Relaxed selection in evolution of genes regulating limb development gives clue to variation in forelimb morphology of cetaceans and other mammals.

Cetaceans have evolved unique limb structures, such as flippers, due to genetic changes during their transition to aquatic life. However, the full understanding of the genetic and evolutionary mechanisms behind these changes is still developing. By examining 25 limb-related protein-coding genes across various mammalian species, we compared genetic changes between aquatic mammals, like whales, and other mammals with unique limb structures such as bats, rodents and elephants. Our findings revealed significant modifications in limb-related genes, including variations in the Hox, GDF5 and Evx genes. Notably, a relaxed selection in several key genes was observed, suggesting a lifting of developmental constraints, which might have facilitated the emergence of morphological innovations in cetacean limb morphology. We also uncovered non-synonymous changes, insertions and deletions in these genes, particularly in the polyalanine tract of HOXD13, which are distinctive to cetaceans or convergent with other aquatic mammals. These genetic variations correlated with the diverse and specialized limb structures observed in cetaceans, indicating a complex interplay of relaxed selection and specific mutations in mammalian limb evolution.

Pervasive relaxed selection in termite genomes.

Genetic changes that enabled the evolution of eusociality have long captivated biologists. More recently, attention has focussed on the consequences of eusociality on genome evolution. Studies have reported higher molecular evolutionary rates in eusocial hymenopteran insects compared with their solitary relatives. To investigate the genomic consequences of eusociality in termites, we analysed nine genomes, including newly sequenced genomes from three non-eusocial cockroaches. Using a phylogenomic approach, we found that termite genomes have experienced lower rates of synonymous substitutions than those of cockroaches, possibly as a result of longer generation times. We identified higher rates of non-synonymous substitutions in termite genomes than in cockroach genomes, and identified pervasive relaxed selection in the former (24-31% of the genes analysed) compared with the latter (2-4%). We infer that this is due to reductions in effective population size, rather than gene-specific effects (e.g. indirect selection of caste-biased genes). We found no obvious signature of increased genetic load in termites, and postulate efficient purging of deleterious alleles at the colony level. Additionally, we identified genomic adaptations that may underpin caste differentiation, such as genes involved in post-translational modifications. Our results provide insights into the evolution of termites and the genomic consequences of eusociality more broadly.

Phased genome assemblies reveal haplotype-specific genetic load in the critically endangered Chinese Bahaba (Teleostei, Sciaenidae).

While haplotype-specific genetic load shapes the evolutionary trajectory of natural and captive populations, mixed-haplotype assembly and genotyping hindered its characterization in diploids. Herein, we produced two phased genome assemblies of the critically endangered fish Chinese Bahaba (Bahaba taipingensis, Sciaenidae, Teleostei) and resequenced 20 whole genomes to quantify population genetic load at a haplotype level. We identified frame-shifting variants as the most deleterious type, followed by mutations in the 5'-UTR, 3'-UTR and missense mutations at conserved amino acids. Phased haplotypes revealed gene deletions and high-impact deleterious variants. We estimated ~1.12% of genes missing or interrupted per haplotype, with a significant overlap of disrupted genes (30.35%) between haplotype sets. Relative proportions of deleterious variant categories differed significantly between haplotypes. Simulations suggested that purifying selection struggled to purge slightly deleterious genetic load in captive breeding compared to genotyping interventions, and that higher inter-haplotypic variance of genetic load predicted more efficient purging by artificial selection. Combining the knowledge of haplotype-resolved genetic load with predictive modelling will be immensely useful for understanding the evolution of deleterious variants and guiding conservation planning.

Comparative gene retention analysis in barley, wild emmer, and bread wheat pangenome lines reveals factors affecting gene retention following gene duplication.

BACKGROUND: Gene duplication is a prevalent phenomenon and a major driving force underlying genome evolution. The process leading to the fixation of gene duplicates following duplication is critical to understand how genome evolves but remains fragmentally understood. Most previous studies on gene retention are based on gene duplicate analyses in single reference genome. No population-based comparative gene retention analysis has been performed to date. RESULTS: Taking advantage of recently published genomic data in Triticeae, we dissected a divergent homogentisate phytyltransferase (HPT2) lineage caught in the middle stage of gene fixation following duplication. The presence/absence of HPT2 in barley (diploid), wild emmer (tetraploid), and bread wheat (hexaploid) pangenome lines appears to be associated with gene dosage constraint and environmental adaption. Based on these observations, we adopted a phylogeny-based orthology inference approach and performed comparative gene retention analyses across barley, wild emmer, and bread wheat. This led to the identification of 326 HPT2-pattern-like genes at whole genome scale, representing a pool of gene duplicates in the middle stage of gene fixation. Majority of these HPT2-pattern-like genes were identified as small-scale duplicates, such as dispersed, tandem, and proximal duplications. Natural selection analyses showed that HPT2-pattern-like genes have experienced relaxed selection pressure, which is generally accompanied with partial positive selection and transcriptional divergence. Functional enrichment analyses showed that HPT2-pattern-like genes are over-represented with molecular-binding and defense response functions, supporting the potential role of environmental adaption during gene retention. We also observed that gene duplicates from larger gene family are more likely to be lost, implying a gene dosage constraint effect. Further comparative gene retention analysis in barley and bread wheat pangenome lines revealed combined effects of species-specific selection and gene dosage constraint. CONCLUSIONS: Comparative gene retention analyses at the population level support gene dosage constraint, environmental adaption, and species-specific selection as three factors that may affect gene retention following gene duplication. Our findings shed light on the evolutionary process leading to the retention of newly formed gene duplicates and will greatly improve our understanding on genome evolution via duplication.

Evolvability Costs of Niche Expansion.

What prevents generalists from displacing specialists, despite obvious competitive advantages of utilizing a broad niche? The classic genetic explanation is antagonistic pleiotropy: genes underlying the generalism produce 'jacks-of-all-trades' that are masters of none. However, experiments challenge this assumption that mutations enabling niche expansion must reduce fitness in other environments. Theory suggests an alternative cost of generalism: decreased evolvability, or the reduced capacity to adapt. Generalists using multiple environments experience relaxed selection in any one environment, producing greater relative lag load. Additionally, mutations fixed by generalist lineages early during their evolution that avoid or compensate for antagonistic pleiotropy may limit access to certain future evolutionary trajectories. Hypothesized evolvability costs of generalism warrant further exploration, and we suggest outstanding questions meriting attention.

Relaxed Selection and the Rapid Evolution of Reproductive Genes.

Evolutionary genomic studies find that reproductive protein genes, those directly involved in reproductive processes, diversify more rapidly than most other gene categories. Strong postcopulatory sexual selection acting within species is the predominant hypothesis proposed to account for the observed pattern. Recently, relaxed selection due to sex-specific gene expression has also been put forward to explain the relatively rapid diversification. We contend that relaxed selection due to sex-limited gene expression is the correct null model for tests of molecular evolution of reproductive genes and argue that it may play a more significant role in the evolutionary diversification of reproductive genes than previously recognized. We advocate for a re-evaluation of adaptive explanations for the rapid diversification of reproductive genes.

Reduced social function in experimentally evolved Dictyostelium discoideum implies selection for social conflict in nature.

Many microbes interact with one another, but the difficulty of directly observing these interactions in nature makes interpreting their adaptive value complicated. The social amoeba Dictyostelium discoideum forms aggregates wherein some cells are sacrificed for the benefit of others. Within chimaeric aggregates containing multiple unrelated lineages, cheaters can gain an advantage by undercontributing, but the extent to which wild D. discoideum has adapted to cheat is not fully clear. In this study, we experimentally evolved D. discoideum in an environment where there were no selective pressures to cheat or resist cheating in chimaeras. Dictyostelium discoideum lines grown in this environment evolved reduced competitiveness within chimaeric aggregates and reduced ability to migrate during the slug stage. By contrast, we did not observe a reduction in cell number, a trait for which selection was not relaxed. The observed loss of traits that our laboratory conditions had made irrelevant suggests that these traits were adaptations driven and maintained by selective pressures D. discoideum faces in its natural environment. Our results suggest that D. discoideum faces social conflict in nature, and illustrate a general approach that could be applied to searching for social or non-social adaptations in other microbes.

Evolutionary consequences of loss of sexual reproduction on male-related traits in parthenogenetic lineages of the pea aphid.

Transition from sexual reproduction to parthenogenesis constitutes a major life-history change with deep evolutionary consequences for sex-related traits, which are expected to decay. The pea aphid Acyrthosiphon pisum shows intraspecific reproductive polymorphism, with cold-resistant cyclically parthenogenetic (CP) lineages that alternate sexual and asexual generations and cold-sensitive obligately parthenogenetic (OP) lineages that produce only asexual females but still males. Here, the genotyping of 219 pea aphid lineages collected in cold-winter and mild-winter regions revealed contrasting population structures. Samples from cold-winter regions consisted mostly of distinct multilocus genotypes (MLGs) usually represented by a single sample (101 different MLGs for 111 samples) and were all phenotyped as CP. In contrast, fewer MLGs were found in mild-winter regions (28 MLGs for 108 samples), all but one being OP. Since the males produced by OP lineages are unlikely to pass on their genes (sexual females being rare in mild-winter regions), we tested the hypothesis that their traits could degenerate due to lack of selection by comparing male production and male reproductive success between OP and CP lineages. Male production was indeed reduced in OP lineages, but a less clear pattern was observed for male reproductive success: females mated with OP males laid fewer eggs (fertilized or not) but OP and CP males fertilized the same proportion of eggs. These differences may stem from the type of selective forces: male production may be counter-selected whereas male performances may evolve under the slower process of relaxed selection. The overall effective reproductive capacity of OP males could result from recent sex loss in OP lineages or underestimated reproductive opportunities.

Two centuries of monarch butterfly collections reveal contrasting effects of range expansion and migration loss on wing traits.

Migratory animals exhibit traits that allow them to exploit seasonally variable habitats. In environments where migration is no longer beneficial, such as oceanic islands, migration-association traits may be selected against or be under relaxed selection. Monarch butterflies are best known for their continent-scale migration in North America but have repeatedly become established as nonmigrants in the tropical Americas and on Atlantic and Pacific Islands. These replicated nonmigratory populations provide natural laboratories for understanding the rate of evolution of migration-associated traits. We measured >6,000 museum specimens of monarch butterflies collected from 1856 to the present as well as contemporary wild-caught monarchs from around the world. We determined 1) how wing morphology varies across the monarch's global range, 2) whether initial long-distance founders were particularly suited for migration, and 3) whether recently established nonmigrants show evidence for contemporary phenotypic evolution. We further reared >1,000 monarchs from six populations around the world under controlled conditions and measured migration-associated traits. Historical specimens show that 1) initial founders are well suited for long-distance movement and 2) loss of seasonal migration is associated with reductions in forewing size and elongation. Monarch butterflies raised in a common garden from four derived nonmigratory populations exhibit genetically based reductions in forewing size, consistent with a previous study. Our findings provide a compelling example of how migration-associated traits may be favored during the early stages of range expansion, and also the rate of reductions in those same traits upon loss of migration.

Contrasting Gene Decay in Subterranean Vertebrates: Insights from Cavefishes and Fossorial Mammals.

Evolution sometimes proceeds by loss, especially when structures and genes become dispensable after an environmental shift relaxes functional constraints. Subterranean vertebrates are outstanding models to analyze this process, and gene decay can serve as a readout. We sought to understand some general principles on the extent and tempo of the decay of genes involved in vision, circadian clock, and pigmentation in cavefishes. The analysis of the genomes of two Cuban species belonging to the genus Lucifuga provided evidence for the largest loss of eye-specific genes and nonvisual opsin genes reported so far in cavefishes. Comparisons with a recently evolved cave population of Astyanax mexicanus and three species belonging to the Chinese tetraploid genus Sinocyclocheilus revealed the combined effects of the level of eye regression, time, and genome ploidy on eye-specific gene pseudogenization. The limited extent of gene decay in all these cavefishes and the very small number of loss-of-function mutations per pseudogene suggest that their eye degeneration may not be very ancient, ranging from early to late Pleistocene. This is in sharp contrast with the identification of several vision genes carrying many loss-of-function mutations in ancient fossorial mammals, further suggesting that blind fishes cannot thrive more than a few million years in cave ecosystems.

How broad is the selfing syndrome? Insights from convergent evolution of gene expression across species and tissues in the Capsella genus.

The shift from outcrossing to selfing is one of the main evolutionary transitions in plants. It is accompanied by profound effects on reproductive traits, the so-called selfing syndrome. Because the transition to selfing also implies deep genomic and ecological changes, one also expects to observe a genomic selfing syndrome. We took advantage of the three independent transitions from outcrossing to selfing in the Capsella genus to characterize the overall impact of mating system change on RNA expression, in flowers but also in leaves and roots. We quantified the extent of both selfing and genomic syndromes, and tested whether changes in expression corresponded to adaptation to selfing or to relaxed selection on traits that were constrained in outcrossers. Mating system change affected gene expression in all three tissues but more so in flowers than in roots and leaves. Gene expression in selfing species tended to converge in flowers but diverged in the two other tissues. Hence, convergent adaptation to selfing dominates in flowers, whereas genetic drift plays a more important role in leaves and roots. The effect of mating system transition is not limited to reproductive tissues and corresponds to both adaptation to selfing and relaxed selection on previously constrained traits.

Novel genome sequence of Chinese cavefish (Triplophysa rosa) reveals pervasive relaxation of natural selection in cavefish genomes.

All cavefishes, living exclusively in caves across the globe, exhibit similar phenotypic traits, including the characteristic loss of eyes. To understand whether such phenotypic convergence shares similar genomic bases, here we investigated genome-wide evolutionary signatures of cavefish phenotypes by comparing whole-genome sequences of three pairs of cavefishes and their surface fish relatives. Notably, we newly sequenced and generated a whole-genome assembly of the Chinese cavefish Triplophysa rosa. Our comparative analyses revealed several shared features of cavefish genome evolution. Cavefishes had lower mutation rates than their surface fish relatives. In contrast, the ratio of nonsynonymous to synonymous substitutions (ω) was significantly elevated in cavefishes compared to in surface fishes, consistent with the relaxation of purifying selection. In addition, cavefish genomes had an increased mutational load, including mutations that alter protein hydrophobicity profiles, which were considered harmful. Interestingly, however, we found no overlap in positively selected genes among different cavefish lineages, indicating that the phenotypic convergence in cavefishes was not caused by positive selection of the same sets of genes. Analyses of previously identified candidate genes associated with cave phenotypes supported this conclusion. Genes belonging to the lipid metabolism functional ontology were under relaxed purifying selection in all cavefish genomes, which may be associated with the nutrient-poor habitat of cavefishes. Our work reveals previously uncharacterized patterns of cavefish genome evolution and provides comparative insights into the evolution of cave-associated phenotypic traits.

Evolution, inactivation and loss of short wavelength-sensitive opsin genes during the diversification of Neotropical cichlids.

Natural variation in the number, expression and function of sensory genes in an organism's genome is often tightly linked to different ecological and evolutionary forces. Opsin genes, which code for the first step in visual transduction, are ideal models for testing how ecological factors such as light environment may influence visual system adaptation. Neotropical cichlid fishes are a highly ecologically diverse group that evolved in a variety of aquatic habitats, including black (stained), white (opaque) and clear waters. We used cross-species exon capture to sequence Neotropical cichlid short wavelength-sensitive (SWS) opsins, which mediate ultraviolet (UV) to blue visual sensitivity. Neotropical cichlid SWS1 opsin (UV-sensitive) underwent a relaxation of selective constraint during the early phases of cichlid diversification in South America, leading to pseudogenization and loss. Conversely, SWS2a (blue-sensitive) experienced a burst of episodic positive selection at the base of the South American cichlid radiation. This burst coincides with SWS1 relaxation and loss, and is consistent with findings in ecomorphological studies characterizing a period of extensive ecological divergence in Neotropical cichlids. We use ancestral sequence reconstruction and protein modelling to investigate mutations along this ancestral branch that probably modified SWS2a function. Together, our results suggest that variable light environments played a prominent early role in shaping SWS opsin diversity during the Neotropical cichlid radiation. Our results also illustrate that long-term evolution under light-limited conditions in South America may have reduced visual system plasticity; specifically, early losses of UV sensitivity may have constrained the evolutionary trajectory of Neotropical cichlid vision.

Genetic load and biological changes to extant humans.

Extant humans are currently increasing their genetic load, which is informing present and future human microevolution. This has been a gradual process that has been rising over the last centuries as a consequence of improved sanitation, nutritional improvements, advancements in microbiology and medical interventions, which have relaxed natural selection. Moreover, a reduction in infant and child mortality and changing societal attitudes towards fertility have led to a decrease in total fertility rates (TFRs) since the 19th century. Generally speaking, decreases in differential fertility and mortality have meant that there is less opportunity for natural selection to eliminate deleterious mutations from the human gene pool. It has been argued that the average human may carry ~250-300 mutations that are mostly deleterious, as well as several hundred less-deleterious variants. These deleterious alleles in extant humans mean that our fitness is being constrained. While such alleles are viewed as reducing human fitness, they may also have had an adaptive function in the past, such as assisting in genetic complexity, sexual recombination and diploidy. Saying this, our current knowledge on these fitness compromising alleles is still lacking.

The cognitive and speech genes are jointly shaped by both positive and relaxed selection in the human lineage.

The emergence of a coordinated network of cognitive and speech genes in the human lineage performing overlapping functions is a great evolutionary puzzle. Prior studies on the speech gene FOXP2 are inconclusive on the nature of selection operating on this gene in the human lineage. Here, I show that the evolution of FOXP2 is accelerated in the human lineage due to relaxation of purifying selection (relaxed selection). Five potential genes associated with human-specific intelligence and speech genes have evolved under the impact of positive selection and three genes including FOXP2 have undergone relaxation of purifying selection in the human lineage. Overall, three evolutionary processes namely positive selection, relaxation of purifying selection and neutral evolution have contributed for the genomic evolution of extraordinary cognitive ability and speech in the hominin lineage. The cognitive and speech genes subjected to natural selection in the human lineage have demonstrated a coevolutionary trend.

Plastomes of eight Ligusticum species: characterization, genome evolution, and phylogenetic relationships.

BACKGROUND: The genus Ligusticum consists of approximately 60 species distributed in the Northern Hemisphere. It is one of the most taxonomically difficult taxa within Apiaceae, largely due to the varied morphological characteristics. To investigate the plastome evolution and phylogenetic relationships of Ligusticum, we determined the complete plastome sequences of eight Ligusticum species using a de novo assembly approach. RESULTS: Through a comprehensive comparative analysis, we found that the eight plastomes were similar in terms of repeat sequence, SSR, codon usage, and RNA editing site. However, compared with the other seven species, L. delavayi exhibited striking differences in genome size, gene number, IR/SC borders, and sequence identity. Most of the genes remained under the purifying selection, whereas four genes showed relaxed selection, namely ccsA, rpoA, ycf1, and ycf2. Non-monophyly of Ligusticum species was inferred from the plastomes and internal transcribed spacer (ITS) sequences phylogenetic analyses. CONCLUSION: The plastome tree and ITS tree produced incongruent tree topologies, which may be attributed to the hybridization and incomplete lineage sorting. Our study highlighted the advantage of plastome with mass informative sites in resolving phylogenetic relationships. Moreover, combined with the previous studies, we considered that the current taxonomy system of Ligusticum needs to be improved and revised. In summary, our study provides new insights into the plastome evolution, phylogeny, and taxonomy of Ligusticum species.

Male phenotypes in a female framework: Evidence for degeneration in sperm produced by male snails from asexual lineages.

How changes in selective regimes affect trait evolution is an important open biological question. We take advantage of naturally occurring and repeated transitions from sexual to asexual reproduction in a New Zealand freshwater snail species, Potamopyrgus antipodarum, to address how evolution in an asexual context-including the potential for relaxed selection on male-specific traits-influences sperm morphology. The occasional production of male offspring by the otherwise all-female asexual P. antipodarum lineages affords a unique and powerful opportunity to assess the fate of sperm traits in a context where males are exceedingly rare. These comparisons revealed that the sperm produced by 'asexual' males are markedly distinct from sexual counterparts. We also found that the asexual male sperm harboured markedly higher phenotypic variation and was much more likely to be morphologically abnormal. Together, these data suggest that transitions to asexual reproduction might be irreversible, at least in part because male function is likely to be compromised. These results are also consistent with a scenario where relaxed selection and/or mutation accumulation in the absence of sex translates into rapid trait degeneration.

The Organelle Genomes in the Photosynthetic Red Algal Parasite Pterocladiophila hemisphaerica (Florideophyceae, Rhodophyta) Have Elevated Substitution Rates and Extreme Gene Loss in the Plastid Genome.

Comparative organelle genome studies of parasites can highlight genetic changes that occur during the transition from a free-living to a parasitic state. Our study focuses on a poorly studied group of red algal parasites, which are often closely related to their red algal hosts and from which they presumably evolved. Most of these parasites are pigmented and some show photosynthetic capacity. Here, we assembled and annotated the complete organelle genomes of the photosynthetic red algal parasite, Pterocladiophila hemisphaerica. The plastid genome is the smallest known red algal plastid genome at 68,701 bp. The plastid genome has many genes missing, including all photosynthesis-related genes. In contrast, the mitochondrial genome is similar in architecture to that of other free-living red algae. Both organelle genomes show elevated mutation rates and significant changes in patterns of selection, measured as dN/dS ratios. This caused phylogenetic analyses, even of multiple aligned proteins, to be unresolved or give contradictory relationships. Full plastid datasets interfered by selected best gene evolution models showed the supported relationship of P. hemisphaerica within the Ceramiales, but the parasite was grouped with support as sister to the Gracilariales when interfered under the GHOST model. Nuclear rDNA showed a supported grouping of the parasite within a clade containing several red algal orders including the Gelidiales. This photosynthetic parasite, which is unable to photosynthesize with its own plastid due to the total loss of all photosynthesis genes, raises intriguing questions on parasite-host organelle genome capabilities and interactions.

Analysis of substitution rates showed that TLR5 is evolving at different rates among mammalian groups.

BACKGROUND: Toll-like receptors (TLRs) are the most widely studied innate immunity receptors responsible for recognition of invading pathogens. Among the TLR family, TLR5 is the only that senses and recognizes flagellin, the major protein of bacterial flagella. TLR5 has been reported to be under overall purifying selection in mammals, with a small proportion of codons under positive selection. However, the variation of substitution rates among major mammalian groups has been neglected. Here, we studied the evolution of TLR5 in mammals, comparing the substitution rates among groups. RESULTS: In this study we analysed the TLR5 substitution rates in Euungulata, Carnivora, Chiroptera, Primata, Rodentia and Lagomorpha, groups. For that, Tajima's relative rate test, Bayesian inference of evolutionary rates and genetic distances were estimated with CODEML's branch model and RELAX. The combined results showed that in the Lagomorpha, Rodentia, Carnivora and Chiroptera lineages TLR5 is evolving at a higher substitution rate. The RELAX analysis further suggested a significant relaxation of selective pressures for the Lagomorpha (K = 0.22, p < 0.01), Rodentia (K = 0.58, p < 0.01) and Chiroptera (K = 0.65, p < 0.01) lineages and for the Carnivora ancestral branches (K = 0.13, p < 0.01). CONCLUSIONS: Our results show that the TLR5 substitution rate is not uniform among mammals. In fact, among the different mammal groups studied, the Lagomorpha, Rodentia, Carnivora and Chiroptera are evolving faster. This evolutionary pattern could be explained by 1) the acquisition of new functions of TLR5 in the groups with higher substitution rate, i.e. TLR5 neofunctionalization, 2) by the beginning of a TLR5 pseudogenization in these groups due to some redundancy between the TLRs genes, or 3) an arms race between TLR5 and species-specific parasites.

Parallel selection on ecologically relevant gene functions in the transcriptomes of highly diversifying salmonids.

BACKGROUND: Salmonid fishes are characterised by a very high level of variation in trophic, ecological, physiological, and life history adaptations. Some salmonid taxa show exceptional potential for fast, within-lake diversification into morphologically and ecologically distinct variants, often in parallel; these are the lake-resident charr and whitefish (several species in the genera Salvelinus and Coregonus). To identify selection on genes and gene categories associated with such predictable diversifications, we analysed 2702 orthogroups (4.82 Mbp total; average 4.77 genes/orthogroup; average 1783 bp/orthogroup). We did so in two charr and two whitefish species and compared to five other salmonid lineages, which do not evolve in such ecologically predictable ways, and one non-salmonid outgroup. RESULTS: All selection analyses are based on Coregonus and Salvelinus compared to non-diversifying taxa. We found more orthogroups were affected by relaxed selection than intensified selection. Of those, 122 were under significant relaxed selection, with trends of an overrepresentation of serine family amino acid metabolism and transcriptional regulation, and significant enrichment of behaviour-associated gene functions. Seventy-eight orthogroups were under significant intensified selection and were enriched for signalling process and transcriptional regulation gene ontology terms and actin filament and lipid metabolism gene sets. Ninety-two orthogroups were under diversifying/positive selection. These were enriched for signal transduction, transmembrane transport, and pyruvate metabolism gene ontology terms and often contained genes involved in transcriptional regulation and development. Several orthogroups showed signs of multiple types of selection. For example, orthogroups under relaxed and diversifying selection contained genes such as ap1m2, involved in immunity and development, and slc6a8, playing an important role in muscle and brain creatine uptake. Orthogroups under intensified and diversifying selection were also found, such as genes syn3, with a role in neural processes, and ctsk, involved in bone remodelling. CONCLUSIONS: Our approach pinpointed relevant genomic targets by distinguishing among different kinds of selection. We found that relaxed, intensified, and diversifying selection affect orthogroups and gene functions of ecological relevance in salmonids. Because they were found consistently and robustly across charr and whitefish and not other salmonid lineages, we propose these genes have a potential role in the replicated ecological diversifications.