Mating unplugged: a model for the evolution of mating plug (dis-)placement

Mating plugs are male-derived structures that may impede female remating by physically obstructing the female genital tract. Although mating plugs exist in many taxa, the forces shaping their evolution are poorly understood. A male can clearly benefit if his mating plug secures his paternity. It is unclear, however, how plug efficacy can be maintained over evolutionary time in the face of counteracting selection on males' ability to remove any plugs placed by their rivals. Here, I present a game-theory model and a simulation model to address this problem. The models predict that evolutionarily stable levels of mating-plug efficacy should be high when (1) the number of mating attempts per female is low; (2) the sex ratio is male-biased, and (3) males are sperm-limited. I discuss these results in the light of empirical data.     

Genital damage in the orb-web spider Argiope bruennichi (Araneae: Araneidae) increases paternity success

The morphology of male genitalia often suggests functions besidessperm transfer that may have evolved under natural or sexualselection. In several species of sexually cannibalistic spiders,males damage their paired genitalia during mating, limitingthem to one copulation per pedipalp. Using a triple-mating experiment,we tested if genital damage in the orb-web spider Argiope bruennichiincreases male fitness either through facilitating his escapefrom an aggressive female or by obstructing the female's inseminationducts against future copulation attempts from other males. Wefound no survival advantage for males damaging their pedipalps;however, copulations into a previously used insemination ductwere significantly shorter when the previous male had left partsof his genitalia inside the insemination duct. Because copulationduration determines paternity in this species, our result suggeststhat male genital damage in A. bruennichi is sexually selected.By breaking off parts of their intromittent organs inside avirgin female, males can reduce sperm competition and therebyincrease their paternity success.     

意大利蝗交配行为观察

通过昆虫行为观测仪与室外饲养观察,分析意大利蝗Calliptamus italicus的交配行为,阐明交配经历和温度对意大利蝗交配持续时间的影响。结果发现:(1)意大利蝗的交配行为包括相遇和交配两个阶段。相遇行为包括避让、打斗和交配3种类型,其中打斗多发生在雄雄相遇、雄虫与一对正在交配成虫相遇情况。避让和交配则多发生在雌雌相遇、雌雄相遇、雌虫与一对正在交配成虫相遇情况。雄虫在交配过程中占据主动。意大利蝗的交配行为包括曲腹、爬背、抱对和交配4个过程,平均时间为11.25±1.40 min。(2)雌雄虫初次交配的持续时间(12.88±0.67 min)显著长于雌雄虫均有交配经历的时间(10.47±0.39 min)(P0.05);初次交配雌虫和有交配经历雄虫的交配持续时间(11.00±0.75 min),与初次交配雄虫和有交配经历雌虫的时间(12.12±0.67 min)无显著差异(P0.05)。(3)随温度升高意大利蝗的交配持续时间缩短,27℃时交配持续时间最长(15.93±2.25 min),且与36℃、42℃时的时间差异显著(P0.05),42℃时的交配持续时间最短,为6.01±0.43 min,与36℃时的时间(7.47±0.52 min)差异不显著(P0.05)。     

Securing paternity: mating plugs in the dwarf spider Oedothorax retusus (Araneae: Erigoninae)

One of the various male strategies to prevent or impede female remating is the production of a mating plug that covers the female genital opening or remains inside of the female genital tract after mating. Such structures have been described for many species in many animal taxa; however, in most cases, we know little or nothing about their specific adaptive value. Our investigations demonstrate that females of the dwarf spider species Oedothorax retusus (Westring, 1851) (Linyphiidae, Erigoninae) exhibit a substance on one or both of her paired genital openings only after copulation. We performed double-mating trials and forced the second male to mate into the previously used or unused spermathecal duct of the female by amputating one of his paired male gonopods (pedipalps). Furthermore, to investigate whether the duration of the first mating has an effect on the size and efficiency of the mating plug, we interrupted first matings after either 1 or 3 min, categorized plug size and recorded mating behaviour of subsequent males. The amount of secretion transferred was larger in long compared to short copulations. A long first copulation successfully prevented subsequent males from mating into the used ducts, whereas mating success after short first matings was similar to matings into unused copulatory ducts of the females. The present study demonstrates that a male O. retusus can prevent a rival from transferring sperm into the same spermatheca by applying a mating plug, but only if he mates for long enough.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 96 , 574–583.     

Differentiation of somatic cells in the ovariuteri of the apoikogenic scorpion Euscorpius italicus (Chelicerata,Scorpiones, Euscorpiidae)

In apoikogenic scorpions, growing oocytes protrude from the gonad (ovariuterus) and develop in follicles exposed to the mesosomal (i.e. hemocoelic) cavity. During subsequent stages of oogenesis (previtellogenesis and vitellogenesis), the follicles are connected to the gonad surface by prominent somatic stalks. The aim of our study was to analyze the origin, structure and functioning of somatic cells accompanying protruding oocytes. We show that these cells differentiate into two morphologically distinct subpopulations: the follicular cells and stalk cells. The follicular cells gather on the hemocoelic (i.e. facing the hemocoel) surface of the oocyte, where they constitute a cuboidal epithelium. The arrangement of the follicular cells on the oocyte surface is not uniform; moreover, the actin cytoskeleton of these cells undergoes significant modifications during oocyte growth. During initial stages of the stalk formation the stalk cells elongate and form F-actin rich cytoplasmic processes by which the stalk cells are tightly connected to each other. Additionally, the stalk cells develop microvilli directed towards the growing oocyte. Our findings indicate that the follicular cells covering hemocoelic surfaces of the oocyte and the stalk cells represent two distinct subpopulations of epithelial cells, which differ in morphology, behavior and function.     

Not just a chastity belt: the functional significance of mating plugs in garter snakes,revisited

During the spring emergence of red‐sided garter snakes (Thamnophis sirtalis parietalis) in Manitoba, Canada, the operational sex ratio is strongly skewed towards males, who scramble to locate and court newly emerged females. A high frequency of multiple paternity litters suggests that the females are promiscuous; the gelatinous copulatory plugs (CPs) deposited by males may confer fitness benefits via passive mate guarding. Because precopulatory female choice is limited in large mating aggregations, sexual conflict may place a premium on preventing females from ejecting male sperm. In snakes, sperm are produced in the testes and delivered through the ductus deferens, and the CP is thought to be produced by the renal sexual segment and conveyed through the ureter. We manipulated the delivery of the two fluids separately by surgically ligating the ducts. Ureter‐ligated males did not produce a CP, causing their sperm to leak out of the female's cloaca immediately after copulation. Contrary to previous suggestions, histology revealed sperm distributed throughout the CP. Thus, the CP may function as a spermatophore: the protein matrix contains the sperm, which are liberated gradually as the plug dissolves. The likelihood of a male depositing a CP fell significantly after his second mating, perhaps limiting his reproductive success. These results challenge the hypothesis that passive mate guarding is the primary function of the CP in T. sirtalis parietalis. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 109 , 893–907.     

Inhibition of female mating by male accessory gland substances in the ground beetle Leptocarabus procerulus

The ground beetle Leptocarabus procerulus (Chaudoir) possesses seminal substances that have a physical function to form mating plugs and a physiological function to induce female refractory behaviour, which act together to hinder female remating. Little is known about the physiological properties of the substances inducing female refractory behaviour, especially with respect to their secretory organ, dose‐dependency, molecular characteristics and the effect of female maturity. By injecting male‐derived substances into females, substances that induce female refractory behaviour are shown to be produced in the male accessory gland but not in the testis. Injection of extracts from the accessory gland increases the female refractory period at moderate doses but not at lower or higher doses. By contrast, injection of extracts from the testis reduces the female refractory period at high doses. The lack of an effect of accessory gland substances at higher doses could be the result of an anomalous effect of unnaturally large doses of seminal products by direct injection, the toxicity of seminal substances that deter female responses, or counteraction by injected substances that promote female receptivity. The accessory gland substances lose their activity when heated, although the testis substances do not. Females without mature eggs tend to reject mating entirely, although variation in the number of mature eggs (one or more) is not associated with the female refractory period, indicating the limited effect of female reproductive maturity. These findings may help to clarify the physiological basis of the evolution of the elaborated male mating behaviour in L. procerulus.     

Effects of two seminal fluid transcripts on post‐mating behaviour in the simultaneously hermaphroditic flatworm Macrostomum lignano

The seminal fluid proteins (SFPs) transferred to mating partners along with sperm often play crucial roles in mediating post‐mating sexual selection. One way in which sperm donors can maximize their own reproductive success is by modifying the partner's (sperm recipient's) post‐copulatory behaviour to prevent or delay re‐mating, thereby decreasing the likelihood or intensity of sperm competition. Here, we adopted a quantitative genetic approach combining gene expression and behavioural data to identify candidates that could mediate such a response in the simultaneously hermaphroditic flatworm Macrostomum lignano. We identified two putative SFPs—Mlig‐pro46 and Mlig‐pro63—linked to both mating frequency and ‘suck’ frequency, a distinctive behaviour, in which, upon ejaculate receipt, the worm places its pharynx over its female genital opening and apparently attempts to remove the received ejaculate. We, therefore, performed a manipulative experiment using RNA interference‐induced knockdown to ask how the loss of Mlig‐pro46 and Mlig‐pro63 expression, singly and in combination, affects mating frequency, partner suck propensity and sperm competitive ability. None of the knockdown treatments impacted strongly on the mating frequency or sperm competitive ability, but knockdown of Mlig‐pro63 resulted in a significantly decreased suck propensity of mating partners. This suggests that Mlig‐pro63 may normally act as a cue in the ejaculate to trigger recipient suck behaviour and—given that other proteins in the ejaculate have the opposite effect—could be one component of an ongoing arms race between donors and recipients over the control of ejaculate fate. However, the adaptive significance of Mlig‐pro46 and Mlig‐pro63 from a donor perspective remains enigmatic.     

Precopulatory but not postcopulatory male reproductive traits diverge in response to mating system manipulation in Drosophila melanogaster

Competition between males creates potential for pre‐ and postcopulatory sexual selection and conflict. Theory predicts that males facing risk of sperm competition should evolve traits to secure their reproductive success. If those traits are costly to females, the evolution of such traits may also increase conflict between the sexes. Conversely, under the absence of sperm competition, one expectation is for selection on male competitive traits to relax thereby also relaxing sexual conflict. Experimental evolution studies are a powerful tool to test this expectation. Studies in multiple insect species have yielded mixed and partially conflicting results. In this study, we evaluated male competitive traits and male effects on female costs of mating in Drosophila melanogaster after replicate lines evolved for more than 50 generations either under enforced monogamy or sustained polygamy, thus manipulating the extent of intrasexual competition between males. We found that in a setting where males competed directly with a rival male for access to a female and fertilization of her ova polygamous males had superior reproductive success compared to monogamous males. When comparing reproductive success solely in double mating standard sperm competition assays, however, we found no difference in male sperm defense competitiveness between the different selection regimes. Instead, we found monogamous males to be inferior in precopulatory competition, which indicates that in our system, enforced monogamy relaxed selection on traits important in precopulatory rather than postcopulatory competition. We discuss our findings in the context of findings from previous experimental evolution studies in Drosophila ssp. and other invertebrate species.     

Repeated copulations as a strategy to maximize fertilization in the fly, Dryomyza anilis (Dryomyzidae)

In the fly, Dryomyza anilis, males copulate repeatedly withthe same female during oviposition. Each copulation bout consistsof intromission followed by several tapping sequences duringwhich the male touches the external genitalia of the femalewith his claspers. The relative fertilization success of thelast male to mate with the female increases with the numberof tapping sequences. In this study, male benefits of repeatedcopulation bouts were examined by comparing matings with thesame number of tapping sequences in one and several copulationbouts. The relative fertilization success for the last maleincreased with successive copulation bouts. Simulated ovipositionpatterns showed that fertilization success in matings with manycopulation bouts was higher than in matings with one bout onlyif eggs were distributed between bouts in a certain way. Thesepredicted oviposition patterns were compared with natural ones.Although males may benefit from repeated copulation, femalesare likely to prefer matings with quick oviposition. This intersexualconflict may also affect natural oviposition patterns. It issuggested that repeated copulation in D. anilis may have evolvedin connection with males trying to secure their paternity becausefemales can discharge sperm at any moment during mating.     

A new species of Euscorpius Thorell, 1876 (Scorpiones, Euscorpiidae) from Turkey

A new species of the genus Euscorpius Thorell, 1876is described based on specimens collected from Dilek Peninsula (Davutlar, Aydın) in Turkey. It is characterized by an oligotrichous trichobothrial pattern (Pv= 7, et= 5/6, eb= 4) and small size. Euscorpius (Euscorpius) avcii sp. n. is the first named species of the subgenus Euscorpius from Turkey.     

The effect of female mating history on sperm precedence in the two-spot ladybird, Adalia bipunctata (Coleoptera, Coccinellidae)

Effects of two different mating regimes on sperm precedencein the two-spot ladybird, Adalia bipunctata, were studied usingthe polymorphic gene for melanism as a marker for paternity.Virgin nonmelanic females (homozygous recessive) were matedto nonmelanic male(s) and then, after laying fertilized eggs,were mated to a melanic male of known genotype. The resultsafter the two successive single matings showed a highly variabledegree of paternity of the second male. Initial multiple matingwith nonmelanic males did not alter the pattern of paternityafter the subsequent single mating with a melanic male, butit had two other effects: (1) the female showed an increasein rejection behavior, and (2) a longer copulation was requiredfor high success of the melanic male. Additional observationsin which families were reared from beetles collected in copulain the field demonstrated that sperm competition also occursunder natural conditions. The outcome of the competition wasvariable with frequent sperm mixing.     

Seasonal variation in genital and body size, sperm displacement ability, female mating rate, and male harassment in two calopterygid damselflies (Odonata: Calopterygidae)

Sperm competition is a pervasive force. One adaptation is the male ability to displace the rivals' sperm that females have stored from previous copulations. In the damselfly, Calopteryx haemorrhoidalis asturica , males with wider aedeagi displace more spermathecal sperm. The present study documents that the same mechanism operates in another damselfly, Hetaerina americana . However, this genital width in both species decreases along the season, but late-emerging females have more sperm displaced than early-emerging females. Because territorial males mated more and were larger in body and genital size than nonterritorial males, late-season females mated with considerably larger males with respect to female size and this produced higher sperm displacement. Assuming female benefits from storing sperm but that such benefit does not prevail if males displace sperm, it is predicted that, along the season, females will mate less and male harassment (in terms of male mating attempts and oviposition duration) will increase. These predictions were corroborated. In H. americana , it was also tested whether spermathecal sperm became less viable along the season. The results obtained did not corroborate this. This is the first evidence indicating that season affects sperm displacement ability and female mating frequency due to changes in male body and genital size.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 96 , 815–829.     

Effect of the mating plug on female chemical attractiveness and mating acceptance in a scorpion

After mating, females may experience a decline in sexual receptivity and attractiveness that may be associated with changes in the production and emission of sex pheromones. In some cases, these changes are produced by chemical substances or structures (e.g., mating plugs) produced by males as a strategy to avoid or reduce sperm competition. In scorpions, sex pheromones may be involved in finding potential mates and starting courtship. Here, we tested the hypothesis that the males of Urophonius brachycentrus, a species that produces a mating plug, use chemical communication (sex pheromones) to detect, localize, and discriminate females according to their mating status (virgin or inseminated), aided by chemical signaling. We also explored the effect of extracting of the mating plug on chemical communication and mating acceptance. We used Y‐maze olfactometers with different stimuli to analyze male choice and exploration time. To evaluate mating acceptance, we measured the attractiveness and receptivity of females of different mating status. We found that chemical communication occurs through volatile pheromones, but not contact pheromones. Males equally preferred sites with virgin or inseminated females with removed mating plug. In turn, females with these mating statuses were more attractive and receptive for males than inseminated females. This study suggests that the mating plug significantly affects female chemical attractiveness with an effect on volatile pheromones and decreasing sexual mating acceptance of females. The decline in the female's sexual receptivity is a complex process that may respond to several non‐exclusive mechanisms imposed by males and strategically modulated by females.     

Coevolution of non-fertile sperm and female receptivity in a butterfly

Sexual conflict can promote rapid evolution of male and female reproductive traits. Males of many polyandrous butterflies transfer nutrients at mating that enhances female fecundity, but generates sexual conflict over female remating due to sperm competition. Butterflies produce both normal fertilizing sperm and large numbers of non-fertile sperm. In the green-veined white butterfly, Pieris napi, non-fertile sperm fill the females'' sperm storage organ, switching off receptivity and thereby reducing female remating. There is genetic variation in the number of non-fertile sperm stored, which directly relates to the female''s refractory period. There is also genetic variation in males'' sperm production. Here, we show that females'' refractory period and males'' sperm production are genetically correlated using quantitative genetic and selection experiments. Thus selection on male manipulation may increase the frequency of susceptible females to such manipulations as a correlated response and vice versa.     

Effects of caffeine on mating behavior and sperm precedence in Tribolium castaneum

Biogenic amines such as dopamine are physiologically neuroactive substances that affect behavioral and physiological traits in invertebrates, and it has long been known that these substances affect mating behavior in insects. Caffeine is a dopamine activator and thus enhances dopamine receptor activity. However, the effects of caffeine intake on insect mating behavior have been largely unexplored. Therefore, we examined the effect of caffeine on mating behavior in the red flour beetle Tribolium castaneum. Caffeine, which activates dopamine, affected the mating behavior of T. castaneum males. Males who orally ingested caffeine courted faster than males who did not, resulting in faster mounting of females and less time to a male's external aedeagus protrusion. However, the present results showed no difference in sperm precedence measured as a P2 value between males fed caffeine and males not fed caffeine. We discuss the effects of caffeine on insect mating and the possibility that caffeine consumption may cause males to mate with more females in the laboratory.     

Patterns of sperm precedence are not affected by female mating history in Drosophila melanogaster

In promiscuously mating species, there is strong selection on males to maximize their share of paternity through both defensive and offensive means. This has been most extensively examined using the Drosophila melanogaster model system. In these studies, sperm competition has been examined by mating a virgin female to two consecutive males and then determining the fertilization success of both the first male (defending, P1) and the second male (offending, P2). Recent evidence suggests that male defense may be influenced by female mating history (i.e., virgin versus nonvirgin). Here, by mating females to males with three different genotypes, we show that female mating history does not affect male defensive or offensive abilities in sperm competition. We also show that, although female lifetime fecundity was not correlated with the number of times that she mated, it was reduced by increased exposure to males. These data indicate that measures of P1 and P2 previously reported in D. melanogaster may be robust to the specific mating history of the females used in these studies.     

Species boundaries and phylogeography of the "Euscorpius carpathicus complex" (Scorpiones: Euscorpiidae) in Italy

Euscorpius tergestinus (C.L. Koch, 1837), Euscorpius concinnus (C.L. Koch, 1837) and Euscorpius sicanus (C.L. Koch, 1837), three presumed closely related species belonging to the "carpathicus group", occur in the Italian peninsula with a largely parapatric distribution and some zones of range overlap. These areas of sympatry represent interesting opportunities to investigate species boundaries in natural populations. Here we report on a study exploring genetic variation in sympatric populations of the three species from central Tuscany. Additional collecting sites, from different localities across Italy, were also included in the analysis in order to explore the phylogeographic structure of the group. Species boundaries and evolutionary relationships were examined by sequence comparison of mitochondrial 16S rRNA and nuclear ITS-1 rRNA gene fragments. DNA sequence data show no evidence of genetic introgression between different evolutionary lineages from the area of range overlap, suggesting the absence of either past or ongoing inter-specific gene flow. It is therefore probable that reproductive barriers exist, preventing gene pools from amalgamating. Furthermore, our results support the recent morphological distinction of E. tergestinus, as traditionally classified, into two different species: E. tergestinus and E. concinnus. Both mitochondrial and nuclear sequence data clearly indicate that the two taxa represent well-supported and deeply divergent lineages. Euscorpius sicanus seems to represent a monophyletic taxon, but the high genetic variability observed within this taxon calls for future investigation. The present distribution patterns across the Italian peninsula were mainly interpreted as the consequence of climatic oscillations.