Heterospecific Males Research Articles

Mate choice in the brain: species differ in how male traits 'turn on' gene expression in female brains.

Mate choice plays a fundamental role in speciation, yet we know little about the molecular mechanisms that underpin this crucial decision-making process. Stickleback fish differentially adapted to limnetic and benthic habitats are reproductively isolated and females of each species use different male traits to evaluate prospective partners and reject heterospecific males. Here, we integrate behavioural data from a mate choice experiment with gene expression profiles from the brains of females actively deciding whether to mate. We find substantial gene expression variation between limnetic and benthic females, regardless of behavioural context, suggesting general divergence in constitutive gene expression patterns, corresponding to their genetic differentiation. Intriguingly, female gene co-expression modules covary with male display traits but in opposing directions for sympatric populations of the two species, suggesting male displays elicit a dynamic neurogenomic response that reflects known differences in female preferences. Furthermore, we confirm the role of numerous candidate genes previously implicated in female mate choice in other species, suggesting evolutionary tinkering with these conserved molecular processes to generatedivergent mate preferences. Taken together, our study adds important new insights to our understanding of the molecular processes underlying female decision-making critical for generating sexual isolation and speciation.

Heterospecific interference modulates the reproductive traits of a ladybird beetle

AbstractReproductive interference between species is vital to understand interspecific interactions along with their ecological and evolutionary patterns in nature. We studied potential reproductive interference between two sympatric species of ladybird beetles Propylea dissecta and Cheilomenes sexmaculata (Coleoptera: Coccinellidae). Our aim was to determine whether heterospecific matings occur between these two species, and if so, how they affect the reproductive output of females of both species. We found that heterospecific mating occurs, with P. dissecta females mating with C. sexmaculata males, but not vice versa. To compare the effect of conspecific and heterospecific males on the reproductive output of P. dissecta females, we used two mixed mating treatments: conspecific mixed (an additional P. dissecta male with conspecific pair of P. dissecta), and heterospecific mixed (a C. sexmaculata male with conspecific pair of P. dissecta). We found that conspecific interference did not affect reproductive parameters. However, the presence of a heterospecific C. sexmaculata male interfered with conspecific matings in P. dissecta through multiple mating attempts, resulting in reduced fitness and reproductive success of P. dissecta. Our results indicate that the presence of heterospecific males significantly affected the reproductive output of P. dissecta, apparently as a result of the greater attempted mating activity of C. sexmaculata males. Our findings may be relevant for effective biocontrol strategies involving these insect species.

An experimental test of the evolutionary consequences of sympatry in Drosophila subquinaria.

Prezygotic isolation is often stronger between sympatric as opposed to allopatric taxa, but the underlying cause can be difficult to infer from comparative studies alone. Experimental evolution, where evolutionary responses to treatments manipulating the presence/absence of heterospecific individuals are tracked, can provide a powerful complementary approach. We used experimental evolution to investigate a naturally occurring pattern of reproductive character displacement in the mushroom-feeding fly, Drosophila subquinaria. In nature, female D. subquinaria from populations sympatric with the closely related Drosophila recens discriminate more strongly against heterospecific males than do females from allopatric populations. Starting with 16 replicate allopatric populations of D. subquinaria, we manipulated the presence/absence of D. recens during mating (experimental sympatry vs. control) and, when present, we allowed hybrids to live or kill them each generation. Across 12 generations, heterospecific offspring production from no-choice mating trials between D. subquinaria females and D. recens males declined in both experimental sympatry treatments relative to the control, suggesting increased sexual isolation. Male cuticular hydrocarbon profiles also evolved, but only in the hybrids killed treatment. Our results strongly imply that the existing reproductive character displacement in wild D. subquinaria populations was an evolutionary response to selection arising from secondary contact with D. recens.

Asymmetry in the reproductive interference between two closely related species of spider mites, Panonychus citri and Panonychus osmanthi (Prostigmata: Tetranychidae).

An incomplete species recognition system may cause reproductive interference. Interspecific mating between Panonychus citri and Panonychus osmanthi produced apparently normal F1 hybrid females. However, the F1 females laid few eggs, and the produced eggs did not hatch (suggesting hybrid breakdown). Several common host plants of P. citri and P. osmanthi have been determined, including Japanese pear, peach, and citrus. However, P. osmanthi has been found only on its particular host plants in the genus Osmanthus. To assess the potential for reproductive interference between P. citri and P. osmanthi, we evaluated the competitiveness (1 ♀ × 2 ♂♂) and preference (2 ♀♀ × 1 ♂) of males in mating and guarding behavior and assessed the effect of second mating on offspring paternity using Japanese pear leaf disks. We found that P. citri males were superior competitors. Moreover, the species recognition ability of P. citri males was lower than that of P. osmanthi males, and frequent occurrence of reproductive interference should therefore be expected. In a mixed population (2 ♀♀ × 2 ♂♂), 41% of females copulated with heterospecific males, although the mating superiority of P. citri was not proven. The ratio of effective second mating with conspecific males after heterospecific mating was higher in the crosses to P. citri females (77.8%) than P. osmanthi females (14.3%). Therefore, P. citri females had more chance than P. osmanthi females to recover from the fitness cost imposed by heterospecific mating through subsequent conspecific mating. To date, P. osmanthi has not been detected on host plants other than Osmanthus species. Reproductive interference by P. citri males may therefore narrow the range of host plants available for P. osmanthi.

Integrative characterization of genetic and phenotypic differentiation in an ant species complex with strong hierarchical population structure and low dispersal abilities.

Low dispersal, occurrence of asexual reproduction and geographic discontinuity increase genetic differentiation between populations, which ultimately can lead to speciation. In this work, we used a multidisciplinary framework to characterize the genetic and phenotypic differentiation between and within two cryptic ant species with restricted dispersal, Cataglyphis cursor and C. piliscapa and used behavioral experiments to test for reproductive isolation. Their distribution is segregated by the Rhône River and they have been traditionally distinguished only by hair numbers, although a statistical assessment is still lacking. We found strong genetic (microsatellites, nuclear and mitochondrial sequences), morphological (number of hairs, tibia length, male genitalia) and chemical (cuticular hydrocarbons) differentiation not only between species but also among localities within species. However, inter-specific differentiation was slightly higher than intra-specific differentiation for most markers. Overall, this pattern could either reflect reproductive isolation or could result from a longer period of geographic isolation between species than among localities within species without necessarily involving reproductive isolation. Interestingly, our behavioral experiments showed an absence of mating between species associated to a higher aggressiveness of workers towards heterospecific males. This suggests that sexual selection may, at least partially, fuel reproductive isolation. We also showed that cuticular hydrocarbons, mtDNA sequences and number of hairs provide reliable criteria allowing species discrimination. Overall, this species complex offers a case study to further investigate varying stages of a speciation continuum by estimating reproductive isolation between pairs of localities varying by their level of genetic differentiation.

Evidence for stronger discrimination between conspecific and heterospecific mating partners in sexual vs. asexual female freshwater snails.

Once-useful traits that no longer contribute to fitness tend to decay over time. Here, we address whether the expression of mating-related traits that increase the fitness of sexually reproducing individuals but are likely less useful or even costly to asexual counterparts seems to exhibit decay in the latter. Potamopyrgus antipodarum is a New Zealand freshwater snail characterized by repeated transitions from sexual to asexual reproduction. The frequent coexistence of sexual and asexual lineages makes P. antipodarum an excellent model for the study of mating-related trait loss. Under the presumption (inherent in the Biological Species Concept) that failure to discriminate between conspecific and heterospecific mating partners represents a poor mate choice, we used a mating choice assay including sexual and asexual P. antipodarum females and conspecific (presumed better choice) vs. heterospecific (presumed worse choice) males to evaluate the loss of behavioral traits related to sexual reproduction. We found that sexual females engaged in mating behaviors with conspecific mating partners more frequently and for a greater duration than with heterospecific mating partners. By contrast, asexual females mated at similar frequency and duration as sexual females, but did not mate more often or for longer duration with conspecific vs. heterospecific males. While further confirmation will require inclusion of a more diverse array of sexual and asexual lineages, these results are consistent with a scenario where selection acting to maintain effective mate discrimination in asexual P. antipodarum is weak or ineffective relative to sexual females and, thus, where asexual reproduction is associated with the evolutionary decay of mating-related traits in this system.

Male toads change their aggregation behaviour when hybridization is favoured

In many species, males form aggregations in which they compete for access to mates. Such aggregations can consist of multiple species, such that hybridization or reproductive interference between species becomes a risk. To enhance mating success with conspecifics, males might preferentially aggregate with conspecifics to minimize the risks of mating or competing with heterospecifics, especially if females are attracted to groups of conspecific males or avoid heterospecific males. Here, we used spadefoot toads (genus Spea ) to evaluate whether males respond to the same environmental cue that females use to facultatively alter their mate choice decisions for conspecific versus heterospecific males. We took advantage of a system in which females facultatively prefer to mate with heterospecifics when doing so is adaptive. We found that males preferentially aggregate with conspecific males when hybridization is disfavoured for females (so that females prefer conspecifics); however, males show no such preference for aggregating with conspecifics when hybridization is favoured (and females are more likely to prefer heterospecifics). We did not find this pattern in a different spadefoot species where females do not adaptively hybridize; males preferentially aggregated with conspecifics in one environment and did not alter their behaviour when the environmental cue was changed. Our results therefore suggest that males track the same environmental cues that females use during mate choice. These findings highlight the potential for males to facultatively adjust their behaviour in response to competitors, as well as the role that male–male interactions might play in the likelihood and consequences of hybridization between species. • Males aggregate with conspecifics when hybridization is disfavoured. • When hybridization is favoured, males spend less time with conspecifics. • This context-dependent behaviour mirrors female mate choice. • We found no such behaviour in a congeneric species lacking adaptive hybridization.

The initial response of females towards congeneric males matches the propensity to hybridise in Ophthalmotilapia

Cichlid radiations often harbour closely related species with overlapping niches and distribution ranges. Such species sometimes hybridise in nature, which raises the question how they can coexist. This also holds for the Tanganyika mouthbrooders Ophthalmotilapia ventralis and O. nasuta. Earlier studies found indications of asymmetrical hybridisation with females of O. ventralis accepting males of O. nasuta, but not the other way around. We hypothesised that this was due to differences in the capacity for species recognition. Given the higher propensity of O. ventralis females towards hybridisation, we expect a reduced ability for species recognition in O. ventralis females, compared to O. nasuta females. We staged two experiments, one focusing on 22 female O. nasuta and one on 21 female O. ventralis. These fish were placed in one half of a tank and briefly exposed to a conspecific or a heterospecific male, a conspecific female, or nothing (control). Female response was evaluated by scoring six tracking parameters and by noting the occurrence of ten discrete behaviours before and during the encounter. Females always responded to the presence of another fish by approaching it. Remarkably, for both O. nasuta and O. ventralis, we did not find a different response between encounters with conspecific males and females. However, in agreement with our hypothesis, females of O. nasuta behaved differently towards conspecific or heterospecific males, whereas females of O. ventralis did not. When presented with a heterospecific male, females of O. nasuta performed a lower number of ‘ram’ behaviours. Additionally, they never displayed the ‘flee’ behaviour, a component of the species’ mating repertoire that was seen in all but one of the presentations with a conspecific male. Our findings show that differences in species recognition at first encounter predict to a large degree the outcome of the mating process, even in the absence of mating behaviour.

Female preferences for conspecific males indicate reproductive isolation between sympatric Labeotropheus Ahl from Lake Malaŵi

AbstractIn sexually reproducing taxa, reproductive isolation is key to speciation. Given the speed with which behavioral reproductive isolation evolves, this form of isolation is of particular importance to rapidly evolving species. Here we examine behavioral reproductive isolation in a sympatric, congeneric pair of rock‐dwelling cichlids from Lake Malaŵi, testing the hypothesis that females should prefer conspecific males. We used a pair of Labeotropheus, L. fuelleborni Ahl and L. trewavasae Fryer, from the Chidunga Rocks in southwestern Lake Malaŵi, to test this hypothesis, with the prediction that females would approach conspecific males more frequently than they would heterospecific males. When given a choice between a male L. fuelleborni and L. trewavasae, we found that females did approach males of their own species more frequently than males of the opposite species, and that differences in length and the number of eggspots on the anal fin between the stimulus males had no effect on a female's choice. These results are consistent with behavioral reproductive isolation between these sympatric species, and add further support to the hypothesis that sexual selection played a key role in the speciation of cichlid fishes from Lake Malaŵi.

Testing potential mechanisms of conspecific sperm precedence in Drosophila pseudoobscura.

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

Male–male interactions select for conspicuous male coloration in damselflies

Male ornamentation, such as conspicuous male coloration, can evolve through female mate choice. Alternatively, in species without overt female mate preference, conspicuous male coloration can evolve as a badge of status via intrasexual selection to avoid costly fights or prevent mating attempts with other males. Here, we investigated the drivers of conspicuous male coloration in an ontogenetic colour-changing damselfly, Xanthagrion erythroneurum, where the juvenile males are yellow and change colour to red upon sexual maturity. We first showed that red males were chromatically and achromatically more conspicuous than the yellow males. We then quantified the condition of the males: red males were larger and in better condition than yellow males. We tested female preference in a choice experiment where we artificially manipulated male colour and found that females did not choose mates based on male coloration. We further tested whether the male coloration affected male–male interactions. We presented red and yellow males at the breeding site and measured behavioural responses from conspecific males and heterospecific ( Ischnura heterosticta ) males. Red males received less aggression from conspecifics and heterospecifics than yellow males. Our study experimentally showed that male conspicuousness is not a target of female mate choice in damselflies. Instead, intra- and interspecific male–male interactions appear to be the driver of its evolution. • X. erythroneurum changes colour from yellow to red during ontogeny. • Red males are more conspicuous and in better condition than yellow males. • Females do not prefer more conspicuous red males to yellow males. • Red males incur less within- and across-species aggression. • Males' conspicuous colour appears to have evolved to reduce male-male interactions.

Hybridization, Behavioral Patterns, and Pre- and Postzygotic Isolation Between Two Recently Derived Species of Walnut-Infesting Rhagoletis Fruit Flies in the Highlands of Mexico.

Episodes of isolation and secondary contact among populations of insects of Nearctic origin during Pleistocene glacial/postglacial climatic cycles had a strong evolutionary influence on the diversity of flies in the genus Rhagoletis in mountainous areas of Mexico. As a series of experiments undertaken to gather support for phylogenetic hypotheses on the origin of three walnut-infesting species in the suavis group, we examined pre- and postzygotic isolation between Rhagoletis completa Cresson, 1929 and R. ramosae Hernández-Ortiz, 1985. Despite morphological, biological, and behavioral differences, these two species were found to be capable of hybridization. Mating experiments in large enclosures revealed asymmetric sexual isolation. There were notable differences in male sexual behavior. While R. ramosae males mated exclusively on host fruit, R. completa males used fruit and alternative mating locations. During fruit-guarding and male-male contests, R. completa and R. ramosae males adopted markedly different wing postures. R. completa females were more reluctant to copulate with heterospecific males than R. ramosae females. During no choice crosses in small enclosures, there was a reduction of egg hatch for the hybrid cross of R. completa males × R. ramosae females. Our results and previous studies on reproductive isolation between other species pairs in the suavis group support a clade in which R. ramosae, R. zoquiBush, 1966, and R. completa are close relatives all still capable of hybridizing.

Heterospecific pollination by an invasive congener threatens the native American bittersweet, Celastrus scandens

Invasive plants have the potential to interfere with native species’ reproductive success through a number of mechanisms, including heterospecific pollination and hybridization. This study investigated reproductive interactions between a native North American woody vine (American bittersweet, Celastrus scandens) and an introduced congener (oriental bittersweet, C. orbiculatus). The decline of C. scandens in the eastern portion of its range is coincident with the introduction and spread of C. orbiculatus, and the two species are known to hybridize. The relationship between proximity and floral production of conspecific and heterospecific males on fertilization and hybridization rates was measured at a field site in northwestern Indiana, USA where both species occur and reproduce. We found that the invasive vine had an extreme advantage in both male and female floral production, producing nearly 200 times more flowers per staminate plant and 65 times more flowers per pistillate plant than the native. Using nuclear microsatellite DNA markers we found that hybridization rates were asymmetric; 39% of the C. scandens seeds tested were hybrids, compared to only 1.6% of C. orbiculatus seeds. The asymmetric hybridization rates were likely not solely due to greater abundance of C. orbiculatus pollen because experimental hand crosses revealed that C. scandens had a higher rate (41%) of heterospecific fertilization than C. orbiculatus (2.4%). We previously reported that few hybrids were observed in the wild, and hybrids had greatly reduced fecundity. Thus, in our system, the threat posed by heterospecific pollen is not replacement by hybrids or introgression, but rather asymmetric reproductive interference. Reproductive interference extended to distances as great as 100 meters, thus, efforts to conserve the native species must reduce its exposure to C. orbiculatus over a relatively large spatial scale.

Reproductive interference hampers species coexistence despite conspecific sperm precedence.

Negative interspecific mating interactions, known as reproductive interference, can hamper species coexistence in a local patch and promote niche partitioning or geographical segregation of closely related species. Conspecific sperm precedence (CSP), which occurs when females that have mated with both conspecific and heterospecific males preferentially use conspecific sperm for fertilization, might contribute to species coexistence by mitigating the costs of interspecific mating and hybridization. We discussed whether two species exhibiting CSP can coexist in a local environment in the presence of reproductive interference. First, using a behaviorally explicit mathematical model, we demonstrated that two species characterized by negative mating interactions are unlikely to coexist because the costs of reproductive interference, such as loss of mating opportunity with conspecific partners, are inevitably incurred when individuals of both species are present. Second, we experimentally examined differences in mating activity and preference in two Harmonia ladybird species known to exhibit CSP. These behavioral differences may lead to local extinction of H. yedoensis because of reproductive interference by H. axyridis. This prediction is consistent with field observations that H. axyridis uses various food sources and habitats whereas H. yedoensis is confined to a less preferred prey item and a pine tree habitat. Finally, by a comparative approach, we observed that niche partitioning or parapatric distribution, but not sympatric coexistence in the same habitat, is maintained between species with CSP belonging to a wide range of taxa, including vertebrates and invertebrates living in aquatic or terrestrial environments. Taken together, it is possible that reproductive interference may destabilize local coexistence even in closely related species that exhibit CSP.

Interspecific Mating Effects on Locomotor Activity Rhythms and Refractoriness of Aedes albopictus (Diptera: Culicidae) Females.

Simple SummaryThe superiority of Aedes albopictus in larval resource competition was originally proposed as the cause of displacements of Aedes aegypti in the USA. However, satyrization, a form of reproductive interference, was later invoked as an alternative or complementary mechanism for the observed displacements of Aedes aegypti. This study tests the hypotheses that Ae. albopictus female activity is not altered by the presence of accessory gland extracts from conspecific and heterospecific males, and Ae. albopictus females remain receptive to conspecific males even after receiving accessory gland (AG) extracts from Ae. aegypti males. We performed experiments with a control group (females injected with saline), a group of females injected with accessory gland extracts of Ae. aegypti males and a group of females injected with accessory gland extracts of Ae. albopictus males and measured the locomotor activity and the ability of inseminated females to copulate with conspecific males. Females injected with conspecific and heterospecific extracts showed significant decreases in total and diurnal activity. Females injected with heterospecific extracts showed significant decreases in nocturnal activity. A total of 83% of females injected with heterospecific and 10% of females injected with conspecific extracts copulated with conspecific males. These results considered together with our previous report on effects of interspecific mating and cross-species injections of AG products on Ae. aegypti females, show consistent depressions of locomotor activities between species, but the loss of sexual receptivity only in Ae. aegypti. We propose that different male seminal fluid proteins control these activities.This study tests the hypotheses that the locomotor activity of Ae. albopictus females is not significantly altered by the presence of accessory gland (AG) extracts from conspecific and heterospecific males, and that Ae. albopictus females remain receptive to mating with conspecific males even after receiving AG of Ae. aegypti males. Virgin Ae. albopictus females were injected with saline (control group), AG extracts of Ae. aegypti males (aegMAG) or AG extracts of Ae. albopictus males (albMAG). Locomotor activity was evaluated under 12 h of light and 12 h of darkness at 25 °C. All live Ae. albopictus females were subsequently exposed to conspecific males for 48 h, and their spermathecae were dissected for the presence of sperm. Females injected with aegMAG and albMAG showed significant decreases in total, diurnal and diurnal without lights-on Period activities. Females injected with aegMAG showed significant decreases in nocturnal and nocturnal without lights-off period activities. Females injected with albMAG showed significant decreases in lights-off activity. A total of 83% of Ae. albopictus females injected with aegMAG and 10% of females injected with albMAG were inseminated by conspecific males. These results, coupled with our previous paper on MAG and interspecific mating effects on female Ae. aegypti, demonstrate contrasting outcomes on locomotor activities and loss of sexual receptivity, both conspecific and heterospecific MAGs capable of sterilizing virgin Ae. aegypti, but only conspecific MAGs sterilizing Ae. albopictus, whereas locomotor activities were depressed in females of both species after heterospecific and conspecific injections or treatments.

Genes underlying species differences in cuticular hydrocarbon production between Drosophila melanogaster and D. simulans.

Surface chemical compounds are key components of survival and reproduction in many species. Cuticular hydrocarbons (CHCs) are chemical compounds produced by all insects that are used for both desiccation resistance and chemical communication, including communication related to mating. In the species pair of Drosophila melanogaster and D. simulans, female CHCs stimulate conspecific males to mate and repel heterospecific males. While CHCs are a critical contributor to both reproductive success within a species and isolation between species, few genes underlying species variation in CHC profiles are known. Here, we use genetic mapping of the 3rd chromosome to test a suite of candidate genes for interspecies variation in CHCs. Candidate gene CG5946 was found to be involved in species differences in the production of 7,11-heptacosadiene and 7-tricosene between D. melanogaster and D. simulans. This is therefore a new candidate locus contributing to species-specific variation in the CHC profile. In the process of mapping genes for CHCs, we also identified 29 candidate genes for the reduced survival or inviability of interspecies hybrids.

Response to Comment on "Females engaging in adaptive hybridization prefer high-quality heterospecifics as mates".

Braun et al contend that we did not account for survival, but we did. Differential survival does not alter our conclusions, which were also robust to removing anomalous families. They ignore the study system's natural history justifying our fitness measures, while failing to account for our behavioral data. We stand by our conclusion that females adaptively choose among heterospecific males.

Sexual isolation between Drosophila simulans and D. mauritiana: D. simulans females do not discriminate against intact-wing D. mauritiana males

Female mate choice plays an important role in sexual isolation. The present study examined sexual isolation using D. simulans and D. mauritiana in conditions where females had no opportunity to compare males versus where females were able to choose males. The sound produced by wing vibration in males (courtship song) affects female receptivity in Drosophila. Females of both species copulated with intact conspecific males more than intact heterospecific males and wingless conspecific and heterospecific males. Drosophila mauritiana females copulated only with intact conspecific males within 30-min observations period without comparing other males, suggesting that absolute criteria are used for decision-making to accept courting males. Females of D. simulans copulated with intact D. mauritiana males as well as wingless D. simulans or D. mauritiana males in no-choice conditions. In a choice situation, D. simulans females copulated with intact D. mauritiana males as well as wingless D. mauritiana males when the females were courted by both types of males, suggesting that D. simulans females accept intact D. mauritiana males as if they are mute. Females of D. simulans copulated with intact D. simulans males as well as intact D. mauritiana males when they were courted by males of either type, whereas they copulated with intact D. simulans males more than intact D. mauritiana males in true choice situations. These results suggest that females make a comparative review of courting males before accepting a male and that conspecific song is a factor in criteria affecting female selectivity.

Host Plant Choice Determined by Reproductive Interference between Closely Related Butterflies

AbstractA number of empirical studies have concluded that reproductive interference (RI) contributes to parapatric species distributions or sexual exclusion. However, the possibility that divergent host plant use in phytophagous insects is due to sexual exclusion has seldom been considered. Here, we present evidence that RI is responsible for different host plant use by two Pierid butterfly species, Pieris napi and Pieris melete. When a novel host species was introduced about 50 years ago, two Pierid butterfly species at first used both the ancestral host species and the novel one. Subsequently, P. napi shifted to use only the novel host, while P. melete shifted to specialize on the ancestral host. To explain these patterns, we investigated whether the two host species differ in suitability for larval growth and survival. Additionally, we tested whether RI occurred between the two butterfly species using large outdoor field cages. Courtship of females by conspecific and heterospecific males reduces the number of eggs laid by approximately half. However, RI is asymmetric and would generate selection on P. melete females to evolve to avoid the more suitable host species preferred by P. napi. Thus, our study suggests that sexual exclusion can explain the shift in host plant use by these two butterfly species.

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

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