Male Pheromone Research Articles

Sex Pheromone Mediates Resource Partitioning Between Drosophila melanogaster and D. suzukii.

The spotted-wing drosophila, Drosophila suzukii and the cosmopolitan vinegar fly D. melanogaster feed on soft fruit and berries and widely overlap in geographic range. The presence of D. melanogaster reduces egg-laying in D. suzukii, possibly because D. melanogaster outcompetes D. suzukii larvae feeding in the same fruit substrate. Flies use pheromones to communicate for mating, but pheromones also serve a role in reproductive isolation between related species. We asked whether a D. melanogaster pheromone also modulates oviposition behaviour in D. suzukii. A dual-choice oviposition assay confirms that D. suzukii lays fewer eggs on blueberries exposed to D. melanogaster flies and further shows that female flies have a stronger effect than male flies. This was corroborated by treating berries with synthetic pheromones. Avoidance of D. suzukii oviposition is mediated by the female D. melanogaster pheromone (Z)-4-undecenal (Z4-11Al). Significantly fewer eggs were laid on berries treated with synthetic Z4-11Al. In comparison, the male pheromone (Z)-11-octadecenyl acetate (cVA) had no effect on D. suzukii oviposition. Z4-11Al is a highly volatile compound that is perceived via olfaction and it is accordingly behaviourally active at a distance from the source. D. suzukii is known to engage in mutual niche construction with the yeast Hanseniaspora uvarum, which strongly attracts flies. Adding Z4-11Al to fermenting H. uvarum significantly decreased D. suzukii flight attraction in a laboratory wind tunnel and a field trapping assay. That a D. melanogaster pheromone regulates oviposition in D. suzukii demonstrates that heterospecific pheromone communication contributes to reproductive isolation and resource partitioning in cognate species. Stimulo-deterrent diversion or push-pull methods, building on combined use of attractant and deterrent compounds, have shown promise for control of D. suzukii. A pheromone that specifically reduces D. suzukii attraction and oviposition adds to the toolbox for D. suzukii integrated management.

A modular circuit coordinates the diversification of courtship strategies

Mate recognition systems evolve rapidly to reinforce the reproductive boundaries between species, but the underlying neural mechanisms remain enigmatic. Here we leveraged the rapid coevolution of female pheromone production and male pheromone perception in Drosophila1,2 to gain insight into how the architecture of mate recognition circuits facilitates their diversification. While in some Drosophila species females produce unique pheromones that act to arouse their conspecific males, the pheromones of most species are sexually monomorphic such that females possess no distinguishing chemosensory signatures that males can use for mate recognition3. We show that Drosophila yakuba males evolved the ability to use a sexually monomorphic pheromone, 7-tricosene, as an excitatory cue to promote courtship. By comparing key nodes in the pheromone circuits across multiple Drosophila species, we reveal that this sensory innovation arises from coordinated peripheral and central circuit adaptations: a distinct subpopulation of sensory neurons has acquired sensitivity to 7-tricosene and, in turn, selectively signals to a distinct subset of P1 neurons in the central brain to trigger courtship. Such a modular circuit organization, in which different sensory inputs can independently couple to parallel courtship control nodes, may facilitate the evolution of mate recognition systems by allowing novel sensory modalities to become linked to male arousal. Together, our findings suggest how peripheral and central circuit adaptations can be flexibly coordinated to underlie the rapid evolution of mate recognition strategies across species.

Influencing and Regulating the Estrous Cycle in Outbred CD1 Laboratory Mice (Mus musculus)

AbstractThe Whitten effect is a widely used tool for manipulating the mouse estrous cycle and generating reproductively active females within the laboratory setting. Typically, peak numbers of sexually receptive mice occur following exposure to male pheromones, resulting in a higher number of successful copulations on the third day after exposure. Although this method has improved efficiencies, the percentage of females mated and subsequently deemed to be pregnant/pseudopregnant remains relatively low, around 50%. In experiment 1, we aimed to 1) further understand cyclicity; 2) determine whether the initial cycle stage plays an importance on day 3 receptivity; and 3) identify any repetitive patterns/cycle stabilization. Mice (n = 27) were assigned to group cages according to cycle stage (proestrus, estrus, metestrus, diestrus). Experiment 2 was developed to determine an optimum treatment to promote receptivity by exposure to various pheromone stimuli. Mice (n = 45) were randomly assigned to 5 treatment groups (PBS-treated sham soiled bedding, male soiled bedding, live male, pregnant females, and lactating females). In both experiments, daily vaginal cytology was performed for 21 days to determine the cycle stage. Results from experiment 1 indicate that the initial cycle stage did not contribute to day 3 receptivity, although synchronization within several groups/cages was noted, and that the greatest numbers of estrous animals were obtained on days 6 and 7. Experiment 2 revealed that exposure to live males and lactating females both significantly improved receptivity compared with the PBS, male soiled bedding, and pregnant female groups. These results indicate that current strategies used for routine synchronization could be further improved through alternative housing regimens without compromising animal welfare.

Male cuticular pheromones stimulate removal of the mating plug and promote re-mating through pC1 neurons in Drosophila females

In birds and insects, the female uptakes sperm for a specific duration post-copulation known as the ejaculate holding period (EHP) before expelling unused sperm and the mating plug through sperm ejection. In this study, we found that Drosophila melanogaster females shortens the EHP when incubated with males or mated females shortly after the first mating. This phenomenon, which we termed male-induced EHP shortening (MIES), requires Or47b+ olfactory and ppk23+ gustatory neurons, activated by 2-methyltetracosane and 7-tricosene, respectively. These odorants raise cAMP levels in pC1 neurons, responsible for processing male courtship cues and regulating female mating receptivity. Elevated cAMP levels in pC1 neurons reduce EHP and reinstate their responsiveness to male courtship cues, promoting re-mating with faster sperm ejection. This study established MIES as a genetically tractable model of sexual plasticity with a conserved neural mechanism.

Male cuticular pheromones stimulate removal of the mating plug and promote re-mating through pC1 neurons in Drosophila females.

In birds and insects, the female uptakes sperm for a specific duration post-copulation known as the ejaculate holding period (EHP) before expelling unused sperm and the mating plug through sperm ejection. In this study, we found that Drosophila melanogaster females shortens the EHP when incubated with males or mated females shortly after the first mating. This phenomenon, which we termed male-induced EHP shortening (MIES), requires Or47b+ olfactory and ppk23+ gustatory neurons, activated by 2-methyltetracosane and 7-tricosene, respectively. These odorants raise cAMP levels in pC1 neurons, responsible for processing male courtship cues and regulating female mating receptivity. Elevated cAMP levels in pC1 neurons reduce EHP and reinstate their responsiveness to male courtship cues, promoting re-mating with faster sperm ejection. This study established MIES as a genetically tractable model of sexual plasticity with a conserved neural mechanism.

Cryptic female choice in response to male pheromones in Drosophila melanogaster

Females control the paternity of their offspring by selectively mating with males they perceive to be of high quality. In species where females mate with multiple males in succession, females may bias offspring paternity by favoring the sperm of one male over another, a process known as cryptic female choice.1 While evidence of cryptic female choice exists in multiple taxa, the mechanisms underlying this process have remained difficult to unravel.2 Understanding cryptic female choice requires demonstration of a female-driven post-mating bias in sperm use and paternity and a causal link between this bias and male cues.3 In this study, we present evidence of cryptic female choice in female Drosophila melanogaster. Through experiments utilizing transgenic males expressing fluorescent sperm, we observed that exposure to attractive males between matings prompts females to expel the ejaculate of their initial mate more rapidly than in the presence of less attractive males. While doing so, females exhibit a bias in sperm storage against their first mate, thereby favoring the paternity of their subsequent mate. Our findings reveal that females adjust the timing of ejaculate expulsion in response to male pheromones in their environment, specifically heptanal and 11-cis-vaccenyl acetate, which are sensed by females through specific odorant receptors. We provide a cryptic female choice mechanism allowing a female to modulate the share of paternity of her first mate depending on the sensing of the quality of potential mates in her environment. These findings showcase that paternity can be influenced by events beyond copulation.

Population Density Affects Drosophila Male Pheromones in Laboratory-Acclimated and Natural Lines.

In large groups of vertebrates and invertebrates, aggregation can affect biological characters such as gene expression, physiological, immunological and behavioral responses. The insect cuticle is covered with hydrocarbons (cuticular hydrocarbons; CHCs) which reduce dehydration and increase protection against xenobiotics. Drosophila melanogaster and D. simulans flies also use some of their CHCs as contact pheromones. In these two sibling species, males also produce the volatile pheromone 11-cis-Vaccenyl acetate (cVa). To investigate the effect of insect density on the production of CHCs and cVa we compared the level of these male pheromones in groups of different sizes. These compounds were measured in six lines acclimated for many generations in our laboratory - four wild-type and one CHC mutant D. melanogaster lines plus one D. simulans line. Increasing the group size substantially changed pheromone amounts only in the four D. melanogaster wild-type lines. To evaluate the role of laboratory acclimation in this effect, we measured density-dependent pheromonal production in 21 lines caught in nature after 1, 12 and 25 generations in the laboratory. These lines showed varied effects which rarely persisted across generations. Although increasing group size often affected pheromone production in laboratory-established and freshly-caught D. melanogaster lines, this effect was not linear, suggesting complex determinants.

Partial role of volatile organic compounds in behavioural responses of mice to bedding from cancer-affected congeners.

Tumours induce changes in body odours. We compared volatile organic compounds (VOCs) in soiled bedding of a lung adenocarcinoma male mouse model in which cancer had (CC) versus had not (NC) been induced by doxycycline at three conditions: before (T0), after 2weeks (T2; early tumour development), after 12 weeks (T12; late tumour development) of the induction. In an earlier study, wild-derived mice behaviourally discriminated between CC and NC soiled bedding at T2 and T12. Here, we sought to identify VOCs present in the same soiled bedding that could have triggered the behavioural discrimination. Solid phase micro-extraction was performed to extract VOCs from 3 g-sample stimuli. While wild-derived mice could discriminate the odour of cancerous mice at a very early stage of tumour development (T2), the present study did not identify VOCs that could explain this behaviour. However, consistent with the earlier behavioural study, four VOCs, including two well-known male mouse sex pheromones, were found to be present in significantly different proportions in soiled bedding of CC as compared to NC at T12. We discuss the potential involvement of non-volatile molecules such as proteins and peptides in behavioural discrimination of early tumour development (T2), and point-out VOCs that could help diagnose cancer.

Accelerated hermaphrodite maturation on male pheromones suggests a general principle of coordination between larval behavior and development.

Environment in general and social signals in particular could alter development. In Caenorhabditis elegans, male pheromones hasten development of hermaphrodite larvae. We show that this involves acceleration of growth and both somatic and germline development during the last larval stage (L4). Larvae exposed to male pheromones spend more time in L3 and less in the quiescent period between L3 and L4. This behavioral alteration improves provision in early L4, likely allowing for faster development. Larvae must be exposed to male pheromones in late L3 for behavioral and developmental effects to occur. Latter portions of other larval stages also contain periods of heightened sensitivity to environmental signals. Behavior during the early part of the larval stages is biased toward exploration, whereas later the emphasis shifts to food consumption. We argue that this organization allows assessment of the environment to identify the most suitable patch of resources, followed by acquisition of sufficient nutrition and salient information for the developmental events in the next larval stage. Evidence from other species indicates that such coordination of behavior and development may be a general feature of larval development.

The people behind the papers - Denis Faerberg and Ilya Ruvinsky.

Male pheromones accelerate the development of hermaphrodite larvae in Caenorhabditis elegans, but the importance of this phenomenon is not well understood. A new paper in Development shows that pheromone exposure during larval stage 3 helps coordinate behaviour and development by modulating the timing of the transition to larval stage 4. To learn more about the story behind the paper, we caught up with first author Denis Faerberg who carried out the work in the lab of the corresponding author Ilya Ruvinsky at Northwestern University, USA.

Pheromone-based communication influences the production of somatic extracellular vesicles in C. elegans

Extracellular vesicles (EVs) are integral to numerous biological processes, yet it is unclear how environmental factors or interactions among individuals within a population affect EV-regulated systems. In Caenorhabditis elegans, the evolutionarily conserved large EVs, known as exophers, are part of a maternal somatic tissue resource management system. Consequently, the offspring of individuals exhibiting active exopher biogenesis (exophergenesis) develop faster. Our research focuses on unraveling the complex inter-tissue and social dynamics that govern exophergenesis. We found that ascr#10, the primary male pheromone, enhances exopher production in hermaphrodites, mediated by the G-protein-coupled receptor STR-173 in ASK sensory neurons. In contrast, pheromone produced by other hermaphrodites, ascr#3, diminishes exophergenesis within the population. This process is regulated via the neuropeptides FLP-8 and FLP-21, which originate from the URX and AQR/PQR/URX neurons, respectively. Our results reveal a regulatory network that controls the production of somatic EV by the nervous system in response to social signals.

Complexity of Chemical Emissions Increases Concurrently with Sexual Maturity in Heliconius Butterflies

Pheromone communication is widespread among animals. Since it is often involved in mate choice, pheromone production is often tightly controlled. Although male sex pheromones (MSPs) and anti-aphrodisiacs have been studied in some Heliconius butterfly species, little is known about the factors affecting their production and release in these long-lived butterflies. Here, we investigate the effect of post-eclosion age on chemical blends from pheromone-emitting tissues in Heliconius atthis and Heliconius charithonia, exhibiting respectively free-mating and pupal-mating strategies that are hypothesised to differently affect the timing of their pheromone emissions. We focus on two different tissues: the wing androconia, responsible for MSPs used in courtship, and the genital tip, the production site for anti-aphrodisiac pheromones that affect post-mating behaviour. Gas chromatography-mass spectrometric analysis of tissue extracts from virgin males and females of both species from day 0 to 8 post-eclosion demonstrates the following. Some ubiquitous fatty acid precursors are already detectable at day 0. The complexity of the chemical blends increases with age regardless of tissue or sex. No obvious difference in the time course of blend production was evident between the two species, but female tissues in H. charithonia were more affected by age than in H. atthis. We suggest that compounds unique to male androconia and genitals and whose amount increases with age are potential candidates for future investigation into their roles as pheromones. While this analysis revealed some of the complexity in Heliconius chemical ecology, the effects of other factors, such as the time of day, remain unknown.

EsigPBP3 Was the Important Pheromone-Binding Protein to Recognize Male Pheromones and Key Eucalyptus Volatiles.

Pheromone-binding proteins (PBPs) are specific odorant-binding proteins that can specifically recognize insect pheromones. Through transcriptional analysis of the antennae of adult Endoclita signifer, EsigPBP3 was discovered and identified, and EsigPBP3 was found to be highly expressed in the antennae of male moths. Based on the binding characteristics and ability of EsigPBP3, we can find the key ligands and binding site to consider as a target to control the key wood bore E. signifier. In this study, the fluorescence competitive binding assays (FCBA) showed that EsigPBP3 had a high binding affinity for seven key eucalyptus volatiles. Molecular docking analysis revealed that EsigPBP3 had the strongest binding affinity for the sexual pheromone component, (3E,7E)-4,7,11-trimethyl-1,3,7,10-dodecatetraene. Furthermore, same as the result of FCBA, the EsigPBP3 exhibited high binding affinities to key eucalyptus volatiles, eucalyptol, α-terpinene, (E)-beta-ocimene, (-)-β-pinene, and (-)-α-pinene, and PHE35, MET7, VAL10, PHE38, ILE52, and PHE118 are key sites. In summary, EsigPBP3 exhibits high binding affinity to male pheromones and key volatile compounds and the crucial binding sites PHE35, MET7, VAL10, PHE38, ILE52, and PHE118 can act as targets in the recognition of E. signifier pheromones.

Sensory trap leads to reliable communication without a shift in nonsexual responses to the model cue.

The sensory trap model of signal evolution suggests that males manipulate females into mating using traits that mimic cues used in a nonsexual context. Despite much empirical support for sensory traps, little is known about how females evolve in response to these deceptive signals. Female sea lamprey (Petromyzon marinus) evolved to discriminate a male sex pheromone from the larval odor it mimics and orient only toward males during mate search. Larvae and males release the attractant 3-keto petromyzonol sulfate (3kPZS), but spawning females avoid larval odor using the pheromone antagonist, petromyzonol sulfate (PZS), which larvae but not males, release at higher rates than 3kPZS. We tested the hypothesis that migratory females also discriminate between larval odor and the male pheromone and orient only to larval odor during anadromous migration, when they navigate within spawning streams using larval odor before they begin mate search. In-stream behavioral assays revealed that, unlike spawning females, migratory females do not discriminate between mixtures of 3kPZS and PZS applied at ratios typical of larval versus male odorants. Our results indicate females discriminate between the sexual and nonsexual sources of 3kPZS during but not outside of mating and show sensory traps can lead to reliable sexual communication without females shifting their responses in the original context.

Replenishment of Drosophila Male Pheromone After Mating.

Insect exocrine gland products can be involved in sexual communication, defense, territory labelling, aggregation and alarm. In the vinegar fly Drosophila melanogaster the ejaculatory bulb synthesizes and releases 11-cis-Vaccenyl acetate (cVa). This pheromone, transferred to the female during copulation, affects aggregation, courtship and male-male aggressive behaviors. To determine the ability of male flies to replenish their cVa levels, males of a control laboratory strain and from the desat1 pheromone-defective mutant strain were allowed to mate successively with several females. We measured mating frequency, duration and latency, the amount of cVa transferred to mated females and the residual cVa in tested males. Mating duration remained constant with multiple matings, but we found that the amount of cVa transferred to females declined with multiple matings, indicating that, over short, biologically-relevant periods, replenishment of the pheromone does not keep up with mating frequency, resulting in the transfer of varying quantities of cVa. Adult responses to cVa are affected by early developmental exposure to this pheromone; our revelation of quantitative variation in the amount of cVa transferred to females in the event of multiple matings by a male suggests variable responses to cVa shown by adults produced by such matings. This implies that the natural role of this compound may be richer than suggested by laboratory experiments that study only one mating event and its immediate behavioral or neurobiological consequences.

Male lake char release taurocholic acid as part of a mating pheromone.

The evolutionary origins of sexual preferences for chemical signals remain poorly understood, due, in part, to scant information on the molecules involved. In the current study, we identified a male pheromone in lake char (Salvelinus namaycush) to evaluate the hypothesis that it exploits a non-sexual preference for juvenile odour. In anadromous char species, the odour of stream-resident juveniles guides migratory adults into spawning streams. Lake char are also attracted to juvenile odour but have lost the anadromous phenotype and spawn on nearshore reefs, where juvenile odour does not persist long enough to act as a cue for spawning site selection by adults. Previous behavioural data raised the possibility that males release a pheromone that includes components of juvenile odour. Using metabolomics, we found that the most abundant molecule released by males was also released by juveniles but not females. Tandem mass spectrometry and nuclear magnetic resonance were used to identify the molecule as taurocholic acid (TCA), which was previously implicated as a component of juvenile odour. Additional chemical analyses revealed that males release TCA at high rates via their urine during the spawning season. Finally, picomolar concentrations of TCA attracted pre-spawning and spawning females but not males. Taken together, our results indicate that male lake char release TCA as a mating pheromone and support the hypothesis that the pheromone is a partial match of juvenile odour.

Courtship and mating behavior of Endoclita signifer (Hepialidae: Lepidoptera) and the male sex pheromones in hairbrushes.

Endoclita signifer Walker is the most destructive wood-boring pest of Eucalyptus in China, causing significant economic and ecological damage. As an insect of the primitive Lepidoptera family Hepialidae, E. signifer fly and mat for only 10-20 min at dusk. The courtship and mating behavior of E. signifer adults and whether male moths release sex pheromones are still unknown, especially since transitory flight survival strategies in primitive moths differ from advanced moths like noctuids. In this study, we first observed the courtship and mating behavior of E. signifer by considering the effects of space and then analyzed extracts of male hairbrushes using gas chromatography-electroantennogram detection. Our results indicated that during the courtship period, flying males form courtship fields, lekking, and chase flying females before mating with them; E. signifer were more successful in mating in larger spaces (Length × Width × Height = 9.6 × 7 × 4 m); 5 compounds in the hairbrushes of the male moths which elicited antennal responses of 2 sexes, despite at high concentrations. Combined with it, indicating that communication between male and female may rely on male sex pheromones. These findings can serve as a basis for studying the mechanisms of sex communication in E. signifer and developing sex pheromone-based trapping techniques.

The application of pheromones extracted from Shami goats' male hair and Awassi sheep male wool during the breeding season on LH and Progesterone concentrations in Awassi ewes in Syria.

The efficiency and profitability of animal farming systems are closely linked to the reproductive success of livestock, which requires effective management through biological, hormonal, and nutritional strategies. The "male effect," referring to the influence of male pheromones on female reproductive hormones, offers several benefits. This leads to improved reproductive management, better timing of breeding, and increased fertility rates. Additionally, using natural pheromones provides a non-invasive and sustainable method for managing reproduction, and it can be a cost-effective strategy, reducing the need for expensive hormonal treatments. Incorporating male pheromones into reproductive management practices can thus boost the efficiency and profitability of animal farming systems. The study's objective is to examine the influence of pheromones, extracted from Awassi ram's wool and Shami goat buck's hair, on the secretion of luteinizing hormone (LH) and Progesterone (P4) in Awassi ewes during the breeding season. The pheromones were extracted using a soaking method with dichloromethane as the solvent. The resulting organic extract was then analyzed using a Chromatec-Crystal 5,000 gas chromatography-mass spectrometry device, equipped with a quadrupole mass spectrometer. The experiment was carried out on three equal groups, each consisting of 17 subjects, during the breeding season. In Group I, 15 ml of the ram's wool extract was applied to a piece of gauze and placed in a specially designed mask that was attached directly to the nose. In the second group, a similar procedure was followed using 15 ml of the buck's hair extract. The third group consisted of Awassi rams. The LH assay was conducted immediately before the treatment and again at 1, 10, and 20 hours post-treatment, and after 5 and 30 days of treatment to assess P4 levels. The study found that pheromones from Awassi rams and Shami goat bucks significantly increased the levels of LH and P4 in female Awassi sheep. The study concludes that the pheromones extracted from the wool of rams and Shami goat bucks can stimulate the response of female Awassi sheep. This finding opens up the possibility of using these pheromones in the reproductive management of female Awassi sheep.

The seminal odorant binding protein Obp56g is required for mating plug formation and male fertility in Drosophila melanogaster.

In Drosophila melanogaster and other insects, the seminal fluid proteins (SFPs) and male sex pheromones that enter the female with sperm during mating are essential for fertility and induce profound post-mating effects on female physiology. The SFPs in D. melanogaster and other taxa include several members of the large gene family known as odorant binding proteins (Obps). Work in Drosophila has shown that some Obp genes are highly expressed in the antennae and can mediate behavioral responses to odorants, potentially by binding and carrying these molecules to odorant receptors. These observations have led to the hypothesis that the seminal Obps might act as molecular carriers for pheromones or other compounds important for male fertility, though functional evidence in any species is lacking. Here, we used functional genetics to test the role of the seven seminal Obps in D. melanogaster fertility and the post-mating response (PMR). We found that Obp56g is required for male fertility and the induction of the PMR, whereas the other six genes are dispensable. We found males lacking Obp56g fail to form a mating plug in the mated female's reproductive tract, leading to ejaculate loss and reduced sperm storage, likely due to its expression in the male ejaculatory bulb. We also examined the evolutionary history of these seminal Obp genes, as several studies have documented rapid evolution and turnover of SFP genes across taxa. We found extensive lability in gene copy number and evidence of positive selection acting on two genes, Obp22a and Obp51a. Comparative RNAseq data from the male reproductive tract of multiple Drosophila species revealed that Obp56g shows high male reproductive tract expression in a subset of taxa, though conserved head expression across the phylogeny. Together, these functional and expression data suggest that Obp56g may have been co-opted for a reproductive function over evolutionary time.

Individual Variation in Male Pheromone Production in Xylocopa sonorina Correlates with size and Gland Color.

Sex pheromones are species-specific chemical signals that facilitate the location, identification, and selection of mating partners. These pheromones can vary between individuals, and act as signals of mate quality. Here, we investigate the variation of male pheromones in the mesosomal glands of the large carpenter bee Xylocopa sonorina, within a Northern California population. We tested the hypothesis that morphological traits are correlated with the observed variation in chemical blend composition of these bees. We also conducted behavioral assays to test whether these male pheromones act as long-range attractants to conspecifics. We found that larger males with darker mesosomal glands have a higher pheromone amount in their glands. Our analysis also suggests that this pheromone blend functions as a long-range attractant to both males and females. We show that both male body size and sexual maturation are important factors influencing pheromone abundance, and that this pheromone blend acts as a long-range attractant. We hypothesize that this recorded variation in male pheromone could be important for female choice.