The function of prolonged copulations in Enchenopa treehoppers (Hemiptera: Membracidae).

Copulations are very brief in many species, sometimes taking only seconds, but in other species they can be quite prolonged. Potential explanations for prolonged copulations include time requirements for the transfer of sperm and/or other ejaculate substances. Ejaculate substances could function to regulate female receptivity to subsequent matings, provide nutritional nuptial gifts, or hasten egg oviposition at a potential survival cost to the female. We investigated prolonged copulation in a member of the Enchenopa binotata complex of treehoppers (Hemiptera: Membracidae), in which females rarely remate and copulation can last several hours. We assigned females to treatments in which we interrupted copulation at different times. We also included a control where copulation was not interrupted. We found that females that experienced shorter copulations were more likely to be subsequently receptive to an attractive male. We also found that few females produced offspring when they engaged in short copulations compared to those with longer copulations. We did not find any differences in female survival. Our results support the sperm transfer and receptivity regulation hypotheses. We discuss potential reasons for why these processes should take so long in a species with low female remating.

On the function of a female-like signal type in the vibrational repertoire of Enchenopa male treehoppers (Hemiptera: Membracidae).

Animals often mimic the behaviours or signals of conspecifics of the opposite sex while courting. We explored the potential functions of a novel female-like signal type in the courtship displays of male Enchenopa treehoppers. In these plant-feeding insects, males produce plant-borne vibrational advertisement signals, to which females respond with their own duetting signals. Males also produce a signal type that resembles the female duetting responses. We experimentally tested whether this signal modifies the behaviour of receivers. First, we tested whether the female-like signal would increase the likelihood of a female response. However, females were as likely to respond to playbacks with or without them. Second, we tested whether the female-like signal would inhibit competing males, but males were as likely to produce displays after playbacks with or without them. Hence, we found no evidence that this signal has an adaptive function, despite its presence in the courtship display, where sexual selection affects signal features. Given these findings, we also explored whether the behavioural and morphological factors of the males were associated with the production of the female-like signal. Males that produced this signal had higher signalling effort (longer and more frequent signals) than males that did not produce it, despite being in worse body condition. Lastly, most males were consistent over time in producing the female-like signal or not. These findings suggest that condition-dependent or motivational factors explain the presence of the female-like signal. Alternatively, this signal might not bear an adaptive function, and it could be a way for males to warm up or practice signalling, or even be a by-product of how signals are transmitted through the plant. We suggest further work that might explain our puzzling finding that a signal in the reproductive context might not have an adaptive function.

Social Plasticity Enhances Signal-Preference Codivergence.

AbstractThe social environment is often the most dynamic and fitness-relevant environment animals experience. Here we tested whether plasticity arising from variation in social environments can promote signal-preference divergence-a key prediction of recent speciation theory but one that has proven difficult to test in natural systems. Interactions in mixed social aggregations could reduce, create, or enhance signal-preference differences. In the latter case, social plasticity could establish or increase assortative mating. We tested this by rearing two recently diverged species of Enchenopa treehoppers-sap-feeding insects that communicate with plant-borne vibrational signals-in treatments consisting of mixed-species versus own-species aggregations. Social experience with heterospecifics (in the mixed-species treatment) resulted in enhanced signal-preference species differences. For one of the two species, we tested but found no differences in the plastic response between sympatric and allopatric sites, suggesting the absence of reinforcement in the signals and preferences and their plastic response. Our results support the hypothesis that social plasticity can create or enhance signal-preference differences and that this might occur in the absence of long-term selection against hybridization on plastic responses themselves. Such social plasticity may facilitate rapid bursts of diversification.

Positive-to-negative behavioural responses suggest hedonic evaluation in treefrog mate choice.

Sexual competition hinges on the ability to impress other conspecifics, to drive them away or attract them. In such cases, the selective environment may be hedonic or affective in nature, as it consists of the evaluations of the individuals making the decisions. This may contribute to the power of sexual selection because evaluations may range from positive to negative rather than simply from positive to neutral. Selection due to mate choice may therefore be stronger than currently appreciated. Further, change in preferred mate types can occur simply by changes (flips) in the evaluation of similar display features, adding to the dynamism of sexual selection as well as its strength. We tested the hypothesis of positive-to-negative behavioural responses in mate choice with a playback experiment using two treefrog species with 'mirror image' structures in their advertisement and aggressive calls. Female treefrog responses ranged from approach to evasion, and the presence of an aversive stimulus tainted evaluation of an attractive stimulus. Further, females in the two species showed flips in approach/evasion of stimuli with comparable signal structure. These results suggest that hedonic evaluation may have an important role in mate choice and showcase how mechanistic analysis can help understand evolutionary processes.

Adaptation without Specialization Early in a Host Shift.

AbstractStudents of speciation debate the role of performance trade-offs across different environments early in speciation. We tested for early performance trade-offs with a host shift experiment using a member of the Enchenopa binotata species complex of treehoppers (Hemiptera: Membracidae). In this clade of plant-feeding insects, different species live on different host plants and exhibit strong behavioral and physiological host specialization. After five generations, the experimental host shifts resulted either in no adaptation or in adaptation without specialization. The latter result was more likely in sympatry; in allopatry, populations on novel host plants were more likely to become extinct. We conclude that in the early stages of speciation, adaptation to novel host plants does not necessarily bring about performance trade-offs on ancestral environments. Adaptation may be facilitated rather than hindered by gene flow, which prevents extinction. Additional causes of specialization and assortative mating may be required if colonization of novel environments is to result in speciation.

Evolutionary novelty in communication between the sexes.

The diversity of signalling traits within and across taxa is vast and striking, prompting us to consider how novelty evolves in the context of animal communication. Sexual selection contributes to diversification, and here we endeavour to understand the initial conditions that facilitate the maintenance or elimination of new sexual signals and receiver features. New sender and receiver variants can occur through mutation, plasticity, hybridization and cultural innovation, and the initial conditions of the sender, the receiver and the environment then dictate whether a novel cue becomes a signal. New features may arise in the sender, the receiver or both simultaneously. We contend that it may be easier than assumed to evolve new sexual signals because sexual signals may be arbitrary, sexual conflict is common and receivers are capable of perceiving much more of the world than just existing sexual signals. Additionally, changes in the signalling environment can approximate both signal and receiver changes through a change in transmission characteristics of a given environment or the use of new environments. The Anthropocene has led to wide-scale disruption of the environment and may thus generate opportunity to directly observe the evolution of new signals to address questions that are beyond the reach of phylogenetic approaches.

Combinatorial Signal Processing in an Insect.

AbstractHuman language is combinatorial: phonemes are grouped into syllables, syllables are grouped into words, and so on. The capacity for combinatorial processing is present, in different degrees, in some mammals and birds. We used vibrational insects, Enchenopa treehoppers, to test the hypothesis of basic combinatorial processing against two competing hypotheses: beginning rule (where the early signal portions play a stronger role in acceptability) and no ordering rule (where the order of signal elements plays no role in signal acceptability). Enchenopa males use plant-borne vibrational signals that consist of a whine followed by pulses. We tested the above hypotheses with vibrational playback experiments in which we presented Enchenopa females with stimuli varying in signal element combinations. We monitored female responses to these playbacks with laser vibrometry. We found strong support for combinatorial processing in Enchenopa: in brief, females preferred natural-combination signals regardless of the beginning element and discriminated against reverse-order signals or individual elements. Finding support for the combinatorial rule hypothesis in insects suggests that this capability represents a common solution to the problems presented by complex communication.

The relationship between a combinatorial processing rule and a continuous mate preference function in an insect.

Mate choice involves processing signals that can reach high levels of complexity and feature multiple components, even in small animals with tiny brains. This raises the question of whether and how such organisms deal with this complexity. One solution involves combinatorial processing, whereby different signal elements are processed as single units. Combinatorial processing has been described in several mammals and birds, and recently in a vibrationally signalling insect, Enchenopa treehoppers. Here, we ask about the relationship between combinatorial rules and mate preferences for continuously varying signal features. Enchenopa male advertisement signals are composed of two elements: a 'whine' followed by a set of pulses. The dominant frequency of the whine and element combination both matter to females. We presented synthetic signals varying in element order (natural [whine-pulses], reverse [pulses-whine]) and in frequency to Enchenopa females and recorded their responses. The reverse combination resulted in a decrease in attractiveness of the signals, and also slightly changed the shape of the preference for frequency. We found that females could be classified into three 'types': females with both a strong preference and a strong combinatorial rule, females with both a weak preference and weak rule, and females with a strong preference but a weak rule. Our results suggest that in Enchenopa signal processing, the mate preference for a continuous signal feature 'takes precedence' over, but also interacts with, the combinatorial rule. The relationship between the preference and the rule could evolve to take different forms according to selection on mate choice decisions. We suggest that exploring the relationship between such preferences and rules in species with more complex signals will bring insight into the evolution of the multi-component communication systems.

On the architecture of mate choice decisions: preference functions and choosiness are distinct traits.

Mate choice is an important cause of sexual selection; it can drive the evolution of extravagant ornaments and displays, and promote speciation through the reproductive isolation generated by rapid divergence of sexual traits. Understanding mate choice requires knowledge of the traits involved in generating mate-choice decisions, and how those traits may interact with each other. It has been proposed that mate-choice decisions are the outcome of two components that vary independently: the preference function (the ranking of the attractiveness of prospective mates) and choosiness (the effort invested in mate assessment). Here we test this hypothesis by examining individual variation in female preference functions and choosiness in green treefrogs ( Hyla cinerea). We show that measures describing preference functions and choosiness are not correlated. We also show that both components are influenced differently by variation in female body size, and that preference function shape (closed and preferring intermediate values or open-ended and preferring extremes) has a strong influence on this relationship: function traits are positively correlated with body size only for individuals with closed functions, while choosiness is positively correlated with body size for individuals with open functions, but negatively for those with closed functions.

Mechanisms of Assortative Mating in Speciation with Gene Flow: Connecting Theory and Empirical Research.

The large body of theory on speciation with gene flow has brought to light fundamental differences in the effects of two types of mating rules on speciation: preference/trait rules, in which divergence in both (female) preferences and (male) mating traits is necessary for assortment, and matching rules, in which individuals mate with like individuals on the basis of the presence of traits or alleles that they have in common. These rules can emerge from a variety of behavioral or other mechanisms in ways that are not always obvious. We discuss the theoretical properties of both types of rules and explain why speciation is generally thought to be more likely under matching rather than preference/trait rules. We furthermore discuss whether specific assortative mating mechanisms fall under a preference/trait or matching rule, present empirical evidence for these mechanisms, and propose empirical tests that could distinguish between them. The synthesis of the theoretical literature on these assortative mating rules with empirical studies of the mechanisms by which they act can provide important insights into the occurrence of speciation with gene flow. Finally, by providing a clear framework we hope to inspire greater alignment in the ways that both theoreticians and empiricists study mating rules and how these rules affect speciation through maintaining or eroding barriers to gene flow among closely related species or populations.

Theory Meets Empiry: A Citation Network Analysis.

According to a recent survey, ecologists and evolutionary biologists feel that theoretical and empirical research should coexist in a tight feedback loop but believe that the two domains actually interact very little. We evaluate this perception using a citation network analysis for two data sets, representing the literature on sexual selection and speciation. Overall, 54%-60% of citations come from a paper's own category, whereas 17%-23% are citations across categories. These cross-citations tend to focus on highly cited papers, and we observe a positive correlation between the numbers of citations a study receives within and across categories. We find evidence that reviews can function as integrators between the two literatures, argue that theoretical models are analogous to specific empirical study systems, and complement our analyses by studying a cocitation network. We conclude that theoretical and empirical research are more tightly connected than generally thought but that avenues exist to further increase this integration.

Nephila clavipes spiders (Araneae: Nephilidae) keep track of captured prey counts: testing for a sense of numerosity in an orb-weaver.

Nephila clavipes golden orb-web spiders accumulate prey larders on their webs and search for them if they are removed from their web. Spiders that lose larger larders (i.e., spiders that lose larders consisting of more prey items) search for longer intervals, indicating that the spiders form memories of the size of the prey larders they have accumulated, and use those memories to regulate recovery efforts when the larders are pilfered. Here, we ask whether the spiders represent prey counts (i.e., numerosity) or a continuous integration of prey quantity (mass) in their memories. We manipulated larder sizes in treatments that varied in either prey size or prey numbers but were equivalent in total prey quantity (mass). We then removed the larders to elicit searching and used the spiders' searching behavior as an assay of their representations in memory. Searching increased with prey quantity (larder size) and did so more steeply with higher prey counts than with single prey of larger sizes. Thus, Nephila spiders seem to track prey quantity in two ways, but to attend more to prey numerosity. We discuss alternatives for continuous accumulator mechanisms that remain to be tested against the numerosity hypothesis, and the evolutionary and adaptive significance of evidence suggestive of numerosity in a sit-and-wait invertebrate predator.

Trees to treehoppers: genetic variation in host plants contributes to variation in the mating signals of a plant-feeding insect.

Community genetics research has demonstrated 'bottom-up' effects of genetic variation within a plant species in shaping the larger community with which it interacts, such as compositions of arthropod faunas. We demonstrate that such cross-trophic interactions also influence sexually selected traits. We used a member of the Enchenopa binotata species complex of treehoppers (Hemiptera: Membracidae) to ask whether male mating signals are influenced by host plant genetic variation. We reared a random sample of the treehoppers on potted replicates of a sample of host plant clone lines. We found that treehopper male signals varied according to the clone line on which they developed, showing that genetic variation in host plants affects male treehoppers' behavioural phenotypes. This is the first demonstration of cross-trophic indirect genetic effects on a sexually selected trait. We discuss how such effects may play an important role in the maintenance of variation and within-population phenotypic differentiation, thereby promoting evolutionary divergence.

Genetic variation in host plants influences the mate preferences of a plant-feeding insect.

Many species spend their lives in close association with other organisms, and the environments provided by those organisms can play an important role as causes of variation in phenotypes. When this is the case, the genotypes of the individuals constituting the environment may influence the phenotypes of individuals living in that environment. When these effects are between heterospecifics, interspecific indirect genetic effects (IIGEs) occur. Several studies have detected IIGEs, but whether IIGEs contribute to variation in sexually selected traits remains virtually unexplored. We assessed how mate preferences in a plant-feeding insect are influenced by the genotype of their host plant. We established clone lines of a sample of host plant genotypes constituting the background biotic environment for a random sample of insects that we reared on them. We found that the insects' mate preferences varied according to the clone line on which they developed. These results demonstrate that genetic variation in host plants has cross-trophic consequences on a trait that has strong effects on fitness and interpopulation dynamics such as diversification in communication systems. We discuss how IIGEs on mate preferences may influence the way in which selection acts, including the maintenance of variation and the promotion of evolutionary divergence.

Male Enchenopa treehoppers (Hemiptera: Membracidae) vary mate-searching behavior but not signaling behavior in response to spider silk.

Finding and attracting mates can impose costs on males in terms of increased encounters with, and attraction of, predators. To decrease the likelihood of predation, males may modify mate-acquisition efforts in two main ways: they may reduce mate-searching efforts or they may reduce mate-attraction efforts. The specific behavior that males change in the presence of predator cues should depend upon the nature of risk imposed by the type of predator present in the environment. For example, sit-and-wait predators impose greater costs to males moving in search of mates. Here, we test whether cues of the presence of a sit-and-wait predator lead to a reduction in mate-searching but not mate-acquisition behavior. We used a member of the Enchenopa binotata complex of treehoppers-a clade of vibrationally communicating insects in which males fly in search of mates and produce mate-attraction signals when they land on plant stems. We tested for changes in mate-searching and signaling behaviors when silk from a web-building spider was present or absent. We found that males delayed flight when spider silk was present but only if they were actively searching for mates. These results suggest that males have been selected to reduce predation risk by adjusting how they move about their environment according to the cues of sit-and-wait predators.

Heuristic test reveals little effect of learning and maturation on early prey capture experiences in a web-building spider.

Behaviors can vary throughout an animal's life and this variation can often be explained by changes associated with learning and/or maturing. Currently, there is little consensus regarding how these processes interact to affect behaviors. Here we proposed a heuristic approach to disentangle the effects of learning and maturation on behavior and applied it to the predatory behaviors of Physocyclus globosus spiderlings. We varied the degree of prey difficulty and familiarity spiderlings received along the first instar and across the molt to the second instar and quantified the time spiderlings spent wrapping prey, as a proxy for prey capture efficiency. We found no overall evidence for learning or maturation. Changes in efficiency were mainly due to the switch from difficult to easy prey, or vice versa. However, there was one treatment where spiderlings improved in efficiency before and after the molt, without a switch in prey type. This provides some indication that difficult prey may offer more opportunity for learning or maturation to impact behavior. Although we found little effect of learning or maturation on prey capture efficiency, we suggest that our heuristic approach is effective and could be useful in investigating these processes in other behaviors and other animals.

Diversification under sexual selection: the relative roles of mate preference strength and the degree of divergence in mate preferences.

The contribution of sexual selection to diversification remains poorly understood after decades of research. This may be in part because studies have focused predominantly on the strength of sexual selection, which offers an incomplete view of selection regimes. By contrast, students of natural selection focus on environmental differences that help compare selection regimes across populations. To ask how this disparity in focus may affect the conclusions of evolutionary research, we relate the amount of diversification in mating displays to quantitative descriptions of the strength and the amount of divergence in mate preferences across a diverse set of case studies of mate choice. We find that display diversification is better explained by preference divergence rather than preference strength; the effect of the latter is more subtle, and is best revealed as an interaction with the former. Our findings cast the action of sexual selection (and selection in general) in a novel light: the strength of selection influences the rate of evolution, and how divergent selection is determines how much diversification can occur. Adopting this view will enhance tests of the relative role of natural and sexual selection in processes such as speciation.

Genetic variation in social influence on mate preferences.

Patterns of phenotypic variation arise in part from plasticity owing to social interactions, and these patterns contribute, in turn, to the form of selection that shapes the variation we observe in natural populations. This proximate-ultimate dynamic brings genetic variation in social environments to the forefront of evolutionary theory. However, the extent of this variation remains largely unknown. Here, we use a member of the Enchenopa binotata species complex of treehoppers (Hemiptera: Membracidae) to assess how mate preferences are influenced by genetic variation in the social environment. We used full-sibling split-families as 'treatment' social environments, and reared focal females alongside each treatment family, describing the mate preferences of the focal females. With this method, we detected substantial genetic variation in social influence on mate preferences. The mate preferences of focal females varied according to the treatment families along with which they grew up. We discuss the evolutionary implications of the presence of such genetic variation in social influence on mate preferences, including potential contributions to the maintenance of genetic variation, the promotion of divergence, and the adaptive evolution of social effects on fitness-related traits.

Vibrational playback by means of airborne stimuli.

Substrate-borne vibrational communication is a common form of communication in animals. Current contact-based playback methods limit the number of substrates that can be stimulated simultaneously and potentially change the transmission properties of the substrate. Here, we explore a solution to these limitations by broadcasting airborne stimuli onto plant substrates to impart vibrational playbacks. We demonstrate that one can effectively compensate for the filtering properties of any individual plant across a range of frequencies. We then address how well both compensated broad-band and pure-tone stimuli for one plant individual apply to other individuals across days. Variation within and between plants was similar across the range tested but was quite variable at certain frequencies. Focusing on a subset of this range, at low frequencies, responses were flat across days and pure-tone frequency stimuli in this range were consistently transmitted despite repositioning of plants relative to the loudspeaker. Our results present a potential solution to researchers interested in exposing large samples of individuals to vibrational signals but also highlight the importance of validating the use of airborne stimuli as vibrational playbacks to the particular substrate type and frequency range of interest.

Genotype × environment interaction is weaker in genitalia than in mating signals and body traits in Enchenopa treehoppers (Hemiptera: Membracidae).

Theory predicts that selection acting across environments should erode genetic variation in reaction norms; i.e., selection should weaken genotype × environment interaction (G × E). In spite of this expectation, G × E is often detected in fitness-related traits. It thus appears that G × E is at least sometimes sustained under selection, a possibility that highlights the need for theory that can account for variation in the presence and strength of G × E. We tested the hypothesis that trait differences in developmental architecture contribute to variation in the expression of G × E. Specifically, we assessed the influence of canalization (robustness to genetic or environmental perturbations) and condition-dependence (association between trait expression and prior resource acquisition or vital cellular processes). We compared G × E across three trait types expected to differ in canalization and condition-dependence: mating signals, body size-related traits, and genitalia. Because genitalia are expected to show the least condition-dependence and the most canalization, they should express weaker G × E than the other trait types. Our study species was a member of the Enchenopa binotata species complex of treehoppers. We found significant G × E in most traits; G × E was strongest in signals and body traits, and weakest in genitalia. These results support the hypothesis that trait differences in developmental architecture (canalization and condition-dependence) contribute to variation in the expression of G × E. We discuss implications for the dynamics of sexual selection on different trait types.