Effect of combined feeding stimuli on ingestion by the nematodeTrichostrongylus colubriformis

Ingestion by the nematodeTrichostrongylus colubriformis was stimulated by serotonin, histamine, and dopamine. Immune serum inhibited feeding by both sexes while octopamine depressed feeding by only male nematodes. However, the combined influence of these excitatory and inhibitory stimuli is unknown. Feeding in vitro by both sexes of the nematodeT. colubriformis was similar in millimolar ratios of histamine and dopamine or histamine and serotonin, but not dopamine and serotonin. Feeding by nematodes that was stimulated by histamine, dopamine, and serotonin was decreased by subsequent exposure to octopamine. Conversely, elevated concentrations of histamine, dopamine, and serotonin decreased the inhibitory action of octopamine on helminth ingestion in vitro. Doses of immune serum also decreased the stimulated feeding of nematodes that was induced by histamine, dopamine or serotonin. However, the addition of elevated concentrations of these biogenic amines to immune serum reversed the inhibitory effect. The feeding activity of nematodes may represent a summation of the inhibitory and excitatory stimuli that occur in the chemical environment of the helminth.Mention of a trademark or proprietary product does not constitute a guarantee or warranty by USDA and does not imply its approval to the exclusion of other suitable products.     

Sex-identifying urine and molt signals in lobster (Homarus americanus)

During courtship, premolt female lobsters,Homarus americanus, choose a male and initiate a pair bond by repeated approaches to his shelter. The male allows such a female to share his shelter for about one week. This knowledge formed the basis to search for quantitative evidence for lobster sex pheromone(s) used in courtship: male cues to allow premolt females to identify a preferred male, and female cues to allow males to identify a premolt mature female. In each of four 1500-liter naturalistic aquaria, the behavioral responses of one female and two male lobsters to male and female lobster urine (0.5 ml) and body odor (20 ml) stimuli were observed. These stimuli were injected once or twice per day into a continuously flowing delivery tube attached to lobster shelters. Habituation to stimulus introduction—a serious problem in earlier experiments—was apparently avoided in the more natural social and physical environment we employed in these experiments. We demonstrated that male and female molt body odors contain different chemical substances: females responded to male molt body odor and males responded to female molt body odor but not vice versa. In general, male and female intermolt urine caused strong responses; however, females responded only weakly to male urine. This suggests that male and female urine are chemically different. Female urine and molt body odor caused a typically male high-on-legs response. These results show that molt body odors and intermolt urine contain sex-specific substances, which may be used in lobster courtship as well as other social interactions.     

Laser microbeam studies of role of amphid receptors in chemosensory behavior of nematodeCaenorhabditis elegans

Amphid sensilla, historically considered the primary chemosensory structures of nematodes, were found to be necessary for the detection of only one of the six chemical stimuli that were tested. Only the attraction to cAMP was eliminated by damaging the two lateral lips, which bear the amphid sensilla. The inner labial sensilla, one of which occurs on each of the six lips, are probably the primary receptor structures for the other chemical stimuli. Damaging all six lips, which should destroy all anterior chemosensory input, not only eliminted the attraction to sodium and chloride ions, but reversed the nematodes' response to them. Nematodes with all six lips destroyed showed reversal behavior when exposed to these attractants. Nematodes with damage to all six lips appeared to recover much of their normal chemosensory function within 24 hr after treatment.     

Pheromone attraction in the soybean cyst nematodeHeterodera glycines race 3

MaleHeterodera glycines responded to female nematodes during in vitro bioassay. The male's response was dosage-dependent and significant with a pheromone source of more than five females. Male responsiveness was influenced by the pheromone diffusion and response times. Males were most responsive at three days after emergence from the host plant, while females were also most attractive at the same age. Light intensities that ranged from dark to bright had no effect on female location by the male, although bioassay in a nitrogen atmosphere eliminated sexual communication. Mate location was not significant below 25 °C and declined slightly at 30 or 33 °C. Bioassay at pHs from 5 to 8.5 showed a bimodal effect, with maximal attraction around pH 6.     

Female sex pheromone-mediated effects on behavior and consequences of male competition in the shore crab (Carcinus maenas)

Exposure to receptive female pheromone elicits guarding behavior in shore crab males (Carcinus maenas), but little is known about the effects of sex pheromone on male competition or if the female plays an active role in mate choice. This study examined whether female pheromone enhanced agonistic behavior between males and what effects visual and chemical cues had on the rules and costs of such contests. We also investigated whether females exhibit a preference for males in terms of size. Under laboratory conditions, solitary male shore crabs engaged males who already had possession of a female. The visual and chemical presence of a receptive female had an impact on contest rules and costs. Fights were costly in terms of duration and of sustaining injury with either one or both crabs incurring injury in 40% of fights. To investigate the metabolic consequences of fighting over a perceived sexual resource (chemical cue only), fights were staged between pairs of size-matched males in the presence of water containing the female sex pheromone, water in which males had been kept, and untreated seawater. The duration and intensity of contests were greater when staged in the presence of the female pheromone compared with the two other treatments. Crabs that fought in the presence of female sex pheromone also had a greater accumulation of L-lactate and a reduction of glycogen stores. Fights were less costly in terms of injury with a single chemical cue compared with enhanced costs with a multiple sexual cue. The importance of female choice was determined by presenting postmolt females with different sized males. Males were kept in a fixed position, and the majority of females approached and performed courtship behavior to the largest males, demonstrating that females may be selective in terms of size.     

High Chemical Diversity in a Wasp Pheromone: a Blend of Methyl 6-Methylsalicylate,Fatty Alcohol Acetates and Cuticular Hydrocarbons Releases Courtship Behavior in the Drosophila Parasitoid Asobara tabida

Wasps of genus Asobara, a larval parasitoid of Drosophila, have become model organisms for the study of host-parasite interactions. However, little is known about the role of pheromones in locating mates and courtship behavior in this genus. In the present study, we aimed to identify the female courtship pheromone in Asobara tabida. The chemical compositions of solvent extracts from male and female wasps were analyzed by GC/MS. These extracts, fractions thereof, and synthetic pheromone candidates were tested for their activity in behavioral bioassays. The results demonstrate that the courtship pheromone of A. tabida is characterized by a remarkable chemical diversity. A multi-component blend of female-specific compounds including methyl 6-methylsalicylate (M6M), fatty alcohol acetates (FAAs), and cuticular hydrocarbons (CHCs) released male courtship behavior. Using a combinatory approach that included both purified natural products and synthetic analogs, it was shown that none of the three chemical classes alone was sufficient to release a full behavioral response in males. However, a blend of M6M and FAAs or combinations of one or both of these with female-derived CHCs resulted in wing-fanning responses by males comparable to those elicited by the crude extract of females. Thus, components from all three chemical classes contribute to the bioactivity of the pheromone, but none of the elements plays a key role or is irreplaceable. The fact that one of the FAAs, vaccenyl acetate, is also used as a kairomone by Asobara females to locate Drosophila hosts suggests that a pre-existing sensory responsiveness to vaccenyl acetate might have been involved in the evolution of the female sex pheromone in Asobara.     

Effect of Male House Mouse Pheromone Components on Behavioral Responses of Mice in Laboratory and Field Experiments

Urine of male house mice, Mus musculus, is known to have primer pheromone effects on the reproductive physiology of female mice. Urine-mediated releaser pheromone effects that trigger certain behavioral responses are much less understood, and no field studies have investigated whether urine deposits by male or female mice, or synthetic mouse pheromone, increase trap captures of mice. In field experiments, we baited traps with bedding soiled with urine and feces of caged female or male mice, and recorded captures of mice in these and in control traps containing clean bedding. Traps baited with female bedding preferentially captured adult males, whereas traps baited with male bedding preferentially captured juvenile and adult females, indicating the presence of male- and female-specific sex pheromones in soiled bedding. Analyses of headspace volatiles emanating from soiled bedding by gas chromatography/mass spectrometry revealed that 3,4-dehydro-exo-brevicomin (DEB) was seven times more prevalent in male bedding and that 2-sec-butyl-4,5-dihydrothiazole (DHT) was male-specific. In a follow-up field experiment, traps baited with DEB and DHT captured 4 times more female mice than corresponding control traps, thus indicating that DEB and DHT are sex attractant pheromone components of house mouse males. Our study provides impetus to identify the sex attractant pheromone of female mice, and to develop synthetic mouse pheromone as a lure to enhance the efficacy of trapping programs for mouse control.     

Male bugs modulate pheromone emission in response to vibratory signals from conspecifics

Male Nezara viridula emit a volatile sex pheromone that acts as a long-range attractant to females. Both sexes communicate through vibrations once they are on the same plant. Males respond to the female calling song by emitting a male courtship song, and they orient to the female calling song on a plant. Simultaneity between vibratory and chemical communication during the last steps of mate finding suggests that pheromone emission might be modulated by signals from conspecifics. Male volatile emissions were collected with fibers for solid-phase microextraction, while male bugs were stimulated with natural and artificial signals. Percentages of males releasing pheromone and collected amounts of pheromone increased when males were stimulated with female calling song. Pheromone emission was stable in males stimulated with male rivalry songs, and it decreased in males stimulated with a 100-Hz artificial signal. The ability of male bugs to modulate their pheromone emission may reduce metabolic costs, reduce parasitism, and offer a better synchronization of sexual activity.     

Conditioned Alarm Behavior in Fathead Minnows (Pimephales promelas) Resulting from Association of Chemical Alarm Pheromone with a Nonbiological Visual Stimulus

Fathead minnows (Pimephales promelas) adopt antipredator (alarm) behavior when they detect alarm pheromone released from an injured conspecific. This is an adaptive response since alarm pheromone is generally released only in the context of a predation event. Alarm reactions may also occur in response to chemical and visual stimuli that minnows learn to associate with release of alarm pheromone. Here, we tested if fathead minnows can learn to associate a nonbiological, visual stimulus with predation risk. Minnows were simultaneously exposed to red light and conspecific alarm pheromone, inducing an alarm reaction. When retested using red light alone, small shoals of minnows displayed an antipredator response: dashing movements and disorganized swimming followed by decreased height in the water column and increased shoal cohesion. This resulted from a single-trial exposure to the combined cues and demonstrates a robust ecological mechanism by which minnows learn to recognize indicators of predation risk that may vary in space and time. However, learning to associate risk with biologically irrelevant stimuli may be an ecological liability. How minnows discern between relevant and irrelevant stimuli in nature is not known.     

Evidence for a Substrate-Borne Sex Pheromone in the Parasitoid Wasp Trichogramma brassicae

While the use of chemical cues in host location is well known in parasitoid insects, little is known about their role in sexual communication. Sex pheromones were investigated in the egg parasitoid Trichogramma brassicae, and the evidence for a substrate-borne pheromone was demonstrated. Mated and unmated males concentrated their walking in an area where virgin females (<1 or 5 days old) had walked. Males were tested 15 min after females left the area. Male response decreased over time but was always significant during the 8 min of tracking. This suggests that females deposit chemicals on the substrate for mate finding. The pheromone is soluble in hexane, as shown by two kinds of extracts: female body rinsing and rinsing of the flask where females had walked. Hexane extracts resulted in a positive response by males. The response to different doses showed a saturation above a threshold of 4–8 female equivalents. The results do not exclude the possibility that a volatile part of the pheromone attracts males over long distances. Substrate-borne pheromones may increase mating chances for virgin females searching for oviposition sites, and may also increase mating chances on emergence sites by reducing male dispersion.     

Partial purification of the aggregation pheromone nippolure from femaleNippostrongylus brasiliensis (Nematoda)

Partial purification of a pheromone of the zooparasitic nematodeNippostrongylus brasiliensis was achieved. The polypeptide pheromone is degraded by Pronase while increased recovery is obtained after treatment with the protease inhibitors phenylmethylsulfonyl fluoride orl-1-tosylamide-2-phenylethyl chloromethyl ketone. The pheromone has a net negative charge, based on retention on DEAE cellulose and an approximate isoelectric point of 7.3. A combination of double treatments with phenylmethylsulfonyl fluoride and gel filtration enables storage up to nine days which yields biological activity similar to fresh, untreated chromatographic fractions.     

Pheromonal Mediation of Intraseasonal Declines in the Attractivity of Female Red-Sided Garter Snakes, Thamnophis sirtalis parietalis

During the breeding season, female red-sided garter snakes (Thamnophis sirtalis parietalis) produce and express a sexual attractiveness pheromone that elicits male courtship behavior. Composed of a homologous series of saturated and monounsaturated methyl ketones, this pheromone is expressed in female skin lipids. Recent studies have shown that the sexual attractivity of unmated female garter snakes declines as the breeding season progresses. Here, we investigated whether temporal changes in the quantity and/or quality of the female sexual attractiveness pheromone are responsible for the observed loss of attractivity. Female red-sided garter snakes were collected immediately following spring emergence and held under natural conditions for the duration of the breeding season. Behavioral experiments confirmed that unmated females become significantly less attractive to males within two weeks of emergence from hibernation. Additionally, these females had lower estradiol concentrations at two weeks post-emergence. Subsequent chemical analyses revealed qualitative variation between the pheromone profiles of newly emerged females and those of females at two weeks post-emergence. Together, these results support the hypothesis that changes in the female sexual attractiveness pheromone are responsible for declining post-emergence female attractivity in garter snakes.     

Video camera-computer tracking of nematodeCaenorhabditis elegans to record behavioral responses

A new method is used to analyze responses to changes in the concentration of two chemical stimuli. Nematodes are allowed to move around on the surface of a thin layer of agar across which a stream of air blows to carry volatile stimuli. Darkfield illumination provides high-contrast images of the worms which are acquired by a video camera and fed to a microcomputer which is programed to simultaneously track and record the movements and changes in direction of as many as 25 animals. The results are reported in real time. The worms respond to an increase in CO₂ concentration by decreasing the number moving and increasing the number of changes of direction. Both responses adapt to steady-state levels in about half a minute. This suggests that they respond by changing the probability of initiating a reversal bout. This observation adds a repellent to the class of stimuli thatC. elegans reponds to by klinokinesis. The resonses to changes in oxygen concentration are somewhat different. Movements and changes in direction both decrease when the oxygen concentration falls and increase when the concentration rises. No adaptation is seen within the one-minute time span observed. This observation provides further evidence that the response to oxygen differs from the response to other chemicals and may be sensed internally. These observations demonstrate that computer tracking is a sensitive method of analyzing animal behavior. It is further demonstrated that a significant response can be detected to a relatively weak stimulus in less than 5 min.     

Signal Honesty through Differential Quantity in the Female-Produced Sex Pheromone of the Moth <Emphasis Type="Italic">Heliothis virescens</Emphasis>

Over the last 50 years, female-produced sex pheromones of moths have been subjected to intensive study. Most work has focused on their role in mate recognition, and little on any role they may have in mate assessment. This is largely because it has been assumed that female, rather than male, moths are “choosy”, and invest larger amounts of carbon in eggs than in pheromone. Recently, we found that pheromone production in the moth Heliothis virescens depended on hemolymph trehalose concentration, and that sugar-stressed females produced less pheromone than unstressed ones. In this paper, we demonstrate, for the first time in moths, that a female-produced pheromone signal can allow H. virescens males to assess sugar resources (quality) of a female. This signal honesty is based on quantitative, rather than qualitative (component ratio), differences in pheromone, produced and released by sugar-stressed and unstressed females. Increasing marginal cost of pheromone production, as sugar resources are depleted, may ensure signal honesty.     

Airborne Chemical Communication in the Wolf Spider Pardosa milvina

Most studies involving chemical communication in spiders focus on contact pheromones attached to spider silk. Here we tested if males of the wolf spider Pardosa milvina use airborne pheromones to identify, locate, and follow females. Using a two-choice olfactometer, we tested the response of adult male P. milvina to a number of potential chemical cues while controlling for concomitant visual and vibratory stimuli. An airborne chemical cue from adult virgin female P. milvina elicited a positive taxis response from the male. We also tested adult male responses to penultimate instar female P. milvina, one adult male P. milvina, and two adult males together. In each case, test males showed no attraction to the stimuli. Additional experiments were run with pitfall traps baited with adult virgin female P. milvina as attractants. Again, we controlled for visual and vibratory cues from females. Pitfall traps containing virgin females captured significantly more males than control traps. Collectively, these experiments demonstrate evidence of an airborne sex pheromone in P. milvina.     

2-Methylthiazolidine and 4-ethylguaiacol,male sex pheromone components of the cockroachNauphoeta cinerea (dictyoptera,blaberidae): A reinvestigation

InNauphoeta cinerea, male calling behavior is associated with sex pheromone release by the sternal glands. The male pheromone that attracts females from a distance is a mixture of 2-methylthiazolidine and 4-ethylguaiacol. It is active at very low concentrations, 0.05 and 0.01 ng, respectively. Two other compounds, 3-hydroxy-2-butanone and 2-methyl-2thiazoline, act at close range, keeping the female in the vicinity of the male. The function of the volatile pheromone and those of previously described contact pheromones are discussed in regard to their possible involvement in the establishment of male dominant-subordinate relationships.     

Male Phyllotreta striolata (F.) Produce an Aggregation Pheromone: Identification of Male-specific compounds and Interaction with Host Plant Volatiles

The chrysomelid beetle Phyllotreta striolata is an important pest of Brassicaceae in Southeast Asia and North America. Here, we identified the aggregation pheromone of a population of P. striolata from Taiwan, and host plant volatiles that interact with the pheromone. Volatiles emitted by feeding male P. striolata attracted males and females in the field. Headspace volatile analyses revealed that six sesquiterpenes were emitted specifically by feeding males. Only one of these, however, elicited an electrophysiological response from antennae of both sexes. A number of host plant volatiles, e.g., 1-hexanol, (Z)-3-hexen-1-ol, and the glucosinolate hydrolysis products allyl isothiocyanate (AITC), 3-butenyl isothiocyanate, and 4-pentenyl isothiocyanate also elicited clear responses from the antenna. The active male-specific compound was identified as (+)-(6R,7S)-himachala-9,11-diene by chiral stationary phase gas-chromatography with coupled mass spectrometry, and by comparison with reference samples from Abies nordmanniana, which is known to produce the corresponding enantiomer. The pheromone compound was synthesized starting from (-)-α-himachalene isolated from Cedrus atlantica. Under field conditions, the activity of the synthetic pheromone required concomitant presence of the host plant volatile allyl isothiocyanate. However, both synthetic (+)-(6R,7S)-himachala-9,11-diene alone and in combination with AITC were attractive in a two-choice laboratory assay devoid of other natural olfactory stimuli. We hypothesize that P. striolata adults respond to the pheromone only if specific host volatiles are present. In the same laboratory set up, more beetles were attracted by feeding males than by the synthetic stimuli. Thus, further research will be necessary to reveal the components of a more complex blend of host or male-produced semiochemicals that might enhance trap attractiveness in the field.     

Pheromone-mediated mating in the aphid parasitoid,Aphidius nigripes (hymenoptera: Aphididae)

Mating in the aphid parasitoid,Aphidius nigripes, is mediated by a female sex pheromone that stimulates both upwind flight and courtship behavior by males. There is no evidence of any overt behavior associated with the release of the pheromone, but bioassay results, using female extracts, suggest that the pheromone is produced in the abdomen, although it may be actively released from the entire body surface. Age does not appear to have a major influence on either pheromone production by the female or on male responsiveness. However, field data suggest the presence of a strong diel periodicity in the emission of and/or receptivity to the pheromone. Laboratory bioassay results indicated diel changes in the male response but more refined techniques are needed to determine if such periodicity exists in pheromone production by females. Following mating, female pheromone production is significantly reduced. The potential use of the parasitoid sex pheromone in integrated management of aphids is discussed.     

Sex pheromone communication in the nematode,Rhabditis pellio

Both males and females ofRhabditis pellio release pheromones that attract the opposite sex prior to copulation. A quantitative bioassay for the female-produced pheromone was designed, based on male movement toward a pheromone source placed at one end of a 10-mm strip of bacterial material maintained on nutrient agar in a petri plate. Females produced pheromone from the age at which they attained the adult stage (3 days following hatching from the egg) and maintained a relatively constant production level until at least the ninth day of life. Similarly, males became responsive to the female pheromone by the third day and remained responsive through the ninth day, although the time required for the males to migrate toward a female pheromone source increased with increasing age. No daily rhythm of pheromone responsiveness by males or pheromone production by females was observed when the nematodes were conditioned to a 1212 h light-dark cycle.     

New Contact Sex Pheromone Components of the German Cockroach, Blattella germanica, Predicted from the Proposed Biosynthetic Pathway

Upon contacting the cuticle of a sexually mature female, a male German cockroach exhibits a characteristic courtship behavior: he turns away from the female and raises his wings, thereby exposing tergal glands. The glandular secretion stimulates the female to mount the male and feed, thus positioning her appropriately for copulation. A multi-component contact sex pheromone produced by females is responsible for eliciting courtship behavior. The most abundant pheromone components are 3,11-dimethylnonacosan-2-one and 3,11-dimethylheptacosan-2-one, oxidation products of the abundant hydrocarbon analogs 3,11-dimethylnonacosane and 3,11-dimethylheptacosane, respectively. The C₂₉-dimethyl ketone is thought to be further metabolized to two less abundant pheromone components, 29-hydroxy-3,11-dimethylnonacosan-2-one and 29-oxo-3,11-dimethylnonacosan-2-one. Based on this proposed biosynthetic pathway of pheromone production, we hypothesized that 3,11-dimethylheptacosan-2-one also would be oxidized to give two candidate pheromone components, 27-hydroxy-3,11-dimethylheptacosan-2-one, and 27-oxo-3,11-dimethylheptacosan-2-one. By using bioassay-guided fractionation and chemical analyses of cuticular extracts of virgin females and synthesis of the (3S,11S)-isomer of each of the two predicted pheromone components, we showed that the epicuticle of the German cockroach does indeed contain these two compounds. The contact sex pheromone of the female German cockroach, thus may consist of at least six biosynthetically related components.