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BackgroundTrypanosomes are single-celled eukaryotic parasites characterised by the unique biology of their mitochondrial DNA. African livestock trypanosomes impose a major burden on agriculture across sub-Saharan Africa, but are poorly understood compared to those that cause sleeping sickness and Chagas disease in humans. Here we explore the potential of the maxicircle, a component of trypanosome mitochondrial DNA to study the evolutionary history of trypanosomes.ResultsWe used long-read sequencing to completely assemble maxicircle mitochondrial DNA from four previously uncharacterized African trypanosomes, and leveraged these assemblies to scaffold and assemble a further 103 trypanosome maxicircle gene coding regions from published short-read data. While synteny was largely conserved, there were repeated, independent losses of Complex I genes. Comparison of pre-edited and non-edited genes revealed the impact of RNA editing on nucleotide composition, with non-edited genes approaching the limits of GC loss. African tsetse-transmitted trypanosomes showed high levels of RNA editing compared to other trypanosomes. The gene coding regions of maxicircle mitochondrial DNAs were used to construct time-resolved phylogenetic trees, revealing deep divergence events among isolates of the pathogensTrypanosoma bruceiandT. congolense.ConclusionsOur data represents a new resource for experimental and evolutionary analyses of trypanosome phylogeny, molecular evolution and function. Molecular clock analyses yielded a timescale for trypanosome evolution congruent with major biogeographical events in Africa and revealed the recent emergence ofTrypanosoma brucei gambienseandT. equiperdum, major human and animal pathogens.

Background Eukaryotic protein-coding genes consist of exons and introns. Exon–intron borders are conserved between species and thus their changes might be observed only on quite long evolutionary distances. One of the rarest types of change, in which intron relocates over a short distance, is called "intron sliding", but the reality of this event has been debated for a long time. The main idea of a search for intron sliding is to use the most accurate genome annotation and genome sequence, as well as high-quality transcriptome data. We applied them in a search for sliding introns in mammals in order to widen knowledge about the presence or absence of such phenomena in this group. Results We didn’t find any significant evidence of intron sliding in the primate group (human, chimpanzee, rhesus macaque, crab-eating macaque, green monkey, marmoset). Only one possible intron sliding event supported by a set of high quality transcriptomes was observed between EIF1AX human and sheep gene orthologs. Also, we checked a list of previously observed intron sliding events in mammals and showed that most likely they are artifacts of genome annotations and are not shown in subsequent annotation versions as well as are not supported by transcriptomic data. Conclusions We assume that intron sliding is indeed a very rare evolutionary event if it exists at all. Every case of intron sliding needs a lot of supportive data for detection and confirmation.

Background Phenotypic convergence between distinct species provides an opportunity to examine the predictability of genetic evolution. Unrelated species sharing genetic underpinnings for phenotypic convergence suggests strong genetic constraints, and thus high predictability of evolution. However, there is no clear big picture of the genomic constraints on convergent evolution. Genome-based phylogenies have confirmed many cases of phenotypic convergence in birds, making them a good system for examining genetic constraints in phenotypic convergence. In this study, we used hierarchical genomic approaches to estimate genetic constraints in three convergent avian traits: nocturnality, raptorial behavior and foot-propelled diving. Results Phylogeny-based hypothesis tests and positive selection tests were applied to compare 16 avian genomes, representing 14 orders, and identify genes with strong convergence signals. We found 43 adaptively convergent genes (ACGs) associated with the three phenotypic convergence cases and assessed genetic constraints in all three cases, from (amino acid) site mutations to genetic pathways. We found that the avian orders shared few site mutations in the ACGs that contributed to the convergent phenotypes, and that these ACGs were not enriched in any genetic pathways. In addition, different pairs of orders with convergent foot-propelled diving or raptorial behaviors shared few ACGs. We also found that closely related orders that shared foot-propelled diving behavior did not share more ACGs than did distinct orders, suggesting that convergence among these orders could not be explained by their initial genomic backgrounds. Conclusions Our analyses of three avian convergence events suggest low constraints for phenotypic convergence across multiple genetic levels, implying that genetic evolution is unpredictable at the phylogenetic level of avian order. Ours is one of first studies to apply hierarchical genomic examination to multiple avian convergent cases to assess the genetic constraints in life history trait evolution.

Background Phylogenetic relationships among the myriapod subgroups Chilopoda, Diplopoda, Symphyla and Pauropoda are still not robustly resolved. The first phylogenomic study covering all subgroups resolved phylogenetic relationships congruently to morphological evidence but is in conflict with most previously published phylogenetic trees based on diverse molecular data. Outgroup choice and long-branch attraction effects were stated as possible explanations for these incongruencies. In this study, we addressed these issues by extending the myriapod and outgroup taxon sampling using transcriptome data. Results We generated new transcriptome data of 42 panarthropod species, including all four myriapod subgroups and additional outgroup taxa. Our taxon sampling was complemented by published transcriptome and genome data resulting in a supermatrix covering 59 species. We compiled two data sets, the first with a full coverage of genes per species (292 single-copy protein-coding genes), the second with a less stringent coverage (988 genes). We inferred phylogenetic relationships among myriapods using different data types, tree inference, and quartet computation approaches. Our results unambiguously support monophyletic Mandibulata and Myriapoda. Our analyses clearly showed that there is strong signal for a single unrooted topology, but a sensitivity of the position of the internal root on the choice of outgroups. However, we observe strong evidence for a clade Pauropoda+Symphyla, as well as for a clade Chilopoda+Diplopoda. Conclusions Our best quartet topology is incongruent with current morphological phylogenies which were supported in another phylogenomic study. AU tests and quartet mapping reject the quartet topology congruent to trees inferred with morphological characters. Moreover, quartet mapping shows that confounding signal present in the data set is sufficient to explain the weak signal for the quartet topology derived from morphological characters. Although outgroup choice affects results, our study could narrow possible trees to derivatives of a single quartet topology. For highly disputed relationships, we propose to apply a series of tests (AU and quartet mapping), since results of such tests allow to narrow down possible relationships and to rule out confounding signal.

BackgroundEP300 is a conserved protein in vertebrates, which serves as a key mediator of cellular homeostasis. Mutations and dysregulation of EP300 give rise to severe human developmental disorders and malignancy.Danio reriois a promising model organism to study EP300 related diseases and drugs; however, the effect of EP300 duplicates derived from teleost-specific whole genome duplication should not just be neglected.ResultsIn this study, we obtained EP300 protein sequences of representative teleosts, mammals and sauropsids, with which we inferred a highly supported maximum likelihood tree. We observed that Ep300 duplicates (Ep300a and Ep300b) were widely retained in teleosts and universally expressed in a variety of tissues. Consensus sequences of Ep300a and Ep300b had exactly the same distribution of conserved domains, suggesting that their functions should still be largely overlapped. We analyzed the molecular evolution of Ep300 duplicates in teleosts, using branch-site models, clade models and site models. The results showed that both duplicates were subject to strong positive selection; however, for an extant species, generally at most one copy was under positive selection. At the clade level, there were evident positive correlations between evolutionary rates, the number of positively selected sites and gene expression levels. In Ostariophysi, Ep300a were under stronger positive selection than Ep300b; in Neoteleostei, another species-rich teleost clade, the contrary was the case. We also modeled 3D structures of zf-TAZ domain and its flanking regions of Ep300a and Ep300b ofD. rerioandOryzias latipesand found that in either species the faster evolving copy had more short helixes.ConclusionsCollectively, the two copies of Ep300 have undoubtedly experienced directional divergence in main teleost clades. The divergence of EP300 between teleosts and mammals should be greater than the divergence between different teleost clades. Further studies are needed to clarify to what extent the EP300 involved regulatory network has diverged between teleosts and mammals, which would also help explain the huge success of teleosts.

The Cyanidiophyceae is an early-diverged red algal class that thrives in extreme conditions around acidic hot springs. Although this lineage has been highlighted as a model for understanding the biology of extremophilic eukaryotes, little is known about the molecular evolution of their mitochondrial genomes (mitogenomes).To fill this knowledge gap, we sequenced five mitogenomes from representative clades of Cyanidiophyceae and identified two major groups, here referred to as Galdieria-type (G-type) and Cyanidium-type (C-type). G-type mitogenomes exhibit the following three features: (i) reduction in genome size and gene inventory, (ii) evolution of unique protein properties including charge, hydropathy, stability, amino acid composition, and protein size, and (iii) distinctive GC-content and skewness of nucleotides. Based on GC-skew-associated characteristics, we postulate that unidirectional DNA replication may have resulted in the rapid evolution of G-type mitogenomes.The high divergence of G-type mitogenomes was likely driven by natural selection in the multiple extreme environments that Galdieria species inhabit combined with their highly flexible heterotrophic metabolism. We speculate that the interplay between mitogenome divergence and adaptation may help explain the dominance of Galdieria species in diverse extreme habitats.

BackgroundThe aragonite shelled, planktonic gastropod family Atlantidae (shelled heteropods) is likely to be one of the first groups to be impacted by imminent ocean changes, including ocean warming and ocean acidification. With a fossil record spanning at least 100 Ma, atlantids have experienced and survived global-scale ocean changes and extinction events in the past. However, the diversification patterns and tempo of evolution in this family are largely unknown.ResultsBased on a concatenated maximum likelihood phylogeny of three genes (cytochromecoxidase subunit 1 mitochondrial DNA, 28S and 18S ribosomal rRNA) we show that the three extant genera of the family Atlantidae,Atlanta, ProtatlantaandOxygyrus,form monophyletic groups. The genusAtlantais split into two groups, one exhibiting smaller, well ornamented shells, and the other having larger, less ornamented shells. The fossil record, in combination with a fossil-calibrated phylogeny, suggests that large scale atlantid extinction was accompanied by considerable and rapid diversification over the last 25 Ma, potentially driven by vicariance events.ConclusionsNow confronted with a rapidly changing modern ocean, the ability of atlantids to survive past global change crises gives some optimism that they may be able to persist through the Anthropocene.

Background The family Labridae made up of 519 species in the world. The functional evolution of the feeding-related jaws leaded to differentiation of species, and the pharyngeal jaw apparatus evolved independently, but evolutionary mechanism still remain unaddressed in wrasses. Mitogenomes data can be used to infer genetic diversification and investigate evolutionary history of wrasses, whereas only eight complete mitogenomes in this family have been sequenced to date. Here, we sequenced the complete mitogenomes of Iniistius trivittatus to investigate genetic differentiation among wrasse species. Results We sequenced the complete mitogenomes of I. trivittatus using a novel PCR strategy. The I. trivittatus mitogenomes is 16,820 bp in length and includes 13 protein -coding genes, 2 ribosomal RNA (rRNA) genes, 22 transfer RNA (tRNA) genes, and a control region. Compared to eight known mitochondrial genome, 2 additional noncoding regions (lengths of 121 and 107 bp), or so-called inserts, are found in the intergenic regions 12S rRNA - tRNAVal - 16S rRNA. The presumed origin of the two rare inserts is from tRNA- related retrotransposons. Compared with cytochrome b gene, the two insert sequences are highly conserved at the intraspecies level, but they showed significant variation and low similarity (I. trivittatus by checking the phylogenetic trees based on the complete mitogenomes of Labrida species. This finding provides evidence that in the mitogenomes, retrotransposon inserts result in intraspecific homoplasmy and interspecific heteroplasmy by natural selection and adaptation to various environments. Conclusions This study found additional mitogenome inserts limited in wrasse species. The rRNA genes with inserts might have experienced a selective pressure for adaptation to feeding modes. Such knowledge can enable a better understanding of molecular mechanism underlying morphological evolution in wrasses.

Background Understanding the structure and variability of adaptive loci such as the major histocompatibility complex (MHC) genes is a primary research goal for evolutionary and conservation genetics. Typically, classical MHC genes show high polymorphism and are under strong balancing selection, as their products trigger the adaptive immune response in vertebrates. Here, we assess the allelic diversity and patterns of selection for MHC class I and class II loci in a threatened shorebird with highly flexible mating and parental care behaviour, the Snowy Plover (Charadrius nivosus) across its broad geographic range. Results We determined the allelic and nucleotide diversity for MHC class I and class II genes using samples of 250 individuals from eight breeding population of Snowy Plovers. We found 40 alleles at MHC class I and six alleles at MHC class II, with individuals carrying two to seven different alleles (mean 3.70) at MHC class I and up to two alleles (mean 1.45) at MHC class II. Diversity was higher in the peptide-binding region, which suggests balancing selection. The MHC class I locus showed stronger signatures of both positive and negative selection than the MHC class II locus. Most alleles were present in more than one population. If present, private alleles generally occurred at very low frequencies in each population, except for the private alleles of MHC class I in one island population (Puerto Rico, lineage tenuirostris). Conclusion Snowy Plovers exhibited an intermediate level of diversity at the MHC, similar to that reported in other Charadriiformes. The differences found in the patterns of selection between the class I and II loci are consistent with the hypothesis that different mechanisms shape the sequence evolution of MHC class I and class II genes. The rarity of private alleles across populations is consistent with high natal and breeding dispersal and the low genetic structure previously observed at neutral genetic markers in this species.

Background Convergent and parallel evolution provide unique insights into the mechanisms of natural selection. Some of the most striking convergent and parallel (collectively recurrent) amino acid substitutions in proteins are adaptive, but there are also many that are selectively neutral. Accordingly, genome-wide assessment has shown that recurrent sequence evolution in orthologs is chiefly explained by nearly neutral evolution. For paralogs, more frequent functional change is expected because additional copies are generally not retained if they do not acquire their own niche. Yet, it is unknown to what extent recurrent sequence differentiation is discernible after independent gene duplications in different eukaryotic taxa. Results We develop a framework that detects patterns of recurrent sequence evolution in duplicated genes. This is used to analyze the genomes of 90 diverse eukaryotes. We find a remarkable number of families with a potentially predictable functional differentiation following gene duplication. In some protein families, more than ten independent duplications show a similar sequence-level differentiation between paralogs. Based on further analysis, the sequence divergence is found to be generally asymmetric. Moreover, about 6% of the recurrent sequence evolution between paralog pairs can be attributed to recurrent differentiation of subcellular localization. Finally, we reveal the specific recurrent patterns for the gene families Hint1/Hint2, Sco1/Sco2 and vma11/vma3. Conclusions The presented methodology provides a means to study the biochemical underpinning of functional differentiation between paralogs. For instance, two abundantly repeated substitutions are identified between independently derived Sco1 and Sco2 paralogs. Such identified substitutions allow direct experimental testing of the biological role of these residues for the repeated functional differentiation. We also uncover a diverse set of families with recurrent sequence evolution and reveal trends in the functional and evolutionary trajectories of this hitherto understudied phenomenon.

Background The distribution of genetic diversity and the underlying processes are important for conservation planning but are unknown for most species and have not been well studied in many regions. In East Asia, the Sichuan Basin and surrounding mountains constitute an understudied region that exhibits a “ring” of high species richness overlapping the eastern edge of the global biodiversity hotspot Mountains of Southwest China. We examine the distributional history and genetic diversification of the Emei mustache toad Leptobrachium boringii, a typical “ring” element characterized by disjunct ranges in the mountains, by integrating time-calibrated gene tree, genetic variability, individual-level clustering, inference of population splitting and mixing from allele frequencies, and paleoclimatic suitability modeling. Results The results reveal extensive range dynamics, including secondary contact after long-term isolation via westward dispersal accompanied by variability loss. They allow the proposal of a model that combines recurrent contractions caused by Quaternary climatic changes and some failed expansions under suitable conditions for explaining the shared disjunct distribution pattern. Providing exceptional low-elevation habitats in the hotspot area, the eastern edge harbors both long-term refugial and young immigrant populations. This finding and a synthesis of evidence from other taxa demonstrate that a certain contributor to biodiversity, one that preserves and receives low-elevation elements of the east in this case, can be significant for only a particular part of a hotspot. By clarifying the low variability of these refugial populations, we show that discordant mitochondrial estimates of diversity can be obtained for populations that experienced admixture, which would have unlikely left proportional immigrant alleles for each locus. Conclusions Dispersal after long-term isolation can explain much of the spatial distribution of genetic diversity in this species, while secondary contact and long-term persistence do not guarantee a large variation. The model for the formation of disjunct ranges may apply to many other taxa isolated in the mountains surrounding the Sichuan Basin. Furthermore, this study provides insights into the heterogeneous nature of hotspots and discordant variability obtained from genome-wide and mitochondrial data.

Background Many animals rely heavily on olfaction to navigate their environment. Among rodents, olfaction is crucial for a wide range of social behaviors. The vomeronasal olfactory system in particular plays an important role in mediating social communication, including the detection of pheromones and recognition signals. In this study we examine patterns of vomeronasal type-1 receptor (V1R) evolution in the house mouse and related species within the genus Mus. We report the extent of gene repertoire turnover and conservation among species and clades, as well as the prevalence of positive selection on gene sequences across the V1R tree. By exploring the evolution of these receptors, we provide insight into the functional roles of receptor subtypes as well as the dynamics of gene family evolution. Results We generated transcriptomes from the vomeronasal organs of 5 Mus species, and produced high quality V1R repertoires for each species. We find that V1R clades in the house mouse and relatives exhibit distinct evolutionary trajectories. We identify putative species-specific gene expansions, including a large clade D expansion in the house mouse. While gene gains are abundant, we detect very few gene losses. We describe a novel V1R clade and highlight candidate receptors for future study. We find evidence for distinct evolutionary processes across different clades, from largescale turnover to highly conserved repertoires. Patterns of positive selection are similarly variable, as some clades exhibit abundant positive selection while others display high gene sequence conservation. Based on clade-level evolutionary patterns, we identify receptor families that are strong candidates for detecting social signals and predator cues. Our results reveal clades with receptors detecting female reproductive status are among the most conserved across species, suggesting an important role in V1R chemosensation. Conclusion Analysis of clade-level evolution is critical for understanding species’ chemosensory adaptations. This study provides clear evidence that V1R clades are characterized by distinct evolutionary trajectories. As receptor evolution is shaped by ligand identity, these results provide a framework for examining the functional roles of receptors.

Background We have described the diversity of complete mtDNA sequences from ‘relic’ groups of the Russian Far East, primarily the Nivkhi (who speak a language isolate with no clear relatedness to any others) and Oroki of Sakhalin, as well as the sedentary Koryak from Kamchatka and the Udegey of Primorye. Previous studies have shown that most of their traditional territory was dramatically reshaped by the expansion of Tungusic-speaking groups. Results Overall, 285 complete mitochondrial sequences were selected for phylogenetic analyses of published, revised and new mitogenomes. To highlight the likely role of Neolithic expansions in shaping the phylogeographical landscape of the Russian Far East, we focus on the major East Eurasian maternal lineages (Y1a, G1b, D4m2, D4e5, M7a2, and N9b) that are restricted to the coastal area. To obtain more insight into autochthonous populations, we removed from the phylogeographic analysis the G2a, G3a2, M8a1, M9a1, and C4b1 lineages, also found within our samples, likely resulting from admixture between the expanding proto-Tungus and the indigenous Paleoasiatic groups with whom they assimilated. Phylogenetic analysis reveals that unlike the relatively diverse lineage spectrum observed in the Amur estuary and northwestern Sakhalin, the present-day subpopulation on the northeastern coast of the island is relatively homogenous: a sole Y1a sublineage, conspicuous for its nodal mutation at m.16189 T > C!, includes different haplotypes. Sharing of the Y1a-m.16189 T > C! sublineages and haplotypes among the Nivkhi, Ulchi and sedentary Koryak is also evident. Aside from Y1a, the entire tree approach expands our understanding of the evolutionary history of haplogroups G1, D4m, N9b, and M7a2. Specifically, we identified the novel haplogroup N9b1 in Primorye, which implies a link between a component of the Udegey ancestry and the Hokkaido Jomon. Conclusions Through a comprehensive dataset of mitochondrial genomes retained in autochthonous populations along the coast between Primorye and the Bering Strait, we considerably extended the sequence diversity of these populations to provide new features based on the number and timing of founding lineages. We emphasize the value of integrating genealogical information with genetic data for reconstructing the population history of indigenous groups dramatically impacted by twentieth century resettlement and social upheavals.

Background Although phylogenomic analyses are increasingly used to reveal evolutionary relationships among ciliates, relatively few nuclear protein-coding gene markers have been tested for their suitability as candidates for inferring phylogenies within this group. In this study, we investigate the utility of the heat-shock protein 90 gene (Hsp90) as a marker for inferring phylogenetic relationships among hypotrich ciliates. Results A total of 87 novel Hsp90 gene sequences of 10 hypotrich species were generated. Of these, 85 were distinct sequences. Phylogenetic analyses based on these data showed that: (1) the Hsp90 gene amino acid trees are comparable to the small subunit rDNA tree for recovering phylogenetic relationships at the rank of class, but lack sufficient phylogenetic signal for inferring evolutionary relationships at the genus level; (2) Hsp90 gene paralogs are recent and therefore unlikely to pose a significant problem for recovering hypotrich clades; (3) definitions of some hypotrich orders and families need to be revised as their monophylies are not supported by various gene markers; (4) The order Sporadotrichida is paraphyletic, but the monophyly of the “core” Urostylida is supported; (5) both the subfamily Oxytrichinae and the genus Urosoma seem to be non-monophyletic, but monophyly of Urosoma is not rejected by AU tests. Conclusions Our results for the first time demonstrate that the Hsp90 gene is comparable to SSU rDNA for recovering phylogenetic relationships at the rank of class, and its paralogs are unlikely to pose a significant problem for recovering hypotrich clades. This study shows the value of careful gene marker selection for phylogenomic analyses of ciliates.

BackgroundThe latest advancements in DNA sequencing technologies have facilitated the resolution of the phylogeny of insects, yet parts of the tree of Holometabola remain unresolved. The phylogeny of Neuropterida has been extensively studied, but no strong consensus exists concerning the phylogenetic relationships within the order Neuroptera. Here, we assembled a novel transcriptomic dataset to address previously unresolved issues in the phylogeny of Neuropterida and to infer divergence times within the group. We tested the robustness of our phylogenetic estimates by comparing summary coalescent and concatenation-based phylogenetic approaches and by employing different quartet-based measures of phylogenomic incongruence, combined with data permutations.ResultsOur results suggest that the order Raphidioptera is sister to Neuroptera + Megaloptera. Coniopterygidae is inferred as sister to all remaining neuropteran families suggesting that larval cryptonephry could be a ground plan feature of Neuroptera. A clade that includes Nevrorthidae, Osmylidae, and Sisyridae (i.e. Osmyloidea) is inferred as sister to all other Neuroptera except Coniopterygidae, and Dilaridae is placed as sister to all remaining neuropteran families. Ithonidae is inferred as the sister group of monophyletic Myrmeleontiformia. The phylogenetic affinities of Chrysopidae and Hemerobiidae were dependent on the data type analyzed, and quartet-based analyses showed only weak support for the placement of Hemerobiidae as sister to Ithonidae + Myrmeleontiformia. Our molecular dating analyses suggest that most families of Neuropterida started to diversify in the Jurassic and our ancestral character state reconstructions suggest a primarily terrestrial environment of the larvae of Neuropterida and Neuroptera.ConclusionOur extensive phylogenomic analyses consolidate several key aspects in the backbone phylogeny of Neuropterida, such as the basal placement of Coniopterygidae within Neuroptera and the monophyly of Osmyloidea. Furthermore, they provide new insights into the timing of diversification of Neuropterida. Despite the vast amount of analyzed molecular data, we found that certain nodes in the tree of Neuroptera are not robustly resolved. Therefore, we emphasize the importance of integrating the results of morphological analyses with those of sequence-based phylogenomics. We also suggest that comparative analyses of genomic meta-characters should be incorporated into future phylogenomic studies of Neuropterida.

[Background]: Cysteine-RIch Secretory Proteins (CRISP) are expressed in the reproductive tract of mammalian males and are involved in fertilization and related processes. Due to their important role in sperm performance and sperm-egg interaction, these genes are likely to be exposed to strong selective pressures, including postcopulatory sexual selection and/or male-female coevolution. We here perform a comparative evolutionary analysis of Crisp genes in mammals. Currently, the nomenclature of CRISP genes is confusing, as a consequence of discrepancies between assignments of orthologs, particularly due to numbering of CRISP genes. This may generate problems when performing comparative evolutionary analyses of mammalian clades and species. To avoid such problems, we first carried out a study of possible orthologous relationships and putative origins of the known CRISP gene sequences. Furthermore, and with the aim to facilitate analyses, we here propose a different nomenclature for CRISP genes (EVAC1–4, “EVolutionarily-analyzed CRISP”) to be used in an evolutionary context., [Results]: We found differing selective pressures among Crisp genes. CRISP1/4 (EVAC1) and CRISP2 (EVAC2) orthologs are found across eutherian mammals and seem to be conserved in general, but show signs of positive selection in primate CRISP1/4 (EVAC1). Rodent Crisp1 (Evac3a) seems to evolve under a comparatively more relaxed constraint with positive selection on codon sites. Finally, murine Crisp3 (Evac4), which appears to be specific to the genus Mus, shows signs of possible positive selection. We further provide evidence for sexual selection on the sequence of one of these genes (Crisp1/4) that, unlike others, is thought to be exclusively expressed in male reproductive tissues., [Conclusions]: We found differing selective pressures among CRISP genes and sexual selection as a contributing factor in CRISP1/4 gene sequence evolution. Our evolutionary analysis of this unique set of genes contributes to a better understanding of Crisp function in particular and the influence of sexual selection on reproductive mechanisms in general., This work was supported by the Spanish Ministry of Economy, Industry and Competitiveness (grants CGL2011–26341 and CGL2016–80577-P)., We acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI).

Background Tree squirrels (Sciuridae, Sciurini), in particular the highly diverse Neotropical lineages, are amongst the most rapidly diversifying branches of the mammal tree of life but also some of the least known. Negligence of this group by systematists is likely a product of the difficulties in assessing morphological informative traits and of the scarcity or unavailability of fresh tissue samples for DNA sequencing. The highly discrepant taxonomic arrangements are a consequence of the lack of phylogenies and the exclusive phenotypic-based classifications, which can be misleading in a group with conservative morphology. Here we used high-throughput sequencing and an unprecedented sampling of museum specimens to provide the first comprehensive phylogeny of tree squirrels, with a special emphasis on Neotropical taxa. Results We obtained complete or partial mitochondrial genomes from 232 historical and modern samples, representing 40 of the 43 currently recognized species of Sciurini. Our phylogenetic analyses—performed with datasets differing on levels of missing data and taxa under distinct analytical methods—strongly support the monophyly of Sciurini and consistently recovered 12 major clades within the tribe. We found evidence that the diversity of Neotropical tree squirrels is underestimated, with at least six lineages that represent taxa to be named or revalidated. Ancestral state reconstructions of number of upper premolars and number of mammae indicated that alternative conditions of both characters must have evolved multiple times throughout the evolutionary history of tree squirrels. Conclusions Complete mitogenomes were obtained from museum specimens as old as 120 years, reinforcing the potential of historical samples for phylogenetic inferences of elusive lineages of the tree of life. None of the taxonomic arrangements ever proposed for tree squirrels fully corresponded to our phylogenetic reconstruction, with only a few of the currently recognized genera recovered as monophyletic. By investigating the evolution of two morphological traits widely employed in the taxonomy of the group, we revealed that their homoplastic nature can help explain the incongruence between phylogenetic results and the classification schemes presented so far. Based on our phylogenetic results we suggest a tentative supraspecific taxonomic arrangement for Sciurini, employing 13 generic names used in previous taxonomic classifications.

Background Parasites may mediate the success of biological invasions through their effect on host fitness and thus, on host population growth and stability. However, a release from the pressure of parasites is strongly related to the genetic differentiation of the host. In invasive host populations, the number of available genetic variants, allowing them to ‘fight’ the infection, are likely to be influenced by founder events and genetic drift. The level standing genetic variation of invasive populations may be crucial in successfully adapting to new environments and resisting diseases. We studied invasive populations of raccoon that experienced a random reduction in genetic diversity during the establishment and evaluated the relationship between host immune genetic diversity and intestinal parasites infection. Results We distinguished two different genetic clusters that are characterized by different sets of functionally relevant MHC-DRB alleles. Both clusters were characterized by considerably different allele-parasite associations and different levels of parasite infection. The specific resistance MHC-DRB alleles explained the lower prevalence of Digenea parasites. An increased infection intensity was related to the presence of two MHC-DRB alleles. One of these alleles significantly decreased in frequency over time, causing a decrease of Digenea abundance in raccoons in consecutive years. Conclusions Our findings suggest that intestinal parasites can exert selective pressure on an invasive host with lowered levels of immune genetic diversity and contribute to promoting local adaptation over time. The random genetic drift that created the two different genetic clusters in the invasive raccoon range imposed completely different MHC-parasite associations, strongly associated with the infection status of populations. Our findings underline the role of standing genetic variation in shaping host-parasite relationships and provide empirical support that functional genetic variation may be, at least partly, responsible for differences in the success of invasive populations.

Background Sympatric sister species provide an opportunity to investigate the genetic mechanisms and evolutionary forces that maintain species boundaries. The persistence of morphologically and genetically distinct populations in sympatry can only occur if some degree of reproductive isolation exists. A pair of sympatric sister species of Primulina (P. depressa and P. danxiaensis) was used to explore the genetic architecture of hybrid male sterility. Results We mapped one major- and seven minor-effect quantitative trait loci (QTLs) that underlie pollen fertility rate (PFR). These loci jointly explained 55.4% of the phenotypic variation in the F2 population. A Bateson–Dobzhansky–Muller (BDM) model involving three loci was observed in this system. We found genotypic correlations between hybrid male sterility and flower morphology, consistent with the weak but significant phenotypic correlations between PFR and floral traits. Conclusions Hybrid male sterility in Primulina is controlled by a polygenic genetic basis with a complex pattern. The genetic incompatibility involves a three-locus BDM model. Hybrid male sterility is genetically correlated with floral morphology and divergence hitchhiking may occur between them.

Background Many organisms are responding to climate change with dramatic range shifts, involving plastic and genetic changes to cope with novel climate regimes found at higher latitudes. Using experimental lineages of the seed beetle Callosobruchus maculatus, we simulated the initial phase of colonisation to progressively cooler and/or more variable conditions, to investigate how adaptation and phenotypic plasticity contribute to shifts in thermal tolerance during colonisation of novel climates. Results We show that heat and cold tolerance rapidly evolve during the initial stages of adaptation to progressively cooler and more variable climates. The evolved shift in cold tolerance is, however, associated with maladaptive plasticity under the novel conditions, resulting in a pattern of countergradient variation between the ancestral and novel, fluctuating thermal environment. In contrast, lineages exposed to progressively cooler, but constant, temperatures over several generations expressed only beneficial plasticity in cold tolerances and no evolved response. Conclusions We propose that thermal adaptation during a range expansion to novel, more variable climates found at high latitudes and elevations may typically involve genetic compensation arising from maladaptive plasticity in the initial stages of adaptation, and that this form of (countergradient) thermal adaptation may represent an opportunity for more rapid and labile evolutionary change in thermal tolerances than via classic genetic assimilation models for thermal tolerance evolution (i.e., selection on existing reaction norms). Moreover, countergradient variation in thermal tolerances may typically mask cryptic genetic variability for these traits, resulting in apparent evolutionary stasis in thermal traits.

BackgroundRecent human transcriptomic analyses revealed a very large number of testis-enriched genes, many of which are involved in spermatogenesis. This comprehensive transcriptomic data lead us to the question whether positive selection was a decisive force influencing the evolution and variability of testis-enriched genes in humans. We used two methodological approaches to detect different levels of positive selection, namely episodic positive diversifying selection (i.e., past selection) in the human lineage within primate phylogeny, potentially driven by sperm competition, and recent positive directional selection in contemporary human populations, which would indicate adaptation to different environments.ResultsIn the human lineage (after correction for multiple testing) we found that only the geneTULP2, for which no functional data are yet available, is subject to episodic positive diversifying selection. Using less stringent statistical criteria (uncorrectedp-values), also the geneSPATA16, which has a pivotal role in male fertility and for which episodes of adaptive evolution have been suggested, also displays a putative signal of diversifying selection in the human branch. At the same time, we found evidence for recent positive directional selection acting on several human testis-enriched genes (MORC1,SLC9B1,ROPN1L,DMRT1,PLCZ1,RNF17,FAM71DandWBP2NL) that play important roles in human spermatogenesis and fertilization. Most of these genes are population-specifically under positive selection.ConclusionEpisodic diversifying selection, possibly driven by sperm competition, was not an important force driving the evolution of testis-enriched genes in the human lineage. Population-specific, recent positive directional selection suggests an adaptation of male reproductive genes to different environmental conditions. Positive selection acts on eQTLS and sQTLs, indicating selective effects on important gene regulatory functions. In particular, the transcriptional diversity regulated by sQTLs in testis-enriched genes may be important for spermatocytes to respond to environmental and physiological stress.

Background Regions within the nuclear ribosomal operon are a major tool for inferring evolutionary relationships and investigating diversity in fungi. In spite of the prevalent use of ribosomal markers in fungal research, central features of nuclear ribosomal DNA (nrDNA) evolution are poorly characterized for fungi in general, including lichenized fungi. The internal transcribed spacer (ITS) region of the nrDNA has been adopted as the primary DNA barcode identification marker for fungi. However, little is known about intragenomic variation in the nrDNA in symbiotic fungi. In order to better understand evolution of nrDNA and the utility of the ITS region for barcode identification of lichen-forming fungal species, we generated nearly complete nuclear ribosomal operon sequences from nine species in the Rhizoplaca melanophthalma species complex using short reads from high-throughput sequencing. Results We estimated copy numbers for the nrDNA operon, ranging from nine to 48 copies for members of this complex, and found low levels of intragenomic variation in the standard barcode region (ITS). Monophyly of currently described species in this complex was supported in phylogenetic inferences based on the ITS, 28S, intergenic spacer region, and some intronic regions, independently; however, a phylogenetic inference based on the 18S provided much lower resolution. Phylogenetic analysis of concatenated ITS and intergenic spacer sequence data generated from 496 specimens collected worldwide revealed previously unrecognized lineages in the nrDNA phylogeny. Conclusions The results from our study support the general assumption that the ITS region of the nrDNA is an effective barcoding marker for fungi. For the R. melanophthalma group, the limited amount of potential intragenomic variability in the ITS region did not correspond to fixed diagnostic nucleotide position characters separating taxa within this species complex. Previously unrecognized lineages inferred from ITS sequence data may represent undescribed species-level lineages or reflect uncharacterized aspects of nrDNA evolution in the R. melanophthalma species complex.

BackgroundDrosophila subobscuraexhibits a rich inversion polymorphism, with some adaptive inversions showing repeatable spatiotemporal patterns in frequencies related to temperature. Previous studies reported increased basal HSP70 protein levels in homokaryotypic strains for a warm-climate arrangement compared to a cold-climate one. These findings do not match the similarhsp70genomic organization between arrangements, where gene expression levels are expected to be similar. In order to test this hypothesis and understand the molecular basis forhsp70expression, we compared basalhsp70mRNA levels in males and females, and analysed the 5′ and 3′ regulatory regions ofhsp70genes in warm- and cold-climate isochromosomal O3 + 4 + 7and OSTlines ofD. subobscura.ResultsWe observed comparable mRNA levels between the two arrangements and a sex-biasedhsp70gene expression. The number of heat-shock elements (HSEs) and GAGA sites on the promoters were identical amongst the OSTand O3 + 4 + 7lines analysed. This is also true for 3′ AU-rich elements where most A and B copies ofhsp70have, respectively, two and one element in both arrangements. Beyond the regulatory elements, the only notable difference between both arrangements is the presence in 3′ UTR of a 14 bp additional fragment after the stop codon in thehsp70A copy in five O3 + 4 + 7lines, which was not found in any of the six OSTlines.ConclusionsThe equivalenthsp70mRNA amounts in OSTand O3 + 4 + 7arrangements provide the first evidence of a parallelism between gene expression and genetic organization inD. subobscuralines having these arrangements. This is reinforced by the lack of important differential features in the number and structure of regulatory elements between both arrangements, despite the genetic differentiation observed when the complete 5′ and 3′ regulatory regions were considered. Therefore, the basal levels ofhsp70mRNA cannot account, in principle, for the adaptive variation of the two arrangements studied. Consequently, further studies are necessary to understand the intricate molecular mechanisms ofhsp70gene regulation inD. subobscura.

Background Gymnosperms represent five of the six lineages of seed plants. However, most sequenced plant mitochondrial genomes (mitogenomes) have been generated for angiosperms, whereas mitogenomic sequences have been generated for only six gymnosperms. In particular, complete mitogenomes are available for all major seed plant lineages except Conifer II (non-Pinaceae conifers or Cupressophyta), an important lineage including six families, which impedes a comprehensive understanding of the mitogenomic diversity and evolution in gymnosperms. Results Here, we report the complete mitogenome of Taxus cuspidata in Conifer II. In comparison with previously released gymnosperm mitogenomes, we found that the mitogenomes of Taxus and Welwitschia have lost many genes individually, whereas all genes were identified in the mitogenomes of Cycas, Ginkgo and Pinaceae. Multiple tRNA genes and introns also have been lost in some lineages of gymnosperms, similar to the pattern observed in angiosperms. In general, gene clusters could be less conserved in gymnosperms than in angiosperms. Moreover, fewer RNA editing sites were identified in the Taxus and Welwitschia mitogenomes than in other mitogenomes, which could be correlated with fewer introns and frequent gene losses in these two species. Conclusions We have sequenced the Taxus cuspidata mitogenome, and compared it with mitogenomes from the other four gymnosperm lineages. The results revealed the diversity in size, structure, gene and intron contents, foreign sequences, and mutation rates of gymnosperm mitogenomes, which are different from angiosperm mitogenomes.

Background Early Cambrian Lagerstätten from China have greatly enriched our perspective on the early evolution of animals, particularly arthropods. However, recent studies have shown that many of these early fossil arthropods were more derived than previously thought, casting uncertainty on the ancestral euarthropod body plan. In addition, evidence from fossilized neural tissues conflicts with external morphology, in particular regarding the homology of the frontalmost appendage. Results Here we redescribe the multisegmented megacheirans Fortiforceps and Jianfengia and describe Sklerolibyon maomima gen. et sp. nov., which we place in Jianfengiidae, fam. nov. (in Megacheira, emended). We find that jianfengiids show high morphological diversity among megacheirans, both in trunk ornamentation and head anatomy, which encompasses from 2 to 4 post-frontal appendage pairs. These taxa are also characterized by elongate podomeres likely forming seven-segmented endopods, which were misinterpreted in their original descriptions. Plesiomorphic traits also clarify their connection with more ancestral taxa. The structure and position of the “great appendages” relative to likely sensory antero-medial protrusions, as well as the presence of optic peduncles and sclerites, point to an overall homology with the anterior head of radiodontans. This is confirmed by our Bayesian phylogeny, which places jianfengiids as the basalmost euarthropods, paraphyletic with other megacheirans, and in contiguity with isoxyids and radiodontans. Conclusions Sklerolibyon and other jianfengiids expand the disparity of megacheirans and suggest that the common euarthropod ancestor possessed a remarkable phenotypic variability associated with the externalized cephalon, as well as endopods that were already heptopodomerous, which differs from previous hypotheses and observations. These animals also demonstrate that the frontalmost pair of arthrodized appendage is homologous between radiodontans and megacheirans, refuting the claim that the radiodontan frontal appendages evolved into the euarthropod labrum, and questioning its protocerebral identity. This evidence based on external anatomy now constitutes a solid benchmark upon which we should address issues of homology, with the help of carefully examined palaeoneurological data.

BackgroundUnderstanding the genomic basis of phenotypic diversity can be greatly facilitated by examining adaptive radiations with hypervariable traits. In this study, we focus on a rapidly diverged species group of mormyrid electric fish in the genusParamormyrops, which are characterized by extensive phenotypic variation in electric organ discharges (EODs). The main components of EOD diversity are waveform duration, complexity and polarity. Using an RNA-sequencing based approach, we sought to identify gene expression correlates for each of these EOD waveform features by comparing 11 specimens ofParamormyropsthat exhibit variation in these features.ResultsPatterns of gene expression amongParamormyropsare highly correlated, and 3274 genes (16%) were differentially expressed. Using our most restrictive criteria, we detected 145–183 differentially expressed genes correlated with each EOD feature, with little overlap between them. The predicted functions of several of these genes are related to extracellular matrix, cation homeostasis, lipid metabolism, and cytoskeletal and sarcomeric proteins. These genes are of significant interest given the known morphological differences between electric organs that underlie differences in the EOD waveform features studied.ConclusionsIn this study, we identified plausible candidate genes that may contribute to phenotypic differences in EOD waveforms among a rapidly diverged group of mormyrid electric fish. These genes may be important targets of selection in the evolution of species-specific differences in mate-recognition signals.

Background Despite decades of research, the horse domestication scenario in East Asia remains poorly understood. Results The study identified 16 haplogroups with fine-scale phylogenetic resolution using mitochondrial genomes of 317 horse samples. The time to the most recent common ancestor of the 16 haplogroups ranges from [0.8–3.1] thousand years ago (KYA) to [7.9–27.1] KYA. With combined analyses of the mitochondrial control region for 35 extant Przewalski’s horses, 3544 modern and 203 ancient horses across the world, researchers provide evidence for that East Asian prevalent haplogroups Q and R were indigenously domesticated or they were involved in numerous distinct genetic components from wild horses in the southern part of East Asia. These events of haplotypes Q and R occurred during 4.7 to 16.3 KYA and 2.1 to 11.5 KYA, respectively. The diffusion of preponderant European haplogroups L from west to East Asia is consistent with the external gene input. Furthermore, genetic differences were detected between northern East Asia and southern East Asia cohorts by Principal Component Analysis, Analysis of Molecular Variance test, the χ2 test and phylogeographic analyses. Conclusions All results suggest a complex picture of horse domestication, as well as geographic pattern in East Asia. Both local origin and external input occurred in East Asia horse populations. And besides, there are at least two different domestication or hybridization centers in East Asia.

BackgroundSecondary contact between closely related lineages can result in a variety of outcomes, including hybridization, depending upon the strength of reproductive barriers. By examining the extent to which different parts of the genome introgress, it is possible to infer the strength of selection and gain insight into the evolutionary trajectory of lineages. Following secondary contact approximately 8000 years ago in the Pacific Northwest, mule deer (Odocoileus hemionus hemionus) and black-tailed deer (O. h. columbianus) formed a hybrid swarm along the Cascade mountain range despite substantial differences in body size (up to two times) and habitat preference. In this study, we examined genetic population structure, extent of introgression, and selection pressures in freely interbreeding populations of mule deer and black-tailed deer using mitochondrial DNA sequences, 9 microsatellite loci, and 95 SNPs from protein-coding genes.ResultsWe observed bi-directional hybridization and classified approximately one third of the 172 individuals as hybrids, almost all of which were beyond the F1 generation. High genetic differentiation between black-tailed deer and mule deer at protein-coding genes suggests that there is positive divergent selection, though selection on these loci is relatively weak. Contrary to predictions, there was not greater selection on protein-coding genes thought to be associated with immune function and mate choice. Geographic cline analyses were consistent across genetic markers, suggesting long-term stability (over hundreds of generations), and indicated that the center of the hybrid swarm is 20-30 km to the east of the Cascades ridgeline, where there is a steep ecological transition from wet, forested habitat to dry, scrub habitat.ConclusionsOur data are consistent with a genetic boundary between mule deer and black-tailed deer that is porous but maintained by many loci under weak selection having a substantial cumulative effect. The absence of clear reproductive barriers and the consistent centering of geographic clines at a sharp ecotone suggests that ecology is a driver of hybrid swarm dynamics. Adaptive introgression in this study (and others) promotes gene flow and provides valuable insight into selection strength on specific genes and the evolutionary trajectory of hybridizing taxa.

BackgroundThe transition from land to sea by the ancestor of cetaceans approximately 50 million years ago was an incredible evolutionary event that led to a series of morphological, physiological, and behavioral adaptations. During this transition, bone microstructure evolved from the typical terrestrial form to the specialized structure found in modern cetaceans. While the bone microstructure of mammals has been documented before, investigations of its genetic basis lag behind. The increasing number of cetaceans with whole-genome sequences available may shed light on the mechanism underlying bone microstructure evolution as a result of land to water transitions.ResultsCetacean bone microstructure is consistent with their diverse ecological behaviors. Molecular evolution was assessed by correlating bone microstructure and gene substitution rates in terrestrial and aquatic species, and by detecting genes under positive selection along ancestral branches of cetaceans. We found that: 1) Genes involved in osteoclast function are under accelerated evolution in cetaceans, suggestive of important roles in bone remodeling during the adaptation to an aquatic environment; 2) Genes in the Wnt pathway critical for bone development and homeostasis show evidence of divergent evolution in cetaceans; 3) Several genes encoding bone collagens are under selective pressure in cetaceans.ConclusionsOur results suggest that evolutionary pressures have shaped the bone microstructure of cetaceans, to facilitate life in diverse aquatic environments.

Background Many living birds exhibit some nocturnal activity, but the genetic basis and evolutionary origins of their nocturnality remain unknown. Results Here, we used a molecular phyloecological approach to analyze the adaptive evolution of 33 phototransduction genes in diverse bird lineages. Our results suggest that functional enhancement of two night-vision genes, namely, GRK1 and SLC24A1, underlies the nocturnal adaption of living birds. Further analyses showed that the diel activity patterns of birds have remained relatively unchanged since their common ancestor, suggesting that the widespread nocturnal activity of many living birds may largely stem from their common ancestor rather than independent evolution. Despite this evolutionary conservation of diel activity patterns in birds, photoresponse recovery genes were found to be frequently subjected to positive selection in diverse bird lineages, suggesting that birds generally have evolved an increased capacity for motion detection. Moreover, we detected positive selection on both dim-light vision genes and bright-light vision genes in the class Aves, suggesting divergent evolution of the vision of birds from that of reptiles and that different bird lineages have evolved certain visual adaptions to their specific light conditions. Conclusions This study suggests that the widespread nocturnality of extant birds has a deep evolutionary origin tracing back to their common ancestor.

Background Triplophysa stoliczkae is the most widespread species in the genus Triplophysa and may have originated from morphological convergence. To understand the evolutionary history of T. stoliczkae, we employed a multilocus approach to investigate the phylogenetics and the morphological evolution of T. stoliczkae on the Qinghai-Tibetan Plateau. Results All phylogenetic analyses (two mitochondrial and five nuclear loci), a genealogical sorting index and species tree inferences suggested that T. stoliczkae consists of distinct lineages that were not closest relatives. The time estimation indicated that the divergence events between “T. stoliczkae” and other Triplophysa species occurred from approximately 0.10 to 4.51 Ma. The ancestral state analyses supported the independent evolution of T. stoliczkae morphology in distinct lineages. The morphometric analysis and convergence estimates demonstrated significant phenotypic convergence among “T. stoliczkae” lineages. Conclusions Triplophysa stoliczkae includes 4 different lineages with similar morphologies. The increasingly harsh environments that have occurred since the Pliocene have driven the occurrences of scrape-feeding fish in the genus Triplophysa. Morphological adaptations associated with scrape-feeding behavior resulted in convergences and the artificial lumping of four different species in the nominal taxon T. stoliczkae. A taxonomic revision for T. stoliczkae is needed. Electronic supplementary material The online version of this article (10.1186/s12862-019-1503-3) contains supplementary material, which is available to authorized users.

BackgroundThe silkmoths and their relatives constitute the ecologically and taxonomically diverse superfamily Bombycoidea, which includes some of the most charismatic species of Lepidoptera. Despite displaying some of the most spectacular forms and ecological traits among insects, relatively little attention has been given to understanding their evolution and the drivers of their diversity.ResultsTo begin to address this problem, we created a new Bombycoidea-specific Anchored Hybrid Enrichment (AHE) probe set and sampled up to 571 loci for 117 taxa across all major lineages of the Bombycoidea, producing a well-supported phylogeny. The tree was overall consistent with prior morphological and molecular studies, although some taxa (e.g.,ArotrosSchaus) were misplaced in the Bombycidae and here formally transferred to Apatelodidae. We identified important evolutionary patterns (e.g., morphology, biogeography, and differences in speciation and extinction), and our analysis of diversification rates highlights the stark increases that exist within the Sphingidae (hawkmoths) and Saturniidae (wild silkmoths).ConclusionsWe postulate that these rate shifts are due to the well-documented bat-moth “arms race” and differences in selective pressures from insectivorous bats. The study establishes a backbone for future evolutionary, comparative, and taxonomic studies, and presents a modified DNA extraction protocol that allows Lepidoptera specimens to be readily sequenced from pinned natural history collections, succeeding in samples up to 30 years old. Our research highlights the flexibility of AHE to generate genomic data from a wide range of museum specimens, both age and preservation method, and will allow researchers to tap into the wealth of biological data residing in natural history collections around the globe.

Background Marine invertebrates are abundant and diverse on the continental shelf in Antarctica, but little is known about their parasitic counterparts. Endoparasites are especially understudied because they often possess highly modified body plans that pose problems for their identification. Asterophila, a genus of endoparasitic gastropod in the family Eulimidae, forms cysts in the arms and central discs of asteroid sea stars. There are currently four known species in this genus, one of which has been described from the Antarctic Peninsula (A. perknasteri). This study employs molecular and morphological data to investigate the diversity of Asterophila in Antarctica and explore cophylogenetic patterns between host and parasite. Results A maximum-likelihood phylogeny of Asterophila and subsequent species-delimitation analysis uncovered nine well-supported putative species, eight of which are new to science. Most Asterophila species were found on a single host species, but four species were found on multiple hosts from one or two closely related genera, showing phylogenetic conservatism of host use. Both distance-based and event-based cophylogenetic analyses uncovered a strong signal of coevolution in this system, but most associations were explained by non-cospeciation events. Discussion The prevalence of duplication and host-switching events in Asterophila and its asteroid hosts suggests that synchronous evolution may be rare even in obligate endoparasitic systems. The apparent restricted distribution of Asterophila from around the Scotia Arc may be an artefact of concentrated sampling in the area and a low obvious prevalence of infection. Given the richness of parasites on a global scale, their role in promoting host diversification, and the threat of their loss through coextinction, future work should continue to investigate parasite diversity and coevolution in vulnerable ecosystems.

Background Diversity at the Major Histocompatibility Complex (MHC) is critical to health and fitness, such that MHC genotype may predict an individual’s quality or compatibility as a competitor, ally, or mate. Moreover, because MHC products can influence the components of bodily secretions, an individual’s body odors may signal its MHC composition and influence partner identification or mate choice. Here, we investigated MHC-based signaling and recipient sensitivity by testing for odor-gene covariance and behavioral discrimination of MHC diversity and pairwise dissimilarity in a strepsirrhine primate, the ring-tailed lemur (Lemur catta). Methods First, we coupled genotyping of the MHC class II gene, DRB, with gas chromatography-mass spectrometry of genital gland secretions to investigate if functional genetic diversity is signaled by the chemical diversity of lemur scent secretions. We also assessed if the chemical similarity between individuals correlated with their MHC-DRB similarity. Next, we assessed if lemurs discriminated this chemically encoded, genetic information in opposite-sex conspecifics. Results We found that both sexes signaled overall MHC-DRB diversity and pairwise MHC-DRB similarity via genital secretions, but in a sex- and season-dependent manner. Additionally, the sexes discriminated absolute and relative MHC-DRB diversity in the genital odors of opposite-sex conspecifics, suggesting that lemur genital odors function to advertise genetic quality. Conclusions In summary, genital odors of ring-tailed lemurs provide honest information about an individual’s absolute and relative MHC quality. Complementing evidence in humans and Old World monkeys, we suggest that reliance on scent signals to communicate MHC quality may be important across the primate lineage. Electronic supplementary material The online version of this article (10.1186/s12862-019-1486-0) contains supplementary material, which is available to authorized users.

Background With an ever-growing number of published genomes, many low levels of the Tree of Life now contain several species with enough molecular data to perform shallow-scale phylogenomic studies. Moving away from using just a few universal phylogenetic markers, we can now target thousands of other loci to decipher taxa relationships. Making the best possible selection of informative sequences regarding the taxa studied has emerged as a new issue. Here, we developed a general procedure to mine genomic data, looking for orthologous single-copy loci capable of deciphering phylogenetic relationships below the generic rank. To develop our strategy, we chose the genus Rosa, a rapid-evolving lineage of the Rosaceae family in which several species genomes have recently been sequenced. We also compared our loci to conventional plastid markers, commonly used for phylogenetic inference in this genus. Results We generated 1856 sequence tags in putative single-copy orthologous nuclear loci. Associated in silico primer pairs can potentially amplify fragments able to resolve a wide range of speciation events within the genus Rosa. Analysis of parsimony-informative site content showed the value of non-coding genomic regions to obtain variable sequences despite the fact that they may be more difficult to target in less related species. Dozens of nuclear loci outperform the conventional plastid phylogenetic markers in terms of phylogenetic informativeness, for both recent and ancient evolutionary divergences. However, conflicting phylogenetic signals were found between nuclear gene tree topologies and the species-tree topology, shedding light on the many patterns of hybridization and/or incomplete lineage sorting that occur in the genus Rosa. Conclusions With recently published genome sequence data, we developed a set of single-copy orthologous nuclear loci to resolve species-level phylogenomics in the genus Rosa. This genome-wide scale dataset contains hundreds of highly variable loci which phylogenetic interest was assessed in terms of phylogenetic informativeness and topological conflict. Our target identification procedure can easily be reproduced to identify new highly informative loci for other taxonomic groups and ranks. Electronic supplementary material The online version of this article (10.1186/s12862-019-1479-z) contains supplementary material, which is available to authorized users.

Background For the understanding of human nature, the evolutionary roots of human moral behaviour are a key precondition. Our question is as follows: Can the altruistic moral rule “Risk your life to save your family members, if you want them to save your life” be evolutionary stable? There are three research approaches to investigate this problem: kin selection, group selection and population genetics modelling. The present study is strictly based on the last approach. Results We consider monogamous and exogamous families, where at an autosomal locus, dominant-recessive alleles determine the phenotypes in a sexual population. Since all individuals’ survival rate is determined by their altruistic family members, we introduce a new population genetics model based on the mating table approach and adapt the verbal definition of evolutionary stability to genotypes. In general, when the resident is recessive, a homozygote is an evolutionarily stable genotype (ESG), if the number of survivors of the resident genotype of the resident homozygote family is greater than that of non-resident heterozygote survivors of the family of the resident homozygote and mutant heterozygote genotypes. Using the introduced genotype dynamics we proved that in the recessive case ESG implies local stability of the altruistic genotype. We apply our general ESG conditions for self-sacrificing life history strategy when the number of new-born offspring does not depend on interactions within the family and the interactions are additive. We find that in this case our ESG conditions give back Hamilton’s rule for evolutionary stability of the self-sacrificing life history strategy. Conclusions In spite of the fact that the kidney transplantations was not a selection factor during the earlier human evolution, nowadays “self-sacrificing” can be observed in the live donor kidney transplantations, when the donor is one of the family members. It seems that selection for self-sacrificing in family produced an innate moral tendency in modulating social cognition in human brain. Electronic supplementary material The online version of this article (10.1186/s12862-019-1478-0) contains supplementary material, which is available to authorized users.

Background Allopatric speciation has played a particularly important role in archipelagic settings where populations evolve in isolation after colonizing different islands. The Indo-Australasian island realm is an unparalleled natural laboratory of biotic diversification. Here we explore how the level of earth-historic isolation has influenced genetic differentiation across the region by investigating phylogeographic patterns in the Pitta sordida species complex. Results We generated a de novo genome and compared population genomics of 29 individuals of Pitta sordida from the entire distributional range and we reconstructed phylogenetic relationship using mitogenomes, a multi-nuclear gene dataset and single nucleotide polymorphisms (SNPs). We found deep divergence between an eastern and a western group of taxa across Indo-Australasia. Within both groups we have identified major lineages that are geographically separated into Philippines, Borneo, western Sundaland, and New Guinea, respectively. Although these lineages are genetically well-differentiated, suggesting a long-term isolation, there are signatures of extensive gene flow within each lineage throughout the Pleistocene, despite the wide geographic range occupied by some of them. We found little evidence of hybridization or introgression among the studied taxa, but forsteni from Sulawesi makes an exception. This individual, belonging to the eastern clade, is genetically admixed between the western and eastern clades. Geographically this makes sense as Sulawesi is not far from Borneo that houses a population of hooded pittas that belongs to the western clade. Conclusions We found that geological vicariance events cannot explain the current genetic differentiation in the Pitta sordida species complex. Instead, the glacial-interglacial cycles may have played a major role therein. During glacials the sea level could be up to 120 m lower than today and land bridges formed within both the Sunda Shelf and the Sahul Shelf permitting dispersal of floral and faunal elements. The geographic distribution of hooded pittas shows the importance of overwater, “stepping-stone” dispersals not only to deep-sea islands, but also from one shelf to the other. The most parsimonious hypothesis is an Asian ancestral home of the Pitta sordida species complex and a colonization from west to east, probably via Wallacea. Electronic supplementary material The online version of this article (10.1186/s12862-019-1481-5) contains supplementary material, which is available to authorized users.

Background Understanding how variation in gene expression contributes to morphological diversity is a major goal in evolutionary biology. Cichlid fishes from the East African Great lakes exhibit striking diversity in trophic adaptations predicated on the functional modularity of their two sets of jaws (oral and pharyngeal). However, the transcriptional basis of this modularity is not so well understood, as no studies thus far have directly compared the expression of genes in the oral and pharyngeal jaws. Nor is it well understood how gene expression may have contributed to the parallel evolution of trophic morphologies across the replicate cichlid adaptive radiations in Lake Tanganyika, Malawi and Victoria. Results We set out to investigate the role of gene expression divergence in cichlid fishes from these three lakes adapted to herbivorous and carnivorous trophic niches. We focused on the development stage prior to the onset of exogenous feeding that is critical for understanding patterns of gene expression after oral and pharyngeal jaw skeletogenesis, anticipating environmental cues. This framework permitted us for the first time to test for signatures of gene expression underlying jaw modularity in convergent eco-morphologies across three independent adaptive radiations. We validated a set of reference genes, with stable expression between the two jaw types and across species, which can be important for future studies of gene expression in cichlid jaws. Next we found evidence of modular and non-modular gene expression between the two jaws, across different trophic niches and lakes. For instance, prdm1a, a skeletogenic gene with modular anterior-posterior expression, displayed higher pharyngeal jaw expression and modular expression pattern only in carnivorous species. Furthermore, we found the expression of genes in cichlids jaws from the youngest Lake Victoria to exhibit low modularity compared to the older lakes. Conclusion Overall, our results provide cross-species transcriptional comparisons of modularly-regulated skeletogenic genes in the two jaw types, implicating expression differences which might contribute to the formation of divergent trophic morphologies at the stage of larval independence prior to foraging. Electronic supplementary material The online version of this article (10.1186/s12862-019-1483-3) contains supplementary material, which is available to authorized users.

Background Current patterns of population genetic variation may have been shaped by long-term evolutionary history and contemporary demographic processes. Understanding the underlying mechanisms that yield those patterns is crucial for informed conservation of endangered species. The critically endangered white-headed langur, Trachypithecus leucocephalus, is endemic to a narrow range in southwest China. This species shows very low genetic diversity in its 2 main relict populations, Fusui and Chongzuo. Whether this has been caused by a short evolutionary history or recent population declines is unknown. Therefore, we investigated the contributions of historical and recent population demographic changes to population genetic diversity by using 15 nuclear microsatellite markers and mitochondrial DNA (mtDNA) control region sequences. Results Using genetic data from 214 individuals we found a total of 9 mtDNA haplotypes in the Fusui population but only 1 haplotype in the Chongzuo population, and we found an overall low genetic diversity (haplotype and nucleotide diversities: h = 0.486 ± 0.036; π = 0.0028 ± 0.0003). The demographic history inferred from mtDNA and microsatellite markers revealed no evidence for historical population size fluctuations or recent population bottlenecks. Simulations of possible population divergence histories inferred by DIYABC analysis supported a recent divergence of the Chongzuo population from the Fusui population and no population bottlenecks. Conclusions Despite severe population declines caused by anthropogenic activities in the last century, the low genetic diversity of the extant white-headed langur populations is most likely primarily due to the species’ shallow evolutionary history and to a recent, local population founder event. Electronic supplementary material The online version of this article (10.1186/s12862-019-1451-y) contains supplementary material, which is available to authorized users.

BackgroundOur ability to investigate processes shaping the evolutionary diversification of corals (Cnidaria: Anthozoa) is limited by a lack of understanding of species boundaries. Discerning species has been challenging due to a multitude of factors, including homoplasious and plastic morphological characters and the use of molecular markers that are either not informative or have not completely sorted. Hybridization can also blur species boundaries by leading to incongruence between morphology and genetics. We used traditional DNA barcoding and restriction-site associated DNA sequencing combined with coalescence-based and allele-frequency methods to elucidate species boundaries and simultaneously examine the potential role of hybridization in a speciose genus of octocoral,Sinularia.ResultsSpecies delimitations using two widely used DNA barcode markers,mtMutSand 28S rDNA, were incongruent with one another and with the morphospecies identifications, likely due to incomplete lineage sorting. In contrast, 12 of the 15 morphospecies examined formed well-supported monophyletic clades in both concatenated RAxML phylogenies and SNAPP species trees of >6,000 RADSeq loci. DAPC and Structure analyses also supported morphospecies assignments, but indicated the potential for two additional cryptic species. Three morphologically distinct species pairs could not, however, be distinguished genetically. ABBA-BABA tests demonstrated significant admixture between some of those species, suggesting that hybridization may confound species delimitation inSinularia.ConclusionsA genomic approach can help to guide species delimitation while simultaneously elucidating the processes generating diversity in corals. Results support the hypothesis that hybridization is an important mechanism in the evolution of Anthozoa, including octocorals, and future research should examine the contribution of this mechanism in generating diversity across the coral tree of life.

Background Gene duplication has played an important role in the evolution and domestication of flowering plants. Yet little is known about how plant duplicate genes evolve and are retained over long timescales, particularly those arising from small-scale duplication (SSD) rather than whole-genome duplication (WGD) events. Results We address this question in the Poaceae (grass) family by analyzing gene expression data from nine tissues of Brachypodium distachyon, Oryza sativa japonica (rice), and Sorghum bicolor (sorghum). Consistent with theoretical predictions, expression profiles of most grass genes are conserved after SSD, suggesting that functional conservation is the primary outcome of SSD in grasses. However, we also uncover support for widespread functional divergence, much of which occurs asymmetrically via the process of neofunctionalization. Moreover, neofunctionalization preferentially targets younger (child) duplicate gene copies, is associated with RNA-mediated duplication, and occurs quickly after duplication. Further analysis reveals that functional divergence of SSD-derived genes is positively correlated with both sequence divergence and tissue specificity in all three grass species, and particularly with anther expression in B. distachyon. Conclusions Our results suggest that SSD-derived grass genes often undergo rapid functional divergence that may be driven by natural selection on male-specific phenotypes. These observations are consistent with those in several animal species, suggesting that duplicate genes take similar evolutionary trajectories in plants and animals. Electronic supplementary material The online version of this article (10.1186/s12862-019-1415-2) contains supplementary material, which is available to authorized users.

Background A major focus of evolutionary biology is the formation of reproductive barriers leading to divergence and ultimately, speciation. Often, it is not clear whether the separation of populations is complete or if there still is ongoing gene flow in the form of rare cases of admixture, known as isolation with migration. Here, we studied the speciation of two fire ant species, Solenopsis invicta and Solenopsis richteri, both native to South America, both inadvertently introduced to North America in the early twentieth century. While the two species are known to admix in the introduced range, in the native range no hybrids were found. Results We conducted a population genomic survey of native and introduced populations of the two species using reduced representation genomic sequencing of 337 samples. Using maximum likelihood analysis over native range samples, we found no evidence of any gene flow between the species since they diverged. We estimated their time of divergence to 190,000 (100,000–350,000) generations ago. Modelling the demographic history of native and introduced S. invicta populations, we evaluated their divergence times and historic and contemporary population sizes, including the original founder population in North America, which was estimated at 26 (10–93) unrelated singly-mated queens. Conclusions We provide evidence for complete genetic isolation maintained between two invasive species in their natïve range, based, for the first time, on large scale genomic data analysis. The results lay the foundations for further studies into different stages in the formation of genetic barriers in dynamic, invasive populations. Electronic supplementary material The online version of this article (10.1186/s12862-019-1437-9) contains supplementary material, which is available to authorized users.

Background RNA interference (RNAi) related pathways provide defense against viruses and transposable elements, and have been implicated in the suppression of meiotic drive elements. Genes in these pathways often exhibit high levels of adaptive substitution, and over longer timescales show gene duplication and loss—most likely as a consequence of their role in mediating conflict with these parasites. This is particularly striking for Argonaute 2 (Ago2), which is ancestrally the key effector of antiviral RNAi in insects, but has repeatedly formed new testis-specific duplicates in the recent history of the obscura species-group of Drosophila. Results Here we take advantage of publicly available genomic and transcriptomic data to identify six further RNAi-pathway genes that have duplicated in this clade of Drosophila, and examine their evolutionary history. As seen for Ago2, we observe high levels of adaptive amino-acid substitution and changes in sex-biased expression in many of the paralogs. However, our phylogenetic analysis suggests that co-duplications of the RNAi machinery were not synchronous, and our expression analysis fails to identify consistent male-specific expression. Conclusions These results confirm that RNAi genes, including genes of the antiviral and piRNA pathways, have undergone multiple independent duplications and that their history has been particularly labile within the obscura group. However, they also suggest that the selective pressures driving these changes have not been consistent, implying that more than one selective agent may be responsible., BMC Evolutionary Biology, 19, ISSN:1471-2148

Background The S100A7 gene, also called psoriasin, was first described as an upregulated protein in psoriatic skin. For the past years, the importance of this protein as a key effector of innate immunity has been clearly established, not only due to its importance protecting against bacteria skin insult in humans, but also because of its important role in amplifying inflammatory processes. Given the importance of S100A7 in host defense, S100A7 genes have been mostly studied in humans. Here we provide a detailed analysis of the evolution of the gene family encoding for the S100A7 protein in mammals. Results Examination of several mammalian genomes revealed an unexpected variation in the copy number of S100A7. Among the most representative mammalian groups, we report that multiple events of duplication, gene loss and high mutation rates are shaping the evolution of this gene family. An unexpected result comes from Myotis species (order Chiroptera), where we found an outstanding S100A7 gene radiation, resulting in more than 10 copies in M. lucifugus and 5 copies in M. brandtii. These findings suggest a unique adaptive road in these species and are suggestive of special role of this protein in their immune system. Conclusions We found different evolutionary histories among different mammalian groups. Overall, our results suggest that this gene family is evolving under the birth-and-death model of evolution. To our knowledge, this work represents the first detailed analysis of phylogenetic relationships of S100A7 within mammals and therefore will pave the way to further clarify their unique function in the immune system. Electronic supplementary material The online version of this article (10.1186/s12862-019-1433-0) contains supplementary material, which is available to authorized users.

Background Recent studies have demonstrated that Bayesian species delimitation based on the multispecies coalescent model can produce inaccurate results by misinterpreting population splits as species divergences. An approach based on the genealogical divergence index (gdi) was shown to be a viable alternative, especially for delimiting allopatric populations where gene flow is low. We implemented these analyses to assess species boundaries in Southeast Asian toads, a group that is understudied and characterized by numerous unresolved species complexes. Results Multilocus phylogenetic analyses showed that deep evolutionary relationships including the genera Sigalegalephrynus, Ghatophryne, Parapelophryne, Leptophryne, Pseudobufo, Rentapia, and Phrynoides remain unresolved. Comparison of genetic divergences revealed that intraspecific divergences among allopatric populations of Pelophyrne signata (Borneo vs. Peninsular Malaysia), Ingerophrynus parvus (Peninsular Malaysia vs. Myanmar), and Leptophryne borbonica (Peninsular Malaysia, Java, Borneo, and Sumatra) are consistent with interspecific divergences of other Southeast Asian bufonid taxa. Conversely, interspecific divergences between Pelophryne guentheri/P. api, Ansonia latiffi/A. leptopus, and I. gollum/I. divergens were low (

Background Animals have a greater diversity of signalling pathways than their unicellular relatives, consistent with the evolution and expansion of these pathways occurring in parallel with the origin of animal multicellularity. However, the genomes of sponges and ctenophores – non-bilaterian basal animals – typically encode no, or far fewer, recognisable signalling ligands compared to bilaterians and cnidarians. For instance, the largest subclass of receptor tyrosine kinases (RTKs) in bilaterians, the Eph receptors (Ephs), are present in sponges and ctenophores, but their cognate ligands, the ephrins, have not yet been detected. Results Here, we use an iterative HMM analysis to identify for the first time membrane-bound ephrins in sponges and ctenophores. We also expand the number of Eph-receptor subtypes identified in these animals and in cnidarians. Both sequence and structural analyses are consistent with the Eph ligand binding domain (LBD) and the ephrin receptor binding domain (RBD) having evolved via the co-option of ancient galactose-binding (discoidin-domain)-like and monodomain cupredoxin domains, respectively. Although we did not detect a complete Eph-ephrin signalling pathway in closely-related unicellular holozoans or in other non-metazoan eukaryotes, truncated proteins with Eph receptor LBDs and ephrin RBDs are present in some choanoflagellates. Together, these results indicate that Eph-ephrin signalling was present in the last common ancestor of extant metazoans, and perhaps even in the last common ancestor of animals and choanoflagellates. Either scenario pushes the origin of Eph-ephrin signalling back much earlier than previously reported. Conclusions We propose that the Eph-LBD and ephrin-RBD, which were ancestrally localised in the cytosol, became linked to the extracellular parts of two cell surface proteins before the divergence of sponges and ctenophores from the rest of the animal kingdom. The ephrin-RBD lost the ancestral capacity to bind copper, and the Eph-LBD became linked to an ancient RTK. The identification of divergent ephrin ligands in sponges and ctenophores suggests that these ligands evolve faster than their cognate receptors. As this may be a general phenomena, we propose that the sequence-structure approach used in this study may be usefully applied to other signalling systems where no, or a small number of, ligands have been identified.

Color and pattern phenotypes have clear implications for survival and reproduction in many species. However, the mechanisms that produce this coloration are still poorly characterized, especially at the genomic level. Here we have taken a transcriptomics-based approach to elucidate the underlying genetic mechanisms affecting color and pattern in a highly polytypic poison frog. We sequenced RNA from the skin from four different color morphs during the final stage of metamorphosis and assembled a de novo transcriptome. We then investigated differential gene expression, with an emphasis on examining candidate color genes from other taxa. Overall, we found differential expression of a suite of genes that control melanogenesis, melanocyte differentiation, and melanocyte proliferation (e.g., tyrpl, lefl, leol, and mitf) as well as several differentially expressed genes involved in purine synthesis and iridophore development (e.g., arfgapl, arfgap2, airc, and gairt). Our results provide evidence that several gene networks known to affect color and pattern in vertebrates play a role in color and pattern variation in this species of poison frog.

Background DNA barcoding has been developed as a useful tool for species discrimination. Several sequence-based species delimitation methods, such as Barcode Index Number (BIN), REfined Single Linkage (RESL), Automatic Barcode Gap Discovery (ABGD), a Java program uses an explicit, determinate algorithm to define Molecular Operational Taxonomic Unit (jMOTU), Generalized Mixed Yule Coalescent (GMYC), and Bayesian implementation of the Poisson Tree Processes model (bPTP), were used. Our aim was to estimate Chinese katydid biodiversity using standard DNA barcode cytochrome c oxidase subunit I (COI-5P) sequences. Results Detection of a barcoding gap by similarity-based analyses and clustering-base analyses indicated that 131 identified morphological species (morphospecies) were assigned to 196 BINs and were divided into four categories: (i) MATCH (83/131 = 64.89%), morphospecies were a perfect match between morphospecies and BINs (including 61 concordant BINs and 22 singleton BINs); (ii) MERGE (14/131 = 10.69%), morphospecies shared its unique BIN with other species; (iii) SPLIT (33/131 = 25.19%, when 22 singleton species were excluded, it rose to 33/109 = 30.28%), morphospecies were placed in more than one BIN; (iv) MIXTURE (4/131 = 5.34%), morphospecies showed a more complex partition involving both a merge and a split. Neighbor-joining (NJ) analyses showed that nearly all BINs and most morphospecies formed monophyletic cluster with little variation. The molecular operational taxonomic units (MOTUs) were defined considering only the more inclusive clades found by at least four of seven species delimitation methods. Our results robustly supported 61 of 109 (55.96%) morphospecies represented by more than one specimen, 159 of 213 (74.65%) concordant BINs, and 3 of 8 (37.5%) discordant BINs. Conclusions Molecular species delimitation analyses generated a larger number of MOTUs compared with morphospecies. If these MOTU splits are proven to be true, Chinese katydids probably contain a seemingly large proportion of cryptic/undescribed taxa. Future amplification of additional molecular markers, particularly from the nuclear DNA, may be especially useful for specimens that were identified here as problematic taxa.

Background Sex-determining systems may profoundly influence the ecology, behaviour and demography of animals, yet these relationships are poorly understood. Here we investigate whether species with temperature-dependent (TSD) and genetic sex determination (GSD) differ in key demographic traits, using data from 181 species representing all major phylogenetic lineages of extant reptiles. Results We show that species with TSD exhibit significantly higher within-species variance in sex ratios than GSD species in three major life stages: birth or hatching, juvenility and adulthood. In contrast, sex differences in adult mortality rates do not differ between GSD and TSD species. However, TSD species exhibit significantly greater sex differences in maturation ages than GSD species. Conclusion These results support the recent theoretical model that evolution of TSD is facilitated by sex-specific fitness benefits of developmental temperatures due to bimaturism. Our findings suggest that different sex-determination systems are associated with different demographic characteristics that may influence population viability and social evolution. Electronic supplementary material The online version of this article (10.1186/s12862-019-1386-3) contains supplementary material, which is available to authorized users.

Background Sex-biased gene expression is thought to drive the phenotypic differences in males and females in metazoans. Drosophila has served as a primary model for studying male-female differences in gene expression, and its effects on protein sequence divergence. However, the forces shaping evolution of sex-biased expression remain largely unresolved, including the roles of selection and pleiotropy. Research on sex organs in Drosophila, employing original approaches and multiple-species contrasts, provides a means to gain insights into factors shaping the turnover and magnitude (fold-bias) of sex-biased expression. Results Here, using recent RNA-seq data, we studied sex-biased gonadal expression in 10,740 protein coding sequences in four species of Drosophila, D. melanogaster, D. simulans, D. yakuba and D. ananassae (5 to 44 My divergence). Using an approach wherein we identified genes with lineage-specific transitions (LSTs) in sex-biased status (amongst testis-biased, ovary-biased and unbiased; thus, six transition types) standardized to the number of genes with the ancestral state (S-LSTs), and those with clade-wide expression bias status, we reveal several key findings. First, the six categorical types of S-LSTs in sex-bias showed disparate rates of turnover, consistent with differential selection pressures. Second, the turnover in sex-biased status was largely unrelated to cross-tissue expression breadth, suggesting pleiotropy does not restrict evolution of sex-biased expression. Third, the fold-sex-biased expression, for both testis-biased and ovary-biased genes, evolved directionally over time toward higher values, a crucial finding that could be interpreted as a selective advantage of greater sex-bias, and sexual antagonism. Fourth, in terms of protein divergence, genes with LSTs to testis-biased expression exhibited weak signals of elevated rates of evolution (than ovary-biased) in as little as 5 My, which strengthened over time. Moreover, genes with clade-wide testis-specific expression (44 My), a status not observed for any ovary-biased genes, exhibited striking acceleration of protein divergence, which was linked to low pleiotropy. Conclusions By studying LSTs and clade-wide sex-biased gonadal expression in a multi-species clade of Drosophila, we describe evidence that interspecies turnover and magnitude of sex-biased expression have been influenced by selection. Further, whilst pleiotropy was not connected to turnover in sex-biased gonadal expression, it likely explains protein sequence divergence. Electronic supplementary material The online version of this article (10.1186/s12862-019-1377-4) contains supplementary material, which is available to authorized users.