Identification of a Sex Pheromone Component for the Blueberry Leafminer, Caloptilia porphyretica

Coupled gas chromatographic-electroantennographic detection (GC-EAD) of both gland extracts and effluvial collections from female blueberry leafminer, Caloptilia porphyretica Braun (Lepidoptera: Gracillariidae), showed that females produced a single EAD-active compound. The amount of the compound collected from virgin female C. porphyretica was below GC and mass spectrometry (MS) detection thresholds, even with highly concentrated gland extracts (approximately 150 female equivalent). (E)-11-Hexadecenal (E11-16:Ald) was determined to be a sex pheromone component mainly by comparison of retention times with authentic standards on both polar and nonpolar capillary columns, microreaction-GC-EAD analyses, and field trapping tests. GC-EAD experiments showed that synthetic E11-16:Ald exhibited extraordinarily high electrophysiological activity, stimulating significant male antennal responses at as low as 10 fg. Traps baited with E11-16:Ald alone were attractive to males. Addition of 1 or 3% of its geometric isomer, Z11-16:Ald, to E11-16:Ald did not significantly increase trap captures, but an inhibitory effect was observed at the 10% level. The influence of two kinds of rubber septa on attraction was also evaluated. Male moth captures were higher in traps baited with red rubber septa than with gray rubber septa at 30-300-microg doses. Monitoring of adult flight activity with 3-microg doses of E11-16:Ald indicated at least three distinct flight periods throughout the 2003 season.     

A Versatile Method for On-Line Analysis of Volatile Compounds from Living Samples

A system especially designed for the on-line analysis of volatiles emitted by living aerobic organisms is described. The instrument consists of a modified thermal desorber and utilizes the purge and cold trap (PCT) technique coupled with gas chromatography–mass spectrometry. The system enables the use of a variety of sample vessels especially adapted to individual test organisms. Quantitative analyses of some volatile semiochemicals revealed that when only nanogram amounts of these compounds are evaporated at room temperature for a period of 10 min, they may be trapped and detected by this device. Emission of volatiles from two insect species and a plant species have been studied in detail by using this system.     

Anal sac secretion of the red fox,Vulpes vulpes; volatile fatty acids and diamines: Implications for a fermentation hypothesis of chemical recognition

Putrescine (1,4-diaminobutane) and cadaverine (1,5-di-aminopentane) were identified in the anal sac secretions of the red fox,Vulpes vulpes, and of the lion, Panthera leo. Anal sac secretion samples obtained over a period of 10 weeks by sampling from within each sac of each of 6 captive foxes were analyzed and putrescine, cadaverine, and volatile fatty acid compositions and secretion pH values recorded. A significant (P < 0.001) negative correlation of pH (range 6.5–9.4) with total volatile fatty acid concentration was observed. Secretion compositions are discussed in the context of a fermentation hypothesis of chemical recognition. Secretion samples could not be unambiguously assigned to particular foxes on the basis of simple comparisons of volatile fatty acid profiles alone. Composition differences were noted between secretions obtained at a given time from corresponding right and left sacs.     

Host recognition and the study of a chemical basis for attraction by cuckoo bumble bees (Hymenoptera: Apidae)

Species ofPsithyrus (Hymenoptera; Apidae) are obligate bumble bee social parasites. In this study, females ofP. vestalis andP. ashtoni were presented with pentane extracts prepared from different body parts of queens of their respective host species,Bombus terrestris andB. terricola. Parasites of both species were capable of distinguishing host bees from other bumble bee species using chemical cues contained within extracts. Among extracts of several body parts presented to parasites, the abdomen produced the greatest behavioral response, with Dufour's gland and terminal tergal segments eliciting the greatest response among abdominal regions. Extracts of these two body parts obtained fromB. terrestris queens shared a number of compounds, identified by GC-MS. Among the identified compounds are a number that have been reported to be of importance in bee sociochemistry.     

Identification and Function of Prothoracic Exocrine Gland Steroids of the Dytiscid Beetles Graphoderus cinereus and Laccophilus minutus

The chemical composition of the prothoracic defensive secretion of the water beetles Graphoderus cinereus and Laccophilus minutus was examined by gas chromatographic–mass spectrometric analysis of trimethylsilylated gland extracts. The main components are 3,11-dihydroxy-5-pregnan-20-one in G. cinereus and 3,12-dihydroxy-5-pregnan20-one in L. minutus. Fully saturated pregnanes have not been found in the prothoracic defensive glands of dytiscids before. Based on tests with synthetic compounds, their biological role was examined in a feeding assay with the minnow Phoxinus phoxinus. Agar pellets containing either steroid strongly deterred feeding.     

Differential metabolism of 1,8-cineole in insects

In order to compare the metabolism of 1,8-cineole in the pyrgo beetle, Paropsisterna tigrina, three other herbivorous insect species, Faex nigroconspersa, Chrysophtharta bimaculata, and Oxyops vitiosa, were fed 1,8-cineole leaf diets. F. nigroconspersa adults excreted predominantly 9-hydroxy-1,8-cineole (36.2% of the volatile constituents) with some 2-hydroxy-1,8-cineole (11.4%). In contrast, larvae excreted predominantly 2-hydroxy-1,8-cineole (27.4%) and smaller proportions of 9-hydroxy-1,8-cineole (5.2%) and 3-hydroxy-1,8-cineole (4.3%). C. bimaculata adults excreted predominantly 3-hydroxy-1,8-cineole (16.5%). Oxyops vitiosa adults, on a lower 1,8-cineole diet, excreted predominantly 2,9-dihydroxy-1,8-cineole (4.2%) and 2-hydroxy-1,8-cineole (3.5%), with smaller proportions of 3-hydroxy-1,8-cineole (1.1%) and 9-hydroxy-1,8-cineole (0.5%). This is the first reported occurrence of a dihydroxycineole as an insect metabolite. Gas chromatographic and mass spectral data for hydroxycineoles are recorded and interspecific metabolite variation discussed.     

Comparative Investigation of the Volatile Urinary Profiles in Different <Emphasis Type="Italic">Phodopus</Emphasis> Hamster Species

Stir bar sorptive extraction method was used for investigation of the urinary volatile profiles in male and female Phodopus campbelli and Phodopus sungorus hamsters. Additionally, female Phodopus roborowsky urinary profiles were characterized. A quantitative analytical approach allowed comparisons of 17 selected compounds in urine. Results showed that campbelli and sungorus species show similar urinary volatile profiles for males and females. Differences appeared only in concentrations. Several unique compounds, such as pyrazine derivatives, were found to be gender- and age-specific. P. roborowsky females exhibited a completely different urinary volatile profile from campbelli and sungorus females, featuring a unique set of substituted quinoxalines.     

The role of olfaction in chemosensory-based predator recognition in the fathead minnow,Pimephales promelas

Solitary fathead minnows (Pimephales promelas) were rendered anosmic and exposed to chemical stimuli from a predatory northern pike (Esox lucius) to determine the role of olfaction in the minnow's ability to recognize predators on the basis of chemical stimuli. Anosmic fish did not respond to the pike stimuli with a typical fright reaction, while control fish, with intact olfactory receptors, did. These results demonstrate that the olfactory system is necessary for the ability of fathead minnows to recognize northern pike as a predator and that the gustatory and single-celled chemosensory systems are not sufficient for this recognition in the absence of olfactory input. Olfactory impairment was behaviorally confirmed by exposing minnows to alarm substance (Schreckstoff).     

Cuticular hydrocarbons of the paper wasp,Polistes fuscatus: A search for recognition pheromones

The cuticular chemicals of 124 individual wasps (foundresses and workers) from 23 colonies ofPolistes fuscatus were analyzed. The compounds identified, all of which were hydrocarbons, were similar to those of other vespid wasps in that the bulk of the hydrocarbons were 23–33 carbons in chain length. However, the hydrocarbon profile ofP. fuscatus differed from those of its congeners in its proportions of straight-chain alkanes, methylalkanes, and alkenes. Three of the 20 identified hydrocarbons, 13- and 15-MeC₃₁, 11,15- and 13,17-diMeC₃₁, and 13-, 15-, and 17-MeC₃₃, had properties postulated for recognition pheromones: colony specificity, efficacy in assigning wasps to the appropriate colony, heritability, lack of differences between foundresses and workers, and distinctive stereochemistry.     

Electrophysiologically and Behaviorally Active Volatiles of Buffalo Gourd Root Powder for Corn Rootworm Beetles

The dried, powdered roots of buffalo gourd, Cucurbita foetidissima, were tested in a cornfield and shown to attract adult northern and southern corn rootworm beetles. Coupled gas chromatography–electroantennography (GC-EAG) analyses of headspace samples of the root powder showed several GC-EAG-active compounds on the antennae of female northern, southern, and western corn rootworms. Among other techniques, solid-phase microextraction and GC-mass spectrometry identified the following GC-EAG-active compounds: hexanol, nonanal, 1-octen-3-ol, benzaldehyde, benzyl alcohol, (E)-3-octen-2-one, (E,E)-3,5-octadien-2-one, and (E,Z)-3,5-octadien-2-one. EAG dose–response studies of several of the identified root powder volatiles also were performed and compared with results from known attractants. Field tests of synthetic root powder volatiles in commercial cornfields showed that northern corn rootworm adults were attracted to (E,E)-3,5-octadien-2-one. The antennae of the Diabrotica species and the field tests showed specificity for different geometrical isomers of 3,5-octadien-2-one, with a behavioral preference for (E,E)-3,5-octadien-2-one. In addition, we have shown that the efficacy of buffalo gourd root powder as a feeding stimulant and arrestant can be enhanced for northern and western corn rootworm adults by augmenting buffalo gourd root powder with additional (E,E)-3,5-octadien-2-one.     

Soldier frontal glands of the termiteMacrotermes subhyalinus: Morphology,chemical composition,and use in defense

Soldiers of the East African fungus-growing termiteMacrotermes subhyalinus (Rambur) (Termitidae: Macrotermitinae) employ both mechanical and chemical defenses. Soldiers release a chemical secretion composed of long-chain saturated and monounsaturated hydrocarbons into wounds inflicted by their powerful mandibles. Chemical analysis of the secretion shows the paraffin fraction to consist primarily ofn-tricosane,n-pentacosane, 3- and 5-methylpentacosane, and 5-methylheptacosane. The major oleflns were identified as (Z)-9-heptacosene and (Z)-9-nonacosene. The secretion originates from the frontal glands of both major and minor soldiers; however, the hypertrophied gland of the major soldiers contains 500-fold more secretion than that of the minor soldiers. This secretion appears to impair the healing of wounds in test ants, and thus could represent a valuable supplement to the mechanical defense mechanism.     

Substrate specificity of acetyltransferase and reductase enzyme systems used in pheromone biosynthesis by Asian corn borer,Ostrinia furnacalis

The substrate specificity of the acetyltransferase and the reductase enzyme systems used byOstrinia furnacalis (Lepidoptera: Pyralidae) in pheromone biosynthesis was studied in vivo by topical application of precursors to pheromone glands. Each of the tetradecenols, varying in double bond position (from 7 to 13) and geometry of the double bond, was converted to the corresponding acetate by the acetyltransferase. The similarity in the conversion rates of all tested fatty alcohols indicated that the acetyltransferase has a low substrate specificity. Most of the corresponding tetradecenoic acids could also be converted to the respective acetates. However, very different conversion rates among the tested fatty acids demonstrated that the reductase system has a higher substrate specificity than the acetyltransferase. The conversion rates of mostE isomers were higher than those of the correspondingZ isomers, except for the ()-11-tetradecenoic acids, in which much moreZ isomer was converted to the product. Saturated tetradecanoic acid was converted to the corresponding acetate at a high rate; the shorter homolog, tridecanoic acid, was converted at a lower rate (56%), and conversion to the respective acetates of the longer homolog, pentadecanoic and hexadecanoic acids, was insignificant (<5%). The results from the present study showed that specificity of pheromone production is to a large extent controlled by the pheromone gland reductase system.Guenée, Lepidoptera: Pyralidae     

Sex Pheromone of the Cranberry Blossom Worm, Epiglaea apiata

The cranberry blossom worm, Epiglaea apiata (Grote) (Lepidoptera: Noctuidae), is a major pest of cranberries in New Jersey. The female sexpheromone of this moth was identified as a blend of (Z)-9-hexadecenyl acetate (Z9-16:Ac), (Z)-9-tetradecenyl acetate (Z9-14:Ac), and tetradecyl acetate (14:Ac) by gas chromatographic–electroantennographic detection and gas chromatography–mass spectrometry. The ratio of the components in extracts of the female pheromone gland was determined to be 65 : 2 : 33 of the Z9-16:Ac, Z9-14:Ac, and 14:Ac, respectively. The double bond positions of the pheromone components were confirmed by dimethyl disulfide derivatization. In addition to the above three components, a mixture of C₄–C₁₀ aliphatic acids was present in both gland extracts and effluvia collections, and the acids elicited significant EAD responses from male moth antennae. However, addition of the C₄–C₁₀ aliphatic acids to the pheromone blend did not significantly increase trap captures. Three-hundred- and 1000-g doses of a synthetic blend containing Z9-16:Ac, Z9-14:Ac, and 14:Ac (65 : 2 : 33), on a rubber septum were more attractive to males than lower doses.     

Investigation of Co-Based DeNOx Catalysts for the Treatment of Natural Gas Lean-Burn Engine Exhaust

The catalytic reduction of nitric oxide with methane in lean conditions has been investigated on cobalt-based powder and monolith catalysts such as Co-exchanged ferrierite, Co-exchanged beta zeolite and Co–ZrO₂. From catalyst laboratory testing the following activity scale was obtained: Co-ferrierite > Co-beta >>> Co–ZrO₂, suggesting a significant role of the support catalytic matrix. In the tests at the engine exhaust, the best results were obtained with Co-ferrierite, after water separation from the exhaust. However, in the presence of the high concentration of water, typical of the engine exhaust, the performances were dramatically reduced.     

Pheromone Components of the Wheat Stem Sawfly: Identification, Electrophysiology, and Field Bioassay

Volatiles collections and cuticular extracts of the wheat stem sawfly, Cephus cinctus Norton (Hymenoptera: Cephidae), were analyzed by coupled gas chromatographic–electroantennographic detection (GC-EAD) in order to detect possible pheromone components of this species. Volatiles collections from male and female sawflies contained the same 13 GC-EAD-active compounds that stimulated both male and female antennae. GC-EAD-active compounds were identified by mass spectrometry, and the identities were verified by acquiring or synthesizing authentic standards with identical GC retention times and mass spectra. 9-Acetyloxynonanal gave the strongest EAD response. Additional GC-EAD-active compounds included 13-acetyloxytridecanal, aldehydes with 9–16 carbon chain lengths, acids with 8–10 carbon chain lengths, and phenylacetic acid. The last was instantly detectable by human nose when several males were allowed to interact, but not from isolated insects. On a per-insect basis, grouped males overall released higher amounts of the GC-EAD-active compounds than grouped females. Notable quantitative differences between the sexes were relatively higher amounts of 9-acetyloxynonanal, phenylacetic acid, and tetradecanal from males and hexadecanal from females. In the absence of an absolute, qualitative difference between sexes, these relative differences might be important for sex recognition. Field bioassay demonstrated the attractiveness of 9-acetyloxynonanal for male and female C. cinctus. The major compounds of male and female cuticular extracts showed no GC-EAD activity, but ozonolysis of extracts gave dramatically increased amounts of GC-EAD-active material. Hence, double-bond-containing cuticular compounds are suggested as precursors of the aldehydic pheromone components.     

Identification of a New Blend of Apple Volatiles Attractive to the Apple Maggot, Rhagoletis pomonella

Solid-phase microextraction (SPME) and gas chromatography coupled with electroantennographic detection (GC-EAD) were used to identify a new blend of volatiles from apples as the key attractants for the apple maggot fly, Rhagoletis pomonella (Walsh). The new five-component blend contains butyl butanoate (10%), propyl hexanoate (4%), butyl hexanoate (37%), hexyl butanoate (44%), and pentyl hexanoate (5%) compared with a previously reported seven-component mix of hexyl acetate (35%), (E)-2-hexen-1-yl acetate (2%), butyl 2-methylbutanoate (8%), propyl hexanoate (12%), hexyl propanoate (5%), butyl hexanoate (28%), and hexyl butanoate (10%). Volatiles from five different varieties of apple elicited reproducible and high EAD responses from R. pomonella antennae to the same five chemicals. In flight-tunnel choice tests involving red sticky spheres with odor sources, the new five-component blend of apple volatiles showed significantly more activity than the previous seven-component blend or the single compound, butyl hexanoate. In a field trial captures with the new five-component blend were better than with butyl hexanoate, which is currently used with commercial apple maggot monitoring spheres.     

Toward the chemical ecology of medicinal plant use in chimpanzees: The case ofVernonia amygdalina,a plant used by wild chimpanzees possibly for parasite-related diseases

The bitter and related constituents have been isolated fromVernonia amygdalina (Compositae), a plant ingested by wild chimpanzees possibly suffering from parasite-related diseases in the Mahale Mountains National Park, Tanzania. Isolated from the plant were four known sesquiterpene lactones, seven new steroid glucosides, and two aglycones of the glucosides. The sesquiterpene lactones showed significant in vitro antischistosomal, plasmodicidal, and leishmanicidal activities. Antischistosomal activity was also found for the major steroid glucoside, vernonioside B₁. A trend in the glucosides to show significant antischistosomal, plasmodicidal, and amebicidal activities when the sugar moiety was removed, was observed. Vernodalin, judged as the most significant constituent for antiparasitic activities in vitro, was tested for in vivo antischistosomal effect. It was, however, highly toxic to the cercaria-infected mouse. Chimpanzees have been only rarely observed to ingest anything but the pith of the young stem. The occurrence of vernonioside B₁ and its aglycone vernoniol B₁, the major constituents among the steroid-related constituents, were detected at significant levels in the pith. However, vernodalin was abundant only in the leaves and bark. Thus, chimpanzees at Mahale were hypothesized to control parasite-related diseases by ingesting the young pith of this tree containing steroid-related constituents.     

Structure-activity relationships of pyrrolizidine alkaloids in insect chemical defense against the orb-weaving spider Nephila clavipes

Pyrrolizidine alkaloids (PAs) are known to protect Arctiidae moths and Danainae and Ithomiinae butterflies against the orb-weaving spider Nephila clavipes (Araneae, Araneidae, Tetragnathinae), which liberates adults of these insects unharmed from its web. We tested against this spider the role of eight PAs and one derived structure [an 89:11 mixture of the 12-membered macrocyclic diester free base integerrimine and senecionine and the respective N-oxide; two hydrolysis products from this mixture (the necine base retronecine, its respective N-oxide, and a mixture of integerriminic and senecionic necic acids); the 12-membered macrocyclic senkirkine; the 9-O-monoester free base senecioylretronecine and its respective N-oxide; and the 9-O-monoester free base callimorphine (a PA biosynthesized only by insects from retronecine)]. The mixture integerrimine–senecionine N-oxide seems to be more active than the respective free base [LibD₅₀ (liberation dose 50) = 0.042 and 0.153 g/dry weight of prey, respectively], but the difference in activity between the N-oxide and free base of the 9-O-monoester senecioylretronecine was slight (LibD₅₀ = 0.167 and 0.104, respectively). Senkirkine, an otonecine base PA that does not form N-oxide and is not found in insects, was the less active, showing the highest LibD₅₀ (0.354). The difference in antipredator activity between N-oxides and free bases from macrocyclic diesters and monoesters may be correlated with physicochemical properties of these molecules in interaction with the Nephila receptors. For the active structures, there was a significant correlation between dosage and antipredator activity. Both forms of retronecine and a necic acid mixture were inactive, supporting the hypothesis that PAs biosynthesized by insects from retronecine were originally produced and stored in order to optimize chemical defense. Comparison of dose/activity data with reported amounts of PAs in butterflies suggested that, in general, PA-specialist insects are protected against predation by Nephila.     

Multiple acceptors for pheromonal enantiomers on single olfactory cells in the Douglas-fir beetle,Dendroctonus pseudotsugae Hopk. (Coleoptera: Scolytidae)

Olfactory perception of pheromonal enantiomers by male and female Douglas-fir beetles,Dendroctonus pseudotsugae Hopk. (Coleoptera: Scolytidae), was investigated by electrophysiological techniques and behavioral bioassays. Electroantennograms (EAGs) and single-cell responses indicated both sexes to be more responsive to racemic frontalin and the (–)-enantiomer at lower dosages. At higher dosages, little difference was noted in responses to either enantiomer. However, response to the racemic mixture at higher dosages was slightly greater than responses to either enantiomer alone. In laboratory behavioral bioassays, responses to low concentrations of (–)-frontalin and the racemic mixture exceeded response to the (+)-enantiomer alone. At a higher concentration, responses to the racemic mixture or either enantiomer alone did not differ. The results indicate that separate enantiomer-specific acceptors may exist on the same pheromone receptor cell.Texas Agricultural Experiment Station paper No. TA-19841. This research was supported in part by Mclntire-Stennis Project 1525, the U.S. Department of Agriculture-sponsored program entitled The Expanded Southern Pine Beetle Research and Applications Program through TAES-SEA-CR grant 904-15-4, and NSF grant PCN 79-21708 to Oregon State University. The findings, opinions and recommendations expressed herein are those of the authors and not necessarily those of the U.S. Department of Agriculture. All programs and information of the Texas Agricultural Experiment Station are available without regard to race, ethnic origin, religion, sex, or age.Mention of a trademark, proprietary product or vendor does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products or vendors that may also be suitable.     

Differential recognition of geometric isomers by the boll weevil,Anthonomus grandis Boh. (Coleoptera: Curculionidae): Evidence for only three essential components in aggregation pheromone

For two decades, the aggregation pheromone of the boll weevil,Anthonomus grandis Boh. (Coleoptera: Curculionidae), was thought to consist of four compounds: I [(+)-(Z)-2-isopropenyl-1-methylcyclobutane ethanol]; II [(Z)-3,3-dimethyl-^I,-cyclohexane ethanol]; III [(Z)-3,3-dimethyl-^1,-cyclohexane acetaldehyde); and IV [(E)-3,3-dimethyl-^1,-cyclohexane acetaldehyde). Evidence is presented from behavioral and electrophysiological studies to show that only three of these components, I, II, and IV, are essential for attraction. Competitive field tests, in which each possible three-component blend was tested against the four-component mixture, demonstrated that omission of I, II. or IV resulted in decreased trap captures (P < 0.01). Trap captures by these blends lacking I, II, or IV resembled those by the hexane solvent alone in a similar experiment. However, omission of III did not significantly alter field attractiveness of the blend. Dosage-response curves constructed from electroantennogram responses of both males and females to serial dilutions of III, IV, and a 5050 mixture of the geometric isomers III and IV showed both sexes to be 10- to 100-fold more sensitive to IV than III. Data from the electrophysiological studies were consistent with a single acceptor type for the (E)-cyclohexylidene aldehyde, IV, for males, and possibly one or two acceptor types for III and IV for females. Possible roles for the (Z)-cyclohexylidene aldehyde, III, and implications for the pheromonal attractant currently used in boll weevil eradication/suppression programs are discussed.