高橋 佑磨

Abstract Organisms have diverse biological clocks synchronised with environmental cycles depending on their habitats. Anticipation of tidal changes has driven the evolution of circatidal rhythms in some marine species. In the freshwater snail, Semisulcospira reiniana, individuals in nontidal areas exhibit circadian rhythms, whereas those in tidal areas exhibit both circadian and circatidal rhythms. We investigated whether the circatidal rhythms are genetically determined or induced by environmental cycles. The exposure to a simulated tidal cycle did not change the intensity of circatidal rhythm in individuals in the nontidal population. However, snails in the tidal population showed different activity rhythms depending on the presence or absence of the exposure. Transcriptome analysis revealed that genes with circatidal oscillation increased due to entrainment to the tidal cycle in both populations and dominant rhythmicity was consistent with the environmental cycle. These results suggest plasticity in the endogenous rhythm in the gene expression in both populations. Note that circatidal oscillating genes were more abundant in the tidal population than in the nontidal population, suggesting that a greater number of genes are associated with circatidal clocks in the tidal population compared to the nontidal population. This increase of circatidal clock–controlled genes in the tidal population could be caused by genetic changes in the biological clock or the experience of tidal cycle in the early life stage. Our findings suggest that the plasticity of biological rhythms may have contributed to the adaptation to the tidal environment in S. reiniana.

Abstract Non-genetic variation is the phenotypic variation induced by the differential expression of a genotype in response to varying environmental cues and is broadly categorized into two types: phenotypic plasticity and developmental noise. These aspects of variation have been suggested to play an important role in adaptive evolution. However, the mechanisms by which these two types of non-genetic variations influence the evolutionary process are currently poorly understood. Using a machine-learning-based phenotyping tool, we independently quantified phenotypic plasticity and developmental noise in the wing morphological traits of the fruit fly Drosophila simulans. Utilizing a rearing experiment, we demonstrated plastic responses in both wing size and shape as well as non-zero heritability of both phenotypic plasticity and developmental noise, which suggests that adaptive phenotypic plasticity can evolve via genetic accommodation in the wing morphology of D. simulans. We found a positive correlation between phenotypic plasticity and developmental noise, while the correlation between the plastic response to three kinds of environmental factors that were examined (nutrient condition, temperature, and light–dark cycle) was poor. These results suggest that phenotypic plasticity and developmental noise contribute to evolvability in a similar manner, however, the mechanisms that underlie the correspondence between these two types of variation remain to be elucidated.

Abstract Seasonal environmental change is one of the most rapid and striking environmental variables. Although relatively rapid adaptation to environmental changes over several years or several decades has been described in many taxa, rapid responses to seasonal environments are delicate, and therefore, the detection of the evolutionary responses requires sensitive methods. In this study, we examined seasonal changes in phenotypes related to thermal tolerance and morphological traits of Drosophila lutescens collected at the spring and autumn periods from a single location. We first demonstrated that flies in the two seasonal periods were almost genetically identical using double-digest restriction site-associated DNA sequencing and analysis. Using an experimental design to eliminate the effect of possible confounding factors that influence phenotypes (i.e., maternal effects and the environmental conditions in which each phenotype was analyzed), we showed that the heat tolerance of D. lutescens was significantly higher in the autumn population than in the spring population. Furthermore, cold tolerance was slightly higher in the spring population than in the autumn one. Although wing length and thorax length did not change significantly between seasons, the ratio of wing length to thorax length changed significantly between them. These results suggest that seasonal environmental heterogeneity induces rapid phenotypic changes within a year. Finally, we discuss the possibility of rapid evolutionary responses to seasonal changes.

Organisms have endogenous timekeeping system(s) to coordinate their biological processes with environmental cycles, allowing adaptation to external rhythmic changes in their environment. The change in endogenous rhythms could contribute to range expansion in a novel rhythmic environment. We hypothesized that populations of the freshwater snail near estuaries show a circatidal rhythm to synchronize with the tidal cycle. We compared the behavioral and gene expression rhythms between non-tidal and tidal populations of the freshwater snail, Semisulcospira reiniana. Individuals inhabiting tidal areas exhibited a rhythmic activity pattern coordinated with the tidal cycle under both field and laboratory conditions, but individuals inhabiting upstream non-tidal areas showed a circadian activity pattern. The proportion of circadian oscillating genes was greater in non-tidal than in tidal individuals, while that of circatidal oscillating genes was greater in tidal than in non-tidal individuals. Additionally, transcriptome-wide population genetic analyses revealed that these two adjacent populations can be clearly distinguished genetically, though the genetic distance was very small. Our results provide evidence of the shift in an endogenous rhythm via range expansion to a novel rhythmic environment. The changes in a small number of genes and/or phenotypic plasticity may contribute to the difference in the endogenous rhythms between non-tidal and tidal populations.

Cities experience changes in abiotic factors, such as warming, increases in noise and light. These changes can lead to phenotypic changes. Several studies have revealed that altered environments change phenotypes in plants and animals in cities. However, limited studies have isolated evolutionary from nongenetic changes. Here, we analyzed the evolution of thermal tolerance and diurnal activity patterns in the urban population of the fruit pest, Drosophila suzukii. Urban and rural isofemale lines were reared under constant conditions. We compared the lower and upper thermal limits (CTmin and CTmax, respectively), and effects of temperature exposure on the thermal limits of urban and rural populations. Common garden experiments showed that urban populations exhibit a lower CTmin than rural populations, suggesting genetic difference in CTmin among populations. On the other hand, the difference in CTmax between urban and rural populations was not significant. Exposure to cold temperature did not affect CTmin in both urban and rural populations. In contrast, exposure to hot temperature increased CTmax especially in urban population, suggesting that urban populations evolved in response to urban heat. We also investigated the daily activity patterns of urban and rural populations and the effect of lifelong artificial light at night on daily activity. We found that night-time light (dim light) reduced the total amount of activity compared to dark night condition. In addition, dim light at night altered the daily rhythm of activity and increased the activity rate at night. The effect of night light on total activity was less in urban than that in rural populations, suggesting that populations in cities evolved to mitigate decreased activity under night light. Our results showed that environmental temperature and artificial light at night evolutionarily and plastically influence ecologically important traits, such as temperature tolerance and diurnal activity.

One of the most important questions in evolutionary biology is how the spatial distribution of species is limited. Asymmetric gene flow from core populations is suggested to increase the number of poorly adapted immigrants in the populations at the range edge. Genetic load due to migration, i.e., migration load, should prevent adaptation to the local habitat, leading to decreases in distribution range via local extinction or the limiting range expansion. However, few experimental studies have examined the effects of immigration on fitness and natural selection within recipient populations. To investigate the influence of migration load on the evolution of distribution range, we performed field and laboratory observations as well as population transcriptomics for the common river snail, Semisulcospira reiniana. This species meets the conditions that migration from source populations can prevent local adaptation in a sink population because they inhabit the broader range of environments, including middle/upper reaches of a river and estuaries within a single river and they may be more vulnerable to being swept away by water currents due to lowered spontaneous (upward) locomotion activity. We found that river steepness was related to the lower distribution limit of S. reiniana, with a narrower distribution range in the steeper river. Population transcriptomic analysis showed that gene flow was heavily asymmetric from the upstream populations to downstream ones in the steep river, suggesting a greater migration load in the steep river. The number of genes putatively involved in adaptation to the local habitat was lower in the steep river than in the gentle river. Gene expression profiles suggested that individuals achieve better local adaptation in the gentle river. Laboratory experiments suggested that evolutionary differences in salinity tolerance among local populations were only found in the gentle river. Our results consistent with the hypothesis that migration load owing to asymmetric gene flow disturbs local adaptation and restricts the distribution range of river snails.

BACKGROUND: Evolutionary processes can cause strong spatial genetic signatures, such as local loss of genetic diversity, or conflicting histories from mitochondrial versus nuclear markers. Investigating these genetic patterns is important, as they may reveal obscured processes and players. The maternally inherited bacterium Wolbachia is among the most widespread symbionts in insects. Wolbachia typically spreads within host species by conferring direct fitness benefits, and/or by manipulating its host reproduction to favour infected over uninfected females. Under sufficient selective advantage, the mitochondrial haplotype associated with the favoured maternally-inherited symbiotic strains will spread (i.e. hitchhike), resulting in low mitochondrial genetic variation across the host species range. METHOD: The common bluetail damselfly (Ischnura elegans: van der Linden, 1820) has recently emerged as a model organism for genetics and genomic signatures of range expansion during climate change. Although there is accumulating data on the consequences of such expansion on the genetics of I. elegans, no study has screened for Wolbachia in the damselfly genus Ischnura. Here, we present the biogeographic variation in Wolbachia prevalence and penetrance across Europe and Japan (including samples from 17 populations), and from close relatives in the Mediterranean area (i.e. I. genei: Rambur, 1842; and I. saharensis: Aguesse, 1958). RESULTS: Our data reveal (a) multiple Wolbachia-strains, (b) potential transfer of the symbiont through hybridization, (c) higher infection rates at higher latitudes, and (d) reduced mitochondrial diversity in the north-west populations, indicative of hitchhiking associated with the selective sweep of the most common strain. We found low mitochondrial haplotype diversity in the Wolbachia-infected north-western European populations (Sweden, Scotland, the Netherlands, Belgium, France and Italy) of I. elegans, and, conversely, higher mitochondrial diversity in populations with low penetrance of Wolbachia (Ukraine, Greece, Montenegro and Cyprus). The timing of the selective sweep associated with infected lineages was estimated between 20,000 and 44,000 years before present, which is consistent with the end of the last glacial period about 20,000 years. CONCLUSIONS: Our findings provide an example of how endosymbiont infections can shape spatial variation in their host evolutionary genetics during postglacial expansion. These results also challenge population genetic studies that do not consider the prevalence of symbionts in many insects, which we show can impact geographic patterns of mitochondrial genetic diversity.

Intrapopulation variation in behaviour, including activity, boldness and aggressiveness, is becoming more widely recognized and is hypothesized to substantially affect ecological and evolutionary dynamics. Although previous studies used candidate-gene approaches and genome-wide association analyses to identify genes correlated with variations in activity and aggressiveness, behavioural variation may not be fully captured in the nuclear genome, as it does not account for mitochondrial genomes. Mitochondrial genes encode products that are key regulators of the cellular energy-producing pathways in metabolic processes and are thought to play a significant role in life-history and reproductive traits. In this study, we considered many isofemale lines of Drosophila immigrans established from two wild populations to investigate whether intrapopulation variation in the mitochondrial genome affected activity level within this species. We identified two major haplogroups in these populations, and activity levels in both larvae and adults differed significantly between the two haplogroups. This result indicated that intrapopulation variation in activity level may be partially controlled by mitochondrial genes, along with the interaction between nuclear and mitochondrial genes and the age of individual organisms.

Odonata species display a remarkable diversity of colour patterns, including intrasexual polymorphisms. In the damselfly ( <italic>Ischnura senegalensis</italic> ), the expression of a sex-determining transcription factor, the <italic>doublesex</italic> ( <italic>Isdsx</italic> ) gene is reportedly associated with female colour polymorphism (CP) (gynomorph for female-specific colour and andromorph for male-mimicking colour). Here, the function of <italic>Isdsx</italic> in thoracic coloration was investigated by electroporation-mediated RNA interference (RNAi). RNAi of the <italic>Isdsx</italic> common region in males and andromorphic females reduced melanization and thus changed the colour pattern into that of gynomorphic females, while the gynomorphic colour pattern was not affected. By contrast, RNAi against the <italic>Isdsx</italic> long isoform produced no changes, suggesting that the <italic>Isdsx</italic> short isoform is important for body colour masculinization in both males and andromorphic females. When examining the expression levels of five genes with differences between sexes and female morphs, two melanin-suppressing genes, <italic>black</italic> and <italic>ebony</italic> , were expressed at higher levels in the <italic>Isdsx</italic> RNAi body area than a control area. Therefore, the <italic>Isdsx</italic> short isoform may induce thoracic colour differentiation by suppressing <italic>black</italic> and <italic>ebony</italic> , thereby generating female CP in <italic>I. senegalensis.</italic> These findings contribute to the understanding of the molecular and evolutionary mechanisms underlying female CP in Odonata.

Latitude is correlated with environmental components that determine the distribution of biodiversity. In combination with geographic factors, latitude-associated environmental variables are expected to influence speciation, but empirical evidence on how those factors interplay is scarce. We evaluated the genetic and environmental variation among populations in the pair of sister species Dioon sonorense-D. vovidesii, two cycads distributed along a latitudinal environmental gradient in northwestern Mexico, to reveal their demographic histories and the environmental factors involved in their divergence. Using genome-wide loci data, we determined the species delimitation, estimated the gene flow, and compared multiple demographic scenarios of divergence. Also, we estimated the variation of climatic variables among populations and used ecological niche models to test niche overlap between species. The effect of geographic and environmental variables on the genetic variation among populations was evaluated using linear models. Our results suggest the existence of a widespread ancestral population that split into the two species ~829 ky ago. The geographic delimitation along the environmental gradient occurs in the absence of major geographic barriers, near the 28th parallel north, where a zonation of environmental seasonality exists. The northern species, D. vovidesii, occurs in more seasonal environments but retains the same niche of the southern species, D. sonorense. The genetic variation throughout populations cannot be solely explained by stochastic processes; the latitudinal-associated seasonality has been an additive factor that strengthened the species divergence. This study represents an example of how speciation can be achieved by the effect of the latitude-associated factors on the genetic divergence among populations.

Genetic diversity within a population, such as polymorphisms and personality, is considered to improve population performance because such intraspecific variations have the potential to alleviate the competition for a limited resource or the risk of predation and sexual harassment at a population level. Variation in the level and rhythm of daily activity in a population could also affect population performance by directly altering ecological, social, and sexual interactions among individuals. However, it remains to be elucidated whether such intra-population variation in the level and rhythms of daily activity exists in a natural population. Here, we investigated the genetic variation in daily activity within a single natural population of Drosophila immigrans. We established 21 isofemale lines from a single natural population and measured larval activity level and the level and daily pattern of adult activity over a 24 hr period. Larval activity level significantly varied among isofemale lines. Likewise, the activity level in the adult stage significantly varied among lines. The significant variation was also found in the daily pattern of adult activity; some lines showed greater activity level in the daytime, and others showed greater activity level in the night. Our results consistently suggest that there is a genetic variation in behavioral activity in a natural population, probably contributing to shaping the population performance.

Studying the mechanisms of the establishment of a population in a novel environment allows us to examine the process of local adaptations and subsequent range expansion. In a river system, detecting genetic or phenotypic differences between a freshwater and brackish water population could contribute to our understanding of the initial process of brackish water adaptation. Here, we investigated behavioural and gene expression responses to salt water in a freshwater and brackish water population of the freshwater snail, Semisulcospira reiniana. Although the individuals in brackish water exhibited significantly higher activity in saltwater than freshwater individuals just after sampling, the activity of freshwater individuals had increased in the second observation after rearing, suggesting that their salinity tolerance was plastic rather than genetic. We found 476 and 1002 differentially expressed genes across salinity conditions in the freshwater and brackish water populations, respectively. The major biological process involved in the salinity response of the freshwater population was the biosynthesis and metabolic processing of nitrogen-containing compounds, but that of the brackish water population was influenced by the chitin metabolic process. These results suggest that phenotypic plasticity induces adaptation to brackish water in the freshwater snail by modifying its physiological response to salinity.

Evolutionary ecological theory suggests that selection arising from interactions with conspecifics, such as sexual and kin selection, may result in evolution of intraspecific conflicts and evolutionary 'tragedy of the commons'. Here, we propose that such an evolution of conspecific conflicts may affect population dynamics in a way that enhances species coexistence. Empirical evidence and theoretical models suggest that more abundant species is more susceptible to invasion of 'selfish' individuals that increase their own reproductive success at the expense of population growth (intraspecific adaptation load). The density-dependent intraspecific adaptation load gives rise to a self-regulation mechanism at the population level, and stabilizes species coexistence at the community level by negative frequency-dependence.

Niche conservatism is the tendency of lineages to retain the same niche as their ancestors. It constrains biological groups and prevents ecological divergence. However, theory predicts that niche conservatism can hinder gene flow, strengthen drift and increase local adaptation: does it mean that it also can facilitate speciation? Why does this happen? We aim to answer these questions. We examined the variation of chloroplast DNA, genome-wide single nucleotide polymorphisms, morphological traits and environmental variables across the Dioon merolae cycad populations. We tested geographical structure, scenarios of demographic history, and niche conservatism between population groups. Lineage divergence is associated with the presence of a geographical barrier consisting of unsuitable habitats for cycads. There is a clear genetic and morphological distinction between the geographical groups, suggesting allopatric divergence. However, even in contrasting available environmental conditions, groups retain their ancestral niche, supporting niche conservatism. Niche conservatism is a process that can promote speciation. In D. merolae, lineage divergence occurred because unsuitable habitats represented a barrier against gene flow, incurring populations to experience isolated demographic histories and disparate environmental conditions. This study explains why cycads, despite their ancient lineage origin and biological stasis, have been able to diversify into modern ecosystems worldwide.

Polymorphisms in a population are expected to increase the growth rate and the stability of the population, leading to the expansion of geographical distribution and mitigation of extinction risk of a species. However, the generality of such ecological consequences of colour polymorphism remains uncertain. Here, via a comparative approach, we assessed whether colour polymorphisms influence climatic niche breadth and extinction risk in some groups of damselflies, butterflies and vertebrates. The climatic niche breadth was greater, and extinction risk was lower in polymorphic species than in monomorphic species in all taxa analysed. The results suggest that colour polymorphism facilitates range expansion and species persistence.

Abstract: Many Odonata species exhibit female-limited polymorphisms, where one morph is similar to the conspecific male in body color and other traits (andromorph), whereas one or more other morphs differ from the male (gynomorphs). Here we investigated the differentially expressed transcripts (DETs) among males and two female morph groups (gynomorphs and andromorphs) using RNA-seq to identify candidate transcripts encoding female-limited polymorphisms in the damselfly Ischnura senegalensis. Seven DETs that had significantly different expression levels between males and gynomorphs, but not between males and andromorphs, were identified. The expression levels of four of these candidate genes, doublesex (dsx), black, ebony, and chaoptin (chp), were selected for further analysis using qRT-PCR. Sequence analysis of the dsx amplicons revealed that this gene produced at least three transcripts. Two short transcripts were mainly expressed in males and andromorphs, whereas the long transcript was specifically expressed in both morph female groups that is, the expression pattern of the dsx splice variants in andromorphs was an intermediate between that of males and gynomorphs. Because the dsx gene functions as a transcription factor that regulates the sex-specific expression of multiple genes, its splice variants in I. senegalensis may explain why the andromorph is female but exhibits some masculinized traits. Because we did not detect different coding sequences of the candidate genes among the different morphs, a diallelic genomic region controlling alternative splicing of dsx, thus determining female-limited polymorphism in I. senegalensis most likely lies in a non-coding region of the dsx gene or in a gene upstream of it.

Although genetic diversity within a population is suggested to improve population-level fitness and productivity, the existence of these effects is controversial because empirical evidence for an ecological effect of genetic diversity and the underlying mechanisms is scarce and incomplete. Here, we show that the natural single-gene behavioural polymorphism (Rover and sitter) in Drosophila melanogaster has a positive effect on population fitness. Our simple numerical model predicted that the fitness of a polymorphic population would be higher than that expected with two monomorphic populations, but only under balancing selection. Moreover, this positive diversity effect of genetic polymorphism was attributable to a complementarity effect, rather than to a selection effect. Our empirical tests using the behavioural polymorphism in D. melanogaster clearly supported the model predictions. These results provide direct evidence for an ecological effect of genetic diversity on population fitness and its condition dependence.

Elucidating the mechanism shaping the spatial variations of traits has long been a central concern of evolutionary biologists. Geographic clines of allele/morph frequencies along environmental gradients are suggested to be established and maintained by the balancing of two opposing evolutionary forces, namely selection that generates spatial differentiation in morph frequencies, and selection and/or stochastic factors that lead to the coexistence of multiple morphs within a population. Thus, testing for both selection and stochastic factors is necessary for a comprehensive understanding of the mechanism underlying clinal variation in morph/allele frequency in natural populations. Here, I identified the evolutionary forces responsible for clinal variation of color morph frequency in Ischnura senegalensis by comparing the population divergence of putatively neutral loci generated by high-throughput next-generation sequencing (F (STn)) with that of the putative color locus (F (STc)). No strong correlation was observed between F (STn) and F (STc), suggesting that stochastic factors contribute less to color-locus population divergence. F (STc) was less than F (STn) between populations exposed to similar environmental conditions, but greater than F (STn) between populations exposed to different environmental conditions, suggesting that both balancing selection and divergent selection act on the color locus. Therefore, two antagonistic selection factors rather than stochastic and historical factors contribute to establishing the clinal variation of morph frequency in I. senegalensis.

Understanding the mechanisms shaping the spatiotemporal distribution of species has long been a central concern of ecology and evolutionary biology. Contemporary patterns of plant assemblies suggest that sexual interactions among species, i.e., reproductive interference, lead to the exclusive distributions of closely related species that share pollinators. However, the fitness consequences and the initial ecological/evolutionary responses to reproductive interference remain unclear in nature, since reproductive isolation or allopatric distribution has already been achieved in the natural community. In Japan, three species of blue-eyed grasses (Sisyrinchium) with incomplete reproductive isolation have recently colonized and occur sympatrically. Two of them are monomorphic with white flowers, whereas the other exhibits heritable color polymorphism (white and purple morphs). Here we investigated the effects of the presence of two monomorphic species on the distribution and reproductive success of color morphs. The frequency and reproductive success of white morphs decreased in area where monomorphic species were abundant, while those of purple morphs did not. The rate of hybridization between species was higher in white morphs than in the purple ones. Resource competition and habitat preference seemed not to contribute to the spatial distribution and reproductive success of two morphs. Our results supported that color-dependent reproductive interference determines the distribution of flower color polymorphism in a habitat, implying ecological sorting promoted by pollinator-mediated reproductive interference. Our study helps us to understand the evolution and spatial structure of flower color in a community.

What limits a species' distribution in the absence of physical barriers? Genetic load due to asymmetric gene flow and the absence of genetic variation due to lack of gene flow are hypothesized to constrain adaptation to novel environments in marginal populations, preventing range expansion. Here, we examined the genetic structure and geographic variation in morphological traits in two damselflies (Ischnura asiatica and I. senegalensis) along a latitudinal gradient in Japan, which is the distribution centre of I. asiatica and the northern limit of I. senegalensis. Genomewide genetic analyses found a loss of genetic diversity at the edge of distribution in I. senegalensis but consistently high diversity in I. asiatica. Gene flow was asymmetric in a south-north direction in both species. Although body size and wing loading showed decreasing latitudinal clines (smaller in north) in I. asiatica in Japan, increasing latitudinal clines (larger in north) in these phenotypic markers were observed in I. senegalensis, particularly near the northern boundary, which coincided well with the location where genetic diversity began a sharp decline. In ectothermic animals, increasing latitudinal cline in these traits was suggested to be established when they failed to adapt to thermal gradient. Therefore, our findings support the possibility that a lack of genetic variation rather than geneflow swamping is responsible for the constraint of adaptation at the margin of geographic distribution.

Odonata (dragonflies and damselflies) present an unparalleled insect model to integrate evolutionary genomics with ecology for the study of insect evolution. Key features of Odonata include their ancient phylogenetic position, extensive phenotypic and ecological diversity, several unique evolutionary innovations, ease of study in the wild and usefulness as bioindicators for freshwater ecosystems worldwide. In this review, we synthesize studies on the evolution, ecology and physiology of odonates, highlighting those areas where the integration of ecology with genomics would yield significant insights into the evolutionary processes that would not be gained easily by working on other animal groups. We argue that the unique features of this group combined with their complex life cycle, flight behaviour, diversity in ecological niches and their sensitivity to anthropogenic change make odonates a promising and fruitful taxon for genomics focused research. Future areas of research that deserve increased attention are also briefly outlined.

Negative frequency-dependent selection derived from positive frequency-dependent foraging is the best-known selection force maintaining genetic polymorphism within a population. However, in flowering plants, positive frequency-dependent foraging by pollinators is expected to accelerate the loss of low-frequency morphs by conferring a fitness advantage to the common morph, leading to monomorphism. In Japan, a non-native species, Sisyrinchium sp., exhibits conspicuous flower color polymorphism within a population comprising both purple morphs (homozygous recessive) and white morphs (heterozygous or homozygous dominant). Here we quantified genotype-specific reproductive success in order to reveal the contribution of overdominant selection on the maintenance of flower color polymorphism in this species. In artificial pollination experiments using individuals with identified genotypes, female reproductive success was higher in the heterozygote than in either homozygote. The frequency of purple morphs in natural populations (ca. 31 %) is similar to the frequency predicted by overdominant selection (25 %). Our results suggest that overdominant selection contributes to the maintenance of color morphs in the natural population of this species.

A continuous spatial gradient in visible traits, which is called a cline, is a natural model system for quantifying the effects of selection and stochastic factors and their relative importance. Geographic clines in phenotypic traits also provide key insights into the evolutionary forces that lead to allopatric speciation in nature. Thus, the underlying mechanisms for establishing clines and their evolutionary consequences remain key topics in evolutionary biology. However, few experimental studies have confirmed the underlying mechanisms of geographic clines in morph/allele frequencies, probably because of the lack of understanding of the theoretical basis of geographic clines in polymorphisms and/or suitable comprehensive tests. Thus, I present a general review of the underlying mechanisms for establishing geographic clines in polymorphisms. I also provide a case study using the female dimorphic damselfly Ischnura senegalensis to illustrate a strategy that confirms the underlying mechanisms of geographic clines in morph frequencies. This review may help to address geographic clines in other polymorphic systems, as well as contribute to a comprehensive understanding of geographic clines in quantitative traits, and thus, their evolutionary consequences in nature.

Animals with color vision use color information in intra-and interspecific communication, which in turn may drive the evolution of conspicuous colored body traits via natural and sexual selection. A recent study found that the transparent wings of small flies and wasps in lower-reflectance light environments display vivid and stable structural color patterns, called "wing interference patterns" (WIPs). Such WIPs were hypothesized to function in sexual selection among small insects with wing displays, but this has not been experimentally verified. Here, to our knowledge we present the first experimental evidence that WIPs in males of Drosophila melanogaster are targets of mate choice from females, and that two different color traits-saturation and hue-experience directional and stabilizing sexual selection, respectively. Using isogenic lines from the D. melanogaster Genetic Reference Panel, we compare attractiveness of different male WIPs against black and white visual backgrounds. We show that males with more vivid wings are more attractive to females than are males with dull wings. Wings with a large magenta area (i.e., intermediate trait values) were also preferred over those with a large blue or yellow area. These experimental results add a visual element to the Drosophila mating array, integrating sexual selection with elements of genetics and evo-devo, potentially applicable to a wide array of small insects with hyaline wings. Our results further underscore that the mode of sexual selection on such visual signals can differ profoundly between different color components, in this case hue and saturation.

The effect of evolutionary changes in traits and phenotypic/genetic diversity on ecological dynamics has received much theoretical attention; however, the mechanisms and ecological consequences are usually unknown. Female-limited colour polymorphism in damselflies is a counter-adaptation to male mating harassment, and thus, is expected to alter population dynamics through relaxing sexual conflict. Here we show the side effect of the evolution of female morph diversity on population performance (for example, population productivity and sustainability) in damselflies. Our theoretical model incorporating key features of the sexual interaction predicts that the evolution of increased phenotypic diversity will reduce overall fitness costs to females from sexual conflict, which in turn will increase productivity, density and stability of a population. Field data and mesocosm experiments support these model predictions. Our study suggests that increased phenotypic diversity can enhance population performance that can potentially reduce extinction rates and thereby influence macroevolutionary processes.

Understanding the relative importance of selection and stochastic factors in population divergence of adaptive traits is a classical topic in evolutionary biology. However, it is difficult to separate these factors and detect the effects of selection when two or more contrasting selective factors are simultaneously acting on a single locus. In the damselfly Ischnura senegalensis, females exhibit color dimorphism and morph frequencies change geographically. We here evaluated the role of selection and stochastic factors in population divergence of morph frequencies by comparing the divergences in color locus and neutral loci. Comparisons between population pairwise F ST for neutral loci and for the color locus did not detect any stochastic factors affecting color locus. Although comparison between population divergence in color and neutral loci using all populations detected only divergent selection, we detected two antagonistic selective factors acting on the color locus, that is, balancing and divergent selection, when considering geographical distance between populations. Our results suggest that a combination of two antagonistic selective factors, rather than stochastic factors, establishes the geographic cline in morph frequency in this system. © 2014 Macmillan Publishers Limited All rights reserved.

Understanding the mechanisms that maintain genetic diversity within a population remains a primary challenge for evolutionary biology. Of the processes capable of maintaining variation, negative frequency-dependent selection (NFDS), under which rare phenotypes (or alleles) enjoy a high fitness advantage, is suggested to be the most powerful. However, few experimental studies have confirmed that this process operates in nature. Although a lot of suggestive evidence has separately been provided in various polymorphic systems, these are not enough to prove the existence of NFDS in each system. Here we present a general review of NFDS and point out some problems with previous works to develop reasonable alternative research strategies for testing NFDS. In the second half of this paper, we focused on NFDS in the common bluetail damselfly, Ischnura senegalensis, that shows female-limited genetic polymorphism. We show (1) the proximate causal mechanisms of the frequency-dependent process, (2) frequency-dependent inter-morph interaction, (3) rare morph advantage and (4) morph frequency oscillations in a natural population. These results provide unequivocal empirical support for NFDS in a natural system.

&lt;title content-type="main"&gt;Abstract Resource partitioning within a species, trophic polymorphism is hypothesized to evolve by disruptive selection when intraspecific competition for certain resources is severe. However, in this study, we reported the secondary partitioning of oviposition resources without resource competition in the damselfly Ischnura senegalensis. In this species, females show color polymorphism that has been evolved as counteradaptation against sexual conflict. One of the female morphs is a blue-green (andromorph, male-like morph), whereas the other morph is brown (gynomorph). These female morphs showed alternative preferences for oviposition resources (plant tissues); andromorphs used fresh (greenish) plant tissues, whereas gynomorphs used decaying (brownish) plants tissues, suggesting that they chose oviposition resources on which they are more cryptic. In addition, the two-color morphs had different egg morphologies. Andromorphs have smaller and more elongated eggs, which seemed to adapt to hard substrates compared with those of gynomorphs. The resource partitioning in this species is achieved by morphological and behavioral differences between the color morphs that allow them to effectively exploit different resources. Resource partitioning in this system may be a by-product of phenotypic integration with body color that has been sexually selected, suggesting an overlooked mechanism of the evolution of resource partitioning. Finally, we discuss the evolutionary and ecological consequences of such resource partitioning.

Time constraints are critical for reproductive success. To understand the spatiotemporal dynamics of morph frequency in the female-dimorphic damselfly Ischnura senegalensis, we compared two different morphs for two important time constraints on female reproductive output, i.e. post-emergence sexual maturation and prolonged copulation. The females of both morphs achieved sexual maturation 4-5days after emergence, suggesting that the rate of sexual mutation does not result in morph-specific fitness. The copulation durations declined with the time of onset of copulation in both morphs. Consequently, all copulations terminated at approximately 12:00hours. Because females show foraging and oviposition activity only after copulation, the copulation duration does not result in morph-specific time constraints. These two important time constraints do not account for morph-specific reproductive success and do not affect the evolutionary equilibrium of morph frequency in I. senegalensis. © 2012 The Entomological Society of Japan.

Mate choice by females is an important component of sexual selection in many species. Theoretically, female sexual traits may be influenced by selection acting on the females via male mate choice, while the evolutionary consequences of male mate preferences are largely unknown, especially in the context of sexual conflict. We tested whether male mate choice affects the evolution of female colour in the damselfly Ischnura senegalensis in which females exhibit dimorphism consisting of a gynomorph that experiences ontogenetic colour change and an andromorph that does not. We first quantitatively confirmed that only gynomorphs change their body colour in relation to sexual maturation. In field experiments, males were unwilling to mate with sexually immature gynomorphs, although they attempted to mate with immature andromorphs. This is because males changed their mating preference for female colour depending on previous copulation experiences with sexually mature females. As a result, immature andromorphs received more male harassment than sexually immature gynomorphs, and then showed decreased food intake. Immature-specific colour functioned to avoid costly male harassment during female prereproductive stages, suggesting that ontogenetic colour changes in females have evolved as an antiharassment strategy of females via male mate choice. (C) 2012 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.

Background: Establishment of geographic morph frequency clines is difficult to explain in organisms with limited gene flow. Balancing selection, such as negative frequency-dependent selection (NFDS), is instead suggested to establish a morph frequency cline on a geographic scale at least theoretically. Here we tested whether a large-scale smooth cline in morph frequency is established by NFDS in the female-dimorphic damselfly, Ischnura senegalensis, where andromorphs and gynomorphs are maintained by NFDS. Results: We found a large-scale latitudinal cline in the morph frequency: andromorph frequency ranged from 0.05 (South) to 0.79 (North). Based on the empirical data on the numbers of eggs, the number of ovariole, abdomen length and latitude, the potential fitness of andromorphs was estimated to be lower than that of gynomorphs in the south, and higher in the north, suggesting the gene-by-environment interaction. From the morph-specific latitudinal cline in potential fitness, the frequency of andromorphs was expected to shift from 0 to 1 without NFDS, because a morph with higher potential fitness wins completely and the two morphs will switch at some point. In contrast, NFDS led to the coexistence of two morphs with different potential fitness in a certain geographic range along latitude due to rare morph advantage, and resulted in a smooth geographic cline of morph frequency. Conclusion: Our results provide suggestive evidence that the combination of NFDS and gene-by-environment interaction, i.e., multi-selection pressure on color morphs, can explain the geographic cline in morph frequency in the current system.

While male mate choice behaviour has been reported in many taxa, little is known about its plasticity and evolutionary consequences. In the damselfly Ischnura senegalensis, females exhibit colour dimorphism (gynomorph and andromorph). The body colour of gynomorphs changed ontogenetically in accordance with sexual maturation, while little change occurred in andromorphs. To test the male mate choice between sexually immature and mature females of both morphs, binary choice experiments were conducted. Virgin males that were reared separately from females after emergence did not show significant preference between sexually immature and mature females for both morphs, indicating that virgin males were unable to discriminate female reproductive status. On the other hand, males that had experienced copulation with gynomorphs preferred sexually mature gynomorphs to sexually immature ones. However, males that had experienced copulation with andromorphs could not discriminate between sexually immature and mature andromorphs, probably due to the absence of significant ontogenetic change in their thoracic colour. Therefore, female body colour is an important cue for males in discriminating between sexual maturation stages. Learned mate discrimination depending on copulation experience might help males to detect potential mates effectively and avoid sexually unreceptive immature female. We finally discuss the adaptive significance of the ontogenetic colour change in females.

Negative frequency-dependent selection (NFDS) is one of the most powerful selective forces maintaining genetic polymorphisms in nature. Recently many prospective cases of polymorphisms by NFDS have been reported. Some of them are very complicated, although strongly supportive of the NFDS. Here we investigate NFDS in wild populations of the dimorphic damselfly Ischnura senegalensis, in which females occur as andromorphs and gynomorphs. Specifically, we (1) test fitness responses to morph frequencies, (2) built a simple population genetic model, and (3) compare the observed and predicted morph-frequency dynamics. Fitnesses of the two morphs are an inverse function of its own frequency in a population, and are about equal when their frequencies are similar. Thus the conditions necessary for NFDS are satisfied. The long-term field surveys show that the morph frequencies oscillate with a period of two generations. Morph frequencies in a small population undergo large oscillations whereas those in a large population do small oscillations. The demographic properties of the observed dynamics agree well with those of our model. This example is one of the simplest confirmed cases of NFDS maintaining genetic polymorphisms in nature.

Females of coenagrionid damselflies exhibit color dimorphism, consisting of an andromorph and a gynomorph. This study compared reproductive traits between the female morphs in both field-captured and laboratory-reared females of the female-dimorphic damselfly Ischnura senegalensis. No difference was found in the onset of egg development between the morphs. The andromorphs developed significantly smaller mature eggs and had significantly more immature eggs than the gynomorphs. These results suggest that the andromorphs are r-strategists (high fecundity with small eggs), whereas the gynomorphs are K-strategists (low fecundity with large eggs). Fecundity and egg size might determine the quantity and quality of the offspring, respectively, indicating that morph-specific reproductive traits would contribute to the overall fitness of each female morph, and consequently be key factors affecting morph frequency in a population.

In animals without any courtship behaviour, persistent mating attempts by males are frequently observed. Male harassment affects female reproductive success in the laboratory, but few studies have evaluated the costs of male harassment in the wild. In the damselfly Ischnura senegalensis, females exhibit colour dimorphism (andromorph and gynomorph), and the morph frequency varies between local populations. In two populations where gynomorphs were common, we found that males harassed more gynomorphs than andromorphs throughout their daily foraging and oviposition activity period. Gynomorphs excreted less faeces than andromorphs, indicating that preferential harassment of gynomorphs decreased their food intake. Gynomorphs also produced fewer eggs than andromorphs. As a result, gynomorphs laid 35% fewer eggs per day than andromorphs, suggesting that male harassment decreased their reproductive success. (C) 2009 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.

Ischnura senegalensis females exhibit color dimorphism, consisting of an andromorph and a gynomorph, which might be maintained under a frequency-dependent process of mating harassment by mate-searching males. Males change their mating preference for female morph depending on prior copulation experience. Binary choice experiments between two female morphs were carried out in four local populations in the early morning (07.00-09.00 hours) and the afternoon (12.00-14.00 hours), times which mark the onset and the end of diurnal mating activity, respectively. According to the line census along the water&apos;s edge, the proportion of andromorphs in the female population varied from 21 to 67% throughout the survey period for four local populations. Males showed non-biased preference for female morphs in the early morning in each local population, while they chose the common morph in the afternoon. Male mating preference for female morphs was positively correlated to the proportion of female morphs in the population. If the selective mating attacks on the common female morphs inhibit their foraging and/or oviposition behavior, frequency-dependent male mating attacks might provide a selective force for maintaining the female color dimorphism in I. senegalensis.