Assortative preferences and discrimination by females against hybrid male song in the grasshoppers Chorthippus brunneus and Chorthippus jacobsi (Orthoptera: Acrididae)

The grasshoppers Chorthippus brunneus and Chorthippus jacobsi are highly differentiated for male mating signals, and form a mosaic hybrid zone in northern Spain. At some sites within this zone, many hybrids are observed. At others, few hybrids are observed. Such bimodal sites may reflect recent contacts between parental genotypes, or local variation in levels of assortative mating or selection against hybrids. Playback of 12 parental and F1 male songs to 296 parental and hybrid females revealed positive assortative preferences in C. brunneus and C. jacobsi females, supporting a direct role of male mating signals in female choice. However, all female genotypic classes showed reduced responsiveness to F1 male songs. Such sexual selection against hybrids is consistent with the narrow cline width observed in the field for song characters relative to morphology. These results have implications for the genetic structure of the hybrid zone and for models of speciation by reinforcement.     

Habitat association and seasonality in a mosaic and bimodal hybrid zone between Chorthippus brunneus and C. jacobsi (Orthoptera: Acrididae)

Understanding why some hybrid zones are bimodal and others unimodal can aid in identifying barriers to gene exchange following secondary contact. The hybrid zone between the grasshoppers Chorthippus brunneus and C. jacobsi contains a mix of allopatric parental populations and inter-mingled bimodal and unimodal sympatric populations, and provides an ideal system to examine the roles of local selection and gene flow between populations in maintaining bimodality. However, it is first necessary to confirm, over a larger spatial scale, previously identified associations between population composition and season and habitat. Here we use cline-fitting of one morphological and one song trait along two valley transects, and intervening mountains, to confirm previously identified habitat associations (mountain versus valley) and seasonal changes in population composition. As expected from previous findings of studies on a smaller spatial scale, C. jacobsi dominated mountain habitats and mixed populations dominated valleys, and C. brunneus became more prevalent in August. Controlling for habitat and incorporating into the analysis seasonal changes in cline parameters and the standard errors of parental trait values revealed wider clines than previous studies (best estimates of 6.4 to 24.5 km in our study versus 2.8 to 4.7 km in previous studies) and increased percentage of trait variance explained (52.7% and 61.5% for transects 1 and 2 respectively, versus 17.6%). Revealing such strong and consistent patterns within a complex hybrid zone will allow more focused examination of the causes of variation in bimodality in mixed populations, in particular the roles of local selection versus habitat heterogeneity and gene flow between differentiated populations.     

Hybrids between Chorthippus brunneus and C. jacobsi (Orthoptera: Acrididae) do not show endogenous postzygotic isolation

Studies of prezygotic and postzygotic isolation in Drosophila have shown in general that species in sympatry tend to evolve prezygotic barriers earlier than do species in allopatry. However, postzygotic barriers tend to evolve at the same evolutionary rate in both sympatric and allopatric species. In contrast to these observations, the grasshoppers Chorthippus parallelus parallelus and C. p. erythropus show complete hybrid male sterility but only limited prezygotic isolation after an estimated 0.5 millions years of divergence. Like their congeners, C. brunneus and C. jacobsi form a hybrid zone where their ranges meet in northern Spain. However, the hybrid zone is mosaic and bimodal and, in contrast to the high levels of postzygotic isolation between C. parallelus subspecies, these two species showed no significant reduction in hybrid fitness in F₁ or backcross generations relative to the parental generations. The level of prezygotic isolation in laboratory tests was comparable to that between C. parallelus subspecies. These results suggest that endogenous postzygotic isolation does not play an important role in the reproductive isolation between C. brunneus and C. jacobsi , or in determining the structure of the hybrid zone. Exogenous postzygotic isolation may be present and should be tested in future studies.  © 2005 The Linnean Society of London, Biological Journal of the Linnean Society , 2005, 84 , 195–203.     

Inheritance of song and stridulatory peg number divergence between Chorthippus brunneus and C. jacobsi, two naturally hybridizing grasshopper species (Orthoptera: Acrididae)

Knowledge of the genetic basis of divergence in mating signal characters that contribute to reproductive isolation is critical to understanding speciation. Here, we describe a semi-automated system for characterizing grasshopper acoustic signals. We used this system to study the genetic basis of divergence in three male calling song components [echeme (EL), syllable (SL) and phrase (PL) lengths] between Chorthippus brunneus and C. jacobsi, two species of grasshoppers that hybridize in northern Spain. We also studied the number of pegs in the stridulatory file. For all characters, additive effects accounted for most of the genetic differentiation between species. However, the three song components also showed small but significant epistatic effects. No sex linkage was detected. Wright-Castle-Lande estimates of the minimum numbers of genetic factors underlying song and peg number divergence were low: peg number (n(e)=5.87+/-5.84), SL (n(e)=2.37+/-4.79) and PL (n(e)=0.87+/-0.86). On the other hand, EL appeared to be controlled by many genes. These results suggest that divergence in SL and PL might be driven by sexual selection whereas EL might not be under selection. This is consistent with experimental results on female song preference in related species. However, the fact that few factors appear to underlie the differences in peg number is surprising. Peg number is not closely related to song characteristics. It often varies between closely related grasshopper species and it has been assumed to be a neutral character. The biometrical approaches used here tend to underestimate the number of factors influencing a trait but provide valuable background for subsequent quantitative trait loci analyses.     

A putative hybrid in nature between Chorthippus brunneus and C. biguttulus(Orthoptera: Acrididae)

The characteristics of the song and morphology of a probable natural hybrid between the two grasshoppers Chorthippus brunneus (Thunberg) and C. biguttulus (L.) are described and compared with those of the parent species. The status of C. eisentrauti (Ramme) is briefly discussed.     

Ants exhibit asymmetric hybridization in a mosaic hybrid zone

Research on hybridization between species provides unparalleled insights into the pre‐ and postzygotic isolating mechanisms that drive speciation. In social organisms, colony‐level incompatibilities may provide additional reproductive barriers not present in solitary species, and hybrid zones offer an opportunity to identify these barriers. Here, we use genotyping‐by‐sequencing to sequence hundreds of markers in a hybrid zone between two socially polymorphic ant species, Formica selysi and Formica cinerea. We characterize the zone, determine the frequency of hybrid workers, infer whether hybrid queens or males are produced and investigate whether hybridization is influenced by colony social organization. We also compare cuticular hydrocarbon profiles and aggression levels between the two species. The hybrid zone exhibits a mosaic structure. The asymmetric distribution of hybrids skewed towards F. cinerea suggests a pattern of unidirectional nuclear gene flow from F. selysi into F. cinerea. The occurrence of backcrossed individuals indicates that hybrid queens and/or males are fertile, and the presence of the F. cinerea mitochondrial haplotype in 97% of hybrids shows that successful F1 hybrids will generally have F. cinerea mothers and F. selysi fathers. We found no evidence that social organization contributes to speciation, because hybrids occur in both single‐queen and multiple‐queen colonies. Strongly differentiated cuticular hydrocarbon profiles and heightened interspecific aggression further reveal that species recognition cues are both present and perceived. The discovery of fertile hybrids and asymmetrical gene flow is unusual in ants, and this hybrid zone will therefore provide an ideal system with which to investigate speciation in social insects.     

Mating signal variation and bimodality in a mosaic hybrid zone between Chorthippus grasshopper species

The grasshoppers Chorthippus brunneus and Chorthippus jacobsi are easily distinguished by male calling song and the number of stridulatory pegs on the hind femur, and form a mosaic hybrid zone in northern Spain. In this paper, we fit a two-dimensional cline to variation in male calling song characters, which are of particular interest as they are likely to be involved in mate choice by females. As with variation in peg number, local habitat makes only a small contribution in explaining deviations in mean song score from clinal expectations. However, the fitted width of the cline for song characters is significantly narrower than for peg number, suggesting that mating signals may be associated with reduced hybrid fitness in the field and that recombination rates are sufficient to allow clines for different characters to diverge in width. Despite this, estimates for the overall elevation in linkage disequilibrium at the zone center, based on covariance between peg and song characters, reveal a substantial overrepresentation of parental genotypes at the cline center relative to the expectations of a tension zone of similar width. Examination of covariance at individual sites reveals that this inflated estimate of linkage disequilibrium is caused by several sites where the distribution of phenotypes is effectively bimodal. This substantial variation in linkage disequilibrium at the cline center could result from local variation in the strength of assortative mating or selection against hybrids, or may reflect the long-distance colonization of empty habitat from outside the hybrid zone, which would continually create new contacts between parental genotypes at the cline center. Hybrid zones like this, in which strong linkage disequilibrium occurs in some situations but not in others, are of particular relevance to speciation research and allow investigation of the spread of combinations of alleles through different genetic and ecological backgrounds.     

Assortative mating across a hybrid zone in Chorthippus parallelus (Orthoptera: Acrididae)

Two subspecies of the grasshopper Chorthippus parallelus meet and form a hybrid zone in the Pyrenees. Both strong hybrid dysfunction and a behavioural difference occur, which would seem to make the zone a suitable candidate for speciation by reinforcement. One of the classic ways of looking for this is to test for increased levels of assortative mating between populations from close to the region of contact. Here we show that, with virgin insects, such assortment decreases as one approaches the centre of the zone in C. parallelus. The pattern is different upon remating, with non-virgins showing a pattern more like that predicted by reinforcement. Overall there is little evidence for reinforcement. We argue that other tests of the model may be more appropriate for stable hybrid zones.     

A hybrid zone between Chorthippus parallelus parallelus and Chorthippus parallelus erythropus (Orthoptera: Acrididae): behavioural characters

Two subspecies of the grasshopper Chorthippus parallelus have ranges that abut in the Pyrenees. Males produce two types of song—calling song and courtship song. We have examined the use and structure of these songs in males from nine sites near Mont-Louis, Pyrénées-Onentales, France. These sites were previously used to identify the position of the hybrid zone on morphological characters. The subspecies differ in the use and structure of both types of song. Clines in these characters correspond in position with the morphological cline. At least one character may be relevant to mate choice but none of the characters show evidence of reinforcement.     

Genetics of behavioural and morphological differences between parapatric subspecies of Chorthippus parallelus (Orthoptera: Acrididae)

The grasshoppers Chorthippus parallelus parallelus and C. p. erythropus form a narrow hybrid zone in the Pyrenees. They differ in several characters of the pattern and structure of male stridulation and in the morphology of the stridulatory file. These characters are considered to be involved in the species' mate recognition system.
Crosses have been made between a Pyrenean C. p. erythropus population and two C. p. parallelus populations, one in the Pyrenees and one in the Massif Central. Reciprocal Fls and backcrosses have been examined for a set of stridulation and associated morphological characters. The crosses confirm that the subspecific differences have a genetic basis and suggest that they are polygenically determined. However the mode of inheritance is not simple. There is evidence for dominance and epistatic effects and for sex-linkage or maternal effects. Genetic correlations exist between some pairs of characters in the backcrosses.
These results are discussed in the context of the hybrid zone and in relation to the general problem of the evolutionary divergence of mate recognition systems.     

Low levels of chromosomal differentiation between the grasshoppers Chorthippus brunneus and Chorthippusjacobsi (Orthoptera; Acrididae) in northern Spain

The grasshopper species Chorthippus brunneus and C. jacobsi (Orthoptera: Acrididae) form a hybrid zone in northern Spain. These species probably diverged while isolated in southern refugia during one of the recent ice ages, and are clearly distinguished by morphology and male calling song. However, in contrast to other Chorthippus taxa that form hybrid zones in Europe, these two species cannot be reliably distinguished on the basis of characteristics of the karyotype such as heterochromatin banding patterns and composition, as revealed by C-banding and fluorochrome staining. Silver staining also reveals the presence of two autosomal nucleolar organiser regions (NORs) in both species. However, differentiation between C. brunneus and C. jacobsi was revealed on the X chromosome using fluorescent in situ hybridisation (FISH). C. brunneus individuals showed additional rDNA sequences on the X chromosome that were not observed in any C. jacobsi individuals. These sequences are not transcribed, indicating either mutational silencing of an ancestral NOR on the X chromosome, or the transposition of non-functional sequences from the autosomes. The implications of these results for the evolution of NOR number in Chorthippus are discussed.     

A hybrid zone between Chorthippus parallelus parallelus and Chorthippus parallelus erythropus (Orthoptera: Acrididae): morphological and electrophoretic characters

Two subspecies of the grasshopper Chorthippus parullelus have distributions that abut in the Pyrenees. We have studied morphological and electrophoretic variation in II populations near Mont Louis, Pyrénées-Orientales, France. The two subspecies differ in several morphological characters and at one enzyme locus, esterase-2, and in this area they hybridize forming a cline in morphology less than 5 km wide. Examination of museum specimens suggests that this hybrid zone extends for most of the length of the Pyrenees, possibly breaking down in the extreme west. As the two subspecies are known to differ in mating behaviour this hybrid zone is well suited to the study of reinforcement.     

Divergence in cuticular hydrocarbons between parapatric subspecies of the meadow grasshopper, Chorthippus parallelus (Orthoptera, Acrididae)

Two subspecies of the grasshopper Chorthippus parallelus (Orthoptera; Acrididae) meet and form a hybrid zone in the Pyrenees. Here we investigate whether the subspecies differ in the blend of cuticular hydrocarbons. Such differences may function as chemical signals, being detected via contact chemoreception and used during mate choice as in other insects. Gas chromatography shows that the subspecies have diverged in the composition of their cuticular hydrocarbons. This difference between subspecies exists over and above variation in cuticular hydrocarbons among individual populations. The majority of the difference can be attributed to the relative amounts of a small group of compounds present in the cuticle. The possible consequences of this chemical divergence for mating encounters between the subspecies are discussed.     

Fitness consequences of potential assortative mating inside and outside a hybrid zone in Chorthippus parallelus (Orthoptera: Acrididae): implications for reinforcement and sexual selection theory

We have examined the fitness consequences of random and potentially non-random matings within two populations taken from inside, and two from outside a hybrid zone in Chorthippus parallelus. When given the opportunity to mate non-randomly, females from all populations laid egg pods more quickly than females obliged to mate at random. A range of fitness parameters measured on the offspring did not show increased fitness following potential non-random mating for any population. However, in non-hybrid populations, the sons of non-randomly mated females had about twice the mating success of the sons of those females forced to mate at random, suggesting the existence of heritable variation for male reproductive success. Hybrid dysfunction did not occur amongst the offspring of randomly mated hybrid females, demonstrating that the lack of dysfunction within these populations is not due to the evolution of assortative mating within them.     

Spatio-temporal dynamics of the Allonemobius fasciatus- A. socius mosaic hybrid zone: a 14-year perspective

Long-term studies of hybrid zones can provide valuable insight into a number of questions that have long attracted the attention of evolutionists. These questions range from the stability and fate of hybrid zones to the relative fitness of hybrids. In this paper we report the results of a 14-year survey of the Allonemobius fasciatus-Allonemobius socius hybrid zone. Populations were collected intensively in 1986 and 1987 and then more sporadically through the end of the 1980s and throughout the 1990s. By documenting changes in the genetic composition of populations near and within the zone during this period of time we assessed: the strength of the reproductive isolation between the two species; the relative growth rates (which can be considered a surrogate of relative fitness) of genotype classes corresponding to hybrids and to pure species individuals; and, the power of single-year and multi-year measurements of relative growth rates to predict changes in the genetic composition of mixed populations through time. In brief, we found very large year-to-year variation in the relative growth rates of pure species and hybrid individuals. This variation may reflect the fact that both species are at the edge of their range and perhaps at the limits of their ability to deal with environmental perturbations. As a consequence of the variation, even multi-year estimates of relative growth rates often provided imprecise predictions regarding the future genotypic composition of mixed populations. Despite our limited ability to predict the dynamics of individual populations, some trends are apparent. A. socius, the southern species, has clearly increased in frequency along a transect through the Appalachian Mountains, indicating that the zone is moving north in this region. In contrast, the zone appeared to be more stable along the East Coast transect. Within mixed populations, character-index profiles are often bimodal and stable through time, indicating relatively strong reproductive isolation between the two species that is not being reinforced, nor is it breaking down.     

How a hybrid zone is maintained: behavioral mechanisms of interbreeding between California and Gambel's quail (Callipepla californica and C. gambelii)

Abstract Hybridizing California and Gambel's Quail ( Callipepla californica and C. gambelii ) are unlike many hybridizing avian species in that pairing primarily occurs within a flock, or covey, that is composed of several families. Coveys in the area of sympatry contain mostly hybrid individuals, relative to parental types. I tested whether individuals perceive covey mates as a single species and whether pairing within the covey causes inbreeding and a loss of reproductive success. Individuals discriminated between the parental species in captivity, but actual pairing in the wild was random with respect to species. Contrary to expectation, coveys were not more inbred than the local population. Results suggest that potential costs of inbreeding may be avoided through sex-biased dispersal and nonrandom pairing. Furthermore, breeding occurred earlier and with greater success in pairs formed within the resident covey, rather than outside it. These findings suggest that fitness benefits gained from pairing early within a mixed-species covey promote interspecific pairing. This study provides evidence that mating behaviors specific to local conditions maintain hybrid zones when genetic costs to interbreeding are small.     

Selection on hybrids of ecologically divergent ecotypes of a marine snail: the relative importance of exogenous and endogenous barriers

Unravelling the form of selection acting on hybrids of ecotypes undergoing ecological speciation is essential to understand the mechanisms behind the evolution of reproductive isolation in the face of gene flow. Shell phenotype is known to be affected by natural selection and is involved in the fitness of the marine snail Littorina saxatilis. Here, we studied the association between shell traits and fitness in hybrids in order to determine the relative role of exogenous and endogenous selection in this hybrid zone of L. saxatilis. We show that directional selection is the predominant mode of selection among hybrids. We also show its heterogeneity, affecting different shell traits, within populations at the level of the microhabitat. Therefore, endogenous selection mechanisms are most probably lacking in this hybrid zone and exogenous barriers (pre‐ and post‐zygotic) are possibly one of the main forces behind the evolution of barriers to gene flow between these ecologically divergent ecotypes. This study shows how this barrier might represent an important type of reproductive isolation within ecological speciation, and this should be taken into account in future studies of speciation in hybrid zones. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 111 , 391–400.     

A fine-scale spatial analysis of the mosaic hybrid zone between Gryllus firmus and Gryllus pennsylvanicus

Abstract.— The pattern of character variation within a hybrid zone, the hybrid zone structure, has been used to infer the processes that maintain hybrid zones. Unfortunately it is difficult to infer process from structure alone because many different processes can produce the same pattern of character variation. Mosaic hybrid zones may be maintained by exogenous selection in a heterogeneous environment and/or endogenous selection against hybrid individuals; habitat preference, premating isolating barriers and/or fertility selection can also contribute. The spatial scale at which a hybrid zone is sampled affects its apparent structure; a hybrid zone may appear clinal at one scale and mosaic at another. Here, we sample the mosaic hybrid zone between two field crickets, Gryllus firmus and G. pennsylvanicus , at a scale that spans the boundaries between individual soil-habitat patches. From our analysis, we find that at fine scales, the mosaic hybrid zone resolves into a set of steep clines across patch boundaries. Both morphological and molecular traits exhibit sharp and generally concordant clines. However, clines for mitochondrial DNA and one anonymous nuclear marker are clearly displaced as a result of current hybridization or past introgression (the "ghost of hybridization past"). Thus, scale is important for the structure of this and probably other hybrid zones. The extremely sharp, concordant clines across patch boundaries indicate that the cricket hybrid zone is undoubtedly structured by selection. However, the detailed mechanisms responsible for the maintenance of the hybrid zone–whether endogenous selection against hybrids, exogenous selection by the environment, and/or behavioral preferences for mates or habitats– remain to be elucidated. Determining these mechanisms will depend on closer inspection of the organisms themselves and their interactions, as is the case for all hybrid zones.