EFFECT OF PBAN ON PHEROMONE PRODUCTION BY MATED <Emphasis Type="Italic">Heliothis virescens</Emphasis> AND <Emphasis Type="Italic">Heliothis subflexa</Emphasis> FEMALES

Mated female Heliothis virescens and H. subflexa were induced to produce sex pheromone during the photophase by injection of pheromone biosynthesis activating neuropeptide (PBAN). When injected with 1 pmol Hez-PBAN, the total amount of pheromone that could be extracted from glands of mated females during the photophase was similar to that extracted from virgin females in the scotophase. The PBAN-induced profile of pheromone components was compared between mated, PBAN-injected females and virgin females during spring and fall. Virgin females exhibited some differences in the relative composition of the pheromone blend between spring and fall, but no such temporal differences were detected in PBAN-injected, mated females. Because the temporal variation in pheromone blend composition was greater for virgin females than for PBAN-injected females, PBAN can be used to determine a females native pheromone phenotype. This procedure has the advantages that pheromone glands can be extracted during the photophase, from mated females that have already oviposited.     

Identification and synthesis of a male-produced sex pheromone from the stink bug Chlorochroa sayi

The reproductive behavior of the stink bug Chlorochroa sayiwas studied in the laboratory. There was a sexual maturation period of about 10 days before bugs began mating. Sexually mature adult bugs engaged in courtship consisting of antennation and head-butting of the female by the male, before the female adopted a receptive posture and copulation occurred. Both sexes mated multiple times during their life-spans, with the mean duration of copulations of virgin bugs (42.3 ± 19.6 min) and experienced bugs (37.3 ± 28.4 min) being similar. Most matings were initiated in the late afternoon or evening, when pheromone production by males was greatest. Males transferred sperm and nutrients constituting about 17% of their body weight to females during mating. Three male-specific components, methyl geranate, methyl citronellate, and methyl (E)-6-2,3-dihydrofarnesoate in a ratio of 100 : 0.45 : 1.6, were first detected in volatiles collected from male bugs on green beans about 9–12 days after the final molt to the adult stage. In vertical Y-tube bioassays, females were attracted to odors from mature male bugs, and to a blend of the three male-produced components. Low numbers of females also were attracted in field trials with the three-component blend. The relatively weak attraction may be a result of other, as yet unknown cues being required in addition to the pheromone, such as visual or substrate-borne vibrational cues.     

Role of semiochemicals in mate location by parasitic sea louse,Lepeophtheirus salmonis

The role of olfaction and diffusible pheromones in mate location behavior of sea lice, Lepeophtheirus salmonis, was assessed with Y-tube behavioral bioassays. The pheromone emitting animals were located in a chamber in one arm of a Y-tube arena, with artificial seawater flowing through both arms. Adult male sea lice displayed both activation and directional responses to seawater conditioned with preadult II virgin females, but were only activated by mated adult female conditioned water. Further, when males were given the choice of preadult II virgin females or mated adult females, a significant number of males chose the arm with the preadult II virgin females. Adult males showed activation responses when presented with water conditioned with adult males but were not attracted to them. When presented with adult males, preadult II virgin females showed only directional responses, but not activation responses. Preadult II virgin female conditioned water was extracted using solid-phase extraction (SPE) protocols pioneered for semiochemical isolation. Adult male sea lice showed significant directional responses to the preadult II virgin female SPE extract. Distillation under vacuum was performed on the extract to give a distillate comprising components with a molecular weight range and physical properties comparable to those of compounds utilized as volatile semiochemicals by terrestrial organisms and a residue comprising components with higher molecular weight range comparable to those utilized as involatile semiochemicals. Adult males were found to be both significantly activated and attracted to the distillate, but not to the residue. This research provides evidence that small, lipophilic organic molecules are used by sea lice as sex pheromone signals to locate a member of the opposite sex.     

Mechanism and Behavioral Context of Male Sex Pheromone Release in <Emphasis Type="Italic">Nasonia vitripennis</Emphasis>

Males of the parasitic wasp Nasonia vitripennis (Hymenoptera: Pteromalidae) attract virgin females by releasing a sex pheromone composed of (4R,5R)- and (4R,5S)-5-hydroxy-4-decanolide (HDL). The pheromone is biosynthesized in the rectal vesicle of males. In the present study, we investigated the mechanism and behavioral context of pheromone release, and determined the range of activity and the longevity of the chemical signal. Our data show that the sex pheromone of N. vitripennis is substrate-borne and is deposited on surfaces by dabbing movements of the abdominal tip, a behavior previously described in N. vitripennis males as ‘abdomen dipping’. Chemical markings deposited by a single male were highly attractive to virgin females. Chemical analyses revealed the presence of HDL in surface washings of marked areas, and HDL amounts correlated with male marking activity. Pheromone deposition occurred spontaneously without any additional cues being present, but marking intensity increased greatly after copulation or after a single contact with a virgin female. In contrast, marking intensity was not influenced by the presence of host puparia. Male pheromone deposits were perceived by females in a still-air olfactometer at distances of up to 4.5 cm and remained attractive for at least 2 h. The function of the substrate-borne sex pheromone is discussed with respect to the mating system of N. vitripennis.     

Pheromone Components from Body Scales of Female <Emphasis Type="Italic">Anarsia lineatella</Emphasis> Induce Contacts by Conspecific Males

Pheromonal communication of adult peach twig borers, Anarsia lineatella Zeller (Lepidoptera: Gelechiidae), was reinvestigated based on recent findings that virgin female-baited traps were more attractive to mate-seeking males than a two-component synthetic sex pheromone consisting of (E)-5-decen-1-yl acetate (1000 μg) and (E)-5-decen-1-ol (100 μg), suggesting that females use additional pheromone components. Hypothesizing that these additional components may be released from body parts other than abdominal sex pheromone glands, we extracted female body scales and analyzed aliquots by coupled gas chromatographic–electroantennographic detection (GC-EAD) and GC–mass spectrometry. Eight straight-chain and four methylated aliphatic hydrocarbons, as well as two acetates, all elicited responses from excised male antennae. In laboratory experiments with synthetic candidate pheromone components, a combination of octadecyl acetate, (R)-11-methyltricosane, and (S)-11-methyltricosane in the presence of gland-derived sex pheromone components were shown to elicit contact of female decoys by males. However, body pheromone components did not enhance attractiveness of sex pheromone components in field trapping experiments, suggesting that they are effective only at close range and that other stimuli are responsible for superior attractiveness of female-baited traps.     

(S)-(+)-linalool,a mate attractant pheromone component in the bee Colletes cunicularius

Enantiomerically pure (S)-(+)-linalool was the main constituent in the extracts of the cephalic secretions of virgin females, mated females, freshly emerged males, and patrolling males of the solitary bee Colletes cunicularius. After copulation, the content of (S)-(+)-linalool emitted by the female was strongly reduced. Electrophysiological experiments revealed that both enantiomers of linalool elicited responses from the antennae of the males. Field tests using the pure enantiomers and the racemate of linalool showed that the number of male bees attracted was highest for (S)-(+)-linalool. The search flight activity in the mating flight area increased dramatically when patrolling males were presented with (S)-(+)-linalool vs (R)-(–)-linalool. Taken together, these data indicate a mate attractant pheromone function of (S)-(+)-linalool.     

Particular Levels of Odors Released by Virgin Females Attract Conspecific Males of the Funnel-Web Spider <Emphasis Type="Italic">Allagelena difficilis</Emphasis>

Female-released chemical signals are crucial clues for mate-searching males to locate and gain sexual receptivity of conspecific females. Abundant behavioral evidence indicates that female spiders release sex pheromones to guide mate-searching behavior of conspecific mature males. However, the chemical nature of spider pheromones is poorly understood. Females of the funnel-web spider, Allagelena difficilis, employ sit-and-wait tactics for mating. Field observations indicate that males leave their retreats to search for potential mates during the breeding season. Therefore, we investigated whether virgin females release a sex attractant to conspecific males and then explored the chemical nature of the female pheromone. Four fatty acids extracted from the female bodies (palmitic acid, linoleic acid, cis-vaccenic acid and stearic acid) constitute a multiple-component sex attractant to conspecific males in A. difficilis. Unexpectedly, mated females also produce the same fatty acids, but at trace levels. Two-choice experiments showed that males were significantly attracted by the blend of the four fatty acids in appropriate concentrations while avoiding the blend consisting of the same acids at very low concentrations, suggesting that mate-searching males are able to discriminate virgin females from mated females by the quantities of female-specific fatty acids in the funnel-web spider A. difficilis.     

Evidence for a sex pheromone in the hide beetle,Dermestes maculatus (De Geer) (Coleoptera: Dermestidae)

Adults of the hide beetle,Dermestes maculatus (De Geer), were shown to have a female-produced sex pheromone which excited males. Male response was positively correlated with increasing age, but females produced a higher level of pheromone at 6–8 days of age than at younger or older ages. Extractable female pheromone and male response varied over the photophase with peak values that occurred during the latter hours. Quantitative bioassay indicated that the 50% male response level (RD₅₀) would be elicited by a pheromone exposure of 0.01 female equivalents (FE). Females extracted 24 hr after mating had a lower level of extractable pheromone than did virgin females of the same age.Mention of a proprietary product does not constitute an endorsement by the Minnesota Agricultural Experiment Station.     

Identification of a Sex Pheromone From Male Yellow MealWorm Beetles, <Emphasis Type="Italic">Tenebrio molitor</Emphasis>

The sex pheromone released by the adult female Tenebrio molitor, 4-methyl-1-nonanol, is well known. In addition, there is evidence that adult males release a pheromone that attracts females. The purpose of the present study was to isolate and identify male-released pheromone(s). Emissions from virgin adult males and females were collected on filter paper and extracted with pentane. Extracts were analyzed by gas chromatography-mass spectrometry. One male-specific compound was detected and identified as (Z)-3-dodecenyl acetate (Z3-12:Ac). In arena bioassays, E3-12:Ac was attractive to females only, at 1 and 10 μg doses. E3-12:Ac was also attractive to females at a 10-μg dose. The presence of both male and female pheromones, each attracting the opposite sex, may contribute to maintaining a high-density population of both sexes.     

Sexual behavior ofMatsucoccus josephi (Homoptera: Margarodidae)

The daily emergence patterns ofMatsucoccus josephi adults and third-instar male larvae raised on artificially infested saplings ofPinus halepensis were determined. A single peak of emergence was found for adult males between 0300 and 0500 hr. Adult females emerged throughout the day, with maximum emergence between 0500 and 0700 hr. Two daily emergence periods were observed in third-instar male larvae, one between 0300 and 0900 hr and the other between 1700 and 2100 hr. Airborne pheromone emitted by adult virgin females was collected using a flow system. No significant differences were recorded in the attraction of the male to crude pheromone collected at different times of day. A single female, virgin or half an hour after mating, was sufficient to attract the males in an olfactometer system. From minor differences in pheromone release throughout the day, and from the lesser degree of attraction by females half an hour after mating, it is assumed that there is no mechanism controlling the daily release of the female sex pheromone. Reduction of pheromone emission after mating is suggested.Contribution from the Agricultural Research Organization, The Volcani Center, Bet Dagan, Israel. No. 2743-E, 1989 series.     

Identification of the sex pheromone of the guernsey carpet beetle,Anthrenus sarnicus Mroczkowski (Coleoptera: Dermestidae)

It has been confirmed that adult virgin females ofAnthrenus sarnicus Mroczkowski exhibit a characteristic headstand posture that is associated with the release of a sex pheromone. Volatiles trapped on filter papers suspended above calling females were attractive to adult virgin males when tested in a two-choice target bioassay. Separate aeration extracts of males and females were analyzed by gas chromatography-mass spectrometry and showed that decanol and decyln-butyrate were released by females only. These components were present in approximately equal amounts and accounted for about 90% of the total area of the chromatogram. Decyl butyrate produced an electroantennogram response with a larger response from males than females. Behaviorally, a mixture of 10g of decanol and 10g of decyl butyrate attracted 88% of males and 10g of decyl butyrate alone attracted 82% of males in the bioassay. The role of decyl butyrate as a sex pheromone is convincing, but this is not the case for decanol.     

Vicia faba–Lygus rugulipennis Interactions: Induced Plant Volatiles and Sex Pheromone Enhancement

The profiles of volatile chemicals emitted by Vicia faba plants damaged by Lygus rugulipennis feeding, and by feeding plus oviposition, were shown to be quantitatively different from those released by undamaged plants. Samples of volatile chemicals collected from healthy plants, plants damaged by males as a consequence of feeding, plants damaged by females as a consequence of feeding and oviposition, plants damaged by feeding with mated males still present, and plants damaged by feeding and oviposition with gravid females still present, showed significant differences in the emission of hexyl acetate, (Z)-β-ocimene, (E)-β-ocimene, (E)-β-caryophyllene, and methyl salicylate. In particular, treatments with mated females present on plants had a significant increase in emission levels of the above compounds, possibly due to eggs laid within plant tissues or active feeding, compared with undamaged plants and plants damaged by males feeding, with or without insects still present. Furthermore, the pheromonal blend released by mated L. rugulipennis females, mainly comprising hexyl butyrate, (E)-2-hexenyl butyrate, and (E)-4-oxo-2-hexenal, was enhanced when females were active on broad bean plants, whereas such an increase was not observed in males. Both sexes gave electroantennogram responses to green leaf volatiles from undamaged plants and to methyl salicylate and (E)-β-caryophyllene emitted by Lygus-damaged plants, suggesting that these compounds may be involved in colonization of host plants by L. rugulipennis. In addition, mated males and females were responsive to hexyl butyrate, (E)-2-hexenyl butyrate, and (E)-4-oxo-2-hexenal released by mated females on V. faba, indicating that these substances could have a dual function as a possible aggregation pheromone in female–female communication, and as a sex pheromone in female–male communication. An erratum to this article can be found at     

(S)-2-pentadecyl acetate and 2-pentadecanone Components of aggregation pheromone ofDrosophila busckii

(S)-2-Pentadecyl acetate and 2-pentadecanone were identified as the major aggregation pheromone components, inDrosophila busckii. Both sexes of flies were attracted equally in a wind-tunnel olfactometer. The flies also responded to racemic 2-pentadecyl acetate but not to the pureR enantiomer. In bioassay, (S)-2-pentadecyl acetate and 2-pentadecanone were each active alone, and a mixture of both increased the number of flies responding ca. twofold. The aggregation pheromone components are found in the ejaculatory bulb of sexually mature males and are transferred primarily to the female cuticle during mating. One third of the pheromone transferred is released by the female to the surrounding environment in a few hours after mating. None of the aggregation pheromone components remained on the mated female's cuticle, leaving two thirds unaccounted for. The same results were obtained when racemic 2-pentadecyl acetate was topically applied to immature and mature virgin males and females. BothD. mulleri andD. busckii were attracted to (S)-2-acetates of 13, 14 and 15 carbons, butD. mulleri preferred (S)-2-tridecyl acetate andD. busckii preferred (S)-2-pentadecyl acetate.     

Pheromone-based Mating and Aggregation in the Sorghum Chafer, <Emphasis Type="Italic">Pachnoda interrupta</Emphasis>

Adults of the sorghum chafer, Pachnoda interrupta Olivier (Coleoptera: Scarabaeidae: Cetoniinae), form aggregations during the mating period in July, but also in October. The beetles aggregate on food sources, e.g., Acacia spp. trees or sorghum with ripe seeds, to feed and mate. During the mating season, field trapping experiments with live beetles as bait demonstrated attraction of males to unmated females, but not to mated females or males, indicating the presence of a female-emitted sex pheromone. Unmated females combined with banana (food source) attracted significantly more males and females than did unmated females alone. Other combinations of beetles with banana were not more attractive than banana alone. Thus, aggregation behavior appears to be guided by a combination of pheromone and host volatiles. Females and males were extracted with hexane during the mating period, and the extracts were compared by using GC-MS. In a field trapping experiment, 19 compounds found only in females were tested, both singly and in a mixture. Traps baited with one of the female-associated compounds, phenylacetaldehyde, caught significantly more beetles than any other treatment. However, the sex ratio of beetles caught in these traps did not differ from that of control traps, and it is possible that other components may be involved in the sex pheromone signal. Furthermore, traps baited with a mixture of all 19 compounds attracted significantly fewer beetles than did phenylacetaldehyde alone.     

Attraction of Male European Tarnished Plant Bug, <Emphasis Type="Italic">Lygus rugulipennis</Emphasis> to Components of the Female Sex Pheromone in the Field

Previous work showed that females of the European tarnished plant bug, Lygus rugulipennis Poppius (Heteroptera: Miridae), produced three chemicals, hexyl butyrate, (E)-2-hexenyl butyrate, and (E)-4-oxo-2-hexenal, and that these were suspected to be components of the female sex pheromone. In field experiments, traps baited with blends of these chemicals dispensed from polyethylene vials and sachets failed to catch significant numbers of males. Here, we report more recent field experiments in which the chemicals were released from glass microcapillary tubes. A blend of hexyl butyrate and (E)-4-oxo-2-hexenal was significantly attractive to male L. rugulipennis. In addition, whereas the mixture of all three components attracted fewer L. rugulipennis males, this tertiary blend captured significantly greater numbers of males of the congeneric species Lygus pratensis than the binary mixture. The possible reasons for the success of the microcapillaries compared with other dispensers are discussed.     

Evidence for a sex pheromone in bark beetle parasitoid Roptrocerus xylophagorum

Male Roptrocerus xylophagorum (Ratzeburg) (Hymenoptera: Pteromalidae) exhibited courtship and mating behaviors including wing fanning, antennation, mounting, and copulation attempts when exposed to glass bulb decoys coated with a whole-body extract of females in hexane, acetone, or methanol. Activity of extract-treated decoys declined gradually over one week. Males responded much less strongly to freeze-killed female cadavers extracted with solvents than to unextracted cadavers; treatment of extracted cadavers with female extract restored male responses. The pheromone was found to be equally present over the surface of both the abdomen and head/thorax of females, and the origin of the pheromone could not be conclusively localized to any single body region. The activity of pheromone on females increased between day 1 and days 3–5 following eclosion; otherwise, pheromone activity was not significantly affected by either female age or mating. Males were arrested within the zone of a glass surface on which females had walked, suggesting that the pheromone might be substrate-borne. Recent exposure to females reduced male responsiveness, but responsiveness was fully restored after a few hours of male isolation from females. When hexane extracts of whole females were fractionated on silica gel, the pheromone's activity was largely recovered with the first, most nonpolar fraction. Female extracts and fractions were analyzed by coupled gas chromatography–mass spectrometry. Cuticular hydrocarbon alkanes were identified as the extract components whose concentrations correlated best with male responses. Evidence of the pheromone's long persistence, low volatility, low polarity, and presence over the insect's entire body surface further supported the hypothesis that the pheromone was composed of one or more cuticular hydrocarbons.     

Homofarnesals: Female Sex Attractant Pheromone Components of the Southern Cowpea Weevil, <Emphasis Type="Italic">Callosobruchus chinensis</Emphasis>

The southern cowpea weevil, Callosobruchus chinensis (Coleoptera: Bruchidae), is a major pest of stored legumes in warm temperate and tropical climates. The female sex attractant pheromone was extracted from filter-paper shelters taken from containers that housed virgin females. The extracts were purified by various chromatographic techniques, and the biologically active components in the fractions were screened by gas chromatographic–electroantennographic detection analysis with male antennae. Two compounds that elicited electrophysiological responses were isolated, and gas chromatography–mass spectrometry, nuclear magnetic resonance, and micro-chemical analyses suggested that the active compounds were homofarnesals, (2Z,6E)- and (2E,6E)-7-ethyl-3,11-dimethyl-2,6,10-dodecatrienals. Males of C. chinensis were significantly attracted to filter paper discs loaded with the synthetic compounds at 0.01–0.1 ng compared to solvent control in a Y-tube olfactometer assay. These pheromone components represent unique chemical structures within the genus Callosobruchus.     

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.     

Role of sex pheromone components in behavioral reproductive isolation betweenAutographa gamma (L.) and eitherTrichoplusia ni (Hübner) ORChrysodeixis chalcites (Esp.) (Lepidoptera: Noctuidae: Plusiinae)

The composition of theAutographa gamma sex pheromone was reexamined and only (Z)-7-dodecenyl acetate and (Z)-7-dodecenol were identified by capillary GC, GC-MS, and dimethyl disulfide derivatization and subsequent GC-MS analysis. The fatty acid content of the pheromone glands was also studied, and a series of saturated and unsaturated acids was identified. However, most of the related pheromonal compounds were not detected. The male response to the pheromone components was studied in a flight tunnel and compared with the response to calling females. The best synthetic baits evoked a response similar to that observed to the virgin females, but males spent significantly more time at calling females than at the synthetic baits. The preferred synthetic baits consisted of (Z)-7-dodecenyl acetate alone or of a blend with 5% (Z)-7-dodecenol. Increasing the relative amount of the alcohol caused a gradual reduction in male response, particularly in the last steps of the courtship sequence. The addition of the minor sex pheromone components of the sympatric Plusiinae species,Trichoplusia ni andChrysodeixis chalcites, to theA. gamma pheromone was also investigated in the flight tunnel. Some of these components exhibited a significantly antagonistic effect on theA. gamma male courtship behavior. The most potent antagonists were (Z)-5-dodecenyl acetate and (Z)-9-tetradecenyl acetate. The response ofA. gamma andT. ni males to conspecific and heterospecific females was also compared in the flight tunnel. WhereasA. gamma males were attracted only to their conspecific females, a small percentage ofT. ni males were also attracted toA. gamma females and 11% performed the whole courtship sequence.Contribution from the Agricultural Research Organization (ARO), No. 3459-E, 1992 series.     

Sex-Specific Trail Pheromone Mediates Complex Mate Finding Behavior in Anoplophora glabripennis

Anoplophora glabripennis (Motsch.) is a polyphagous member of the Cerambycidae, and is considered, worldwide, to be one of the most serious quarantine pests of deciduous trees. We isolated four chemicals from the trail of A. glabripennis virgin and mated females that were not present in trails of mature males. These compounds were identified as 2-methyldocosane and (Z)-9-tricosene (major components), as well as (Z)-9-pentacosene and (Z)-7-pentacosene (minor components); every trail wash sample contained all four chemical components, although the amounts and ratios changed with age of the female. Males responded to the full pheromone blend, regardless of mating status, but virgin females chose the control over the pheromone, suggesting that they may use it as a spacing pheromone to avoid intraspecific competition and maximize resources. Virgin, but not mated, males also chose the major pheromone components in the absence of the minor components, over the control. Taken together, these results indicate that all four chemicals are components of the trail pheromone. The timing of production of the ratios of the pheromone blend components that produced positive responses from males coincided with the timing of sexual maturation of the female.