Speciation is rarely observable directly. A way forward is to compare pairs of ecotypes that evolved in parallel in similar contexts but have reached different degrees of reproductive isolation. Such comparisons are possible in the marine snail Littorina saxatilis by contrasting barriers to gene flow between parallel ecotypes in Spain and Sweden. In both countries, divergent ecotypes have evolved to withstand either crab predation or wave action. Here, we explore transects spanning contact zones between the Crab and the Wave ecotypes using low-coverage whole-genome sequencing, morphological and behavioural traits. Despite parallel phenotypic divergence, distinct patterns of differentiation between the ecotypes emerged: a continuous cline in Sweden indicating a weak barrier to gene flow, but two highly genetically and phenotypically divergent, and partly spatially overlapping clusters in Spain suggesting a much stronger barrier to gene flow. The absence of Spanish early-generation hybrids supported strong isolation, but a low level of gene flow is evident from molecular data. In both countries, highly differentiated loci were located in both shared and country-specific chromosomal inversions but were also present in collinear regions. Despite being considered the same species and showing similar levels of phenotypic divergence, the Spanish ecotypes are much closer to full reproductive isolation than the Swedish ones. Barriers to gene flow of very different strengths between ecotypes within the same species might be explained by dissimilarities in the spatial arrangement of habitats, the selection gradients or the ages of the systems.

Polymorphic short insertions and deletions (INDELs ≤ 50 bp) are abundant, although less common than single nucleotide polymorphisms (SNPs). Evidence from model organisms shows INDELs to be more strongly influenced by purifying selection than SNPs. Partly for this reason, INDELs are rarely used as markers for demographic processes or to detect balancing or divergent selection. Here, we compared INDELs and SNPs in the intertidal snail Littorina saxatilis, focusing on hybrid zones between ecotypes. Using capture sequencing data, we computed INDEL and SNP site frequency spectra (SFS) to compare the impact of purifying, positive and balancing selection on these variant types. Because signatures of selection may be confounded by GC-biased gene conversion and polarization errors, we also examined their influence. We assessed the impact of divergent selection by analysing allele frequency clines across habitat boundaries. We show evidence that short INDELs are affected more by purifying selection and less by positive selection than SNPs, but part of the observed SFS difference can be attributed to GC-biased gene conversion and polarization errors. We did not find a difference in the impact of balancing or divergent selection between short INDELs and SNPs. Short INDELs and SNPs were similarly distributed across the genome and so are likely to respond to indirect selection in the same way. A few regions likely affected by divergent selection were revealed by INDELs and not by SNPs. Short INDELs can be useful genetic markers helping to identify genomic regions under selective constraints or important for adaptation and population divergence.

Speciation is a key evolutionary process that is not yet fully understood. Combining population genomic and ecological data from multiple diverging pairs of marine snails (Littorina) supports the search for speciation mechanisms. Placing pairs on a one-dimensional speciation continuum, from undifferentiated populations to species, obscured the complexity of speciation. Adding multiple axes helped to describe either speciation routes or reproductive isolation in the snails. Divergent ecological selection repeatedly generated barriers between ecotypes, but appeared less important in completing speciation while genetic incompatibilities played a key role. Chromosomal inversions contributed to genomic barriers, but with variable impact. A multidimensional (hypercube) approach supported framing of questions and identification of knowledge gaps and can be useful to understand speciation in many other systems.

Key innovations are fundamental to biological diversification, but their genetic basis is poorly understood. A recent transition from egg-laying to live-bearing in marine snails ( Littorina spp.) provides the opportunity to study the genetic architecture of an innovation that has evolved repeatedly across animals. Individuals do not cluster by reproductive mode in a genome-wide phylogeny, but local genealogical analysis revealed numerous small genomic regions where all live-bearers carry the same core haplotype. Candidate regions show evidence for live-bearer–specific positive selection and are enriched for genes that are differentially expressed between egg-laying and live-bearing reproductive systems. Ages of selective sweeps suggest that live-bearer–specific alleles accumulated over more than 200,000 generations. Our results suggest that new functions evolve through the recruitment of many alleles rather than in a single evolutionary step.

Predicting the outcomes of adaptation is a major goal of evolutionary biology. When temporal changes in the environment mirror spatial gradients, it opens up the potential for predicting the course of adaptive evolution over time based on patterns of spatial genetic and phenotypic variation. We assessed this approach in a 30-year transplant experiment in the intertidal snail Littorina saxatilis. In 1992, snails were transplanted from a predation-dominated environment to one dominated by wave action. On the basis of spatial patterns, we predicted transitions in shell size and morphology, allele frequencies at positions throughout the genome, and chromosomal rearrangement frequencies. Observed changes closely agreed with predictions and transformation was both dramatic and rapid. Hence, adaptation can be predicted from knowledge of the phenotypic and genetic variation among populations.

Inversions are structural mutations that reverse the sequence of a chromosome segment and reduce the effective rate of recombination in the heterozygous state. They play a major role in adaptation, as well as in other evolutionary processes such as speciation. Although inversions have been studied since the 1920s, they remain difficult to investigate because the reduced recombination conferred by them strengthens the effects of drift and hitchhiking, which in turn can obscure signatures of selection. Nonetheless, numerous inversions have been found to be under selection. Given recent advances in population genetic theory and empirical study, here we review how different mechanisms of selection affect the evolution of inversions. A key difference between inversions and other mutations, such as single nucleotide variants, is that the fitness of an inversion may be affected by a larger number of frequently interacting processes. This considerably complicates the analysis of the causes underlying the evolution of inversions. We discuss the extent to which these mechanisms can be disentangled, and by which approach.

A growing body of research shows that chromosomal inversions, where each arrangement is associated with a certain environment and maintains a set of adaptive alleles, make an important contribution to local adaptation. However, inversions often remain unexplored across large geographical scales. It is unclear whether inversions contribute to adaptation across species ranges, which environmental factors affect arrangement frequencies, and whether the adaptive content of the same arrangement varies between locations. Here, we discuss ideas of how allele frequency data (e.g. pool-sequencing data) can be used to learn about inversions in a simple and cost-effective way. If populations connected by migration differ in arrangement frequency, plotting their SNP allele frequencies against each other will reveal a parallelogram whose corners reflect arrangement frequencies. We demonstrate the usefulness of this approach in locally-adapted populations of the intertidal snail Littorina saxatilis (Olivi). For twelve inversions, we estimate arrangement frequencies in 20 populations across the European species range. While roughly half of the inversions contribute to adaptation to the well-studied contrast between wave-exposed and crab-infested habitats, the other half likely contribute to adaptation to shore height. We also find evidence for geographical variation in arrangement content, suggesting variation in adaptive role.

Speciation typically occurs in a time frame too long to be observed directly. This issue can be over-come by studying pairs of populations at different points in the speciation continuum, ideally within clades so that patterns are not confounded by differences among taxa. Such comparisons are possible in the marine snail Littorina saxatilis because it shows repeated occurrence of ecotypes adapted to either crab predation or wave action that differ in age and environmental context. Here, we explored transects spanning hybrid zones between the crab and wave ecotypes to contrast barriers to gene flow in Spain and Sweden, using low coverage whole genome sequencing, shell features, and behavioural traits. The two countries showed parallel divergence but distinct patterns of differentiation between the ecotypes: a continuous cline in Sweden but two highly genetically and phenotypically divergent, and partly spatially-overlapping clusters in Spain. Spanish early-generation hybrids were not observed but a low level of gene flow still seems to occur. In both countries, highly differentiated loci are clustered in genomic regions covered by chromosomal inversions but also occur in collinear regions. Despite being the same species and showing similar levels of phenotypic divergence, the Spanish ecotypes are closer to full reproductive isolation than the Swedish ecotypes. We discuss potential mechanisms contributing to the evolution of these different levels of reproductive isolation, particularly the age of the population, the strength of selection, the spatial context, and the role of assortative mating.

Understanding population divergence that eventually leads to speciation is essential for evolutionary biology. High species diversity in the sea was regarded as a paradox when strict allopatry was considered necessary for most speciation events because geographical barriers seemed largely absent in the sea, and many marine species have high dispersal capacities. Combining genome-wide data with demographic modelling to infer the demographic history of divergence has introduced new ways to address this classical issue. These models assume an ancestral population that splits into two subpopulations diverging according to different scenarios that allow tests for periods of gene flow. Models can also test for heterogeneities in population sizes and migration rates along the genome to account, respectively, for background selection and selection against introgressed ancestry. To investigate how barriers to gene flow arise in the sea, we compiled studies modelling the demographic history of divergence in marine organisms and extracted preferred demographic scenarios together with estimates of demographic parameters. These studies show that geographical barriers to gene flow do exist in the sea but that divergence can also occur without strict isolation. Heterogeneity of gene flow was detected in most population pairs suggesting the predominance of semipermeable barriers during divergence. We found a weak positive relationship between the fraction of the genome experiencing reduced gene flow and levels of genome-wide differentiation. Furthermore, we found that the upper bound of the 'grey zone of speciation' for our dataset extended beyond that found before, implying that gene flow between diverging taxa is possible at higher levels of divergence than previously thought. Finally, we list recommendations for further strengthening the use of demographic modelling in speciation research. These include a more balanced representation of taxa, more consistent and comprehensive modelling, clear reporting of results and simulation studies to rule out nonbiological explanations for general results.

Understanding the factors that have shaped the current distributions and diversity of species is a central and longstanding aim of evolutionary biology. The recent inclusion of genomic data into phylogeographic studies has dramatically improved our understanding in organisms where evolutionary relationships have been challenging to infer. We used whole-genome sequences to study the phylogeography of the intertidal snail Littorina saxatilis, which has successfully colonized and diversified across a broad range of coastal environments in the Northern Hemisphere amid repeated cycles of glaciation. Building on past studies based on short DNA sequences, we used genome-wide data to provide a clearer picture of the relationships among samples spanning most of the species natural range. Our results confirm the trans-Atlantic colonization of North America from Europe, and have allowed us to identify rough locations of glacial refugia and to infer likely routes of colonization within Europe. We also investigated the signals in different datasets to account for the effects of genomic architecture and non-neutral evolution, which provides new insights about diversification of four ecotypes of L. saxatilis (the crab, wave, barnacle, and brackish ecotypes) at different spatial scales. Overall, we provide a much clearer picture of the biogeography of L. saxatilis, providing a foundation for more detailed phylogenomic and demographic studies.

Chromosomal rearrangements (CRs) have been known since almost the beginning of genetics. While an important role for CRs in speciation has been suggested, evidence primarily stems from theoretical and empirical studies focusing on the microevolutionary level (i.e., on taxon pairs where speciation is often incomplete). Although the role of CRs in eukaryotic speciation at a macroevolutionary level has been supported by associations between species diversity and rates of evolution of CRs across phylogenies, these findings are limited to a restricted range of CRs and taxa. Now that more broadly applicable and precise CR detection approaches have become available, we address the challenges in filling some of the conceptual and empirical gaps between micro- and macroevolutionary studies on the role of CRs in speciation. We synthesize what is known about the macroevolutionary impact of CRs and suggest new research avenues to overcome the pitfalls of previous studies to gain a more comprehensive understanding of the evolutionary significance of CRs in speciation across the tree of life.

Key innovations are fundamental to biological diversification, but their genetic architecture is poorly understood. A recent transition from egg-laying to live-bearing in Littorina snails provides the opportunity to study the architecture of an innovation that has evolved repeatedly in animals. Samples do not cluster by reproductive mode in a genome-wide phylogeny, but local genealogical analysis revealed numerous genomic regions where all live-bearers carry the same core haplotype. Associated regions show evidence for live-bearer-specific positive selection, and are enriched for genes that are differentially expressed between egg-laying and live-bearing reproductive systems. Ages of selective sweeps suggest live-bearing alleles accumulated gradually, involving selection at different times in the past. Our results suggest that innovation can have a polygenic basis, and that novel functions can evolve gradually, rather than in a single step.

Kerstin Johannesson is a marine ecologist and evolutionary biologist based at the Tjärnö Marine Laboratory of the University of Gothenburg, which is situated in the beautiful Kosterhavet National Park on the Swedish west coast. Her work, using marine periwinkles (especially Littorina saxatilis and L. fabalis) as main model systems, has made a remarkable contribution to marine evolutionary biology and our understanding of local adaptation and its genetic underpinnings. Kerstin was predestined to study marine gastropods: Her childhood address was Snäckskalsvägen (“Snail shell road”) and her parents bought a summer house almost next door to the Tjärnö Laboratory in 1970, when Kerstin was 15 years old. Nevertheless, Kerstin faced serious challenges when her interests in marine biology began to emerge. For example, in high school, when Kerstin explained her marine interest to the student counsellor for guidance about future education, the counsellor strongly recommended Kerstin a career as a hairdresser instead. And the barriers did not stop at university. Kerstin managed to secure a Master's project about Littorina – but when she decided that she wanted to pursue a PhD project about this system, the professor advising her at the Department of Zoology in Gothenburg was completely negative and explained that “everything was already known about Littorina” and that it was a trivial research question. This almost killed Kerstin's “snail career” but, fortunately for us, she persevered. This must have been difficult for a young student, but Kerstin can be very determined and convincing! Kerstin now sometimes jokes that she is still working on her Master's project on Littorina, but she actually obtained her PhD from the University of Gothenburg in 1986, mentored by Dave Raffaelli and Bob Ward, who taught her a lot about ecology and genetics. Throughout her career, Kerstin has been based at the Tjärnö Laboratory near her childhood summer house. Briefly, from 1986 to 1998, she held positions funded by competitive grants from the Swedish Science Research Council, and then she was appointed Professor of Marine Ecology at the University of Gothenburg in 1999. Kerstin has repeatedly served as the director of the Tjärnö Laboratory. Kerstin's molecular ecology research began in the allozyme era. She provided important evidence for non-neutral behaviour of some allozyme loci, particularly arginine kinase (Ak), which shows a strong and repeatable allele frequency cline from sheltered to more exposed habitats in the flat periwinkle (L. fabalis) (Tatarenkov & Johannesson, 1994), and aspartate aminotransferase (Aat), which shows a shore height cline in the rough periwinkle (L. saxatilis) (Johannesson & Johannesson, 1989). Kerstin's partner Bo was an important early collaborator. A toxic algal bloom in 1988 killed the great majority of snails in Kerstin's study area but, typically, Kerstin saw the opportunities in what many would have considered a disaster. One was the chance to observe the rapid re-formation of the Aat cline, greatly strengthening the evidence for its maintenance by selection (Johannesson et al., 1995). Kerstin also used the fact that snails were completely removed from many tiny rocky islands (“skerries”) in the Koster sea for a series of experiments, including monitoring natural recolonization (Johannesson & Johannesson, 1995), introducing rare colour morphs to study their response to selection (Johannesson & Butlin, 2017) and introducing snails adapted to high-predation, low wave-exposure environments to this low-predation, high-exposure environment, an experiment that she is still monitoring now, more than 30 years later! In addition to allozymes, Kerstin's early work addressed phenotypic variation and the selection pressures driving local adaptation. A particular focus was (and still is) on the forms of L. saxatilis now known as the “crab” and “wave” ecotypes, which occur in crab-infested boulder fields versus on nearby wave-exposed cliffs. Much of Kerstin's early work analysed the differences between these ecotypes, including differences in shell size and shape (e.g., Janson, 1983; Janson & Sundberg, 1983; published under Kerstin's maiden name). Kerstin and collaborators found these ecotypes in numerous locations, for example in Sweden and in Spain, providing an excellent potential example of parallel evolution (Johannesson et al., 2010). This repeated adaptive differentiation over very short distances is enabled by the short mean dispersal distance of L. saxatilis, but its brooding reproductive mode and long tail of dispersal distances also make it an excellent colonizer. This led Kerstin to solve the “paradox of Rockall”, the observation that brooders have a wider distribution than species with planktonic larvae (and thus higher dispersal capabilities) (Johannesson, 1988). High average dispersal may allow other species to reach extremely isolated habitat patches, like the remote islet of Rockall, but it also reduces colonization success because offspring disperse away from the patch. Strong local adaptation and phenotypic differentiation in the periwinkles led to questions about speciation and assortative mating. Strong phenotypic differentiation of a barnacle-dwelling snail (“L. neglecta”) from L. saxatilis crab and wave ecotypes turned out to be associated with little evidence of reduced gene flow, based on allozymes, leading to a debate about species status (e.g., Johannesson & Johannesson, 1993). On the other hand, with Emilio Rolán-Alvarez (University of Vigo), Kerstin showed strong prezygotic isolation between Spanish populations of the crab and wave ecotypes (Rolán-Alvarez et al., 1999). This line of work resulted in an important review on the topic of parallel sympatric divergence and what it tells us about adaptation and speciation (Johannesson, 2001), issues for which Kerstin's work on the very highly-replicated crab-wave ecotype differentiation has become a classic case study. With the emergence of new molecular tools, Kerstin (reluctantly!) got involved in projects using DNA markers. These new approaches particularly allowed for a better understanding of the history of L. saxatilis and its ecotypes. Mitochondrial markers revealed insights into the postglacial recolonization history (Panova et al., 2011), and mitochondrial and nuclear sequences and microsatellites confirmed that the crab and wave ecotypes did evolve repeatedly in different geographical locations, as opposed to a single divergence event (Butlin et al., 2014; Panova et al., 2006). Over the last decade or so, Kerstin has been instrumental in adopting genomic tools to understand the Littorina system, as well as other marine models. She has been a driving force in the Centre for Marine Evolutionary Biology, CeMEB (Johannesson et al., 2022), which formed in 2008 with one aim being to assemble multiple marine genomes. The increasing availability of genomic data allowed for many new insights in L. saxatilis. Genome scans and hybrid zone analyses showed that the genetic basis of ecotype divergence is, at least to some extent, shared between geographical locations, especially on small geographical scales (Morales et al., 2019; Ravinet et al., 2016; Westram et al., 2021). Genomic data also allowed for the mapping of various phenotypic traits to genomic regions (Koch et al., 2021). Recently, thanks to L. saxatilis, Kerstin has become fascinated by chromosomal inversions, which appear to contribute strongly to local adaptation in this system (Faria et al., 2019) and are likely to be partly responsible for the parallel phenotypic patterns in different countries (Morales et al., 2019). But despite these exciting genomic results, Kerstin has always maintained a perspective on the organismal level, studying, for example, the role of embryo abortion rates (Johannesson, Zagrodzka, et al., 2020) and assortative mating (Perini et al., 2020) in adaptive divergence and speciation. Genomic data did not stop Kerstin from looking back fondly to the “good old days” of allozyme work. She has remained convinced that many questions could already be answered with those simpler techniques. And indeed, to Kerstin's joy, the allozyme loci she studied decades ago pop up as strong outliers in genome scans for divergent selection and can now be explored further, for example the Ak locus showing clinal variation between sheltered and exposed habitats in L. fabalis (Le Moan et al., 2022). While Littorina snails have been Kerstin's main focus throughout her career, she has also made great contributions to our understanding of other marine systems. For example, she has been involved in studying the evolution of the seaweeds Fucus vesiculosus and F. radicans, demonstrating pervasive clonal reproduction in marginal populations (Johannesson, Johansson, et al., 2011; Tatarenkov et al., 2007) and analysing population genetic structure (e.g., Ardehed et al., 2016). A main interest has been evolution in the Baltic Sea, a marginal, species-poor sea with low salinity. Kerstin and collaborators showed that Baltic Sea populations of various species are differentiated from conspecific populations in the Atlantic and often show reduced genetic diversity (Johannesson & Andre, 2006), raising concerns about their conservation under environmental change (Johannesson, Smolarz, et al., 2011). The Baltic Sea also enabled one of few multispecies analyses of contact zones and clines across an environmental gradient (here, salinity) (Johannesson, Le Moan, et al., 2020). Despite the “distraction” of other systems, the work on Littorina has always been closest to Kerstin's heart—and this intense, long-term focus on one system from various angles is one aspect that makes her work so outstanding. Her work on Littorina has been versatile, including field surveys and field experiments, genetic and genomic analyses, behavioural assays in the laboratory, crossing experiments (for which she still does all the laborious maintenance herself!), but also collaborations with theoreticians and modellers (e.g., Rafajlovic et al., 2013). Combining her detailed ecological knowledge about the system with state-of-the-art genetic and genomic methods, Kerstin has really done the “ecology” in “molecular ecology” justice and made a major contribution to our understanding of adaptation in natural populations. It is to a large extent due to Kerstin that L. saxatilis has become a well-known model system for local adaptation and parallel evolution. The Littorina community has substantially grown over the decades, not least due to Kerstin's success in securing large Swedish and European grants and establishing various collaborations, and Kerstin is a central figure in the regular meetings of the international Littorina and Littorinid community. Apart from her scientific achievements in the narrow sense, Kerstin has, more than most other researchers, engaged in outreach, teaching, support of governmental agencies, membership of various boards, and marine conservation. For example, she was strongly involved in establishing the co-management of local shrimp fisheries together with local fishermen, biologists and politicians. She was able to persuade the fishermen to learn some marine biology, a rare achievement based on respect for their knowledge. This co-management strategy was later crucial for the establishment of the Kosterhavet National Park in the area, the first marine national park in Sweden. At the moment, Kerstin is in charge of developing a national programme for population genetic monitoring of key marine and freshwater species together with Linda Laikre (Stockholm University). Kerstin has also greatly contributed to communicating evolutionary and marine biology and conservation to the general public, and has received multiple prizes for this; for example, the Swedish “Kunskapspriset” for communicating science to society in 2016. She has written numerous articles for popular science books and journals, and has frequently been represented in the media. For example, she led a children's TV programme about marine biology for the Swedish TV (“Havsforskarna”). Kerstin has also been involved in educating school teachers, providing them with “hands-on” experiences with biological experiments at the Tjärnö Marine Laboratory. But her most impressive “outreach” activity was that she used a dinner with the Swedish king to tell him about snails! To us, what is most outstanding about Kerstin is her inspiring personality. Despite her obvious successes (including membership in the Royal Swedish Academy of Sciences), she has remained down-to-earth and is clearly driven by great curiosity and the joy of scientific work, rather than a desire for prestige. Kerstin is remarkably calm among her seemingly countless jobs. She is always willing to discuss and ask questions, without worrying how others might judge her. Kerstin spends a lot of time doing fieldwork (including driving the boat) and performing seemingly menial tasks like crosses and phenotyping assays in the laboratory, still making new discoveries while doing so. Her rejection of hierarchy and elitism make Kerstin easily approachable for more junior researchers, many of whom she has strongly supported over many years. We believe that especially in times of increasing competition and pressure in science, her balanced curiosity-centred approach makes her an outstanding role model for young (and old) scientists. Personally, we have greatly enjoyed our long-term collaborations with Kerstin and have certainly learned much more than snail facts. We cannot think of a better recipient for the 2022 Molecular Ecology Prize.

Hybridization is a common evolutionary process with multiple possible outcomes. In vertebrates, interspecific hybridization has repeatedly generated parthenogenetic hybrid species. However, it is unknown whether the generation of parthenogenetic hybrids is a rare outcome of frequent hybridization between sexual species within a genus or the typical outcome of rare hybridization events. Darevskia is a genus of rock lizards with both hybrid parthenogenetic and sexual species. Using capture sequencing, we estimate phylogenetic relationships and gene flow among the sexual species, to determine how introgressive hybridization relates to the origins of parthenogenetic hybrids. We find evidence for widespread hybridization with gene flow, both between recently diverged species and deep branches. Surprisingly, we find no signal of gene flow between parental species of the parthenogenetic hybrids, suggesting that the parental pairs were either reproductively or geographically isolated early in their divergence. The generation of parthenogenetic hybrids in Darevskia is, then, a rare outcome of the total occurrence of hybridization within the genus, but the typical outcome when specific species pairs hybridize. Our results question the conventional view that parthenogenetic lineages are generated by hybridization in a window of divergence. Instead, they suggest that some lineages possess specific properties that underpin successful parthenogenetic reproduction.

ABSTRACTConflict over reproduction between females and males exists because of anisogamy and promiscuity. Together they generate differences in fitness optima between the sexes and result in antagonistic coevolution of female and male reproductive traits. Mounting duration is likely to be a compromise between male and female interests whose outcome depends on the intensity of sexual selection. The timing of sperm transfer during mounting is critical. For example, mountings may be interrupted before sperm is transferred as a consequence of female or male choice, or they may be prolonged to function as mate guarding. In the highly promiscuous intertidal snail Littorina saxatilis, mountings vary substantially in duration, from less than a minute to more than an hour, and it has been assumed that mountings of a few minutes do not result in any sperm being transferred. Here, we examined the timing of sperm transfer, a reproductive trait that is likely affected by sexual conflict. We performed time-controlled mounting trials using L. saxatilis males and virgin females, aiming to examine indirectly when the transfer of sperm starts. We observed the relationship between mounting duration and the proportion of developing embryos out of all eggs and embryos in the brood pouch. Developing embryos were observed in similar proportions in all treatments (i.e. 1, 5 and 10 or more minutes at which mountings were artificially interrupted), suggesting that sperm transfer begins rapidly (within 1 min) in L. saxatilis and very short matings do not result in sperm shortage in the females. We discuss how the observed pattern can be influenced by predation risk, population density, and female status and receptivity.

Some say that when the title of a paper is phrased as a question, the authors will not be able to give a clear answer. Our article ‘What is reproductive isolation?’ (Westram et al., 2022a,b) is no exception: noticing that the term ‘Reproductive isolation’ (RI) is frequently used in the literature, yet almost never clearly defined, we tried to come up with a definition— but we quickly ran into numerous complexities that kept us from finding a simple answer. The complexity of the issue is reflected by the commentaries to our article, which highlight the diversity of views among speciation researchers with different interests, different empirical research experiences and different favourite species concepts [a diversity also highlighted by Rosales (2022)]. Some of the commentaries discuss the conceptual (Moyle et al., 2022) and practical (Stuckert & Matute, 2022) limitations of definitions of RI based on gene flow, whereas Planidin et al. (2022) extend the concept to aspects not explicitly covered by us. Rosales (2022) discusses how scientific definitions are developed and what they mean for the community and analyses our work in this context. In this response, we elaborate on why we think a definition based on gene flow is appropriate if we want to ‘make explicit and further elaborate’ (Rosales, 2022) the already existing concept of RI. In addition, we consider other important axes along which speciation could be quantified that were highlighted by the commentaries and discuss how they relate to RI. Numerous other interesting points brought up by the commentaries can unfortunately not be discussed here for reasons of space.

Reproductive isolation (RI) is a core concept in evolutionary biology. It has been the central focus of speciation research since the modern synthesis and is the basis by which biological species are defined. Despite this, the term is used in seemingly different ways, and attempts to quantify RI have used very different approaches. After showing that the field lacks a clear definition of the term, we attempt to clarify key issues, including what RI is, how it can be quantified in principle, and how it can be measured in practice. Following other definitions with a genetic focus, we propose that RI is a quantitative measure of the effect that genetic differences between populations have on gene flow. Specifically, RI compares the flow of neutral alleles in the presence of these genetic differences to the flow without any such differences. RI is thus greater than zero when genetic differences between populations reduce the flow of neutral alleles between populations. We show how RI can be quantified in a range of scenarios. A key conclusion is that RI depends strongly on circumstances-including the spatial, temporal and genomic context-making it difficult to compare across systems. After reviewing methods for estimating RI from data, we conclude that it is difficult to measure in practice. We discuss our findings in light of the goals of speciation research and encourage the use of methods for estimating RI that integrate organismal and genetic approaches.

Local adaptation leads to differences between populations within a species. In many systems, similar environmental contrasts occur repeatedly, sometimes driving parallel phenotypic evolution. Understanding the genomic basis of local adaptation and parallel evolution is a major goal of evolutionary genomics. It is now known that by preventing the break-up of favourable combinations of alleles across multiple loci, genetic architectures that reduce recombination, like chromosomal inversions, can make an important contribution to local adaptation. However, little is known about whether inversions also contribute disproportionately to parallel evolution. Our aim here is to highlight this knowledge gap, to showcase existing studies, and to illustrate the differences between genomic architectures with and without inversions using simple models. We predict that by generating stronger effective selection, inversions can sometimes speed up the parallel adaptive process or enable parallel adaptation where it would be impossible otherwise, but this is highly dependent on the spatial setting. We highlight that further empirical work is needed, in particular to cover a broader taxonomic range and to understand the relative importance of inversions compared to genomic regions without inversions. This article is part of the theme issue ‘Genomic architecture of supergenes: causes and evolutionary consequences’.

Chromosomal inversions have been shown to play a major role in a local adaptation by suppressing recombination between alternative arrangements and maintaining beneficial allele combinations. However, so far, their importance relative to the remaining genome remains largely unknown. Understanding the genetic architecture of adaptation requires better estimates of how loci of different effect sizes contribute to phenotypic variation. Here, we used three Swedish islands where the marine snail Littorina saxatilis has repeatedly evolved into two distinct ecotypes along a habitat transition. We estimated the contribution of inversion polymorphisms to phenotypic divergence while controlling for polygenic effects in the remaining genome using a quantitative genetics framework. We confirmed the importance of inversions but showed that contributions of loci outside inversions are of similar magnitude, with variable proportions dependent on the trait and the population. Some inversions showed consistent effects across all sites, whereas others exhibited site-specific effects, indicating that the genomic basis for replicated phenotypic divergence is only partly shared. The contributions of sexual dimorphism as well as environmental factors to phenotypic variation were significant but minor compared to inversions and polygenic background. Overall, this integrated approach provides insight into the multiple mechanisms contributing to parallel phenotypic divergence.