Genomic regions repeatedly involved in divergence among plant‐specialized pea aphid biotypes

Understanding the genetic bases of biological diversification is a long‐standing goal in evolutionary biology. Here, we investigate whether replicated cases of adaptive divergence involve the same genomic regions in the pea aphid, Acyrthosiphon pisum, a large complex of genetically differentiated biotypes, each specialized on different species of legumes. A previous study identified genomic regions putatively involved in host‐plant adaptation and/or reproductive isolation by performing a hierarchical genome scan in three biotypes. This led to the identification of 11 F_ST outliers among 390 polymorphic microsatellite markers. In this study, the outlier status of these 11 loci was assessed in eight biotypes specialized on other host plants. Four of the 11 previously identified outliers showed greater genetic differentiation among these additional biotypes than expected under the null hypothesis of neutral evolution (α < 0.01). Whether these hotspots of genomic divergence result from adaptive events, intrinsic barriers or reduced recombination is discussed.     

Reproductive barriers between two sympatric beetle species specialized on different host plants

Knowledge on interspecific pre‐ and post‐zygotic isolation mechanisms provides insights into speciation patterns. Using crosses (F₁ and backcrosses) of two closely related flea beetles species, Altica fragariae and A. viridicyanea, specialized on different hosts in sympatry, we measured: (a) the type of reproductive isolation and (b) the inheritance mode of preference and host‐specific performance, using a joint‐scaling test. Each species preferred almost exclusively its host plant, creating strong prezygotic isolation between them, and suggesting that speciation may occur at least partly in sympatry. Reproductive isolation was intrinsic between females of A. fragariae and either A. viridicyanea or F₁ males, whereas the other crosses showed ecologically dependent reproductive isolation, suggesting ecological speciation. The genetic basis of preference and performance was at least partially independent, and several loci coded for preference, which limits the possibility of sympatric speciation. Hence, both ecological and intrinsic factors may contribute to speciation between these species.     

Multiple sources of reproductive isolation in a bimodal butterfly hybrid zone

An important evolutionary question concerns whether one or many barriers are involved in the early stages of speciation. We examine pre‐ and post‐zygotic reproductive barriers between two species of butterflies (Heliconius erato chestertonii and H. e. venus) separated by a bimodal hybrid zone in the Cauca Valley, Colombia. We show that there is both strong pre‐ and post‐mating reproductive isolation, together leading to a 98% reduction in gene flow between the species. Pre‐mating isolation plays a primary role, contributing strongly to this isolation (87%), similar to previous examples in Heliconius. Post‐mating isolation was also strong, with absence of Haldane’s rule, but an asymmetric reduction in fertility (< 11%) in inter‐specific crosses depending on maternal genotype. In summary, this is one of the first examples of post‐zygotic reproductive isolation playing a significant role in early stages of parapatric speciation in Heliconius and demonstrates the importance of multiple barriers to gene flow in the speciation process.     

Natural selection contributes to geographic patterns of thermal plasticity in Plantago lanceolata

A long‐standing debate in evolutionary biology concerns the relative importance of different evolutionary forces in explaining phenotypic diversification at large geographic scales. For example, natural selection is typically assumed to underlie divergence along environmental gradients. However, neutral evolutionary processes can produce similar patterns. We collected molecular genetic data from 14 European populations of Plantago lanceolata to test the contributions of natural selection versus neutral evolution to population divergence in temperature‐sensitive phenotypic plasticity of floral reflectance. In P. lanceolata, reflectance plasticity is positively correlated with latitude/altitude. We used population pairwise comparisons between neutral genetic differentiation (F_ST and Jost's D) and phenotypic differentiation (P_ST) to assess the contributions of geographic distance and environmental parameters of the reproductive season in driving population divergence. Data are consistent with selection having shaped large‐scale geographic patterns in thermal plasticity. The aggregate pattern of P_ST versus F_ST was consistent with divergent selection. F_ST explained thermal plasticity differences only when geographic distance was not included in the model. Differences in the extent of cool reproductive season temperatures, and not overall temperature variation, explained plasticity differences independent of distance. Results are consistent with the hypothesis that thermal plasticity is adaptive where growing seasons are shorter and cooler, that is, at high latitude/altitude.     

Lack of parallel genetic patterns underlying the repeated ecological divergence of beach and stream‐spawning kokanee salmon

Recent progress in methods for detecting adaptive population divergence in situ shows promise for elucidating the conditions under which selection acts to generate intraspecific diversity. Rapid ecological diversification is common in fishes; however, the role of phenotypic plasticity and adaptation to local environments is poorly understood. It is now possible to investigate genetic patterns to make inferences regarding phenotypic traits under selection and possible mechanisms underlying ecotype divergence, particularly where similar novel phenotypes have arisen in multiple independent populations. Here, we employed a bottom‐up approach to test for signatures of directional selection associated with divergence of beach‐ and stream‐spawning kokanee, the obligate freshwater form of sockeye salmon (Oncorhynchus nerka). Beach‐ and stream‐spawners co‐exist in many post‐glacial lakes and exhibit distinct reproductive behaviours, life‐history traits and spawning habitat preferences. Replicate ecotype pairs across five lakes in British Columbia, Canada were genotyped at 57 expressed sequence tag‐linked and anonymous microsatellite loci identified in a previous genome scan. Fifteen loci exhibited signatures of directional selection (high F_ST outliers), four of which were identified in multiple lakes. However, the lack of parallel genetic patterns across all lakes may be a result of: 1) an inability to detect loci truly under selection; 2) alternative genetic pathways underlying ecotype divergence in this system; and/or 3) phenotypic plasticity playing a formative role in driving kokanee spawning habitat differences. Gene annotations for detected outliers suggest pathogen resistance and energy metabolism as potential mechanisms contributing to the divergence of beach‐ and stream‐spawning kokanee, but further study is required.     

Patterns of reproductive isolation in a haplodiploid – strong post‐mating,prezygotic barriers among three forms of a social spider mite

In speciation research, much attention is paid to the evolution of reproductive barriers, preventing diverging groups from hybridizing back into one gene pool. The prevalent view is that reproductive barriers evolve gradually as a by‐product of genetic changes accumulated by natural selection and genetic drift in groups that are segregated spatially and/or temporally. Reproductive barriers, however, can also be reinforced by natural selection against maladaptive hybridization. These mutually compatible theories are both empirically supported by studies, analysing relationships between intensity of reproductive isolation and genetic distance in sympatric taxa and allopatric taxa. Here, we present the – to our knowledge – first comparative study in a haplodiploid organism, the social spider mite Stigmaeopsis miscanthi, by measuring premating and post‐mating, pre‐ and post‐zygotic components of reproductive isolation, using three recently diverged forms of the mite that partly overlap in home range. We carried out cross‐experiments and measured genetic distances (mitochondrial DNA and nuclear DNA) among parapatric and allopatric populations of the three forms. Our results show that the three forms are reproductively isolated, despite the absence of premating barriers, and that the post‐mating, prezygotic component contributes most to reproductive isolation. As expected, the strength of post‐mating reproductive barriers positively correlated with genetic distance. We did not find a clear pattern of prezygotic barriers evolving faster in parapatry than in allopatry, although one form did show a trend in line with the ecological and behavioural relationships between the forms. Our study advocates the versatility of haplodiploid animals for investigating the evolution of reproductive barriers.     

Patterns of reproductive isolation within and between two Lygaeus species characterized by sexual conflicts over mating

Theory suggests that, under some circumstances, sexual conflict over mating can lead to divergent sexually antagonistic coevolution among populations for traits associated with mating, and that this can promote reproductive isolation and hence speciation. However, sexual conflict over mating may also select for traits (e.g. male willingness to mate) that enhance gene flow between populations, limiting population divergence. In the present study, we compare pre‐ and post‐mating isolation within and between two species characterized by male–female conflict over mating rate. We quantify sexual isolation among five populations of the seed bug Lygaeus equestris collected from Italy and Sweden, and two replicates of a population of the sister‐species Lygaeus simulans, also collected from Italy. We find no evidence of reproductive isolation amongst populations of L. equestris, suggesting that sexual conflict over mating has not led to population divergence in relevant mating traits in L. equestris. However, there was strong asymmetric pre‐mating isolation between L. equestris and L. simulans: male L. simulans were able to mate successfully with female L. equestris, whereas male L. equestris were largely unable to mate with female L. simulans. We found little evidence for strong post‐mating isolation between the two species, however, with hybrid F₂ offspring being produced. Our results suggest that sexual conflict over mating has not led to population divergence, and indeed perhaps supports the contrary theoretical prediction that male willingness to mate may retard speciation by promoting gene flow.     

Seascape genomics reveals adaptive divergence in a connected and commercially important mollusc,the greenlip abalone (Haliotis laevigata), along a longitudinal environmental gradient

Populations of broadcast spawning marine organisms often have large sizes and are exposed to reduced genetic drift. Under such scenarios, strong selection associated with spatial environmental heterogeneity is expected to drive localized adaptive divergence, even in the face of connectivity. We tested this hypothesis using a seascape genomics approach in the commercially important greenlip abalone (Haliotis laevigata). We assessed how its population structure has been influenced by environmental heterogeneity along a zonal coastal boundary in southern Australia linked by strong oceanographic connectivity. Our data sets include 9,109 filtered SNPs for 371 abalones from 13 localities and environmental mapping across ~800 km. Genotype–environment association analyses and outlier tests defined 8,786 putatively neutral and 323 candidate adaptive loci. From a neutral perspective, the species is better represented by a metapopulation with very low differentiation (global F_ST = 0.0081) and weak isolation by distance following a stepping‐stone model. For the candidate adaptive loci, however, model‐based and model‐free approaches indicated five divergent population clusters. After controlling for spatial distance, the distribution of putatively adaptive variation was strongly correlated to selection linked to minimum sea surface temperature and oxygen concentration. Around 80 candidates were annotated to genes with functions related to high temperature and/or low oxygen tolerance, including genes that influence the resilience of abalone species found in other biogeographic regions. Our study includes a documented example about the uptake of genomic information in fisheries management and supports the hypothesis of adaptive divergence due to coastal environmental heterogeneity in a connected metapopulation of a broadcast spawner.     

Post‐zygotic hybrid viability in sympatric species pairs: a case study from European lampreys

Ecological speciation mechanisms are widely assumed to play an important role in the early stages of divergence between incipient species, and this especially true of fishes. In the present study, we tested explicitly for post‐zygotic barriers to gene flow between a sympatric, recently diverged lamprey species pair that likely arose through ecological divergence. Experimental in vitro hybridization between anadromous parasitic Lampetra fluviatilis and resident nonparasitic Lampetra planeri resulted in a high proportion of embryos capable of attaining the burrowing pro‐larval stage, strongly indicating no post‐zygotic barriers to gene flow between these species. A sympatric, locally‐adapted resident parasitic form of L. fluviatilis was also found to successfully hybridize with both members of this species pair. The consequences of these findings are discussed in the context of lamprey speciation. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 108 , 378–383.     

Genome divergence and diversification within a geographic mosaic of coevolution

Despite substantial interest in coevolution's role in diversification, examples of coevolution contributing to speciation have been elusive. Here, we build upon past studies that have shown both coevolution between South Hills crossbills and lodgepole pine (Pinus contorta), and high levels of reproductive isolation between South Hills crossbills and other ecotypes in the North American red crossbill (Loxia curvirostra) complex. We used genotyping by sequencing to generate population genomic data and applied phylogenetic and population genetic analyses to characterize the genetic structure within and among nine of the ecotypes. Although genome‐wide divergence was slight between ecotypes (F_ST = 0.011–0.035), we found evidence of relative genetic differentiation (as measured by F_ST) between and genetic cohesiveness within many of them. As expected for nomadic and opportunistic breeders, we detected no evidence of isolation by distance. The one sedentary ecotype, the South Hills crossbill, was genetically most distinct because of elevated divergence at a small number of loci rather than pronounced overall genome‐wide divergence. These findings suggest that mechanisms related to recent local coevolution between South Hills crossbills and lodgepole pine (e.g. strong resource‐based density dependence limiting gene flow) have been associated with genome divergence in the face of gene flow. Our results further characterize a striking example of coevolution driving speciation within perhaps as little as 6000 years.     

Asymmetric Assortative Mating Behaviour Reflects Incomplete Pre‐zygotic Isolation in the Nasonia Species Complex

Preference of con‐ over heterospecific mates leading to assortative mating can substantially contribute to pre‐zygotic reproductive isolation and prevent fitness losses if post‐zygotic hybridization barriers already exist. The jewel wasp genus Nasonia displays quite strong and well‐studied post‐zygotic reproductive isolation due to a ubiquitous Wolbachia infection causing cytoplasmic incompatibility between different species. Pre‐zygotic isolation, however, has received far less research attention in this model organism, especially concerning the mechanisms and criteria of mate choice. In the present study, we analysed mate rejection and mate acceptance rates in cross‐comparisons between all four Nasonia species. We put emphasis on observing which sex is more likely to interrupt interspecific matings and how discriminatory behaviour varies across the different species in all possible combinations. We found an asymmetric distribution of assortative mating among the four Nasonia species that appears to be highly influenced by the respective combinations of sex and species. Females appeared to be the main discriminators against heterospecific mating partners, but interestingly, we could also detect mate discrimination and rejection behaviour in males, a widely neglected factor in research on mating behaviour in general and on Nasonia in particular. Moreover, the asymmetry in the assortative mating behaviour was partially reflective of sym‐ or allopatric distributions of natural Nasonia populations.     

Evolution of pre‐zygotic and post‐zygotic barriers to gene flow among three cryptic species within the Anastrepha fraterculus complex

Tropical tephritids are ideally suited for studies on population divergence and speciation because they include species groups undergoing rapid radiation, in which morphologically cryptic species and sister species are abundant. The fraterculus species group in the Neotropical genus Anastrepha is a case in point, as it is composed of a complex of up to seven A. fraterculus morphotypes proposed to be cryptic species. Here, we document pre‐ and post‐zygotic barriers to gene flow among adults of the Mexican A. fraterculus morphotype and three populations (Argentina, Brazil, and Peru) belonging to two separate morphotypes (Brazilian 1 and Peruvian). We unveiled three forms of pre‐zygotic reproductive isolation resulting in strong assortative mating. In field cages, free‐ranging male and female A. fraterculus displayed a strong tendency to form couples with members of the opposite sex belonging to their own morphotype, suggesting that male pheromone emission, courtship displays, or both intervene in shaping female choice before actual contact and coupling. In addition, males and females of the Peruvian morphotype became receptive and mated significantly later than adults of the Mexican and Brazilian 1 morphotypes. After contact, Mexican females exhibited greater mating discrimination than males when facing adults of the opposite sex belonging to either the Peruvian or the Brazilian 1 morphotype as evidenced by vigorous resistance to penetration once they had been forcefully mounted by heterotypic males. Forced copulations resulted in production of F1 hybrids that were either less viable (and partially fertile) than parental crosses or even sterile. Our results suggest that the Mexican morphotype is a distinct biological entity and that pre‐zygotic reproductive isolation through divergence in courtship or male‐produced pheromone and other mechanisms appear to evolve faster than post‐zygotic isolation in the fraterculus species group.     

The role of hybridization during ecological divergence of southwestern white pine (Pinus strobiformis) and limber pine (P. flexilis)

Interactions between extrinsic factors, such as disruptive selection and intrinsic factors, such as genetic incompatibilities among loci, often contribute to the maintenance of species boundaries. The relative roles of these factors in the establishment of reproductive isolation can be examined using species pairs characterized by gene flow throughout their divergence history. We investigated the process of speciation and the maintenance of species boundaries between Pinus strobiformis and Pinus flexilis. Utilizing ecological niche modelling, demographic modelling and genomic cline analyses, we illustrated a divergence history with continuous gene flow. Our results supported an abundance of advanced generation hybrids and a lack of loci exhibiting steep transition in allele frequency across the hybrid zone. Additionally, we found evidence for climate‐associated variation in the hybrid index and niche divergence between parental species and the hybrid zone. These results are consistent with extrinsic factors, such as climate, being an important isolating mechanism. A build‐up of intrinsic incompatibilities and of coadapted gene complexes is also apparent, although these appear to be in the earliest stages of development. This supports previous work in coniferous species demonstrating the importance of extrinsic factors in facilitating speciation. Overall, our findings lend support to the hypothesis that varying strength and direction of selection pressures across the long lifespans of conifers, in combination with their other life history traits, delays the evolution of strong intrinsic incompatibilities.     

Habitat preference and flowering‐time variation contribute to reproductive isolation between diploid and autotetraploid Anacamptis pyramidalis

Tetraploid lineages are typically reproductively isolated from their diploid ancestors by post‐zygotic isolation via triploid sterility. Nevertheless, polyploids often also exhibit ecological divergence that could contribute to reproductive isolation from diploid ancestors. In this study, we disentangled the contribution of different forms of reproductive isolation between sympatric diploid and autotetraploid individuals of the food‐deceptive orchid Anacamptis pyramidalis by quantifying the strength of seven reproductive barriers: three prepollination, one post‐pollination prezygotic and three post‐zygotic. The overall reproductive isolation between the two cytotypes was found very high, with a preponderant contribution of two prepollination barriers, that is phenological and microhabitat differences. Although the contribution of post‐zygotic isolation (triploid sterility) is confirmed in our study, these results highlight that prepollination isolation, not necessarily involving pollinator preference, can represent a strong component of reproductive isolation between different cytotypes. Thus, in the context of polyploidy as quantum speciation, that generates reproductive isolation via triploid sterility, ecological divergence can strengthen the reproductive isolation between cytotypes, reducing the waste of gametes in low fitness interploidy crosses and thus favouring the initial establishment of the polyploid lineage. Under this light, speciation by polyploidy involves ecological processes and should not be strictly considered as a nonecological form of speciation.     

Detecting adaptive trait loci in nonmodel systems: divergence or admixture mapping?

Mapping adaptive trait loci (ATL) underlying ecological divergence is an essential step towards understanding the processes that generate phenotypic diversity. Technological advances have made it possible to sequence exomes in nonmodel systems, providing an efficient means of analysing functional genetic variants. Divergence scans of genetic markers for outlier loci, or ‘divergence mapping’, have been used to map locally adapted genes, but this approach is likely to be underpowered when background divergence is elevated. Genotype–phenotype association tests in admixed populations, or ‘admixture mapping’, may provide a useful approach for mapping locally adapted loci when neutral divergence is high. To determine the power and limits of divergence mapping, we simulated exomes containing a single ATL across two parental populations of varying neutral divergence, estimated divergence and quantified the power to identify the ATL. We found that divergence mapping had very high power when background F_ST is <0.2, but decreased dramatically above this level. To evaluate the utility of admixture mapping, we simulated exomes from admixed populations, then simulated phenotypes, conducted genotype–phenotype association tests and found that even two generations of random mating after admixture could provide high mapping power in scenarios where pure divergence mapping was ineffective (F_ST = 0.35). Moreover, admixture mapping had high power across all levels of divergence after 20 generations since admixture. Together with high‐throughput exome sequencing, admixture mapping could be used to map ATL in systems such as Heliconius butterflies or Gryllus crickets when experimental design and analytical approach are chosen accordingly.     

Population genomics of local adaptation versus speciation in coral reef fishes (Hypoplectrus spp,Serranidae)

Are the population genomic patterns underlying local adaptation and the early stages of speciation similar? Addressing this question requires a system in which (i) local adaptation and the early stages of speciation can be clearly identified and distinguished, (ii) the amount of genetic divergence driven by the two processes is similar, and (iii) comparisons can be repeated both taxonomically (for local adaptation) and geographically (for speciation). Here, we report just such a situation in the hamlets (Hypoplectrus spp), brightly colored reef fishes from the wider Caribbean. Close to 100,000 SNPs genotyped in 126 individuals from three sympatric species sampled in three repeated populations provide genome‐wide levels of divergence that are comparable among allopatric populations (F_st estimate = 0.0042) and sympatric species (F_st estimate = 0.0038). Population genetic, clustering, and phylogenetic analyses reveal very similar patterns for local adaptation and speciation, with a large fraction of the genome undifferentiated (F_st estimate ≈ 0), a very small proportion of F_st outlier loci (0.05–0.07%), and remarkably few repeated outliers (1–3). Nevertheless, different loci appear to be involved in the two processes in Hypoplectrus, with only 7% of the most differentiated SNPs and outliers shared between populations and species comparisons. In particular, a tropomyosin (Tpm4) and a previously identified hox (HoxCa) locus emerge as candidate loci (repeated outliers) for local adaptation and speciation, respectively. We conclude that marine populations may be locally adapted notwithstanding shallow levels of genetic divergence, and that from a population genomic perspective, this process does not appear to differ fundamentally from the early stages of speciation.     

Role of seed germination in adaptation and reproductive isolation in Arabidopsis lyrata

Seed germination is an important developmental and life history stage. Yet, the evolutionary impact of germination has mainly been studied in the context of dormancy, or for its role in reproductive isolation between species. Here, we aim to examine multiple consequences of genetic divergence on germination traits between two Arabidopsis lyrata subspecies: ssp. petraea (Eurasia) and ssp. lyrata (North America). Postdormancy germination time, a potentially adaptive trait, showed differentiation between the populations, and quantitative trait loci (QTL) mapping revealed that the trait variation is mainly controlled by two antagonistic loci. These QTL areas contain several candidate genes with known function in postdormancy germination in A. thaliana. The sequence variation of three genes was consistent with differential selection, and they also included fixed nonsynonymous substitutions with potential to account for the phenotypic differentiation. We further show that the divergence between the subspecies has led to a slight but significant reduction in hybrid germination proportions, indicating incipient reproductive isolation. Comparison of reciprocal F₁ and F₂ progenies suggests that Bateson–Dobzhansky–Muller incompatibilities likely act through uniparentally inherited factors. Examination of genomewide transmission ratio distortion further revealed that cytonuclear interactions cause substantial pregermination inviability in the hybrids. These results confirm that seed germination has adaptive potential beyond the dormancy stage and that hybrid seed inviability can be one of the first reproductive barriers to arise during divergence.     

SEX CHROMOSOME LINKAGE OF MATE PREFERENCE AND COLOR SIGNAL MAINTAINS ASSORTATIVE MATING BETWEEN INTERBREEDING FINCH MORPHS

Assortative mating is a key aspect in the speciation process because it is important for both initial divergence and maintenance of distinct species. However, it remains a challenge to explain how assortative mating evolves when diverging populations are undergoing gene flow (e.g., during hybridization). Here I experimentally test how assortative mating is maintained with frequent gene flow between diverged head‐color morphs of the Gouldian finch (Erythrura gouldiae). Contrary to the predominant view on the development of sexual preferences in birds, cross‐fostered offspring did not imprint on the phenotype of their conspecific (red or black morphs) or heterospecific (Bengalese finch) foster parents. Instead, the mating preferences of F₁ and F₂ intermorph‐hybrids are consistent with inheritance on the Z chromosomes, which are also the location for genes controlling color expression and the genes causing low fitness of intermorph‐hybrids. Genetic associations between color signal and preference loci on the sex chromosomes may prevent recombination from breaking down these associations when the morphs interbreed, helping to maintain assortative mating in the face of gene flow. Although sex linkage of reproductively isolating traits is theoretically expected to promote speciation, social and ecological constraints may enforce frequent interbreeding between the morphs, thus preventing complete reproductive isolation.     

Incipient ecological speciation between successional varieties of a dominant tree involves intrinsic postzygotic isolating barriers

Whereas disruptive selection imposed by heterogeneous environments can lead to the evolution of extrinsic isolating barriers between diverging populations, the evolution of intrinsic postzygotic barriers through divergent selection is less certain. Long‐lived species such as trees may be especially slow to evolve intrinsic isolating barriers. We examined postpollination reproductive isolating barriers below the species boundary, in an ephemeral hybrid zone between two successional varieties of the landscape‐dominant Hawaiian tree, Metrosideros polymorpha, on volcanically active Hawai'i Island. These archipelago‐wide sympatric varieties show the weakest neutral genetic divergence of any taxon pair on Hawai'i Island but significant morphological and ecological differentiation consistent with adaptation to new and old lava flows. Cross‐fertility between varieties was high and included heterosis of F₁ hybrids at the seed germination stage, consistent with a substantial genetic load apparent within varieties through low self‐fertility and a lack of self‐pollen discrimination. However, a partial, but significant, barrier was observed in the form of reduced female and male fertility of hybrids, especially backcross hybrids, consistent with the accumulation of genetic incompatibilities between varieties. These results suggest that partial intrinsic postzygotic barriers can arise through disruptive selection acting on large, hybridizing populations of a long‐lived species.     

Genomic landscape of reproductive isolation in Lucania killifish: The role of sex loci and salinity

Adaptation to different environments can directly and indirectly generate reproductive isolation between species. Bluefin killifish (Lucania goodei) and rainwater killifish (L. parva) are sister species that have diverged across a salinity gradient and are reproductively isolated by habitat, behavioural, extrinsic and intrinsic post‐zygotic isolation. We asked if salinity adaptation contributes indirectly to other forms of reproductive isolation via linked selection and hypothesized that low recombination regions, such as sex chromosomes or chromosomal rearrangements, might facilitate this process. We conducted QTL mapping in backcrosses between L. parva and L. goodei to explore the genetic architecture of salinity tolerance, behavioural isolation and intrinsic isolation. We mapped traits relative to a chromosome that has undergone a centric fusion in L. parva (relative to L. goodei). We found that the sex locus appears to be male determining (XX‐XY), was located on the fused chromosome and was implicated in intrinsic isolation. QTL associated with salinity tolerance were spread across the genome and did not overly co‐localize with regions associated with behavioural or intrinsic isolation. This preliminary analysis of the genetic architecture of reproductive isolation between Lucania species does not support the hypothesis that divergent natural selection for salinity tolerance led to behavioural and intrinsic isolation as a by‐product. Combined with previous studies in this system, our work suggests that adaptation as a function of salinity contributes to habitat isolation and that reinforcement may have contributed to the evolution of behavioural isolation instead, possibly facilitated by linkage between behavioural isolation and intrinsic isolation loci on the fused chromosome.