The Nasonov pheromone of the honeybeeApis mellifera L. (Hymenoptera,Apidae). Part II. Bioassay of the components using foragers

The Nasonov pheromone of the honeybee comprises seven components, (Z)-citral, nerol, geraniol, nerolic acid, geranic acid, and (E,E)-farnesol. Bioassay of individual components showed each attracted foraging bees. A mixture of components in proportions present in the honeybee was as attractive as the natural secretion, and each component contributed to the attractiveness of the mixture. Honeybees responded anemotactically to the source of Nasonov odor. The presence of footprint pheromone enhanced the attractiveness of the synthetic Nasonov mixture. Nasonov and footprint pheromones may prove useful in attracting honeybees to crops needing pollination.     

Attraction of reproductive honey bee swarms to artificial nests by Nasonov pheromone

A crossover experimental design was established to test the attractancy of Nasonov pheromone to reproductive swarms of honey bees. Nineteen swarms were attracted to artificial nest cavities containing a slow-release blend of the Nasonov components citral, geraniol, and nerolic + geranic acids, and only four swarms were attracted to pheromone-free artificial nests. The results indicate that Nasonov pheromone plays a key role in the attraction of honey bee swarms to nest cavities.     

Nasonov pheromone of the honeybee.Apis mellifera L. (Hymenoptera,Apidae)

Electroantennogram (EAG) responses from worker honeybee antennae were obtained for each Nasonov component. Response amplitudes to 10 g of components correlated well with reported relative abilities to attract foragers in the field. EAG responses of worker, queen, and drone antennae to natural pheromone were consistently greater than to synthetic pheromone, a difference only partly explained by enzymic conversion of geraniol to (E)-citral during preparation of natural extracts.     

Nasonov pheromone of the honey bee,Apis mellifera L. (Hymenoptera,Apidae). part III.

GC and GC-MS analyses of the multicomponent Nasonov pheromone of the honey bee, and of the air above insects releasing the pheromone, show that constant composition is maintained during release, despite differing volatilities of the components. The regulating mechanism may involve a specific enzyme process, detected in excised Nasonov glands, which converts the major component geraniol into the more volatile (E)-citral. Analysis of honey bees of known ages and at different times of year shows that maximum secretion occurs when foraging is most likely.     

Olfactory Responses of Neotropical Short-Tailed Singing Mice, <Emphasis Type="Italic">Scotinomys teguina</Emphasis>, to Odors of the Mid-Ventral Sebaceous Gland: Discrimination of Conspecifics,Gender, and Female Reproductive Condition

We examined the olfactory communication of the Neotropical short-tailed singing mouse, Scotinomys teguina, by investigating whether S. teguina responded to odors produced by the mid-ventral sebaceous gland of conspecifics. Females spent significantly more time investigating male odor than an odorless stimulus or a female odor. Males spent significantly more time investigating female odor than an odorless stimulus, but not that of a male odor. This latter result does not seem to be explained by differences in age or sexual experience of test subjects, but may be influenced by reproductive condition of the female odor donor. Male S. teguina spent significantly more time (1) investigating and (2) in total proximity to odors of estrous than non-estrous females. Males spent more time (1) investigating and (2) in total proximity to odors of males than non-estrous females. Finally, given the choice between odor of males vs proestrous females, males showed no preference. Thus, the mid-ventral gland in S. teguina seems to convey information about conspecifics, sex, and female reproductive condition. Male odor, compared with proestrous female odor, is equally interesting to males, suggesting that the gland also plays an important role in male–male communication. Sexual dimorphism in the size of the gland and in the amount of secretion produced by the gland may be related to either male–male competition or female choice.     

Behavioral responses to food volatiles by two species of stored-product coleoptera,Sitophilus oryzae (curculionidae) andTribolium castaneum (tenebrionidae)

Laboratory experiments were conducted to study the behavioral activity of grain-derived volatiles as attractants and pheromone synergists forSitophilus oryzae, an internal-feeding pest of sound grain, andTribolium castaneum, an external-feeding pest of damaged grains and flour. Behavioral studies with two-choice pitfall bioassays determined that the fresh grain volatiles valeraldehyde, maltol, and vanillin were attractive toS. oryzae at various doses, butT. castaneum were not attracted to any dose of any of these three compounds. When oils from pressed grains were bioassayed, sesame oil was significantly repellent and oat and wheat germ oils were attractive toS. oryzae. However, rice, soybean, oat, wheat germ, and corn oils were all attractive toT. castaneum. A commercial food product composed primarily of soybean oil and wheat germ was highly attractive toT. castaneum, but elicited no response fromS. oryzae. A combination of the three grain volatiles valeraldehyde, maltol, and vanillin with the synthetic pheromone sitophinone was more attractive toS. oryzae than either the pheromone alone or the tripartite grain volatile mix. Similarly, a combination of the commercial food product with the pheromone 4,8-dimethyldecanal was more attractive toT. castaneum than either food alone or pheromone alone. Behavioral responses to grain volatiles may reflect the ecological niche of the granivore:S. oryzae colonizes sound grain and is attracted to volatiles characteristic of fresh grain, whileT. castaneum utilizes damaged or deteriorated grains and responds best to oils characteristic of damaged or fungus-infested grain. Synergism of food odors and pheromones suggests that more effective traps can be devised for management of these pest insects.     

Pheromone-Induced Priming of a Defensive Response in Western Flower Thrips

The Western flower thrips Frankliniella occidentalis produces conspicuous anal droplets that function as a direct defense against various predators. These droplets also function in pheromonal communication in that they contain a mixture of decyl acetate and dodecyl acetate, which acts as an alarm. Exposure of thrips to synthetic pheromone is known to promote takeoff or refuge seeking, but the effect of the natural pheromone has not yet been studied. Here, we not only studied the response to natural pheromone, but also tested the new hypothesis that the alarm pheromone primes a defensive response in thrips. This test was carried out by measuring the reaction time to a simulated predator attack after exposure to synthetic or natural alarm pheromone (against a control with no pheromone at all). The reaction was quantified in terms of the time it takes a thrips larva to produce a droplet after attack. We found that thrips larvae produce droplets of alarm pheromone faster when cues associated with danger are present. There were no significant differences in reaction times of responses to synthetic pheromone, natural pheromone, or odors from a patch with a predator attacking a thrips larva. This implies that the synthetic pheromone mimics the natural pheromone, and that other cues emanating from the predator play a minor role. We conclude that the alarm pheromone increases the vigilance of the thrips, and this may promote its survival.An erratum to this article can be found at     

Nasonov pheromone of the honey bee,Apis mellifera L. (Hymenoptera: Apidae)

Composition of the Nasonov pheromone of the honey bee has been reexamined using new procedures, including analysis of pheromone from single live insects by capillary column GC-MS. Two new components have been identified, nerol and (E,E)-farnesol, and the presence of components proposed previously has been confirmed. Absolute amounts or relative proportions of components in the pheromonal secretion have been determined.     

Alert odor from skin gland in deer

Black-tailed deer (Odocoileus hemionus columbianus) discharge an odor from the metatarsal (MT) gland, located on the hindleg, when disturbed or alarmed. Freely moving, captive deer were exposed to the MT odor by means of a remote-release apparatus. Responses by males and females to MT odor from both sexes were recorded with a coding system and a video camera. When the odor was present, females became more alert and left the site more often than in the presence of control odors, odorless air, or deer urine. It is concluded that the MT secretion provides an alert signal, placing the odor into the class of alarm pheromones. There is no evidence that the closely related white-tailed deer,O. virginianus, possesses this alert (or alarm) odor system to the same degree.     

An investigation of human apocrine gland secretion for axillary odor precursors

Recently completed studies from our laboratories have demonstrated that the characteristic human male axillary odors consist of C₆ to C₁₁ normal, branched, and unsaturated aliphatic acids, with (E)-3-methyl-2-hexenoic acid being the most abundant. To investigate the mechanism by which the odor is formed, it is necessary to determine the nature of the odorless precursor(s) found in the apocrine secretion which is converted by the cutaneous microorganisms to the characteristic axillary odor. Pooled apocrine secretion was obtained from several male volunteers by intracutaneous injection of epinephrine. Partitioning this secretion into aqueous and organic soluble fractions was followed by hydrolysis of each fraction with NaOH or incubation with axillary microorganisms (cutaneous lipophilic corynebacterium). Analysis by gas chromatography/mass spectrometry (GC/MS) revealed the presence of (E)- and (Z)-3-methyl-2-hexenoic acid in the aqueous phase hydrolysate and aqueous phase incubated with bacteria; however, only a trace amount was seen in the resultant organic phase mixtures. These results suggest that a water-soluble precursor(s) is converted by the axillary flora to the characteristic axillary odors.     

Attraction of female mediterranean fruit flies to the five major components of male-produced pheromone in a laboratory flight tunnel

Attraction and pheromonal activity of five major identified components of the male-produced sex pheromone of the Mediterranean fruit flyCeratitis capitata to virgin laboratory-reared females was assessed in a laboratory flight tunnel. Dual-choice competitive assays were run to establish a baseline response of virgin females to live male pheromone, individual components, and an ensemble of all five compounds alone (air control) and competitively against one another. Approximately 50% of the females released in the tunnel were captured on leaf models emitting pheromonal odors from five live males. Over 37% of released females responded to an ensemble of five major identified components presented in individual capillaries. Response of females to individual components was less than 10%. Competitive assays showed the live male-produced pheromone to be more attractive than either the five major component ensemble (FMCE) or individual components. Further research is likely to identify other male-produced compounds with pheromonal activity that could improve development of a pheromone-based trap for monitoring Mediterranean fruit fly populations.     

Cross-Attraction of Carpophilus humeralis to Pheromone Components of Other Carpophilus Species

The pineapple beetle, Carpophilus humeralis, is known from field tests to be more attracted to baits containing the pheromone blends of other Carpophilus species and food odors than to food odors alone, but a pheromone is not yet known for C. humeralis. Wind-tunnel bioassays were used to determine specifically which of the Carpophilus pheromone components were the most attractive for C. humeralis. Nine tetraene and triene components normally used in field studies, plus eight additional male-specific compounds from other Carpophilus species, and one additional analog, were used in these experiments. At the 10-ng level, 15 of 18 compounds tested with a food-related coattractant (propyl acetate) were more attractive for C. humeralis than the coattractant alone. With decreasing doses (1, 0.1, and 0.01 ng) the number of attractive compounds declined. Only one compound, (2E,4E,6E,8E)-3,5,7-trimethyl-2,4,6,8-decatetraene (1), had significant attraction for C. humeralis at 0.01 ng, close to the lower threshold level of pheromone attraction for other Carpophilus species. Responses to mixtures of compounds were explored. For example, 1 was synergistic with 4-ethyl-2-methoxyphenol (E), 2,5-diisopropylpyrazine (D), and 2-phenylethanol (P), which are known attractants for C. humeralis; a mixture of 1 plus E, D, and P was 20 times more attractive than 1 alone and five times more than E, D, plus P. The specificity of C. humeralis for certain tetraenes was similar to that exhibited by C. hemipterus, while the specificity for trienes was similar to that of C. freemani. The behavior of C. humeralis to these pheromone components suggests that it may have a pheromone like that of other Carpophilus species or that it could possibly use these compounds as host finding kairomones.     

Male olive fruit fly attraction to synthetic sex pheromone components in laboratory and field tests

Male olive fruit fly attraction to the four synthetic components of the female sex attractant pheromone was studied under laboratory and field conditions. In laboratory tests males responded to all four components tested separately. Component I, (1,7-dioxaspiro[5.5]undecane) was more attractive than any of the remaining three components alone, but a combination of all four was more attractive than component I alone. In field tests with polyethylene vials as pheromone dispensers, the complete mixture, although not statistically significant, was constantly more attractive to males than component I alone. A tendency of enhancement of attraction of component I by combining it with component II (-pinene) or III (n-nonanal) was also observed. In field tests with rubber septa as pheromone dispensers only component I was attractive. Mixtures containing component I were also attractive but not more attractive than component I alone. Evaporation rate and ratio of components as they come out of the dispenser appear to be critical for male response.     

Identification of Queen Sex Pheromone Components of the Bumblebee Bombus terrestris

We investigated the origin and chemical composition of the queen sex pheromone of the primitively eusocial bumblebee, Bombus terrestris (Apidae). Physiologically and behaviorally active compounds were identified by coupled gas chromatography electroantennography (GC-EAD), gas chromatography-mass spectrometry (GC-MS), and laboratory behavioral tests. In the behavioral assays, virgin queens frozen previously at −20°C were highly attractive to males. Dummies impregnated with surface and cephalic extracts obtained from virgin queens that had been frozen at −50°C were more attractive to males than odorless dummies. Male mating behavior was stimulated by components of cephalic secretions that are smeared onto the cuticle surface by the queen. Overall, 21 compounds present in surface and cephalic extracts evoked electroantennographic responses in male antennae. These included saturated and unsaturated fatty acids, ethyl- and methyl esters of the fatty acids, heptacosene, 2-nonanone, and geranyl geraniol. A blend of synthetic versions of these compounds elicited typical male mating behavior. Since solvent-impregnated dummies were approached by the males, but did not release copulatory behavior, visual cues may be important in the initial step of stimulating male mating behavior. Close-range olfactory signals are more important for releasing male mating behavior as well as for species recognition. In further behavioral assays, the attractiveness of a frozen virgin queen decreased as the storage time at −20°C increased from 2 hr to 1 d. Therefore, the chemical composition of the sex pheromone may change during freezing as behaviorally active compounds may decompose.     

Modification of responses byCampylomma verbasci (Heteroptera: Miridae) to pheromone blends in atmospheres permeated with synthetic sex pheromone or individual components

Trapping experiments were conducted in orchards to test the hypothesis that exposure of the mullein bug,Campylomma verbasci (Meyer), to atmospheres permeated with its synthetic sex pheromone, a 94:6 blend of butyl butyrate (BB) and (E)-crotyl butyrate (CB), or BB or CB alone, would alter the pheromone responses of males toward off-ratio blends. Exposure to the natural 94:6 blend shifted the response preference away from the natural ratio to blends enriched in BB, including a 99:1 blend, which is normally significantly less attractive than the natural ratio. In an atmosphere permeated with CB, male mullein bugs were as responsive to blends containing 20, 33, and 43% CB, up to seven times the natural percentage, as they were to the 94:6 blend. In an atmosphere permeated with BB, responses to five blends ranging from 94:6 to 99:1 BB:CB were not significantly different, whereas in an untreated atmosphere blends of 98:2 and 99:1 were significantly less attractive than the 94:6 blend. Individual components appeared to disrupt pheromone communication primarily through the creation of a sensory imbalance and modified interpretation of pheromone blend ratios, whereas disruption by the complete pheromone blend apparently involves several mechanisms, including false-trail following and camouflage. Disruption of pheromone communication was most effective with the natural 94:6 sex pheromone blend.     

Identification of odors from overripe mango that attract vinegar flies,Drosophila melanogaster

Bioassays with a variety of overripe fruits, including mango, plum, pear, and grape, and their extracts showed that odors from overripe mango were most attractive to adult vinegar flies, Drosophila melanogaster. Combined gas chromatography–electroantennographic detection (GC-EAD) analyses of solid-phase microextraction (SPME) and Tenax extracts of overripe mango odors showed that several volatile compounds, including ethanol, acetic acid, amyl acetate, 2-phenylethanol, and phenylethyl acetate elicited significant EAG responses from antennae of female flies. Most of the volatile compounds in the extracts were identified by mass spectral and retention index comparisons with synthetic standards. In cage bioassays, lures with a blend of ethanol, acetic acid, and 2-phenylethanol in a ratio of 1:22:5 attracted six times more flies than any single EAG-active compound. This blend also attracted four times more flies than traps baited with overripe mango or unripe mango. However, in field trials, the blend was not as attractive as suggested by the laboratory bioassay.     

Drones of the Dwarf Honey Bee <Emphasis Type="Italic">Apis Florea</Emphasis> Are Attracted to (2E)-9-Oxodecenoic Acid and (2E)-10-Hydroxydecenoic Acid

The queen mandibular gland component (2E)-9-oxodecenoic acid (9-ODA) has been suggested to function as the major sex pheromone component in all honey bee species. In contrast to this hypothesis, chemical analyses showed that in the Asian dwarf honey bee species, Apis florea, a different decenoic acid, (2E)-10-hydroxydecenoic acid (10-HDA), is the major component in the mandibular gland secretion. We show here that A. florea drones are attracted to 9-ODA as well as to 10-HDA. However, 10-HDA attracted higher numbers of drones at lower dosages than 9-ODA, and also was more attractive when directly compared to 9-ODA in a dual attraction experiment. We conclude that 10-HDA has to be viewed as the major sex pheromone in A. florea. The result that both pheromone components are capable of attracting drones when presented alone was unexpected with regard to existing sex pheromone attraction experiments in honey bees.     

Volatile compounds induced by herbivory act as aggregation kairomones for the Japanese beetle (Popillia japonica Newman)

The Japanese beetle is a polyphagous insect that typically aggregates on preferred host plants in the field. We studied the response of Japanese beetles to artificial damage, fresh feeding damage, and overnight feeding damage to test the hypothesis that beetles are attracted to feeding-induced volatiles. Crabapple leaves that had been damaged overnight by Japanese beetles or fall webworms attracted significantly more Japanese beetles than did undamaged leaves. Artificially damaged leaves or leaves freshly damaged by Japanese beetles, however, were not significantly more attractive than undamaged leaves. Leaves that had been damaged overnight by Japanese beetles or fall webworms produced a complex mixture of aliphatic compounds, phenylpropanoid-derived compounds, and terpenoids. In comparison, artificially damaged leaves or leaves with fresh Japanese beetle feeding damage generated a less complex blend of volatiles, mainly consisting of green-leaf odors. Feeding-induced odors may facilitate host location and/or mate finding by the Japanese beetle.     

Disruption of male spruce budworm orientation to calling females in a wind tunnel by synthetic pheromone

Male spruce budworm [Chorisloneura fumiferana (Clem.)] moths were held for 3 hr in a wind tunnel and subjected to various concentrations of background synthetic pheromone. They were then exposed to calling females and their response was recorded. The background pheromone was presented either as discrete turbulent plumes or as a uniform permeation throughout the tunnel. The numbers of males wing-fanning and flying in response to the calling females decreased as the concentration of background pheromone increased. Of the males which flew, a higher proportion progressed upwind in the discrete plumes than in the uniform permeation, an indication that structure in the pheromone cloud is necessary for upwind progression. In both discrete plumes and uniform permeation fewer males were able to locate the females (i.e., disruption was greater) as the concentration of synthetic pheromone increased, but for the same total release rates, disruption was greater when the synthetic pheromone was released in discrete plumes rather than in a uniform permeation. This implies that disruption which involves luring males to sources of synthetic pheromone is more effective than masking female plumes by uniform permeation and suggests that it is more efficient to release pheromone from a few potent sources than from numerous low-potency sources.     

Aggregation pheromone ofCarpophilus antiquus (Coleoptera: Nitidulidae) and kairomonal use ofC. lugubris pheromone byC. antiquus

Males ofCarpophilus antiquus Melsheimer (Coleoptera: Nitidulidae) emit an aggregation pheromone that was found to be a novel hydrocarbon, (3E,5E,7E,9E)-6,8-diethyl-4-methyl-3,5,7,9-dodecatetraene. A synthetic scheme and spectra (mass and proton NMR) are given for the compound. Beetles produced the pheromone when feeding on a variety of media, including the brewer's yeast-based artificial diet, fermenting whole-wheat bread dough, corn, and prunes; live baker's yeast was generally added to the food media. Males held individually produced, on average, 25 × more pheromone per beetle than males held in groups of 10 or more. Pheromone was not produced until males were at least 5 days old but was still detected from the oldest beetles tested (47 days). In field tests, the pheromone was attractive to both sexes ofC. antiquus, and it was synergized by food volatiles: A combination of pheromone and fermenting whole wheat dough attracted 2.5× more beetles than pheromone alone, but dough by itself was not significantly more attractive than the control. Semiochemical interactions were studied amongC. antiquus and two other sympatric species for which pheromones are known,C. lugubris Murray andC. freemani Dobson.C. antiquus responded readily to the pheromone ofC. lugubris, but all other interspecific responses to the pheromones were weak. In a sample of naturally infested corn ears, the presence ofC. antiquus was strongly associated with the presence ofC. lugubris, as would be expected if the pheromone ofC. lugubris serves as a kairomone forC. antiquus.