult females of the generalist herbivore Epiphyas postvittana were investigated. In a binary choice test in a small (35-cm-diam.) arena, females discriminated between a plant and a laminated card model, laying many more eggs on the plant. Observations of females showed that the greater number of eggs laid on the plant were primarily due to on-plant behaviors, with females spending significantly more time per visit on the plant than on the model. Interestingly, females landed a similar number of times on the plant as on the model, suggesting that volatile chemicals from the plant did not influence host-finding. This observation was further supported by wind-tunnel studies, in which females showed little or no upwind flight activity in response to plants and laid similar numbers of eggs on upwind and downwind plants. Leaf surface texture, a combination of smoothness and fine structure (consisting of the midvein and other raised leaf veins), was found to stimulate egg laying by females. Methanol, dichloromethane, and pentane extracts were made of the waxes on the leaf surface. Of these three extracts, only the nonpolar (pentane) one stimulated egg laying by females. Although we did not find a role for volatile plant chemicals in host finding, they appeared to stimulate increased egg laying when the female was on the plant. It appears likely that these chemical and tactile stimuli in C. japonica are general stimuli, which may be found in a large number of plants and, in combination with the feeding preferences of the larva, may explain the generalist herbivorous nature of this pest.     

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.     

Oviposition Responses of Gravid Female <Emphasis Type="Italic">Culex quinquefasciatus</Emphasis> to Egg Rafts and Low Doses of Oviposition Pheromone Under Semifield Conditions

Semifield experiments were conducted to study the oviposition of gravid Culex quinquefasciatus females in response to one or 10 egg rafts, or 3.0 μg of synthetic oviposition pheromone, (-)-(5R,6S)-6-acetoxy-5-hexadecanolide, a dose equivalent to 10 egg rafts. These treatments were added to the small bowls filled with either hay infusion or water in a small (0.3-m spacing) or a large-square design (3.4-m spacing). Oviposition choice was more pronounced in the “small square” assays. Mean number of egg rafts laid in response to a single egg raft in an oviposition jar filled with hay infusion was significantly greater than with all other treatments. When the oviposition pheromone dose was increased from one to 10 rafts or when 3.0 μg synthetic oviposition pheromone were dispensed on a floating receptacle, synergistic effects were observed between the oviposition pheromone and the hay infusion at both distances. This study is the first demonstration that the amount in a single raft induces oviposition of gravid Cx. quinquefasciatus females under semi-natural conditions.     

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.