Neural Circuits of Sexual Behavior in Caenorhabditis elegans.

The recently determined connectome of the Caenorhabditis elegans adult male, together with the known connectome of the hermaphrodite, opens up the possibility for a comprehensive description of sexual dimorphism in this species and the identification and study of the neural circuits underlying sexual behaviors. The C. elegans nervous system consists of 294 neurons shared by both sexes plus neurons unique to each sex, 8 in the hermaphrodite and 91 in the male. The sex-specific neurons are well integrated within the remainder of the nervous system; in the male, 16% of the input to the shared component comes from male-specific neurons. Although sex-specific neurons are involved primarily, but not exclusively, in controlling sex-unique behavior-egg-laying in the hermaphrodite and copulation in the male-these neurons act together with shared neurons to make navigational choices that optimize reproductive success. Sex differences in general behaviors are underlain by considerable dimorphism within the shared component of the nervous system itself, including dimorphism in synaptic connectivity.

Metabolic pathways that mediate the effects of food deprivation on reproductive behavior in female Drosophila melanogaster.

In most species studied, energy deficits inhibit female reproductive behavior, but the location and nature of energy sensors and how they affect behavior are unknown. Progress has been facilitated by using Drosophila melanogaster, a species in which reproduction and food availability are closely linked. Adult males and females were either fed or food deprived (FD) and then tested in an arena with a fed, opposite-sex conspecific with no food in the testing arena. Only FD females (not FD males) significantly decreased their copulation rate and increased their copulation latency, and the effects of FD were prevented in females fed either yeast alone or glucose alone, but not sucralose alone, cholesterol alone, or amino acids alone. It is well-known that high-fat diets inhibit copulation rate in this species, and the effects of FD on copulation rate were mimicked by treatment with an inhibitor of glucose but not free fatty acid oxidation. The availability of oxidizable glucose was a necessary condition for copulation rate in females fed either yeast alone or fed a nutritive fly medium, which suggests that the critical component of yeast for female copulation rate is oxidizable glucose. Thus, female copulation rate in D. melanogaster is sensitive to the availability of oxidizable metabolic fuels, particularly the availability of oxidizable glucose or substrates/byproducts of glycolysis.NEW & NOTEWORTHY Copulation rate was decreased in food-deprived female but not in male adults when tested without food in the testing arena. Copulation rate was 1) maintained by feeding glucose alone, yeast alone, nutritive medium lacking yeast, but not sucralose, amino acids, or cholesterol alone; 2) decreased by inhibition of glycolysis in females fed either nutritive medium or yeast alone; and 3) not affected by inhibition of fatty acid oxidation. Thus, female copulation rate was linked to glycolytic status.

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.

Sniffing behavior of semi free-ranging Barbary macaques (Macaca sylvanus).

Olfaction is one of the evolutionarily oldest senses and plays a fundamental role in foraging and social interactions across mammals. In primates, the role of olfaction is now well recognized, but better investigated in strepsirrhine and platyrrhine primates than in catarrhines. We observed the sniffing behavior of semi-free ranging Barbary macaques, Macaca sylvanus, at Affenberg Salem, Germany, to assess how frequently macaques sniff and in which contexts, and how sniffing is affected by sex and age. Focal observations of 24 males and 24 females aged 1-25 years showed that Barbary macaques sniffed, on average, 5.24 times per hour, with more than 80% of sniffs directed at food. Irrespective of the context, younger individuals sniffed more often than older ones. Females' sniffs were more often directed at food than male sniffs, while males sniffed more often than females in a social context. Sniffs at conspecifics occurred primarily in a sexual context, with 70% of social sniffs directed at female anogenital swellings performed by males. Of the observed 176 anogenital inspections, 51 involved sniffing of the swelling. Olfactory inspections were followed by copulation significantly less often than merely visual inspections, suggesting that anogenital odors may play a role in male mating decisions, but the role of olfaction in sexual interactions warrants further investigations. In sum, results show that Barbary macaques routinely use olfaction during feeding, but also in a socio-sexual context, corroborating the relevance of the olfactory sense in the lives of catarrhine primates.

Notes on the Courtship and Captive Reproduction of the Southern Vietnam Box Turtle, Cuora picturata, at the Turtle Conservation Center in Cuc Phuong National Park, Vietnam.

Endemic to only three known provinces in South Central Vietnam, Cuora picturata have faced extensive collection pressures for the food, medicinal, and pet trades. Further exacerbating their decline is the lack of protected areas where wild populations exist, with only one known population occurring within a protected area. With threats to wild populations persisting, the development of an assurance colony has been prioritized for C. picturata. The Asian Turtle Program of Indo-Myanmar Conservation has been operating the Turtle Conservation Center (TCC) in Cuc Phuong National Park since 1998. From 2020 to 2022, 24 C. picturata were acquired by the TCC from the illegal trade. With no ideal repatriation plan, these animals provided an opportunity for developing a captive assurance colony at the TCC to safeguard the species from extinction. As semi-aquatic species, box turtles in the genus Cuora are a unique group that shows variations in habitat use and behavior between species. Herein, we documented some specific courtship behaviors not yet described for C. picturata, providing insight into the unique reproductive behaviors of this species.

Postcopulatory Behavior of Tephritid Flies.

Mating produces profound changes in the behavior of female flies, such as an increase in oviposition, reduction in sexual receptivity, increase in feeding, and even excretion. Many of these changes are produced by copulation, sperm, and accessory gland products that males transfer to females during mating. Our knowledge on the function of the male ejaculate and its effect on female insects is still incipient. In this article, we review peri- and postcopulatory behaviors in tephritid flies. We address the effects of male copulatory behavior; copula duration; and the male ejaculate, sperm, and accessory gland products on female remating behavior. Many species from these families are pests of economic importance; thus, understanding male mating effects on female behavior contributes to both developing more effective environmentally friendly control methods and furthering our understanding of evolutionary implications of intersexual competition and sexual conflict.

Multiple mechanisms for inbreeding avoidance used simultaneously in a wild ape.

Mating with close kin can have considerable negative fitness consequences, which are expected to result in selective pressure for inbreeding avoidance mechanisms, such as dispersal, mate choice and post-copulatory biases. Captive studies have suggested that inbreeding avoidance through mate choice is far less widespread than expected and may be absent where other mechanisms already limit inbreeding. However, few studies have examined multiple mechanisms of inbreeding avoidance simultaneously, particularly in the wild. We use 13 years of detailed dispersal, copulation and paternity data from mountain gorillas to examine inbreeding avoidance. We find that partial dispersal of both sexes results in high kinship in multimale groups, but that copulations between close kin occur 40% less than expected. We find strong kin discrimination in mate choice, with significant avoidance of maternal kin but more limited avoidance of paternal kin. We find no evidence for post-copulatory inbreeding avoidance. Our analyses support familiarity-based mechanisms of kin identification and age-based avoidance that limits mating between fathers and daughters in their natal group. Our findings demonstrate that multiple complementary mechanisms for inbreeding avoidance can evolve in a single species and suggest that inbreeding avoidance through mate choice may enable more flexible dispersal systems to evolve.

Alternative reproductive tactics of unflanged and flanged male orangutans revisited.

In many slowly developing mammal species, males reach sexual maturity well before they develop secondary sexual characteristics. Sexually mature male orangutans have exceptionally long periods of developmental arrest. The two male morphs have been associated with behavioral alternative reproductive tactics, but this interpretation is based on cross-sectional analyses predominantly of Northwest Sumatran populations. Here we present the first longitudinal analyses of behavioral changes of 10 adult males that have been observed in both unflanged and flanged morph. We also analyzed long-term behavioral data on an additional 143 individually identified males from two study sites, Suaq (Sumatra, Pongo abelii) and Tuanan (Borneo, Pongo pygmaeus wurmbii), to assess male mating tactics cross-sectionally in relation to population, male morph (unflanged and flanged), and other socio-ecological factors. Both our longitudinal and cross-sectional results confirm and refine previous cross-sectional accounts of the differences in mating tactics between the unflanged and the flanged male morphs. In the unflanged morph, males exhibit higher sociability, particularly with females, and higher rates of both copulation and sexual coercion than in the flanged morph. Based on our results and those of previous studies showing that females prefer flanged males, and that flanged males have higher reproductive success, we conclude that unflanged males face a trade-off between avoiding male-male contest competition and gaining mating access to females, and thus follow a "best-of-a-bad-job" mating strategy.

Effects of different diets on Aedes aegypti adults: improving rearing techniques for sterile insect technique.

The aim was to evaluate the effect of different energy diets available in adulthood on the longevity, dispersal capacity and sexual performance of Aedes aegypti produced under a mass-rearing system. To evaluate the effects of diets in relation to the survival of the adult male insects of Ae. aegypti, six treatments were used: sucrose at a concentration of 10%, as a positive control (sack10); starvation, as a negative control (starvation); sucrose at a concentration of 20% associated with 1 g/l of ascorbic acid (sac20vitC); wild honey in a concentration of 10% (honey10); demerara sugar in a 10% concentration (demerara10); and sucrose at a concentration of 20% associated with 1 g/l of ascorbic acid and 0.5 g/l of amino acid proline (sac20vitCPr). Each treatment had 16 cages containing 50 adult males. For the tests of flight ability and propensity to copulation, five treatments were used (saca10; sac20vitC; mel10; demerara10; and sac20vitCPr), with males each for flight ability and females copulated by a single male for copulation propensity. The diet composed of sucrose at a concentration of 20% associated with ascorbic acid, as an antioxidant, improved the survival, flight ability and propensity to copulate in Ae. aegypti males under mass-rearing conditions, and may be useful to enhance the performance of sterile males, thus improving the success of sterile insect technique programmes.

Latrophilin-1 drives neuron morphogenesis and shapes chemo- and mechanosensation-dependent behavior in C. elegans via a trans function.

Latrophilins are highly conserved Adhesion GPCRs playing essential roles in the mammalian nervous system and are associated with severe neurological disorders. Recently, it has been shown that murine Latrophilins mediate classical G-protein signals to drive synaptogenesis. However, there is evidence that Latrophilins in the nematode Caenorhabditis elegans can also function independently of their seven-transmembrane domain and C terminus (trans function). Here, we show that Latrophilin-1 acts in trans to mediate morphogenesis of sensory structures in the C. elegans nervous system. This trans function is physiologically relevant in copulation behavior. Detailed expression and RNA-Seq analyses revealed specific LAT-1-positive neurons and first insights into the genetic network that is modulated by the receptor function. We conclude that 7TM-independent functions of Latrophilins are essential for neuronal physiology, possibly complementing canonical functions via G protein-mediated signaling.

The genetic basis of variation in sexual aggression: Evolution versus social plasticity.

Male sexual aggression towards females is a form of sexual conflict that can result in increased fitness for males through forced copulations (FCs) or coercive matings at the cost of female lifetime fitness. We used male fruit flies (Drosophila melanogaster) as a model system to uncover the genomic contributions to variation in FC, both due to standing variation in a wild population, and due to plastic changes associated with variation in social experience. We used RNAseq to analyse whole-transcriptome differential expression (DE) in male head tissue associated with evolved changes in FC from lineages previously selected for high and low FC rate and in male flies with varying FC rates due to social experience. We identified hundreds of genes associated with evolved and plastic variation in FC, however only a small proportion (27 genes) showed consistent DE due to both modes of variation. We confirmed this trend of low concordance in gene expression effects across broader sets of genes significant in either the evolved or plastic analyses using multivariate approaches. The gene ontology terms neuropeptide hormone activity and serotonin receptor activity were significantly enriched in the set of significant genes. Of seven genes chosen for RNAi knockdown validation tests, knockdown of four genes showed the expected effect on FC behaviours. Taken together, our results provide important information about the apparently independent genetic architectures that underlie natural variation in sexual aggression due to evolution and plasticity.

Sperm competition intensity affects sperm precedence patterns in a polyandrous gift-giving spider.

Sperm competition drives traits that enhance fertilization success. The amount of sperm transferred relative to competitors is key for attaining paternity. Female reproductive morphology and male mating order may also influence fertilization, however the outcome for sperm precedence under intense sperm competition remains poorly understood. In the polyandrous spider Pisaura mirabilis, males offer nuptial gifts which prolong copulation and increase sperm transfer, factors proposed to alter sperm precedence patterns under strong sperm competition. First, we assessed the degree of female polyandry by genotyping wild broods. A conservative analysis identified up to four sires, with a mean of two sires per brood, consistent with an optimal mating female rate. Then we asked whether intense sperm competition shifts sperm precedence patterns from first male priority, as expected from female morphology, to last male advantage. We varied sexual selection intensity experimentally and determined competitive fertilization outcome by genotyping broods. In double matings, one male monopolised paternity regardless of mating order. A mating order effect with first male priority was revealed when females were mated to four males, however this effect disappeared when females were mated to six males, probably due to increased sperm mixing. The proportion of males that successfully sired offspring drastically decreased with the number of competitors. Longer copulations translated into higher paternity shares independently of mating order, reinforcing the advantage of traits that prolong copulation duration under intense competition, such as the nuptial gift. Sperm competition intensity enhances the impact of competitive sexual traits and imposes multiple effects on paternity.

Prolactin promotes parental responses and alters reproductive axis gene expression, but not courtship behaviors, in both sexes of a biparental bird.

Prolactin, a hormone involved in vertebrate parental care, is hypothesized to inhibit reproductive hypothalamic-pituitary-gonadal (HPG) axis activity during parenting, thus maintaining investment in the current brood as opposed to new reproductive efforts. While prolactin underlies many parental behaviors in birds, its effects on other reproductive behaviors, such as courtship, remain unstudied. How prolactin affects neuropeptide and hormone receptor expression across the avian HPG axis also remains unknown. To address these questions, we administered ovine prolactin (oPRL) or a vehicle control to both sexes in experienced pairs of the biparental rock dove (Columba livia), after nest removal at the end of incubation. We found that oPRL promoted parental responses to novel chicks and stimulated crop growth compared to controls, consistent with other studies. However, we found that neither courtship behaviors, copulation rates nor pair maintenance differed with oPRL treatment. Across the HPG, we found oPRL had little effect on gene expression in hypothalamic nuclei, but increased expression of FSHB and hypothalamic hormone receptor genes in the pituitary. In the gonads, oPRL increased testes size and gonadotropin receptor expression, but did not affect ovarian state or small white follicle gene expression. However, the oviducts of oPRL-treated females were smaller and had lower estrogen receptor expression compared with controls. Our results highlight that some species, especially those that show multiple brooding, may continue to express mating behavior despite elevated prolactin. Thus, mechanisms may exist for prolactin to promote investment in parental care without concurrent inhibition of reproductive function or HPG axis activity.

The first observation of copulation in Andean catfish Astroblepus ubidiai (Siluriformes, Astroblepidae), in Lago San Pablo, Imbabura, Ecuador.

The reproductive behaviour of the Andean catfish Astroblepus ubidiai was observed directly and with an underwater camera in a spring located next to Lago San Pablo in the province of Imbabura, Ecuador, in late 2015 and early 2016. Five mating attempts were observed, four of which were video-taped, and two of which resulted in complete copulation. During copulation, the male grabs the female from the top of the head with its buccal disc and pelvic fins and, from this position, bends its body forming a "C" to reach the genital pore of the female and introduce its copulatory organ. Once copulation is complete, the female swims away rapidly. Details of the copulation are described here through illustrations.

Female wound records suggest mating periods for the Caribbean reef shark at an insular marine protected area from the Equatorial Atlantic Ocean.

Despite being one of the most abundant, economically significant, reef-associated shark species, little is known about the reproductive aspects of the Caribbean reef shark (Carcharhinus perezi). In the present study the authors report the first evidence of mating wounds and scars in female Caribbean reef sharks at the Fernando de Noronha Archipelago, a remote marine protected area in the South Atlantic Ocean. Data from four females suggest this species mates mainly during the austral summer, between February and March. Given that the archipelago has been previously described as a nursery ground for the Caribbean reef shark, these results add information about the reproductive cycle of this species in the equatorial Atlantic Ocean.

Glans inflation morphology and female cloaca copulatory interactions of the male American alligator phallus†.

The phallic glans of the American alligator (Alligator mississippiensis) is the distal termination of the semen-conducting sulcus spermaticus and during copulation has the closest, most intimate mechanical interactions with the female urodeum, the middle cloacal chamber that contains the opening to the vaginal passages and oviducts. However, the details of this interface leading to insemination and gamete uptake are unclear. Here, we: (1) histologically characterize the underlying tissue types and morphologically quantify the shape changes associated with glans inflation into the copulatory conformation, (2) digitally reconstruct from MRI the 3D shape of functional tissue compartments, and (3) diffusible iodine-based contrast-enhanced computed tomography image the copulatory fit between male phallus and female cloaca. We discuss these results in relation to tissue type material properties, the transfer on intromittent forces, establishing potential copulatory lock, inflated glans volume scaling with body mass/length, the mechanics of semen targeting and insemination, and potential female cryptic choice impacting multiple clutch paternity. In part, this study further clarifies the phallic morphological variation observed among crocodylians and begins to investigate the role(s) these divergent male forms play during copulation interacting with female cloacal forms to increase reproductive success.

Testing potential mechanisms of conspecific sperm precedence in Drosophila pseudoobscura.

Drosophila pseudoobscura females that co-occur with sister species D. persimilis show elevated fertilization by conspecific sperm when they mate with both a heterospecific and a conspecific male. This phenomenon, known as conspecific sperm precedence (CSP), has evolved as a mechanism to avoid maladaptive hybridization with D. persimilis. In this study, we assessed pericopulatory (during mating) and postcopulatory (after mating) traits in crosses with sympatric or allopatric D. pseudoobscura females and conspecific or heterospecific males to evaluate potential mechanisms of CSP in this system. We observed shorter copulation duration in crosses with sympatric females, but found no difference in quantity of sperm transferred or female reproductive tract toxicity between sympatry and allopatry. Our data show some support for the hypothesis that parasperm, a short, sterile sperm morph, can protect fertile eusperm from the D. pseudoobscura female reproductive tract, though it is unclear how this might affect patterns of sperm use in sympatry vs. allopatry. Overall, these results suggest that copulation duration could potentially contribute to the elevated CSP observed in sympatry.

Functional morphology of immature mating in a widow spider.

BACKGROUND: Mating generally occurs after individuals reach adulthood. In many arthropods including spiders, the adult stage is marked by a final moult after which the genitalia are fully developed and functional. In several widow spider species (genus Latrodectus), however, immature females may mate a few days before they moult to adulthood, i.e. in their late-subadult stage. While the "adult" mating typically results in cannibalism, males survive the "immature" mating. During both "immature" and "adult" matings, males leave parts of their paired copulatory organs within female genitalia, which may act as mating plugs. To study potential costs and benefits of the two mating tactics, we investigated female genital morphology of the brown widow spider, L. geometricus. Light microscopy, histology and micro-computed tomography of early-subadult, late-subadult and adult females were conducted to determine the overall pattern of genital maturation. We compared genitalia of mated late-subadult and adult females to reveal potential differences in the genitalic details that might indicate differential success in sperm transfer and different environments for sperm storage and sperm competition. RESULTS: We found that the paired sperm storage organs (spermathecae) and copulatory ducts are developed already in late-subadult females and host sperm after immature mating. However, the thickness of the spermathecal cuticle and the staining of the secretions inside differ significantly between the late-subadult and adult females. In late-subadult females mating plugs were found with higher probability in both spermathecae compared to adult females. CONCLUSIONS: Sperm transfer in matings with late-subadult females follows the same route as in matings with adult females. The observed differences in the secretions inside the spermathecae of adult and late-subadult females likely reflect different storage conditions for the transferred sperm which may lead to a disadvantage under sperm competition if the subadult female later re-mates with another male. However, since males mating with late-subadult females typically transfer sperm to both spermathecae they might benefit from numerical sperm competition as well as from monopolizing access to the female sperm storage organs. The assessment of re-mating probability and relative paternity will clarify the costs and benefits of the two mating tactics in light of these findings.

Rapid stress hardening in the Antarctic midge improves male fertility by increasing courtship success and preventing decline of accessory gland proteins following cold exposure.

Rapid hardening is a process that quickly improves an animal's performance following exposure to potentially damaging stress. In this study of the Antarctic midge, Belgica antarctica (Diptera, Chironomidae), we examined how rapid hardening in response to dehydration (RDH) or cold (RCH) improves male pre- and post-copulatory function when the insects are subsequently subjected to a damaging cold exposure. Neither RDH nor RCH improved survival in response to lethal cold stress, but male activity and mating success following sublethal cold exposure were enhanced. Egg viability decreased following direct exposure of the mating males to sublethal cold but improved following RCH and RDH. Sublethal cold exposure reduced the expression of four accessory gland proteins, while expression remained high in males exposed to RCH. Though rapid hardening may be cryptic in males, this study shows that it can be revealed by pre- and post-copulatory interactions with females.

Reproductive interference between Aedes albopictus and Aedes flavopictus at a place of their origin.

Aedes (Stegomyia) albopictus and Aedes (Stegomyia) flavopictus are related species that have overlapping distributions from southern to central Japan. To understand how they interact, we studied reproductive interference between them, particularly focusing on the body size difference between the mating pair. Here, we examined the effects of conspecific, heterospecific and double mating (i.e. heterospecific mating followed by conspecific mating) on copulation duration, egg production and hatchability of eggs using mosquitoes that varied in body size. Females mated only with heterospecific males produced few viable eggs, indicating that post-mating isolation is almost complete. When mated with heterospecific males before conspecific mating, the production of viable eggs was lower than when mated only with conspecific males, revealing the occurrence of reproductive interference. The degree of reproductive interference was larger in Ae. flavopictus than in Ae. albopictus when the female size was small but did not differ between them when the female size was large. Aedes albopictus females appear to be able to distinguish Ae. flavopictus males from conspecific males and larger females are more successful in the rejection of heterospecific males. On the other hand, Ae. flavopictus were not able to discriminate between conspecific and heterospecific males.