Knockout of the delta11-desaturase SfruDES1 disrupts sex pheromone biosynthesis, mating and oviposition in the fall armyworm, Spodoptera frugiperda.

Moth insects rely on sex pheromones for long distance attraction and searching for sex partners. The biosynthesis of moth sex pheromones involves the catalytic action of multiple enzymes, with desaturases playing a crucial role in the process of carbon chain desaturation. However, the specific desaturases involved in sex pheromone biosynthesis in fall armyworm (FAW), Spodoptera frugiperda, have not been clarified. In this study, a Δ11 desaturase (SfruDES1) gene in FAW was knocked out using the CRISPR/Cas9 genome editing system. A homozygous mutant of SfruDES1 was obtained through genetic crosses. The gas chromatography-mass spectrometry (GC-MS) analysis results showed that the three main sex pheromone components (Z7-12:Ac, Z9-14:Ac, and Z11-16:Ac) and the three minor components (Z9-14:Ald, E11-14:Ac and Z11-14:Ac) of FAW were not detected in homozygous mutant females compared to the wild type. Furthermore, behavioral assay demonstrated that the loss of SfruDES1 resulted in a significant reduction in the attractiveness of females to males, along with disruptions in mating behavior and oviposition. Additionally, in a heterologous expression system, recombinant SfruDES1 could introduce a cis double bond at the Δ11 position in palmitic acid, which resulted in the changes in components of the synthesized products. These findings suggest desaturase plays a key role in the biosynthesis of sex pheromones, and knockout of the SfruDES1 disrupts sex pheromone biosynthesis and mating behavior in FAW. The SfruDES1 could serve as tool to develop a control method for S. frugiperda.

C. elegans males optimize mate-preference decisions via sex-specific responses to multimodal sensory cues.

For sexually reproducing animals, selecting optimal mates is important for maximizing reproductive fitness. In the nematode C. elegans, populations reproduce largely by hermaphrodite self-fertilization, but the cross-fertilization of hermaphrodites by males also occurs. Males' ability to recognize hermaphrodites involves several sensory cues, but an integrated view of the ways males use these cues in their native context to assess characteristics of potential mates has been elusive. Here, we examine the mate-preference behavior of C. elegans males evoked by natively produced cues. We find that males use a combination of volatile sex pheromones (VSPs), ascaroside sex pheromones, surface-associated cues, and other signals to assess multiple features of potential mates. Specific aspects of mate preference are communicated by distinct signals: developmental stage and sex are signaled by ascaroside pheromones and surface cues, whereas the presence of a self-sperm-depleted hermaphrodite is likely signaled by VSPs. Furthermore, males prefer to interact with virgin over mated, and well-fed over food-deprived, hermaphrodites; these preferences are likely adaptive and are also mediated by ascarosides and other cues. Sex-typical mate-preference behavior depends on the sexual state of the nervous system, such that pan-neuronal genetic masculinization in hermaphrodites generates male-typical social behavior. We also identify an unexpected role for the sex-shared ASH sensory neurons in male attraction to ascaroside sex pheromones. Our findings lead to an integrated view in which the distinct physical properties of various mate-preference cues guide a flexible, stepwise behavioral program by which males assess multiple features of potential mates to optimize mate preference.

Pheromone-mediated command from the female to male clock induces and synchronizes circadian rhythms of the moth Spodoptera littoralis.

To extract any adaptive benefit, the circadian clock needs to be synchronized to the 24-h day-night cycles. We have investigated if it is a general property of the brain's circadian clock to recognize social interactions as external time givers. Sociosexual interactions with the opposite sex are universal, prevalent even in the lives of solitary animals. The solitary adult life of the Spodoptera littoralis moth is singularly dedicated to sex, offering an ideal context for exploring the impact of sociosexual cues on circadian timekeeping. We have identified specific olfactory cues responsible for social entrainment, revealing a surprisingly strong influence of pheromone-mediated remote sociosexual interactions on circadian rhythms. Males' free-running rhythms are induced and synchronized by the sex pheromone that the female releases in a rhythmic fashion, highlighting a hierarchical relation between the female and male circadian oscillators. Even a single pulse of the sex pheromone altered clock gene expression in the male brain, surpassing the effect of light on the clock. Our finding of a daytime-dependent, lasting impact of pheromone on male's courtship efficacy indicates that circadian timing in moths is a trait under sexual selection. We have identified specific components of the sex-pheromone blend that lack mate-attractive property but have powerful circadian effects, providing rationale for their continued retention by the female. We show that such volatiles, when shared across sympatric moth species, can trigger communal synchronization. Our results suggest that the sex pheromone released by female moths entrains males' behavioral activity rhythm to ensure synchronized timing of mating.

Behavioural consequences of intraspecific variability in a mate recognition signal.

Mate recognition is paramount for sexually reproducing animals, and many insects rely on cuticular hydrocarbons (CHCs) for close-range sexual communication. To ensure reliable mate recognition, intraspecific sex pheromone variability should be low. However, CHCs can be influenced by several factors, with the resulting variability potentially impacting sexual communication. While intraspecific CHC variability is a common phenomenon, the consequences thereof for mate recognition remain largely unknown. We investigated the effect of CHC variability on male responses in a parasitoid wasp showing a clear-cut within-population CHC polymorphism (three distinct female chemotypes, one thereof similar to male profiles). Males clearly discriminated between female and male CHCs, but not between female chemotypes in no-choice assays. When given a choice, a preference hierarchy emerged. Interestingly, the most attractive chemotype was the one most similar to male profiles. Mixtures of female CHCs were as attractive as chemotype-pure ones, while a female-male mixture negatively impacted male responses, indicating assessment of the entire, complex CHC profile composition. Our study reveals that the evaluation of CHC profiles can be strict towards 'undesirable' features, but simultaneously tolerant enough to cover a range of variants. This reconciles reliable mate recognition with naturally occurring variability.

Novel pheromone-mediated reproductive behaviour in the stag beetle, Lucanus cervus.

The iconic European stag beetle (Lucanus cervus) (Coleoptera: Lucanidae) is one of the largest terrestrial beetles in Europe. Due to decreasing population numbers, thought to be a consequence of habitat loss, this beetle has become a near-threatened species across much of Europe, and a reliable monitoring system is required to measure its future population trends. As part of a programme aimed at conserving UK populations, we have investigated the chemical ecology of the beetle, with a view to developing an efficient semiochemical-based monitoring system. Such a scheme will be beneficial not only in the UK but across the European range of the species, where the beetle is of conservation concern. Here, we report on a surprising discovery of a male-produced pheromone, which provokes initial sexual receptivity in females, and which has not been previously identified in the animal kingdom. Furthermore, we assign sex pheromone function to a previously described female-specific compound.

Pheromone traps and climate variations influence populations of Sahlbergella singularis (Hemiptera: Miridae) and associated damage of cocoa in Cameroon.

Knowledge of insect pest ecology and biology is important for maximizing crop protection and reducing crop losses. Currently, we lack an efficient control program for the cocoa mirid Sahlbergella singularis Haglund (Hemiptera: Miridae), the principal insect pest of cocoa in West and Central Africa. A 2-yr study was conducted in 11 plantations across Ayos and Konye, two of the largest cocoa growing areas of Cameroon. We evaluated the effects of mirid sex pheromone and climatic variations on mirid population dynamics and their associated cocoa damage. Sex pheromone traps caught 1.5-fold higher mirids in Ayos than in Konye, with more overall counts in 2015 than in 2016. Cocoa pod counts were also significantly higher in 2015 than in 2016 and were negatively correlated with temperature and relative humidity. In both localities, mirid populations and associated cocoa pod damage were suppressed in plantations where sex pheromone traps were used. Damage incidence was positively correlated with mirid counts, confirming that the cocoa pod is the preferential site for mirid feeding and reproduction. As such, damage incidence could be used as proxy for comparative mirid population level due to the mirid's cryptic habit. Of the recorded weather variables, only relative humidity was correlated (negatively) with damage severity. Our data on the relationships between damage caused by mirids and their population dynamics and sex pheromone trap catches suggest that an effective control strategy using mass trapping could be developed for mirid management in cocoa plantations.

Total synthesis of (6R,12R)-6,12-dimethylpentadecan-2-one, the sex pheromone of Diabrotica balteata LeConte.

Diabrotica balteata LeConte is one of the most important polyphagous agricultural pests. The sex pheromone of this pest was synthesized using Evans asymmetric alkylation, ring-opening reaction of (R)-2-methyloxirane, SN 2 alkylation of secondary tosylate, and coupling of chiral tosylate with Grignard reagent as central strategies. The sex pheromone prepared herein would be useful to control D. balteata.

Stored alcohol and fatty acid intermediates and the biosynthesis of sex pheromone aldehyde in the moth Chloridea virescens.

In most species of moths, the female produces and releases a volatile sex pheromone from a specific gland to attract a mate. Biosynthesis of the most common type of moth sex pheromone component (Type 1) involves de novo synthesis of hexadecanoate (16:Acyl), followed by modification to various fatty acyl intermediates, then reduction to a primary alcohol, which may be acetylated or oxidized to produce an acetate ester or aldehyde, respectively. Our previous work on the moth Chloridea virescens (Noctuidae) showed that females produce 90% of the major pheromone component, (Z)-11-hexadecenal (Z11-16:Ald), via a direct and rapid route of de novo biosynthesis with highly labile intermediates, and ca. 10% from an indirect route that likely mobilizes a pre-synthesized 16-carbon skeleton, possibly, (Z)-11-hexadecenoate (Z11-16:Acyl) or hexadecanoate (16:Acyl). In this paper, we use stable isotope tracer/tracee techniques to study the dynamics of the precursor alcohol (Z)-11-hexadecenol (Z11-16:OH) and stores of Z11-16:Acyl and 16:Acyl to determine their roles in biosynthesis of Z11-16:Ald. We found: (i) that intracellular Z11-16:OH is synthesized at roughly the same rate as Z11-16:Ald, indicating that translocation and oxidation of this moiety does not rate limit biosynthesis of Z11-16:Ald, (ii) intracellular Z11-16:OH consists of two pools, a highly labile one rapidly translocated out of the cell and converted to Z11-16:Ald, and a less labile one that mostly remains in gland cells, (iii) during pheromone biosynthesis, net stores of Z11-16:Acyl increase, suggesting it is not the source of Z11-16:Ald produced by the indirect route, and (iv) no evidence for the gland synthesizing stored 16:Acyl prior to (up to 2 days before eclosion), or after, synthesis of pheromone commenced, suggesting the bulk of this stored moiety is synthesized elsewhere and transported to the gland prior to gland maturation. Thus, the pheromone gland of C. virescens produces very little stored fat over its functional lifetime, being optimized to produce sex pheromone.

(R)-(+)-γ-Decalactone is Conserved in North America as a Pheromone Component of Osmoderma eremicola (Coleoptera: Scarabaeidae) and a Kairomone of Elater abruptus (Coleoptera: Elateridae).

The scarab genus Osmoderma (Coleoptera: Scarabaeidae) includes several large species called hermit beetles that develop within dead and decaying hardwood trees. Males of at least three Palearctic species produce the aggregation-sex pheromone (R)-(+)-γ-decalactone, including the endangered O. eremita (Scopoli). However, hermit beetles have received less attention in the western hemisphere, resulting in a large gap in our knowledge of the chemical ecology of Nearctic species. Here, we identify (R)-( +)-γ-decalactone as the primary component of the aggregation-sex pheromone of the North American species Osmoderma eremicola (Knoch). Field trials at sites in Wisconsin and Illinois revealed that both sexes were attracted to lures containing (R)-(+)-γ-decalactone or the racemate, but only males of O. eremicola produced the pheromone in laboratory bioassays, alongside an occasional trace of the chain-length analog γ-dodecalactone. Females of the congener O. scabra (Palisot de Beauvois) were also significantly attracted by γ-decalactone, suggesting further conservation of the pheromone, as were females of the click beetle Elater abruptus Say (Coleoptera: Elateridae), suggesting that this compound may have widespread kairomonal activity. Further research is needed to explore the behavioral roles of both lactones in mediating behavioral and ecological interactions among these beetle species.

Structure, Synthesis, and Bioassays of Sex Pheromone for Pyrausta machaeralis (Lepidoptera: Crambidae).

The leaf skeletonizer, Pyrausta machaeralis (Lepidoptera: Crambidae), is a serious insect pest of teak (Tectona grandis) in China. The application of insect pheromones is widely applied as an environmentally friendly technology for integrated pest management (IPM). In the present study, crude extracts of sex pheromone glands of calling P. machaeralis females were collected and then analyzed using gas chromatography/electroantennographic detection (GC/EAD) and gas chromatography-mass spectrometry (GC-MS). The combination of infrared spectroscopy (IR) and nuclear magnetic resonance (NMR) spectrometry was used for structure identification. Afterward, their electrophysiological and behavioral activity was evaluated in the laboratory and field. Herein, we eventually determined two active components, E-11-tetradecenyl acetate (E11-14:Ac) and Z-11-tetradecenyl acetate (Z11-14:Ac), at a ratio of 96:4, as the sex pheromone of P. machaeralis. The identification of sex pheromones would facilitate the development of efficient strategies for monitoring and controlling the field populations of P. machaeralis.

Identification of the female sex pheromone of Bastilla arctotaenia (Lepidoptera: Erebidae).

(3Z,6Z,9Z)-3,6,9-henicosatriene was identified as a major component of female sex pheromone of Bastilla arctotaenia (Lepidoptera: Erebidae), a pest of cultivated roses, by gas chromatograph-electroantennographic detector( GC-EAD) and gas chromatograph/mass spectrometry (GC/MS) analyses. The single (3Z,6Z,9Z)-3,6,9-henicosatriene (1.0 mg/lure) successfully attracted B. arctotaenia males in the field.

Pheromonal variation and mating between two mitotypes of fall armyworm (Spodoptera frugiperda) in Africa.

In the Americas, the fall armyworm (Spodoptera frugiperda) exists in two genetically distinct strains, the corn (C) and rice (R) strains. Despite their names, these strains are not associated with host plant preferences but have been shown to vary in pheromone composition and male responses. Recently, S. frugiperda was detected in Africa as an invasive species, but knowledge about variation in strain types, pheromone composition and inter-strain mating of populations of the pest in the continent has not been fully examined. Therefore, this study aimed to investigate variations, if any in the pheromone composition of female moths, male moth responses, and mating between C and R mitotypes of S. frugiperda populations in Kenya, as well as their geographic distribution. Strains (mitotypes) of S. frugiperda were identified using mitochondrial DNA (mtDNA) markers, and their pheromonal composition determined by coupled gas chromatography-mass spectrometric (GC-MS) analysis. Male moth responses to these compounds were evaluated using GC-electroantennographic detection (EAD), electroantennogram (EAG), and wind tunnel assays. Oviposition assays were used to determine whether R and C mitotype moths could mate and produce eggs. The results showed that both the R and C mitotypes were present, and there were no statistically significant differences in their distribution across all sampled locations. Five pheromone compounds including (Z)-7-dodecenyl acetate (Z7-12:OAc), (Z)-7-tetradecenyl acetate (Z7-14:OAc), (Z)-9-tetradecenyl acetate (Z9-14:OAc), (Z)-11-tetradecenyl acetate (Z11-14:OAc) and (Z)-11-hexadecenyl acetate (Z11-16:OAc), were detected in the pheromone glands of female moths of both mitotypes, with Z9-14:OAc being the most abundant. The relative percentage composition of Z9-14:OAc was similar in both mitotypes. However, the R mitotype had a 2.7 times higher relative percentage composition of Z7-12:OAc compared to the C mitotype moth, while the C mitotype moth had a 2.4 times higher relative percentage composition of Z11-16:OAc than the R mitotype moth. Male moths of both mitotypes exhibited similar responses to the pheromone compounds, showing the strongest responses to Z9-14:OAc and Z7-12:OAc in electrophysiological and behavioural assays. There was mating between R and C mitotypes with egg production comparable to mating within the same mitotype. Our results revealed that differences between the two S. frugiperda mitotypes are characterized by female moth pheromone composition rather than male moth responses to the pheromones, and that this does not prevent hybridisation between the mitotypes, which may have implications for their management.

Circabidian rhythm of sex pheromone reception in a scarab beetle.

Animals have endogenous clocks that regulate their behavior and physiology. These clocks rely on environmental cues (time givers) that appear approximately every 24 h due to the Earth's rotation; thus, most insects exhibit a circadian rhythm. One notable exception is the scarab beetle, Holotrichia parallela, a severe agricultural pest in China, Japan, South Korea, and India. Females emerge from the soil every other night, reach the canopy of host plants, evert an abdominal gland, and release a pheromone bouquet comprising l-isoleucine methyl ester (LIME) and l-linalool. To determine whether this circa'bi'dian rhythm affects the olfactory system, we aimed to identify H. parallela sex pheromone receptor(s) and study their expression patterns. We cloned 14 odorant receptors (ORs) and attempted de-orphanizing them in the Xenopus oocyte recording system. HparOR14 gave robust responses to LIME and smaller responses to l-linalool. Structural modeling, tissue expression profile, and RNAi treatment followed by physiological and behavioral studies support that HparOR14 is a sex pheromone receptor-the first of its kind discovered in Coleoptera. Examination of the HparOR14 transcript levels throughout the adult's life showed that on sexually active days, gene expression was significantly higher in the scotophase than in the photophase. Additionally, the HparOR14 expression profile showed a circabidian rhythm synchronized with the previously identified pattern of sex pheromone emission. 48 h of electroantennogram recordings showed that responses to LIME were abolished on non-calling nights. In contrast, responses to the green leaf volatile (Z)-3-henexyl acetate remained almost constant throughout the recording period.

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.

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.

The greening-causing agent alters the behavioral and electrophysiological responses of the Asian citrus psyllid to a putative sex pheromone.

The Asian Citrus Psyllid (ACP), Diaphorina citri, is a vector of the pathological bacterium Candidatus Liberibacter asiaticus (CLas), which causes the most devastating disease to the citrus industry worldwide, known as greening or huanglongbing (HLB). Earlier field tests with an acetic acid-based lure in greening-free, 'Valencia' citrus orange groves in California showed promising results. The same type of lures tested in São Paulo, Brazil, showed unsettling results. During the unsuccessful trials, we noticed a relatively large proportion of females in the field, ultimately leading us to test field-collected males and females for Wolbachia and CLas. The results showed high rates of Wolbachia and CLas infection in field populations. We then compared the olfactory responses of laboratory-raised, CLas-free, and CLas-infected males to acetic acid. As previously reported, CLas-uninfected males responded to acetic acid at 1 µg. Surprisingly, CLas-infected males required 50 × higher doses of the putative sex pheromone, thus explaining the failure to capture CLas-infected males in the field. CLas infection was also manifested in electrophysiological responses. Electroantennogram responses from CLas-infected ACP males were significantly higher than those obtained with uninfected males. To the best of our knowledge, this is the first report of a pathogen infection affecting a vector's response to a sex attractant.

Characterization of the pheromone receptors in Mythimna loreyi reveals the differentiation of sex pheromone recognition in Mythimna species.

Pheromone receptors (PRs) are key proteins in the molecular mechanism of pheromone recognition, and exploring the functional differentiation of PRs between closely related species helps to understand the evolution of moth mating systems. Pheromone components of the agricultural pest Mythimna loreyi have turned into (Z)-9-tetradecen-1-yl acetate (Z9-14:OAc), (Z)-7-dodecen-1-yl acetate (Z7-12:OAc), and (Z)-11-hexadecen-1-yl acetate, while the composition differs from that of M. separata in the genus Mythimna. To understand the molecular mechanism of pheromone recognition, we sequenced and analyzed antennal transcriptomes to identify 62 odorant receptor (OR) genes. The expression levels of all putative ORs were analyzed using differentially expressed gene analysis. Six candidate PRs were quantified and functionally characterized in the Xenopus oocytes system. MlorPR6 and MlorPR3 were determined to be the receptors of major and minor components Z9-14:OAc and Z7-12:OAc. MlorPR1 and female antennae (FA)-biased MlorPR5 both possessed the ability to detect pheromones of sympatric species, including (Z,E)-9,12-tetradecadien-1-ol, (Z)-9-tetradecen-1-ol, and (Z)-9-tetradecenal. Based on the comparison of PR functions between M. loreyi and M. separata, we analyzed the differentiation of pheromone recognition mechanisms during the evolution of the mating systems of 2 Mythimna species.

Evidence for a role of SNMP2 and antennal support cells in sensillum lymph clearance processes of moth pheromone-responsive sensilla.

In insect antenna, following the activation of olfactory sensory neurons, odorant molecules are inactivated by enzymes in the sensillum lymph. How the inactivation products are cleared from the sensillum lymph is presently unknown. Here we studied the role of support cells (SCs) and the so-called sensory neuron membrane protein 2 (SNMP2), a member of the CD36 family of lipid transporters abundantly expressed in SCs, in sensillum lymph clearance processes in the moths Heliothis virescens and Bombyx mori. In these species, the sex pheromone components are inactivated to long-chain fatty acids. To approach a role of SNMP2 in the removal of such inactivation products, we analyzed the uptake of a fluorescent long-chain fatty acid analog into a newly generated HvirSNMP2-expressing cell line. We found an increased uptake of the analog into SNMP2-cells compared to control cells, which could be blocked by the CD36 protein inhibitor, SSO. Furthermore, analyses of sensilla from antenna treated with the fatty acid analog indicated that SNMP2-expressing SCs are able to take up fatty acids from the sensillum lymph. In addition, sensilla from SSO-pretreated antenna of B. mori showed reduced removal of the fluorescent analog from the sensillum lymph. Finally, we revealed that SSO pretreatment of male silkmoth antenna significantly prolonged the duration of the female pheromone-induced wing-fluttering behavior, possibly as a result of impaired lymph clearance processes. Together our findings in H. virescens and B. mori support a pivotal role of olfactory SCs in sensillum lymph maintenance processes and suggest an integral role of SNMP2 in the removal of lipophilic "waste products" such as fatty acids resulting from sex pheromone inactivation.

Gymnandrosoma aurantianum (Lepidoptera: Tortricidae): Calling Rhythm and Effect of Age on Female Sex Pheromone Titer and Male Antennal Responses.

The macadamia nut borer, Gymnandrosoma aurantianum Lima (Lepidoptera: Tortricidae), is the main pest of Macadamia integrifolia Maiden & Betche (Proteales: Proteaceae) in Central America. Despite its importance, there is little information on its reproductive behavior. We studied the calling behavior and effect of age on female sex pheromone production and male antennal response to the pheromone. Calling behavior of G. aurantianum females occurred at the onset of scotophase, between 17:00 and 20:00 h. The mean time spent calling was 64 min. It was observed that 30% of the 50 females exhibited a calling posture characterized by extrusion of the pheromone gland from the tip of the abdomen and they spread their wings slightly. Female age had no influence on the sex pheromone production, while male antennal response increased with age and pheromone dosage. The information obtained in this work contributes to understanding the sexual behavior of G. aurantianum and thus to the creation of strategies to control this important pest in macadamia.

Molecular mechanism of sex pheromone perception in male Mythimna loreyi revealed by in vitro system.

BACKGROUND: Mythimna loreyi is an important agricultural pest with a sensitive sex pheromone communication system. To clarify the pheromone binding proteins (PBPs) and pheromone receptors (PRs) involved in sex pheromone perception is important for both understanding the molecular olfactory mechanism and developing a new pest control strategy in M. loreyi. RESULTS: First, the electroantennogram (EAG) assay showed that male M. loreyi displayed the highest response to the major sex pheromone component Z9-14:Ac, and higher responses to two minor components, Z7-12:Ac and Z11-16:Ac. Second, the fluorescence competition binding assay showed that PBP1 bound all three pheromones and other tested compounds with high or moderate affinity, while PBP2 and PBP3 each bound only one pheromone component and few other compounds. Third, functional study using the Xenopus oocyte system demonstrated that, of the six candidate PRs, PR2 was weakly sensitive to the major pheromone Z9-14:Ac, but was strongly sensitive to pheromone analog Z9-14:OH; PR3 was strongly and specifically sensitive to a minor component Z7-12:Ac; PR4 and OR33 were both weakly sensitive to another minor component, Z11-16:Ac. Finally, phylogenetic relationship and ligand profiles of PRs were compared among six species from two closely related genera Mythimna and Spodoptera, suggesting functional shifts of M. loreyi PRs toward Spodoptera PRs. CONCLUSION: Functional differentiations were revealed among three PBPs and six PRs in sex pheromone perception, laying an important basis for understanding the molecular mechanism of sex pheromone perception and for developing new control strategies in M. loreyi. © 2023 Society of Chemical Industry.