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A New Spiralian Phylogeny Places the Enigmatic Arrow Worms among Gnathiferans
Abstract
Chaetognaths (arrow worms) are an enigmatic group of marine animals whose phylogenetic position remains elusive, in part because they display a mix of developmental and morphological characters associated with other groups [1, 2]. In particular, it remains unclear whether they are a sister group to protostomes [1, 2], one of the principal animal superclades, or whether they bear a closer relationship with some spiralian phyla [3, 4]. Addressing the phylogenetic position of chaetognaths and refining our understanding of relationships among spiralians are essential to fully comprehend character changes during bilaterian evolution [5]. To tackle these questions, we generated new transcriptomes for ten chaetognath species, compiling an extensive phylogenomic dataset that maximizes data occupancy and taxonomic representation. We employed inference methods that consider rate and compositional heterogeneity across taxa to avoid limitations of earlier analyses [6]. In this way, we greatly improved the resolution of the protostome tree of life. We find that chaetognaths cluster together with rotifers, gnathostomulids, and micrognathozoans within an expanded Gnathifera clade and that this clade is the sister group to other spiralians [7, 8]. Our analysis shows that several previously proposed groupings are likely due to systematic error, and we propose a revised organization of Lophotrochozoa with three main clades: Tetraneuralia (mollusks and entoprocts), Lophophorata (brachiopods, phoronids, and ectoprocts), and a third unnamed clade gathering annelids, nemerteans, and platyhelminthes. Consideration of classical morphological, developmental, and genomic characters in light of this topology indicates secondary loss as a fundamental trend in spiralian evolution.
... The asterisk indicates the position of the mouth. 1: Zrzavý et al. (1998) [27]; 2: Halanych (1996) [28], Peterson & Eernisse (2001) [29]; 3: Paps et al. (2009) [30], Marlétaz et al. 2019 [26]; 4: Mallat & Winchell (2002) [31]; 5: Littlewood et al. (1998) [32]; 6: Matus et al. (2006) [33], Papillon et al. (2004) [34], Dunn et al. (2008) [35], Philippe et al. (2011) [36]; 7: Giribet et al. (2000) [37], Helfenbein et al. (2004) [38], Marlétaz et al. (2006) [39]; 8: Telford & Holland (1993) [40]; Papillon et al. (2003) [41]; cc, corona ciliata; cg, cerebral ganglion; cn: caudal nerve; dlm, dorsal longitudinal muscle; eg, esophageal ganglion; es, esophagus; epi: epidermis; ey: eye; hd: head; in, instestine; lsc, lateral somata clusters; mc: main connective; mvs, medioventral somata clusters; np, trunk neuropil; seg, subesophageal ganglion; so, ciliary sensory organ; tl: tail; tr: trunk; vlm, ventral longitudinal muscle, vsg, vestibular ganglion. Scale bars equal 50 µm (a) and 130 µm (b) portion of the global ocean zooplankton [21]. ...
... Arrow worms exhibit genetic and embryonic developmental features that resemble those of protostomes and deuterostomes, resulting in varying phylogenetic positions within Bilateria ( Fig. 1c; reviewed in [22][23][24][25]). However, based on a comparison of some bilaterian genome-wide transcriptomes, chaetognaths have recently been suggested to belong to the clade Gnathifera (Ahlrichs, 1995), which includes animals with internal chitinous jaw-like structures such as gnathostomulids (jaw worms) and rotifers (wheel animals) [26]. ...
... To further explore the molecular aspects of nervous system development in chaetognaths, we performed an in-depth characterization of evolutionarily highly conserved genes involved in bilaterian neurogenesis in the early post-embryonic stages of Spadella cephaloptera. We determined whether these axial patterning genes are regionally expressed along the mediolateral axis of the developing nerve cord similar to what is observed in diverse bilaterian representatives identified in [16,26]. We first screened for gene orthologs of foxA, nkx2.2, pax6, pax3/7, msx, ChAT, VAChT, and elav in the draft transcriptome of S. cephaloptera [47] and examined their spatial expression using in situ hybridization on hatchlings and early juveniles. ...
Background
Chaetognaths are a clade of marine worm-like invertebrates with a heavily debated phylogenetic position. Their nervous system superficially resembles the protostome type, however, knowledge regarding the molecular processes involved in neurogenesis is lacking. To better understand these processes, we examined the expression profiles of marker genes involved in bilaterian neurogenesis during post-embryonic stages of Spadella cephaloptera. We also investigated whether the transcription factor encoding genes involved in neural patterning are regionally expressed in a staggered fashion along the mediolateral axis of the nerve cord as it has been previously demonstrated in selected vertebrate, insect, and annelid models.
Methods
The expression patterns of genes involved in neural differentiation (elav), neural patterning (foxA, nkx2.2, pax6, pax3/7, and msx), and neuronal function (ChAT and VAChT) were examined in S. cephaloptera hatchlings and early juveniles using whole-mount fluorescent in situ hybridization and confocal microscopy.
Results
The Sce-elav⁺ profile of S. cephaloptera hatchlings reveals that, within 24 h of post-embryonic development, the developing neural territories are not limited to the regions previously ascribed to the cerebral ganglion, the ventral nerve center (VNC), and the sensory organs, but also extend to previously unreported CNS domains that likely contribute to the ventral cephalic ganglia. In general, the neural patterning genes are expressed in distinct neural subpopulations of the cerebral ganglion and the VNC in hatchlings, eventually becoming broadly expressed with reduced intensity throughout the CNS in early juveniles. Neural patterning gene expression domains are also present outside the CNS, including the digestive tract and sensory organs. ChAT and VAChT domains within the CNS are predominantly observed in specific subpopulations of the VNC territory adjacent to the ventral longitudinal muscles in hatchlings.
Conclusions
The observed spatial expression domains of bilaterian neural marker gene homologs in S. cephaloptera suggest evolutionarily conserved roles in neurogenesis for these genes among bilaterians. Patterning genes expressed in distinct regions of the VNC do not show a staggered medial-to-lateral expression profile directly superimposable to other bilaterian models. Only when the VNC is conceptually laterally unfolded from the longitudinal muscle into a flat structure, an expression pattern bearing resemblance to the proposed conserved bilaterian mediolateral regionalization becomes noticeable. This finding supports the idea of an ancestral mediolateral patterning of the trunk nervous system in bilaterians.
... Moreover, the internal relationships of spiralians remain disputed, despite the availability of transcriptome and even genomic data in more and more lineages [10][11][12], particularly, because several clades (gastrotrichs, gnathostomulids, platyhelminthes) are prone to long-branch attraction due to their fast rates of evolution [13]. Recently, the clade Gnathifera, gathering chaetognaths, rotifers and other jaw-bearing groups, has been established [11,12]. ...
... Moreover, the internal relationships of spiralians remain disputed, despite the availability of transcriptome and even genomic data in more and more lineages [10][11][12], particularly, because several clades (gastrotrichs, gnathostomulids, platyhelminthes) are prone to long-branch attraction due to their fast rates of evolution [13]. Recently, the clade Gnathifera, gathering chaetognaths, rotifers and other jaw-bearing groups, has been established [11,12]. The relationships among lophotrochozoans taxa however remain contentious, particularly that of annelids, molluscs, platyhelminthes and lophophorates ( Figure 1). ...
... In this review, we refer to the sister-group of Gnathifera as Lophotrochozoa for convenience and clarity, but we acknowledge that some authors might hold other views [14]. For instance, platyhelminthes have moved from representing a relatively early diverging lineage among spiralians together with gastrotrichs (the 'Rouphozoa' hypothesis [15]) to possibly be close relatives to nemerteans and annelids (the 'Vermizoa' clade [11]) ( Figure 1). Similarly, the monophyly of lophophorates has been repeatedly questioned and reaffirmed, with bryozoans sometimes excluded from it [10][11][12]. ...
Spiralians represent the least studied superclade of bilaterian animals, despite exhibiting the widest diversity of organisms. Although spiralians include iconic organisms, such as octopus, earthworms and clams, a lot remains to be discovered regarding their phylogeny and biology. Here, we review recent attempts to apply single-cell transcriptomics, a new pioneering technology enabling the classification of cell types and the characterisation of their gene expression profiles, to several spiralian taxa. We discuss the methodological challenges and requirements for applying this approach to marine organisms and explore the insights that can be brought by such studies, both from a biomedical and evolutionary perspective. For instance, we show that single-cell sequencing might help solve the riddle of the homology of larval forms across spiralians, but also to better characterise and compare the processes of regeneration across taxa. We highlight the capacity of single-cell to investigate the origin of evolutionary novelties, as the mollusc shell or the cephalopod visual system, but also to interrogate the conservation of the molecular fingerprint of cell types at long evolutionary distances. We hope that single-cell sequencing will open a new window in understanding the biology of spiralians, and help renew the interest for these overlooked but captivating organisms.
... Spiralian genomics holds the potential to offer unique insights into the evolutionary origins of animal body plans [13] and cell types [14], thereby advancing our understanding of animal evolution [15]. [19] for Gnathifera, Lu et al. [18] for Mesozoa and Rouphozoa and Laumer et al. [17] and Luo et al. [6] for Lophotrochozoa. Note that Micrognathozoa contains only one species and is ranked as a class in the NCBI. ...
... Silhouettes of representative species from each spiralian phylum were adapted from PhyloPic. (B) Notable hypothesized clades within Spiralia, including Parenchymia supported by Marlétaz et al. [19]. Four major subclades are highlighted in bold. ...
... monophyletic clade of non-ecdysozoan protostomes. In general, Spiralia contains four major subclades: Gnathifera, Mesozoa, Rouphozoa and Lophotrochozoa sensu stricto (hereafter referred to as Lophotrochozoa) [16][17][18][19][20] (Figure 1). ...
Recent developments in sequencing technologies have greatly improved our knowledge of phylogenetic relationships and genomic architectures throughout the tree of life. Spiralia, a diverse clade within Protostomia, is essential for understanding the evolutionary history of parasitism, gene conversion, nervous systems and animal body plans. In this review, we focus on the current hypotheses of spiralian phylogeny and investigate the impact of long-read sequencing on the quality of genome assemblies. We examine chromosome-level assemblies to highlight key genomic features that have driven spiralian evolution, including karyotype, synteny and the Hox gene organization. In addition, we show how chromosome rearrangement has influenced spiralian genomic structures. Although spiralian genomes have undergone substantial changes, they exhibit both conserved and lineage-specific features. We recommend increasing sequencing efforts and expanding functional genomics research to deepen insights into spiralian biology.
... Our intention here is not to attempt to resolve the long-standing phylogenetic problem but instead to examine the relative utility of DNA versus protein sequences for inferring deep phylogenies. Studies of metazoan phylogenies have almost exclusively relied on protein sequences (Philippe et al., 2011;Laumer et al., 2015;Telford et al., 2015;Kocot et al., 2017;Cannon et al., 2016;Laumer et al., 2019;Marlétaz et al., 2019;Philippe et al., 2019;Kapli and Telford, 2020;Kapli et al., 2021), and no corresponding DNA alignments for the protein sequences analyzed in those studies are easily available. As a result, we have very limited knowledge of whether DNA sequences would yield similar phylogenetic results to protein sequences. ...
... The two PhyloBayes analyses differed in the recovered relationships of the Lophotrochozoa lineages with the exception of the flatworm (Macrostomum lignano). Overall, the differences among all inferred trees concern parts of the phylogeny that had low bootstrap support or posterior probability and are known controversial parts of the animal phylogeny (Philippe et al., 2011;Laumer et al., 2015;Telford et al., 2015;Kocot et al., 2017;Cannon et al., 2016;Laumer et al., 2019;Marlétaz et al., 2019;Philippe et al., 2019;Kapli and Telford, 2020;Kapli et al., 2021). ...
... Deuterostomes have been a long-trusted clade in the animal phylogeny. However, it was not supported in recent phylogenomic studies (Marlétaz et al., 2019;Philippe et al., 2019), and was hypothesized to be an artefact of model misspecification . Similarly, the placement of Xenacoelomorpha is uncertain, either sister to Nephrozoa (Lophotrochozoa, Ecdysozoa, Ambulacraria, Chordata) (Cannon et al., 2016) or to Ambulacraria (Philippe et al., 2019;Kapli and Telford, 2020). ...
Inference of deep phylogenies has almost exclusively used protein rather than DNA sequences, based on the perception that protein sequences are less prone to homoplasy and saturation or to issues of compositional heterogeneity than DNA sequences. Here we analyze a model of codon evolution under an idealized genetic code and demonstrate that those perceptions may be misconceptions. We conduct a simulation study to assess the utility of protein versus DNA sequences for inferring deep phylogenies, with protein-coding data generated under models of heterogeneous substitution processes across sites in the sequence and among lineages on the tree, and then analyzed using nucleotide, amino acid, and codon models. Analysis of DNA sequences under nucleotide-substitution models (possibly with the third codon positions excluded) recovered the correct tree at least as often as analysis of the corresponding protein sequences under modern amino acid models. We also applied the different data-analysis strategies to an empirical dataset to infer the metazoan phylogeny. Our results from both simulated and real data suggest that DNA sequences may be as useful as proteins for inferring deep phylogenies and should not be excluded from such analyses. Analysis of DNA data under nucleotide models has a major computational advantage over protein-data analysis, potentially making it feasible to use advanced models that account for among-site and among-lineage heterogeneity in the nucleotide-substitution process in inference of deep phylogenies.
... Cohen proposed that Phoronida should be treated as a subphylum Phoroniformea within brachiopods (Cohen & Weydmann, 2005;Cohen, 2000Cohen, , 2013Santagata & Cohen, 2009). However, later transcriptome-based analyses were rather favoring a sister relation of phoronids and bryozoans (Figure 1e; Laumer et al., 2015Laumer et al., , 2019Marlétaz et al., 2019;Nesnidal et al., 2013;Zverkov et al., 2019), with which they seem to share some potential morphological and genetic synapomorphies (Temereva, 2017b(Temereva, , 2019bWernström et al., 2022), while brachiopods were retrieved as the more distant branch within monophyletic Lophophorata. Recently, the increased sampling of the bryozoan transcriptomes once again challenged the monophyly of Lophophorates resulting in the topologies in which phoronids and brachiopods form sister groups, while bryozoans, together with kamptozoans, form clade Polyzoa only distantly related to the other lophophorates ( Figure 1f; Drábková et al., 2022;Khalturin et al., 2022). ...
... Recently, the increased sampling of the bryozoan transcriptomes once again challenged the monophyly of Lophophorates resulting in the topologies in which phoronids and brachiopods form sister groups, while bryozoans, together with kamptozoans, form clade Polyzoa only distantly related to the other lophophorates ( Figure 1f; Drábková et al., 2022;Khalturin et al., 2022). As for today, the close relatedness of phoronids and brachiopods seems well-established (Drábková et al., 2022;Dunn et al., 2008;Khalturin et al., 2022;Kocot et al., 2017;Laumer et al., 2015Laumer et al., , 2019Luo et al., 2018;Marlétaz et al., 2019;Zverkov et al., 2019), however, their relation to bryozoans remains disputable. Therefore, the immediate sister group of phoronids is still effectively unknown. ...
... The new, more comprehensive phylogenetic analysis that took into account those newly discovered camenellid fossils, suggested that Eccentrotheca is not a sister taxon of phoronids but instead, it belongs to the brachiopod lineage of tommotiids (Figure 4; Guo et al., 2022). The same analysis showed that phoronids are more closely related to bryozoan than to any other known group of fossil tommotiids-a topology reminiscent of most of the modern molecular phylogenies (Laumer et al., 2015(Laumer et al., , 2019Marlétaz et al., 2019;Nesnidal et al., 2013;Zverkov et al., 2019). ...
Phoronids, together with brachiopods and bryozoans, form the animal clade Lophophorata. Modern lophophorates are quite diverse-some can biomineralize while others are soft-bodied, they could be either solitary or colonial, and they develop through various eccentric larval stages that undergo different types of metamorphoses. The diversity of this clade is further enriched by numerous extinct fossil lineages with their own distinct body plans and life histories. In this review, I discuss how data on phoronid development, genetics, and morphology can inform our understanding of lophophorate evolution. The actinotrocha larvae of phoronids is a well documented example of intercalation of the new larval body plan, which can be used to study how new life stages emerge in animals with biphasic life cycle. The genomic and embryonic data from phoronids, in concert with studies of the fossil lophophorates, allow the more precise reconstruction of the evolution of lophophorate biomineralization. Finally, the regenerative and asexual abilities of phoronids can shed new light on the evolution of coloniality in lophophorates. As evident from those examples, Phoronida occupies a central role in the discussion of the evolution of lophophorate body plans and life histories.
... Nonetheless, recent phylogenomic studies have found several meiofaunal phyla branching off at the deeper nodes of Bilateria and Spiralia/Lophotrochozoa ( Fig. 1.1). These include the acoelomorph worms, which have been recovered as sister group to the remaining Bilateria (Cannon et al. 2016), as well as the Gnathifera consistently recovered as sister group to the remainder of Spiralia (Struck et al. 2014;Laumer et al. 2015aLaumer et al. , 2019Marlétaz et al. 2019). Moreover, within Ecdysozoa several microscopic phyla exist that subtend macroscopic branches (Laumer et al. 2015a(Laumer et al. , 2019Borner et al. 2014;Campbell et al. 2011). ...
... Moreover, the embryology is still only known from a restricted number of representatives, some of which show deviation from the spiral cleavage pattern (Hejnol 2010;Martín-Durán and Marlétaz 2020). However, Spiralia are also wellsupported in phylogenomic analyses (Laumer et al. 2015a(Laumer et al. , 2019Marlétaz et al. 2019), and there are no reasons to question its monophyly. The basal branching order in Spiralia has been a topic of intense debate during the last decade, but we are moving closer to a consensus by consistently identifying the Gnathifera as the sister group to all other Spiralia. ...
... However, recent studies revealed that the Hox protein gene MedPost, that so far had been considered unique for chaetognaths, is present in rotifers as well, suggesting a closer relationship (Fröbius and Funch, 2017). This affinity was confirmed by a subsequent transcriptomic phylogenetic study, which substantiated the relationship between Chaetognatha and the gnathiferan taxa (Marlétaz et al. 2019;Laumer et al. 2019) (Fig. 1.1). Additional evidence is provided by the fossil record with studies suggesting that the Cambrian taxon Amiskwia was equipped with internal, pharyngeal hard parts that could be homologized with the gnathiferan jaws (Caron and Cheung 2019; Vinther and Parry 2019) and, thus, representing a taxon with a combination of gnathiferan and chaetognath characters (see further discussion below in Sect. ...
Meiofaunal life forms are found all over the animal tree of life, and miniaturization seems to have occurred within otherwise macrofaunal clades multiple times. While sponges, comb jellies and cnidarians suggest a macroscopic ancestry for Metazoa, several phyla are exclusively meiofaunal, however, and may evidence a wider microscopic ancestry of some major groups, such as Ecdysozoa and Spiralia/Lophotrochozoa. This is an unsolved debate, which should be tackled from a synthesis of zoomorphological, palaeontological, molecular and phylogenetic approaches to test alternative scenarios. Advances in microscopic techniques have led to a renaissance in anatomical studies that allows for new and detailed examination of both extant and extinct meiofauna, revealing an unseen wealth of information. Likewise, the rapid development in genomic sequencing and analytical tools makes detailed reconstructions of meiofauna genomes feasible. The anticipated flood of new morphological and molecular data on meiofauna will broaden integrative and comparative studies and hopefully allow scientists of this generation to answer the long-debated questions of how the animal kingdom evolved and ramified into today's amazing diversity of life. In this enormously complex tree of life, what is the significance of minute creatures represented by meiofauna?
... In Spiralia, the expression of opsins and other photoreceptor genes has so far been studied in mollusks (Vöcking et al., 2015(Vöcking et al., , 2017Wollesen et al., 2019), annelids (Arendt et al., 2004), platyhelminths (Rawlinson et al., 2019), and brachiopods (Passamaneck & Martindale, 2013). Gnathifera have recently been identified as the sister taxon to all other Lophotrochozoa (Marlétaz et al., 2019). They comprise the arrow worms (Chaetognatha), wheel animals (Rotifera), jaw worms (Gnathostomulida), and the tiny Micrognathozoa and have not yet been investigated with respect to genes underlying photoreceptor formation and function (Figure 1; Marlétaz et al., 2019). ...
... Gnathifera have recently been identified as the sister taxon to all other Lophotrochozoa (Marlétaz et al., 2019). They comprise the arrow worms (Chaetognatha), wheel animals (Rotifera), jaw worms (Gnathostomulida), and the tiny Micrognathozoa and have not yet been investigated with respect to genes underlying photoreceptor formation and function (Figure 1; Marlétaz et al., 2019). Chaetognaths are a rather small taxon of marine, torpedo-shaped coelomic animals with horizontally projecting fins, and cuticular grasping spines to catch prey (Shinn, 1997). ...
... As a major macroplanktonic component without respiratory or circulatory systems, chaetognaths exhibit traits reminiscent of deuterostomes (e.g., aspects of the gastrulation process) as well as of protostomes (e.g., nervous system development). Hence, their phylogenetic position had been contentious until phylogenetic analyses recently placed them within the Gnathifera (summarized by Harzsch & Wanninger, 2010;Harzsch et al., 2015;Marlétaz et al., 2019). ...
The phylogenetic position of chaetognaths, or arrow worms, has been debated for decades, however recently they have been grouped into the Gnathifera, a sister clade to all other Spiralia. Chaetognath photoreceptor cells are anatomically unique by exhibiting a highly modified cilium and are arranged differently in the eyes of the various species. Studies investigating eye development and underlying gene regulatory networks are so far missing. To gain insights into the development and the molecular toolkit of chaetognath photoreceptors and eyes a new transcriptome of the epibenthic species Spadella cephaloptera was searched for opsins. Our screen revealed two copies of xenopsin and a single copy of peropsin. Gene expression analyses demonstrated that only xenopsin1 is expressed in photoreceptor cells of the developing lateral eyes. Adults likewise exhibit two xenopsin1 + photoreceptor cells in each of their lateral eyes. Beyond that, a single cryptochrome gene was uncovered and found to be expressed in photoreceptor cells of the lateral developing eye. In addition, cryptochrome is also expressed in the cerebral ganglia in a region in which also peropsin expression was observed. This condition is reminiscent of a nonvisual photoreceptive zone in the apical nervous system of the annelid Platynereis dumerilii that performs circadian entrainment and melatonin release. Cryptochrome is also expressed in cells of the corona ciliata, an organ in the posterior dorsal head region, indicating a role in circadian entrainment. Our study highlights the importance of the Gnathifera for unraveling the evolution of photoreceptors and eyes in Spiralia and Bilateria.
... Interestingly, there are three different gland cell types in the apical organ in early rudiment hoplonemertean larvae (Magarlamov et al., 2020) and in Müller's larvae (Figure 4a According to recent molecular phylogenies, Platyhelminthes and Nemertea are possible sister groups (Marlétaz et al., 2019, Philippe et al., 2019; this clade has been called Parenchymia (Nielsen, 1995). An interesting parallel in polyclad larvae and larvae of hoplonemerteans is that in both groups the multiciliated cells with a lumen, and the ciliated tuft cells of the apical organ are intimately associated with gland cells (Magarlamov et al., 2020). ...
... Nerves associated with the apical organ drawn in red, glands in green and purple. (e) Spiralian larvae in a tree afterMarlétaz et al. (2019) with apical organs highlighted. Yellow circle marks presence of ciliated sensory cells, pink circle marks presence of ampullary cells. ...
The tiger flatworm Prostheceraeus crozieri (Polycladida) develops via an eight-lobed, and three-eyed planktonic Müller's larva. This larva has an apical organ, ultrastructural details of which remain elusive due to a scarcity of studies. The evolution and possible homology of the polyclad larva with other spiralian larvae is still controversial. Here, we provide ultrastructural data and three-dimensional reconstructions of the apical organ of P. crozieri. The apical organ consists of an apical tuft complex and a dorso-apical tuft complex. The apical tuft complex features a central tuft of five long cilia, which emerge from four or five individual cells that are themselves encircled by two anchor cells. The necks of six multibranched gland cells are sandwiched between ciliated tuft cell bodies and anchor cells. The proximal parts of the ciliated cell bodies are in contact with the lateral brain neuropil via gap junctions. Located dorsally of the apical tuft complex, the dorso-apical tuft complex is characterized by several long cilia of sensory neurons, these emerge from an epidermal lumen and are closely associated with several gland cells that form a crescent apically around the dorsal anchor cell, and laterally touch the brain neuropil. Such ciliated sensory neurons emerging from a ciliated lumen are reminiscent of ampullary cells of mollusc and annelid larvae; a similar cell type can be found in the hoplonemertean decidula larva. We hypothesize that the ampullary-like cells and the tuft-forming sensory cells in the apical organs of these spiralian larvae could be homologous.
... Bilateria have long been split into two lineages, Protostomia (Ecdysozoa + Spiralia [Lophotrochozoa]) and Deuterostomia (traditionally: Chordata + Ambulacraria [= Hemichordata + Echinodermata]) [31], historically based on the different origins of the mouth and other features during development [32]. However, recent phylogenomic studies challenged the monophyly of Deuterostomia and recovered paraphyletic deuterostomes in conjunction with Xenambulacraria [33,34]. This combination of results, if confirmed, would have substantial implications for our understanding of the last common ancestor of all Bilateria, which might then have been a fairly large organism, with pharyngeal gill slits and other traits previously thought to represent apomorphies of Deuterostomia ([see 34] for an in-depth discussion). ...
... With reference to the phylogenetic placement of the Xenoacoelomorpha, our analyses favour the Nephrozoa hypothesis. However, the debate on the placement of the Xenoacolemorpha is much less developed [4, 11, 28-30, 34, 62], with some key new hypotheses (e.g., the non-monophyly of Deuterostomia) recently emerging [33,34]. Clearly, more studies, using different datasets and methods, as well as the development of more sophisticated evolutionary models for the analysis of gene content data, are necessary to more firmly establish the relationships at the root of the Bilateria. ...
An accurate phylogeny of animals is needed to clarify their evolution, ecology, and impact on shaping the biosphere. Although datasets of several hundred thousand amino acids are nowadays routinely used to test phylogenetic hypotheses, key deep nodes in the metazoan tree remain unresolved: the root of animals, the root of Bilateria, and the monophyly of Deuterostomia. Instead of using the standard approach of amino acid datasets, we performed analyses of newly assembled genome gene content and morphological datasets to investigate these recalcitrant nodes in the phylogeny of animals. We explored extensively the choices for assembling the genome gene content dataset and model choices of morphological analyses. Our results are robust to these choices and provide additional insights into the early evolution of animals, they are consistent with sponges as the sister group of all the other animals, the worm-like bilaterian lineage Xenacoelomorpha as the sister group of the other Bilateria, and tentatively support monophyletic Deuterostomia.
... Finally, recent work proposed new models at the base of Holozoa, with Ichtyosporea and Corallochytrea/Pluriformea forming Teretosporea, which together with Filozoa give rise to Holozoa (Torruella et al., 2015;Arroyo et al., 2020). Accordingly, we assembled a dendrogram for holozoans based on consensus information extracted from these and other recent publications (Ryan et al., 2010;Ruggiero et al., 2015;Torruella et al., 2015;Cannon et al., 2016;Kocot et al., 2017;Lu et al., 2017;Sebé-Pedrós et al., 2017;Whelan et al., 2017;Adl et al., 2019;Giribet et al., 2019;Laumer et al., 2019;Marlétaz et al., 2019;Sogabe et al., 2019;Schoch et al., 2020;Schulze and Kawauchi, 2021). ...
... Classification of holozoans. Dendrogram depicting the herein proposed hierarchical clustering of phyla within holozoans assembled based on current literature(Ryan et al., 2010; Ruggiero et al., 2015; Torruella et al., 2015;Cannon et al., 2016;Lu et al., 2017;Sebé-Pedrós et al., 2017;Whelan et al., 2017;Adl et al., 2019;Giribet et al., 2019;Laumer et al., 2019;Marlétaz et al., 2019;Sogabe et al., 2019;Hickman et al., 2020;Schoch et al., 2020; ...
The astacins are a family of metallopeptidases (MPs) that has been extensively described from animals. They are multidomain extracellular proteins, which have a conserved core architecture encompassing a signal peptide for secretion, a prodomain or prosegment and a zinc-dependent catalytic domain (CD). This constellation is found in the archetypal name-giving digestive enzyme astacin from the European crayfish Astacus astacus . Astacin catalytic domains span ∼200 residues and consist of two subdomains that flank an extended active-site cleft. They share several structural elements including a long zinc-binding consensus sequence (HEXXHXXGXXH) immediately followed by an EXXRXDRD motif, which features a family-specific glutamate. In addition, a downstream SIMHY-motif encompasses a “Met-turn” methionine and a zinc-binding tyrosine. The overall architecture and some structural features of astacin catalytic domains match those of other more distantly related MPs, which together constitute the metzincin clan of metallopeptidases. We further analysed the structures of PRO-, MAM, TRAF, CUB and EGF-like domains, and described their essential molecular determinants. In addition, we investigated the distribution of astacins across kingdoms and their phylogenetic origin. Through extensive sequence searches we found astacin CDs in > 25,000 sequences down the tree of life from humans beyond Metazoa, including Choanoflagellata, Filasterea and Ichtyosporea. We also found < 400 sequences scattered across non-holozoan eukaryotes including some fungi and one virus, as well as in selected taxa of archaea and bacteria that are pathogens or colonizers of animal hosts, but not in plants. Overall, we propose that astacins originate in the root of Holozoa consistent with Darwinian descent and that the latter genes might be the result of horizontal gene transfer from holozoan donors.
... However, in order to include the full set of 31 phyla considered for this study, other works that specifically addressed the placement of certain phyla were taken into account, which was the case for Kinorhyncha, Loricifera, and Nematomorpha (56), and for Dicyemida and Orthonectida (57). Certain nodes that are proposed only in new studies, supported by relatively low bootstrap values, or that are in contradiction between different analyses in the literature were highlighted as still disputed/problem atic (58)(59)(60)(61)(62)(63), as this tree is presented for guidance only. ...
Invertebrates constitute the majority of animal species on Earth, including most disease-causing agents or vectors, with more diverse viromes when compared to vertebrates. Recent advancements in high-throughput sequencing have significantly expanded our understanding of invertebrate viruses, yet this knowledge remains biased toward a few well-studied animal lineages. In this study, we analyze invertebrate DNA and RNA viromes for 31 phyla using 417 publicly available RNA-Seq data sets from diverse environments in the marine-terrestrial and marine-freshwater gradients. This study aims to (i) estimate virome compositions at the family level for the first time across the animal tree of life, including the first exploration of the virome in several phyla, (ii) quantify the diversity of invertebrate viromes and characterize the structure of invertebrate-virus infection networks, and (iii) investigate host phylum and habitat influence on virome differences. Results showed that a set of few viral families of eukaryotes, comprising Retroviridae, Flaviviridae, and several families of giant DNA viruses, were ubiquitous and highly abundant. Nevertheless, some differences emerged between phyla, revealing for instance a less diverse virome in Ctenophora compared to the other animal phyla. Compositional analysis of the viromes showed that the host phylum explained over five times more variance in composition than its habitat. Moreover, significant similarities were observed between the viromes of some phylogenetically related phyla, which could highlight the influence of co-evolution in shaping invertebrate viromes.
IMPORTANCE
This study significantly enhances our understanding of the global animal virome by characterizing the viromes of previously unexamined invertebrate lineages from a large number of animal phyla. It showcases the great diversity of viromes within each phylum and investigates the role of habitat shaping animal viral communities. Furthermore, our research identifies dominant virus families in invertebrates and distinguishes phyla with analogous viromes. This study sets the road toward a deeper understanding of the virome across the animal tree of life.
... This tentacular organ is regarded as a synapomorphy of three animal phyla, which are gathered into a united clade called Lophophorata: Phoronida, Bryozoa, and Brachiopoda (see review in . The monophyly of the lophophorates was supported by some authors (Jang & Hwang, 2009;Marlétaz et al., 2019;Nesnidal et al., 2013;Zverkov et al., 2019). The lophophore tentacles surround the mouth, but not the anus (Hyman, 1959) and hereby the lophophore differs from other tentacle organs; for example, from the entoprocts' tentacle apparatus, in which tentacles surround the anus as well. ...
Brachiopods have the most complex lophophore in comparison with other lophophorates, i.e., phoronids and bryozoans. However, at early ontogenetic stages, brachiopods have a lophophore of simple morphology, which consists of the oral tentacles. Data on the ultrastructure of the oral tentacles is mostly missing. Nonetheless, it has recently been suggested that the structure of oral tentacles is ancestral for all lophophorates in general, and for brachiopods in particular. The fine structure of the oral tentacles in the brachiopod Hemithiris psittacea is studied using light microscopy, transmission and scanning electron microscopy, cytochemistry and confocal laser scanning microscopy. The oral tentacles have a round shape in transverse section, and four ciliary zones, i.e., one frontal, two lateral, and one abfrontal. Latero‐frontal sensory cells occur among the frontal epithelium. Four basiepithelial nerves in the ciliary epithelium are colocalized with ciliary zones. Lophophores of simple morphology in phoronids and brachiopods are characterized by non‐specified round forms of tentacles. In phoronids and bryozoans, tentacles have additional latero‐frontal ciliary zones that function as a sieve during filtration. In most brachiopods, lateral cilia are involved in the capture of food particles, whereas latero‐frontal cells are retained in the frontal zone as sensory elements. The oral tentacles of H. psittacea contain a coelomic canal and have distinct frontal and abfrontal longitudinal muscles, which are separated from each other by peritoneal cells. A similar structure of tentacle muscles occurs in all bryozoans, whereas in phoronids, the frontal and abfrontal tentacle muscles are not separated by peritoneal cells. We suggest that the lophophorates' ancestor had tentacles, which were similar to the tentacles of some phoronids with lophophore of simple morphology. We also assume that the structure of the oral tentacles is ancestral for all brachiopods and the specialization of brachiopod tentacles correlates with the appearance of the double row of tentacles.
... While the absence of grasping spines in Amiskwia led to a rejection of a relationship with chaetognaths, recent studies demonstrated the presence of an internal jaw apparatus (11,12), similar to that possessed by gnathiferans. Recent molecular phylogenetic studies have found that chaetognaths and gnathiferans may form a clade (13)(14)(15), Chaetognathifera (16). The presence of a gnathostomulid-like jaw in Amiskwia has led to competing interpretations of its position in the tree of life, with the lateral fins representing either shared plesiomorphies (12,16) or convergences (11) with those in chaetognaths. ...
Chaetognaths, with their characteristic grasping spines, are the oldest known pelagic predators, found in the lowest Cambrian (Terreneuvian). Here, we describe a large stem chaetognath, Timorebestia koprii gen. et sp. nov., from the lower Cambrian Sirius Passet Lagerstätte, which exhibits lateral and caudal fins, a distinct head region with long antennae and a jaw apparatus similar to Amiskwia sagittiformis . Amiskwia has previously been interpreted as a total-group chaetognathiferan, as either a stem-chaetognath or gnathostomulid. We show that T. koprii shares a ventral ganglion with chaetognaths to the exclusion of other animal groups, firmly placing these fossils on the chaetognath stem. The large size (up to 30 cm) and gut contents in T. koprii suggest that early chaetognaths occupied a higher trophic position in pelagic food chains than today.
... Furthermore, the lack of Post1 and Scr genes in bryozoans is shared with phoronids which might indicate that these genes were lost in their last common ancestor. A sister-group relationship between phoronids and bryozoans has been proposed by several phylogenomic studies [35][36][37][38] and our finding regarding the shared missing Hox genes in bryozoans and phoronids are supportive of this hypothesis. However, a recent phylogenomic analyses recovered a sister group relationship between Kamptozoa (= Entoprocta) and Bryozoa, which contradicts the monophyly of Lophophorata [39]. ...
Background
Bryozoans are mostly sessile aquatic colonial invertebrates belonging to the clade Lophotrochozoa, which unites many protostome bilaterian phyla such as molluscs, annelids and brachiopods. While Hox and ParaHox genes have been extensively studied in various lophotrochozoan lineages, investigations on Hox and ParaHox gene complements in bryozoans are scarce.
Results
Herein, we present the most comprehensive survey of Hox and ParaHox gene complements in bryozoans using four genomes and 35 transcriptomes representing all bryozoan clades: Cheilostomata, Ctenostomata, Cyclostomata and Phylactolaemata. Using similarity searches, phylogenetic analyses and detailed manual curation, we have identified five Hox genes in bryozoans (pb, Dfd, Lox5, Lox4 and Post2) and one ParaHox gene (Cdx). Interestingly, we observed lineage-specific duplication of certain Hox and ParaHox genes (Dfd, Lox5 and Cdx) in some bryozoan lineages.
Conclusions
The bryozoan Hox cluster does not retain the ancestral lophotrochozoan condition but appears relatively simple (includes only five genes) and broken into two genomic regions, characterized by the loss and duplication of serval genes. Importantly, bryozoans share the lack of two Hox genes (Post1 and Scr) with their proposed sister-taxon, Phoronida, which suggests that those genes were missing in the most common ancestor of bryozoans and phoronids.
... A great deal is known about these genetic and developmental mechanisms in a few speci c vertebrate and ecdysozoan organisms (e.g., frog, mouse, chick, y, and C. elegans) (15)(16)(17)(18)). Yet little is understood about the genetic mechanisms underlying development in the third major clade of bilaterians, Spiralia (Lophotrophozoa sensu lato) (19)(20)(21). Beyond increasing taxonomic diversity to understand the evolution and development of phenotypes, Spiralia includes a great diversity of morphology and life histories within annelids, molluscs, brachiopods, rotifers, and platyhelminths (22). ...
Background: The evolution of centralized nervous systems (CNSs) a fascinating and complex topic; further work is needed to understand the genetic and developmental homology between organisms with a CNS. Research into a limited number of species suggests that CNSs may be homologous across Bilateria. This hypothesis is based in part on similar functions of BMP signaling in establishing fates along the dorsal-ventral (D-V) axis including limiting neural specification to one region of ectoderm. From an evolutionary-developmental perspective, the best way to understand a system is to explore it in a wide range of organisms to create a full picture.
Methods: Here we expand our understanding of BMP signaling in Spiralia, the third major clade of bilaterians, by examining phenotypes after expression of a dominant-negative BMP Receptor 1 and after knock-out of the putative BMP antagonist Chordin-like using CRISPR/Cas9 gene editing in the annelid Capitella teleta (Pleistoannelida).
Results: Ectopic expression of the dominant-negative Cte-BMPR1 did not increase CNS tissue or alter overall D-V axis formation in the trunk. Instead, we observed a unique asymmetric phenotype: a distinct loss of left tissues including the left eye, brain, foregut, and trunk mesoderm. Adding ectopic BMP4 early during cleavage stages reversed the dominant-negative Cte-BMPR1 phenotype, leading to a similar loss or reduction of right tissues instead. Surprisingly, a similar asymmetric loss of left tissues was evident from CRISPR knock-out of Cte-Chordin-like but concentrated in the trunk rather than the episphere.
Conclusions: We further solidify the hypothesis that the function of BMP signaling during establishment of the D-V axis and CNS is fundamentally different in at least Pleistoannelida, possibly in Spiralia, and is not required for nervous system delimitation in this group. Our results support hypotheses of either multiple evolutionary origins of CNSs across Bilateria or divergence in the molecular mechanisms of CNS specification and D-V axis formation in annelids.
... We identified orthologous sequences using a conservative bioinformatic pipeline and conducted phylogenomic analyses on five datasets including a 92% occupancy (i.e., each gene was sampled for at least 92% of the taxa) supermatrix composed of 663 genes (92_pct), a 75% occupancy supermatrix (3,825 genes; 75_pct), a 50% occupancy supermatrix (6,430 genes; 50_pct), and matrices with the top 250 and top 500 genes scored by genesortR (24). Analyses were performed under multiple concatenation-based and coalescent-aware methodologies, with the former analyzed using both maximum likelihood (ML) and Bayesian inference (BI) implementations of site-heterogeneous and site-homogeneous models, as these approaches are known to differ in their susceptibility to model violations (25,26). Results were remarkably stable across all datasets and methods, with all nodes identically resolved, including support for placement of The Diasoma-Cyrtosoma hypothesis proposing a Scaphopoda-Bivalvia (Diasoma) clade, which is supported by similarities in the weakly developed head, pedal morphology, formation of the mantle and shell, and lateral compression of the body with the Paleozoic group Rostroconchia (13)(14)(15)(16). ...
The almost simultaneous emergence of major animal phyla during the early Cambrian shaped modern animal biodiversity. Reconstructing evolutionary relationships among such closely spaced branches in the animal tree of life has proven to be a major challenge, hindering understanding of early animal evolution and the fossil record. This is particularly true in the species-rich and highly varied Mollusca where dramatic inconsistency among paleontological, morphological, and molecular evidence has led to a long-standing debate about the group’s phylogeny and the nature of dozens of enigmatic fossil taxa. A critical step needed to overcome this issue is to supplement available genomic data, which is plentiful for well-studied lineages, with genomes from rare but key lineages, such as Scaphopoda. Here, by presenting chromosome-level genomes from both extant scaphopod orders and leveraging complete genomes spanning Mollusca, we provide strong support for Scaphopoda as the sister taxon of Bivalvia, revitalizing the morphology-based Diasoma hypothesis originally proposed 50 years ago. Our molecular clock analysis confidently dates the split between Bivalvia and Scaphopoda at ~520 Ma, prompting a reinterpretation of controversial laterally compressed Early Cambrian fossils, including Anabarella , Watsonella, and Mellopegma, as stem diasomes. Moreover, we show that incongruence in the phylogenetic placement of Scaphopoda in previous phylogenomic studies was due to ancient incomplete lineage sorting (ILS) that occurred during the rapid radiation of Conchifera. Our findings highlight the need to consider ILS as a potential source of error in deep phylogeny reconstruction, especially in the context of the unique nature of the Cambrian Explosion.
... The distribution of trochophore larvae in the animal tree of life shows a strong correlation with phylogenetic position. Within Bilateria, the animals showing bilateral body plans and constituting the majority of animals since the Cambrian period, three major clades have been firmly recognized, namely, Spiralia, Ecdysozoa, and Deuterostomia (Laumer et al., 2019;Marlétaz et al., 2019). Trochophore larvae are widely and exclusively distributed in spiralian lineages, including Mollusca, Annelida, Entoprocta, and likely Nemertea (Maslakova et al., 2004;Nielsen, 2018) (Figure 1a). ...
The body patterning of trochophore larvae is important for understanding spiralian evolution and the origin of the bilateral body plan. However, considerable variations are observed among spiralian lineages, which have adopted varied strategies to develop trochophore larvae or even omit a trochophore stage. Some spiralians, such as patellogastropod mollusks, are suggested to exhibit ancestral traits by producing equal-cleaving fertilized eggs and possessing "typical" trochophore larvae. In recent years, we developed a potential model system using the patellogastropod Lottia peitaihoensis (= Lottia goshimai). Here, we introduce how the species were selected and establish sources and techniques, including gene knockdown, ectopic gene expression, and genome editing. Investigations on this species reveal essential aspects of trochophore body patterning, including organizer signaling, molecular and cellular processes connecting the various developmental functions of the organizer, the specification and behaviors of the endomeso-derm and ectomesoderm, and the characteristic dorsoventral decoupling of Hox expression. These findings enrich the knowledge of trochophore body patterning and have important implications regarding the evolution of spiralians as well as bilateral body plans. K E Y W O R D S Lottia, spiralia, trochophore
... We identified orthologous gene families among our metazoan proteomes using the OrthoFinder package (Emms and Kelly, 2019) with default parameters and a user generated species tree as input (Additional File 11). Our user generated species tree topology was based on recent metazoan phylogenies [110][111][112][113], the MolluscDB phylogeny provided on the website [47], and recent Mollusca [114] and nudibranch [115][116][117] phylogenetic analyses. We then analyzed orthologous groups using the program KinFin v1.0.3 [118] to determine which predicted genes in Berghia stephanieae are clade-specific (meaning that they only cluster with sequences from a particular clade). ...
How novel phenotypes originate from conserved genes, processes, and tissues remains a major question in biology. Research that sets out to answer this question often focuses on the conserved genes and processes involved, an approach that explicitly excludes the impact of genetic elements that may be classified as clade-specific, even though many of these genes are known to be important for novel, or clade-restricted, phenotypes. This is especially true for understudied phyla such as mollusks, where limited genomic and functional biology resources for members of this phylum has long hindered assessments of genetic homology and function. To address this gap, we constructed a chromosome-level genome for the gastropod Berghia stephanieae (Valdés, 2005) to investigate the expression of clade-specific genes across both novel and conserved tissue types in this species. The final assembled and filtered Berghia genome is comparable to other high quality mollusk genomes in terms of size (1.05 Gb) and number of predicted genes (24,960 genes), and is highly contiguous. The proportion of upregulated, clade-specific genes varied across tissues, but with no clear trend between the proportion of clade-specific genes and the novelty of the tissue. However, more complex tissue like the brain had the highest total number of upregulated, clade-specific genes, though the ratio of upregulated clade-specific genes to the total number of upregulated genes was low. Our results, when combined with previous research on the impact of novel genes on phenotypic evolution, highlight the fact that the complexity of the novel tissue or behavior, the type of novelty, and the developmental timing of evolutionary modifications will all influence how novel and conserved genes interact to generate diversity.
... Rotifers are microscopic invertebrates found globally in aquatic habitats; at times, they are the most abundant animals in some freshwater ecosystems. Rotifers comprise a phylum within the protostome clade Gnathifera [1,2] and are by far the most experimentally tractable of this early branching example of metazoan evolution [3]. Monogonont rotifers, one of the 2 major groups of Rotifera, have long been used in studies of evolution, limnology, ecology, and toxicology [4,5]. ...
Rotifers have been studied in the laboratory and field for over 100 years in investigations of microevolution, ecological dynamics, and ecotoxicology. In recent years, rotifers have emerged as a model system for modern studies of the molecular mechanisms of genome evolution, development, DNA repair, aging, life history strategy, and desiccation tolerance. However, a lack of gene editing tools and transgenic strains has limited the ability to link genotype to phenotype and dissect molecular mechanisms. To facilitate genetic manipulation and the creation of reporter lines in rotifers, we developed a protocol for highly efficient, transgenerational, CRISPR-mediated gene editing in the monogonont rotifer Brachionus manjavacas by microinjection of Cas9 protein and synthetic single-guide RNA into the vitellaria of young amictic (asexual) females. To demonstrate the efficacy of the method, we created knockout mutants of the developmental gene vasa and the DNA mismatch repair gene mlh3. More than half of mothers survived injection and produced offspring. Genotyping these offspring and successive generations revealed that most carried at least 1 CRISPR-induced mutation, with many apparently mutated at both alleles. In addition, we achieved precise CRISPR-mediated knock-in of a stop codon cassette in the mlh3 locus, with half of injected mothers producing F2 offspring with an insertion of the cassette. Thus, this protocol produces knockout and knock-in CRISPR/Cas9 editing with high efficiency, to further advance rotifers as a model system for biological discovery.
... Morphological and molecular evidence support the position of Priapulida within the clade Ecdysozoa (Giribet and Edgecombe 2020). Although further genomic data from these groups are necessary to robustly test their relationships, recent phylogenomic analyses have placed priapulans as the putative sister group to all other ecdysozoans (Laumer et al. 2019;Marlétaz et al. 2019). Due to this phylogenetic position, increasing the genomic data available for priapulans can help us to better understand the evolution of key gene families associated with the ecdysozoan success, often attributed to their ability to molt. ...
Genomic data for priapulans are limited to a single species, restricting broad comparative analyses, and thorough interrogation of questions spanning phylogenomics, ecdysozoan physiology, and development. To help fill this void, we present here a high-quality priapulan genome for the meiofaunal species Tubiluchus corallicola. Our assembly combines Nanopore and Illumina sequencing technologies, and makes use of a whole genome amplification, to generate enough DNA to sequence this small meiofaunal species. We generated a moderately contiguous assembly (2547 scaffolds), with a high level of completeness (Metazoan BUSCOs n = 954, single copy complete = 89.6%, duplicated = 3.9%, fragmented = 3.5%, missing = 3.0%). We then screened the genome for homologs of the Halloween genes, key genes implicated in the ecdysis (moulting) pathway of arthropods, recovering a putative homolog of shadow. The presence of a shadow orthologue in two priapulan genomes suggests that the Halloween genes may not have evolved in a stepwise manner in Panarthropoda, as previously thought, but may have a deeper origin at the base of Ecdysozoa.
... This assumes that, though there may be a great diversity of sites within the alignment, these will be relatively functionally conserved as the site still corresponds to a similar functional purpose. 6-state Dayhoff recoding is the most used of these recoding tables (Feuda et al., 2017;Foster et al., 2009Foster et al., , 2022Hernandez & Ryan, 2019;Marlétaz et al., 2019), compressing the 20-state amino acid alphabet into 6, but 2-state nucleotide recoding is also used. 2-state nucleotide recoding regards only transversions, changes between purines and pyrimidines, to be relevant (Smith et al., 2009). ...
The availability of phylogenetic data has greatly expanded in recent years. As a result, a new era in phylogenetic analysis is dawning—one in which the methods we use to analyse and assess our data are the bottleneck to producing valuable phylogenetic hypotheses, rather than the need to acquire more data. This makes the ability to accurately appraise and evaluate new methods of phylogenetic analysis and phylogenetic artefact identification more important than ever. Incongruence in phylogenetic reconstructions based on different datasets may be due to two major sources: biological and methodological. Biological sources comprise processes like horizontal gene transfer, hybridization and incomplete lineage sorting, while methodological ones contain falsely assigned data or violations of the assumptions of the underlying model. While the former provides interesting insights into the evolutionary history of the investigated groups, the latter should be avoided or minimized as best as possible. However, errors introduced by methodology must first be excluded or minimized to be able to conclude that biological sources are the cause. Fortunately, a variety of useful tools exist to help detect such misassignments and model violations and to apply ameliorating measurements. Still, the number of methods and their theoretical underpinning can be overwhelming and opaque. Here, we present a practical and comprehensive review of recent developments in techniques to detect artefacts arising from model violations and poorly assigned data. The advantages and disadvantages of the different methods to detect such misleading signals in phylogenetic reconstructions are also discussed. As there is no one‐size‐fits‐all solution, this review can serve as a guide in choosing the most appropriate detection methods depending on both the actual dataset and the computational power available to the researcher. Ultimately, this informed selection will have a positive impact on the broader field, allowing us to better understand the evolutionary history of the group of interest.
... However, other phylogenomic studies identified XAN as sister to the Ambulacraria (echinoderms and hemichordates; Xenambulacraria hypothesis), which sparked the War of the Worms (Telford and Copley, 2016). These studies recover (1) Xenambulacraria as sister to chordates, together forming the deuterostomes or, intriguingly, (2) Xenambulacraria as sister to all the other bilaterians, formed by chordates as sister to protostomes (Marlétaz, 2019;Marlétaz et al., 2019;Philippe et al., 2019). The later would effectively mean the demise of deuterostomes, although these topologies usually show low statistical support. ...
The Animal Kingdom is an astonishingly diverse group. Together with plants and fungi is one of the three major lineages of multicellular eukaryotes. Due to anthropocentrism and/or genuine scientific interest, their origin and diversification are pivotal to modern evolutionary biology. In the last few decades, dramatic technological advances in molecular biology and computational power have generated new phylogenetic proposals, as well as new tools to compare genomes or study cell type evolution. These new approaches complement the insights from fields such as comparative morphology, evodevo, or palaeontology, which all together provide an integrative view of animal evolution, including major evolutionary transitions such as the origin of animals or the emergence of animals with bilateral symmetry. In this paper, we review recent developments in animal phylogenetics, comparative genomics, and cell type evolution related to these two transitions, and we compare animals to another major lineage of multicellular eukaryotes, plants.
... For example, several crucial phylogenetic relationships among higher taxonomic levels of deuterostome are now consistently embedded in the literature, with echinoderms and hemichordates grouped in a clade named ambulacraria and cephalochordate being the sister group to the olfactories (tunicates and vertebrates), (e.g., Swalla and Smith, 2008;Peterson and Eernisse, 2016;Nanglu et al., 2023). In protostome, we are now more confident about the phylogenetic position not only of the common animals such as arthropods, molluscs, annelids, brachiopods, and bryozoans but of some small cryptic groups such as chaetognaths, gnathifers, tardigrades, xenoturbellids, aceolomorphs (Laumer et al., 2019), albeit final agreements are far from being reached about the inner relationship in either ecdysozoans (e.g., Giribet and Edgecombe, 2017) or lophotrochozoans (e.g., Kocot, 2016;Marlétaz et al., 2019). ...
Genic and genomic data have been reshaping our understanding of the earliest radiation event of metazoans, the well-known Cambrian Evolutionary Radiation, not only from the respects of reshuffling the phylogenetic topologies of some animal phyla but by deciphering the deep homologies of many morphological features. These advances, together with the continuing discoveries of the Ediacaran-Cambrian fossils, are unveiling the cladogenetic process of the early metazoans and the patterns of morphologic evolution during this biological radiation event. In this review, I focus on a small but challenging field, the problematic fossils from the early Cambrian fossil Lagerstätten, such as the Chengjiang biota, mainly on the controversies concerning their interpretation and the consequent impacts on understanding the early evolution of animals. The bizarre body plans of the early Cambrian problematica alone do not account for the difficulties in studying their biology and affinity. Instead, it is the combined action of the taphonomic artifacts and the uncertainty in homologizing the preserved characters that impede generating plausible interpretations. Despite all these issues, a testable and repeatable method for interpreting fossils has emerged and is becoming more practicable. The integration of an evolutionary-grade conceptual frame is beneficial to the interpretation of the Cambrian problematic fossils. Together with the focus on taphonomic alternation and homologic assessment, the Cambrian problematic fossils are becoming more informative nodes in the “parsing tree” of early animal evolution.
... Selected transcriptomes were assembled using Trinity and translated using transdecoder (v5.5.0) (Haas et al., 2008). We built HMM profiles using Hmmer (v3.1b2) for each orthologue family and extracted orthologues for phylogenetic reconstruction using the same approach as in (Marlétaz et al., 2019). Subsequent sequences were aligned using Msaprobs (Liu et al., 2010), mistranslated stretches filtered out using HmmCleaner (Di Franco et al., 2019) and diverging regions intractable for phylogenetic analysis removed using BMGE (-g 0.9) (Criscuolo and Gribaldo, 2010). ...
As the only surviving lineages of jawless fishes, hagfishes and lampreys provide a critical window into early vertebrate evolution. Here, we investigate the complex history, timing, and functional role of genome-wide duplications in vertebrates in the light of a chromosome-scale genome of the brown hagfish Eptatretus atami. Using robust chromosome-scale (paralogon-based) phylogenetic methods, we confirm the monophyly of cyclostomes, document an auto-tetraploidization (1RV) that predated the origin of crown group vertebrates ~517 Mya, and establish the timing of subsequent independent duplications in the gnathostome and cyclostome lineages. Some 1RV gene duplications can be linked to key vertebrate innovations, suggesting that this early genomewide event contributed to the emergence of pan-vertebrate features such as neural crest. The hagfish karyotype is derived by numerous fusions relative to the ancestral cyclostome arrangement preserved by lampreys. These genomic changes were accompanied by the loss of genes essential for organ systems (eyes, osteoclast) that are absent in hagfish, accounting in part for the simplification of the hagfish body plan; other gene family expansions account for hagfishes' capacity to produce slime. Finally, we characterise programmed DNA elimination in somatic cells of hagfish, identifying protein-coding and repetitive elements that are deleted during development. As in lampreys, the elimination of these genes provides a mechanism for resolving genetic conflict between soma and germline by repressing germline/pluripotency functions. Reconstruction of the early genomic history of vertebrates provides a framework for further exploration of vertebrate novelties.
... We performed gene family reconstruction using OMA (v.2.4.1) 81 between selected vertebrate species to identify single-copy orthologues. These orthologues were used to infer gene phylogeny after processing as described previously 82 : HMM profiles were built for each orthologous gene family and searched against translated transcriptomes using the HMMer tool (v.3.1b2) 83 . ...
Skates are cartilaginous fish whose body plan features enlarged wing-like pectoral fins, enabling them to thrive in benthic environments1,2. However, the molecular underpinnings of this unique trait remain unclear. Here we investigate the origin of this phenotypic innovation by developing the little skate Leucoraja erinacea as a genomically enabled model. Analysis of a high-quality chromosome-scale genome sequence for the little skate shows that it preserves many ancestral jawed vertebrate features compared with other sequenced genomes, including numerous ancient microchromosomes. Combining genome comparisons with extensive regulatory datasets in developing fins—including gene expression, chromatin occupancy and three-dimensional conformation—we find skate-specific genomic rearrangements that alter the three-dimensional regulatory landscape of genes that are involved in the planar cell polarity pathway. Functional inhibition of planar cell polarity signalling resulted in a reduction in anterior fin size, confirming that this pathway is a major contributor to batoid fin morphology. We also identified a fin-specific enhancer that interacts with several hoxa genes, consistent with the redeployment of hox gene expression in anterior pectoral fins, and confirmed its potential to activate transcription in the anterior fin using zebrafish reporter assays. Our findings underscore the central role of genome reorganization and regulatory variation in the evolution of phenotypes, shedding light on the molecular origin of an enigmatic trait.
... However, to reconstruct the developmental mode of the ancestral flatworm, and inform on homology of polyclad and trochophore larvae, future phylogenetic and ancestral state reconstruction analyses would require the inclusion of many more non-polyclad flatworms and multiple species from sister clades to the flatworms (e.g. nemerteans and annelids [2,45,46]). At present, transcriptomic data for an increasing diversity of these taxa is available [47,48], but data on mode of development for many of these species is lacking. ...
Platyhelminthes (flatworms) are a diverse invertebrate phylum useful for exploring life-history evolution. Within Platyhelminthes, only two clades develop through a larval stage: free-living polyclads and parasitic neodermatans. Neodermatan larvae are considered evolutionarily derived, whereas polyclad larvae are hypothesized to be ancestral due to ciliary band similarities among polyclad and other spiralian larvae. However, larval evolution has been challenging to investigate within polyclads due to low support for deeper phylogenetic relationships. To investigate polyclad life-history evolution, we generated transcriptomic data for 21 species of polyclads to build a well-supported phylogeny for the group. The resulting tree provides strong support for deeper nodes, and we recover a new monophyletic clade of early branching cotyleans. We then used ancestral state reconstructions to investigate ancestral modes of development within Polycladida and more broadly within flatworms. In polyclads, we were unable to reconstruct the ancestral state of deeper nodes with significant support because early branching clades show diverse modes of development. This suggests a complex history of larval evolution in polyclads that likely includes multiple losses and/or multiple gains. However, our ancestral state reconstruction across a previously published platyhelminth phylogeny supports a direct developing prorhynchid/polyclad ancestor, which suggests that a larval stage in the life cycle evolved along the polyclad stem lineage or within polyclads.
... Lophotrochozoa is one of the largest metazoan clades and includes annelids, molluscs, nemerteans, bryozoans, brachiopods, and platyhelminths (Luo et al. 2018;Marlétaz et al. 2019). Species of this group occupy many ecological niches in aquatic and terrestrial environments and exhibit diverse photoreceptive visual structures, from simple pigmented areas to sophisticated camera-type eyes (Serb and Eernisse 2008;Bok et al. 2017;Rawlinson et al. 2019). ...
Opsins are G-coupled receptors playing a key role in metazoan visual processes. While many studies enriched our understanding of opsin diversity in several animal clades, the opsin evolution in Lophotrochozoa, one of the major metazoan groups, remains poorly understood. Using recently developed phylogenetic approaches, we investigated the opsin evolution in 74 lophotrochozoan genomes. We found that the common ancestor of Lophotrochozoa possessed at least seven opsin paralog groups that underwent divergent evolutionary history in the different phyla. Furthermore, we showed for the first time opsin-related molecules in Bilateria that we named pseudopsins, which may prove critical in uncovering opsin evolution.
The marine annelid Platynereis dumerilii is a model organism used in many research areas including evolution and development, neurobiology, ecology and regeneration. Here we present the genomes of P. dumerilii and of the closely related P. massiliensis and P. megalops, to facilitate comparative genomic approaches and help explore Platynereis biology. We used long-read sequencing technology and chromosomal-conformation capture along with extensive transcriptomic resources to obtain and annotate a draft genome assembly of ~1.47 Gbp for P. dumerilii, of which more than half represent repeat elements. We predict around 29,000 protein-coding genes, with relatively large intron sizes, over 38,000 non-coding genes, and 580 miRNA loci. We further explore the high genetic variation (~3% heterozygosity) within the Platynereis species complex. Gene ontology reveals the most variable loci to be associated with pigmentation, development and immunity. The current work sets the stage for further development of Platynereis genomic resources.
We present the counterpart to Chap. 4 in Chap. 5, where we focus on the role of “permanent” exceptions and their importance in scientific knowledge and biology. These exceptions can be analyzed at any scale and are often not appreciated beyond anecdotal mentions. They can be aberrant or teratological groups, taxonomic groups unique for some features of their biology, “intermediate exceptions” with minorities of species or clades with certain characteristics compared to the majority, or organisms with exceptional distributions in space or time. In this chapter, we reviewed groups that have been difficult to classify because of their rarity, rarities that can be used for human benefit, exceptional groups that make us rethink phylogenetic relationships, rare and incredible biological phenomena, and evolutionary and ecological aspects of rare species. Our goal is to vindicate rarities and minorities, to highlight their importance for the understanding of evolution, and to begin to make these cases visible and treasured in the teaching of biology.
In animals, three main RNA interference mechanisms have been described so far, which respectively maturate three types of small noncoding RNAs (sncRNAs): miRNAs, piRNAs and endo-siRNAs. The diversification of these mechanisms is deeply linked with the evolution of the Argonaute gene superfamily since each type of sncRNA is typically loaded by a specific Argonaute homolog. Moreover, other protein families play pivotal roles in the maturation of sncRNAs, like the DICER ribonuclease family, whose DICER1 and DICER2 paralogs maturate respectively miRNAs and endo-siRNAs. Within Metazoa, the distribution of these families has been only studied in major groups, and there are very few data for clades like Lophotrochozoa. Thus, we here inferred the evolutionary history of the animal Argonaute and DICER families including 43 lophotrochozoan species. Phylogenetic analyses along with newly sequenced sncRNA libraries suggested that in all Trochozoa the proteins related to the endo-siRNA pathway have been lost, a part of them in some phyla (i.e., Nemertea, Bryozoa, Entoprocta), while all of them in all the others. On the contrary, early diverging phyla, Platyhelminthes and Syndermata, showed a complete endo-siRNA pathway. On the other hand, miRNAs were revealed the most conserved and ubiquitous mechanism of the metazoan RNA interference machinery, confirming their pivotal role in animal cell regulation.
Genome-wide information has so far been unavailable for ribbon worms of the clade Hoplonemertea, the most species-rich class within the phylum Nemertea. While species within Pilidiophora, the sister clade of Hoplonemertea, possess a pilidium larval stage and lack stylets on their proboscis, Hoplonemertea species have a planuliform larva and are armed with stylets employed for the injection of toxins into their prey. To further compare these developmental, physiological, and behavioral differences from a genomic perspective, the availability of a reference genome of a Hoplonemertea species is crucial. To this end, we herein present the annotated chromosome-level genome assembly for Emplectonema gracile (Nemertea; Hoplonemertea; Monostilifera; Emplectonematidae), an easily collected nemertean well-suited for laboratory experimentation. The genome is 157.9 Mbp in span. Hi-C scaffolding yielded 15 putative chromosomes with a scaffold N50 of 10.0 Mbp and a BUSCO completeness score of 95.3%. Structural annotation predicted 20,684 protein-coding genes. The high-quality reference genome reaches an Earth BioGenome standard level of 7.C.Q50. These data will be highly useful for future investigations towards a better understanding of the evolution, development, morphology, and toxicology of Nemertea.
Significance
The genome of Emplectonema gracile is highly contiguous, well annotated, and shorter than those of the other two ribbon worm species sequenced to date. This genome is a valuable resource for studies on molecular ecology, venom evolution, and regeneration in marine invertebrates.
Acoel flatworms possess epidermal sensory-receptor cells on their body surfaces and exhibit behavioral repertoires such as geotaxis and phototaxis. Acoel epidermal sensory receptors should be mechanical and/or chemical receptors; however, the mechanisms of their sensory reception have not been elucidated. We examined the three-dimensional relationship between epidermal sensory receptors and their innervation in an acoel flatworm, Praesagittifera naikaiensis. The distribution of the sensory receptors was different between the ventral and dorsal sides of worms. The nervous system was mainly composed of a peripheral nerve net, an anterior brain, and three pairs of longitudinal nerve cords. The nerve net was located closer to the body surface than the brain and the nerve cords. The sensory receptors have neural connections with the nerve net in the entire body of worms. We identified five homologs of polycystic kidney disease (PKD): PKD1-1, PKD1-2, PKD1-3, PKD1-4, and, PKD2, from the P. naikaiensis genome. All of these PKD genes were implied to be expressed in the epidermal sensory receptors of P. naikaiensis. PKD1-1 and PKD2 were dispersed across the entire body of worms. PKD1-2, PKD1-3, and PKD1-4 were expressed in the anterior region of worms. PKD1-4 was also expressed around the mouth opening. Our results indicated that P. naikaiensis possessed several types of epidermal sensory receptors to convert various environmental stimuli into electrical signals via the PKD channels and transmit the signals to afferent nerve and/or effector cells.
As the only surviving lineages of jawless fishes, hagfishes and lampreys provide a crucial window into early vertebrate evolution1–3. Here we investigate the complex history, timing and functional role of genome-wide duplications4–7 and programmed DNA elimination8,9 in vertebrates in the light of a chromosome-scale genome sequence for the brown hagfish Eptatretus atami. Combining evidence from syntenic and phylogenetic analyses, we establish a comprehensive picture of vertebrate genome evolution, including an auto-tetraploidization (1RV) that predates the early Cambrian cyclostome–gnathostome split, followed by a mid–late Cambrian allo-tetraploidization (2RJV) in gnathostomes and a prolonged Cambrian–Ordovician hexaploidization (2RCY) in cyclostomes. Subsequently, hagfishes underwent extensive genomic changes, with chromosomal fusions accompanied by the loss of genes that are essential for organ systems (for example, genes involved in the development of eyes and in the proliferation of osteoclasts); these changes account, in part, for the simplification of the hagfish body plan1,2. Finally, we characterize programmed DNA elimination in hagfish, identifying protein-coding genes and repetitive elements that are deleted from somatic cell lineages during early development. The elimination of these germline-specific genes provides a mechanism for resolving genetic conflict between soma and germline by repressing germline and pluripotency functions, paralleling findings in lampreys10,11. Reconstruction of the early genomic history of vertebrates provides a framework for further investigations of the evolution of cyclostomes and jawed vertebrates.
Background
Obtaining adequate DNA for long-read genome sequencing remains a roadblock to producing contiguous genomes from small-bodied organisms. Multiple displacement amplification (MDA) leverages Phi29 DNA polymerase to produce micrograms of DNA from picograms of input. Few genomes have been generated using this approach, due to concerns over biases in amplification related to GC and repeat content and chimera production. Here, we explored the utility of MDA for generating template DNA for PacBio HiFi sequencing using Caenorhabditis elegans (Nematoda) and Lepidodermella squamata (Gastrotricha).
Results
HiFi sequencing of libraries prepared from MDA DNA produced highly contiguous and complete genomes for both C. elegans (102 Mbp assembly; 336 contigs; N50 = 868 Kbp; L50 = 39; BUSCO_nematoda: S:92.2%, D:2.7%) and L. squamata (122 Mbp assembly; 157 contigs; N50 = 3.9 Mb; L50 = 13; BUSCO_metazoa: S: 78.0%, D: 2.8%). Amplified C. elegans reads mapped to the reference genome with a rate of 99.92% and coverage of 99.75% with just one read (of 708,811) inferred to be chimeric. Coverage uniformity was nearly identical for reads from MDA DNA and reads from pooled worm DNA when mapped to the reference genome. The genome of Lepidodermella squamata , the first of its phylum, was leveraged to infer the phylogenetic position of Gastrotricha, which has long been debated, as the sister taxon of Platyhelminthes.
Conclusions
This methodology will help generate contiguous genomes of microscopic taxa whose body size precludes standard long-read sequencing. L. squamata is an emerging model in evolutionary developmental biology and this genome will facilitate further work on this species.
The clade Syndermata includes the endoparasitic Acanthocephala, the epibiotic Seisonidea, and the free-living Bdelloidea and Monogononta. The phylogeny of Syndermata is highly debated, hindering the understanding of the evolution of morphological features, reproductive modes, and lifestyles within the group. Here, we use publicly available whole-genome data to re-evaluate syndermatan phylogeny and assess the credibility of alternative hypotheses, using a new combination of phylogenomic methods. We found that the Hemirotifera and Pararotatoria hypotheses were recovered under combinations of datasets and methods with reduced possibility of systematic error in concatenation-based analyses. In contrast, the Seisonidea-sister and Lemniscea hypotheses were recovered under dataset combinations with increased possibility of systematic error. Hemirotifera was further supported by whole-genome microsynteny analyses and species-tree methods that use multi-copy orthogroups after removing distantly related outgroups. Pararotatoria was only partially supported by microsynteny-based phylogenomic reconstructions. Hence, Hemirotifera and partially Pararotatoria were supported by independent phylogenetic methods and data-evaluation approaches. These two hypotheses have important implications for the evolution of syndermatan morphological features, such as the gradual reduction of locomotory ciliation from the common ancestor of Syndermata in the stem lineage of Pararotatoria. Our study illustrates the importance of combining various types of evidence to resolve difficult phylogenetic questions.
Background
How novel phenotypes originate from conserved genes, processes, and tissues remains a major question in biology. Research that sets out to answer this question often focuses on the conserved genes and processes involved, an approach that explicitly excludes the impact of genetic elements that may be classified as clade-specific, even though many of these genes are known to be important for many novel, or clade-restricted, phenotypes. This is especially true for understudied phyla such as mollusks, where limited genomic and functional biology resources for members of this phylum have long hindered assessments of genetic homology and function. To address this gap, we constructed a chromosome-level genome for the gastropod Berghia stephanieae (Valdés, 2005) to investigate the expression of clade-specific genes across both novel and conserved tissue types in this species.
Results
The final assembled and filtered Berghia genome is comparable to other high-quality mollusk genomes in terms of size (1.05 Gb) and number of predicted genes (24,960 genes) and is highly contiguous. The proportion of upregulated, clade-specific genes varied across tissues, but with no clear trend between the proportion of clade-specific genes and the novelty of the tissue. However, more complex tissue like the brain had the highest total number of upregulated, clade-specific genes, though the ratio of upregulated clade-specific genes to the total number of upregulated genes was low.
Conclusions
Our results, when combined with previous research on the impact of novel genes on phenotypic evolution, highlight the fact that the complexity of the novel tissue or behavior, the type of novelty, and the developmental timing of evolutionary modifications will all influence how novel and conserved genes interact to generate diversity.
Tissue inhibitors of metalloproteinase (TIMPs) play a pivotal role in regulating extracellular matrix (ECM) dynamics and have been extensively studied in vertebrates. However, understanding their evolution across invertebrate phyla is limited. Utilizing the high-quality Pteria penguin genome, we conducted phylogenomic orthology analyses across metazoans, revealing the emergence and distribution of the TIMP gene family. Our findings show that TIMP repertoires originated during eumetazoan radiation, experiencing independent duplication events in different clades, resulting in varied family sizes. Particularly, Pteriomorphia bivalves within Mollusca exhibited the most significant expansion and displayed the most diverse TIMP repertoires among metazoans. These expansions were attributed to multiple gene duplication events, potentially driven by the demands for functional diversification related to multiple adaptive traits, contributing to the adaptation of Pteriomorphia bivalves as stationary filter feeders. In this context, Pteriomorphia bivalves offer a promising model for studying invertebrate TIMP evolution.
Background Unlike most free-living platyhelminths, catenulids, the sister group to all remaining flatworms,
do not have eyes. Instead, the most prominent sensory structures in their heads are statocysts or sensory pits. The latter, found in the family Stenostomidae, are concave depressions located laterally on the head that represent one of the taxonomically important traits of the family. In the past, the sensory pits of flatworms have been homologized with the cephalic organs of nemerteans, a clade that occupies a sister position to platyhelminths in some recent phy- logenies. To test for this homology, we studied morphology and gene expression in the sensory pits of the catenulid Stenostomum brevipharyngium.
Results We used confocal and electron microscopy to investigate the detailed morphology of the sensory
pits, as well as their formation during regeneration and asexual reproduction. The most prevalent cell type
within the organ is epidermally-derived neuron-like cells that have cell bodies embedded deeply in the brain
lobes and long neurite-like processes extending to the bottom of the pit. Those elongated processes are adorned with extensive microvillar projections that fill up the cavity of the pit, but cilia are not associated with the sensory pit. We also studied the expression patterns of some of the transcription factors expressed in the nemertean cephalic organs during the development of the pits. Only a single gene, pax4/6, is expressed in both the cerebral organs
of nemerteans and sensory pits of S. brevipharyngium, challenging the idea of their deep homology.
Conclusions Since there is no morphological or molecular correspondence between the sensory pits of Stenosto- mum and the cerebral organs of nemerteans, we reject their homology. Interestingly, the major cell type contribut- ing to the sensory pits of stenostomids shows ultrastructural similarities to the rhabdomeric photoreceptors of other flatworms and expresses ortholog of the gene pax4/6, the pan-bilaterian master regulator of eye development. We suggest that the sensory pits of stenostomids might have evolved from the ancestral rhabdomeric photoreceptors that lost their photosensitivity and evolved secondary function. The mapping of head sensory structures on plathel- minth phylogeny indicates that sensory pit-like organs evolved many times independently in flatworms.
Many animals undergo indirect development, where their embryogenesis produces an intermediate life stage, or larva, that is often free‐living and later metamorphoses into an adult. As their adult counterparts, larvae can have unique and diverse morphologies and occupy various ecological niches. Given their broad phylogenetic distribution, larvae have been central to hypotheses about animal evolution. However, the evolution of these intermediate forms and the developmental mechanisms diversifying animal life cycles are still debated. This review focuses on Spiralia, a large and diverse clade of bilaterally symmetrical animals with a fascinating array of larval forms, most notably the archetypical trochophore larva. We explore how classic research and modern advances have improved our understanding of spiralian larvae, their development, and evolution. Specifically, we examine three morphological features of spiralian larvae: the anterior neural system, the ciliary bands, and the posterior hyposphere. The combination of molecular and developmental evidence with modern high‐throughput techniques, such as comparative genomics, single‐cell transcriptomics, and epigenomics, is a promising strategy that will lead to new testable hypotheses about the mechanisms behind the evolution of larvae and life cycles in Spiralia and animals in general. We predict that the increasing number of available genomes for Spiralia and the optimization of genome‐wide and single‐cell approaches will unlock the study of many emerging spiralian taxa, transforming our views of the evolution of this animal group and their larvae.
Meiofauna-a collective term to define microscopic animals-represent a numerically important component of biodiversity in most of Earth's ecosystems and play a crucial role in biogeochemical cycles. Meiofauna have also been used as models to understand fundamental adaptive processes, have contributed to a better understanding of the animal's Tree of Life, and are believed to be a treasure trove for future genomic studies. To celebrate the diversity of research topics brought to us by the term "meiofauna", we gathered a multidisciplinary team of 42 ecologists, taxonomists, morphologists, biogeographers, molecular biologists, and scientific disseminators to list 194 fundamental questions in meiofaunal research. Then, through an online survey, 251 scientists, administrators, students, and stakeholders assisted us in reducing this list to 50 top-priority questions. Applied topics related to anthropogenic impact and climate change received the highest scores, whereas questions related to areas in development such as genomics or adaptations, received less attention. Whereas we might not be exploiting meiofauna's full potential yet, more and more integrative approaches and technological developments will create opportunities to employ these fascinating organisms to answer broad and important questions, despite their impediments related to their small body size. Meiofauna research agenda should balance amongst investigating general questions, addressing more specialized research topics, and generating primary data on distribution, taxonomy, traits, and DNA sequences. The geographical and taxonomic biases that have historically affected meiofaunal research can be alleviated by promoting international cooperation, open data sharing, and an increase effort in education, taxonomic training, as well as scientific communication. We hope that this will get both researchers and the general public intrigued by those small critters that constantly lurk unseen in front of us.
Few other invertebrates captivate our attention as cephalopods do. Octopods, cuttlefish, and squids amaze with their behavior and sophisticated body plans that belong to the most intriguing among mollusks. Little is, however, known about their body plan formation and the role of Hox genes. The latter homeobox genes pattern the anterior–posterior body axis and have only been studied in a single decapod species so far. Here, we study developmental Hox and ParaHox gene expression in Octopus vulgaris.Hox genes are expressed in a near-to-staggered fashion, among others in homologous organs of cephalopods such as the stellate ganglia, the arms, or funnel. As in other mollusks Hox1 is expressed in the nascent octopod shell rudiment. While ParaHox genes are expressed in an evolutionarily conserved fashion, Hox genes are also expressed in some body regions that are considered homologous among mollusks such as the cephalopod arms and funnel with the molluscan foot. We argue that cephalopod Hox genes are recruited to a lesser extent into the formation of non-related organ systems than previously thought and emphasize that despite all morphological innovations molecular data still reveal the ancestral molluscan heritage of cephalopods.
Spiralians, one of the major clades of bilaterians, share a unique development known as spiralian development, characterized by the formation of tiers of cells called quartets, which exhibit different developmental potentials along the animal-vegetal axis. Recently, spiralian-specific TALE-type homeobox genes (SPILE) have been identified, some of which show zygotic and staggered expression patterns along the animal-vegetal axis and function in quartet specification in mollusks. However, it is unclear which maternal molecular components control the zygotic expression of these transcription factors. In this study, we focused on SPILE-E, a maternal transcription factor, and investigated its expression and function in mollusks. We found that the maternal and ubiquitous expression of SPILE-E in the cleavage stages is conserved in molluskan species, including limpets, mussels, and chitons. We knocked down SPILE-E in limpets and revealed that the expression of transcription factors specifically expressed in the part of the first quartet (1q2 ; foxj1b) and second quartet (2q; SPILE-B) was abolished, whereas the macromere-quartet marker (SPILE-C) was ectopically expressed in 1q2 in SPILE-E morphants. Moreover, we showed that the expression of SPILE-A, which upregulates SPILE-B but represses SPILE-C expression, decreased in SPILE-E morphants. Consistent with changes in the expression pattern of the above transcription factors, SPILE-E-morphant larvae exhibited patchy or complete loss of expression of marker genes of ciliated cells and shell fields, possibly reflecting incomplete specification of 1q2 and 2q. Our results provide a molecular framework for quartet specification and highlight the importance of maternal lineage-specific transcription factors in the development and evolution of spiralians. This article is protected by copyright. All rights reserved.
Conserved cis-regulatory elements (CREs) control Engrailed-, Pax2-, and dachshund-related gene expression networks directing the formation and function of corresponding midbrain circuits in arthropods and vertebrates. Polarized outgroup analyses of 31 sequenced metazoan genomes representing all animal clades reveal the emergence of Pax2- and dachshund-related CRE-like sequences in anthozoan Cnidaria. The full complement, including Engrailed-related CRE-like sequences, is only detectable in spiralians, ecdysozoans, and chordates that have a brain; they exhibit comparable genomic locations and extensive nucleotide identities that reveal the presence of a conserved core domain, all of which are absent in non-neural genes and, together, distinguish them from randomly assembled sequences. Their presence concurs with a genetic boundary separating the rostral from caudal nervous systems, demonstrated for the metameric brains of annelids, arthropods, and chordates and the asegmental cycloneuralian and urochordate brain. These findings suggest that gene regulatory networks for midbrain circuit formation evolved within the lineage that led to the common ancestor of protostomes and deuterostomes.
While glycans underlie many biological processes, such as protein folding, cell adhesion and cell–cell recognition, deep evolution of glycosylation machinery remains an understudied topic. N-linked glycosylation is a conserved process in which mannosidases are key trimming enzymes. One of them is the glycoprotein endo-α-1,2-mannosidase which participates in the initial trimming of mannose moieties from an N-linked glycan inside the cis-Golgi. It is unique as the only endo-acting mannosidase found in this organelle. Relatively little is known about its origins and evolutionary history; so far it was reported to occur only in vertebrates. In this work, a taxon-rich bioinformatic survey to unravel the evolutionary history of this enzyme, including all major eukaryotic clades and a wide representation of animals, is presented. The endomannosidase was found to be more widely distributed in animals and other eukaryotes. The protein motif changes in context of the canonical animal enzyme were tracked. Additionally, the data show the two canonical vertebrate endomannosidase genes, MANEA and MANEAL, arose at the second round of the two vertebrate genome duplications and one more vertebrate paralog, CMANEAL, is uncovered. Finally, a framework where N-glycosylation co-evolved with complex multicellularity is described. A better understanding of the evolution of core glycosylation pathways is pivotal to understanding biology of eukaryotes in general, and the Golgi apparatus in particular. This systematic analysis of the endomannosidase evolution is one step towards this goal.
A number of well‐preserved fossil taxa from the early Cambrian Chengjiang biota of South China have played important roles in the construction and analysis of the early evolutionary tree of metazoans. Certain other Chengjiang taxa, erected based on limited characters, have yielded little or no information concerning key aspects of their palaeobiology, and consequently their phylogenetic significance has been difficult to assess. One such taxon is the large conical animal Glossolites magnus. Exceptionally preserved new material of this soft‐bodied species herein, bearing oral tentacles and internal structures, indicates that G. magnus is not, as previously proposed, a hyolith. This work provides further information about possible biological affinities and palaeoautecology, which still remain open questions.
While glycans underlie many biological processes, such as protein folding, cell adhesion and cell-cell recognition, deep evolution of glycosylation machinery remains an understudied topic. N-linked glycosylation is a conserved process in which mannosidases are key trimming enzymes. One of them is the glycoprotein endo‑α‑1,2‑mannosidase which participates in the initial trimming of mannose moieties from an N-linked glycan inside the cis-Golgi. It is unique as the only endo-acting mannosidase found in this organelle. Relatively little is known about its origins and evolutionary history; so far it was thought to occur only in vertebrates. Here I perform a taxon-rich bioinformatic survey to unravel the evolutionary history of this enzyme, including all major eukaryotic clades and a wide representation of animals. I found the endomannosidase to be vastly more widely distributed in animals than previously thought and in fact present in almost all eukaryotic clades. I tracked protein motif changes in context of the canonical animal enzyme. Additionally, my data show that the two canonical versions of endomannosidase in vertebrates, MANEA and MANEAL, arose at the second round of the two vertebrate genome duplications and indicate presence of a third protein, named here CMANEAL. Finally, I describe a framework where N-glycosylation co-evolved with complex multicellularity. A better understanding of the evolution of core glycosylation pathways is pivotal to understanding biology of eukaryotes in general, and the Golgi apparatus in particular. This systematic analysis of the endomannosidase evolution is one step towards this goal.
Asymmetries are essential for proper organization and function of organ systems. Genetic studies in bilaterians have shown signaling through the Nodal/Smad2 pathway plays a key, conserved role in the establishment of body asymmetries. However, while the main molecular players in the network for the establishment of left-right asymmetry (LRA) have been deeply described in deuterostomes, little is known about the regulation of Nodal signaling in spiralians. Here, we identified orthologs of the egf-cfc gene, a master regulator of the Nodal pathway in vertebrates, in several invertebrate species, which includes the first evidence of its presence in non-deuterostomes. Our functional experiments indicate that despite being present, egf-cfc does not play a role in the establishment of LRA in gastropods. However, experiments in zebrafish suggest that a single amino acid mutation in the egf-cfc gene in at least the common ancestor of chordates was the necessary step to induce a gain-of-function in LRA regulation. This study shows that the egf-cfc gene likely appeared in the ancestors of deuterostomes and protostomes, before being adopted as a master mechanism to regulate the Nodal pathway and the establishment of LRA in some lineages of deuterostomes.
Os conhecimentos taxonômico, filogenético e ecológico sobre a biodiversidade possuem lacunas para várias espécies, gerando ausência de estratégias de conservação para elas. Dentro da diversidade de organismos, alguns são de difícil acesso, o que os tornam complexos para exploração em sala de aula. Assim, o presente trabalho teve como objetivo abordar características morfológicas gerais sobre micrognatozoários por meio de mapa conceitual e explicitando dados informativos sobre o processo de construção dos dispositivos gráficos. Os conhecimentos biológicos acerca do táxon e sobre a técnica de mapeamento de conceitos foram pesquisados em bibliografias relacionadas a esses temas e os procedimentos metodológicos foram efetivados. A construção do mapa conceitual explicitou, visualmente, informações acerca do táxon Micrognathozoa, com parentesco próximo aos Rotifera, dentro da linhagem dos Gnathifera. As informações ilustradas proporcionam visibilidade mais didática para leitura, constituindo-se elementos potenciais para o ensino, ao abordar sobre a diversidade biológica. Com a elaboração e divulgação de um escrito fundamentado cientificamente, acerca do esquema visual criado, oportunidades são dadas para a conservação da biodiversidade. Nesse sentido, o fomento às publicações envolvendo mapas conceituais dinamiza o processo de ensino, tornando mais eficaz a aprendizagem sobre os grupos de animais enigmáticos.
Background:
Multiple RNA samples are frequently processed together and often mixed before multiplex sequencing in the same sequencing run. While different samples can be separated post sequencing using sample barcodes, the possibility of cross contamination between biological samples from different species that have been processed or sequenced in parallel has the potential to be extremely deleterious for downstream analyses.
Results:
We present CroCo, a software package for identifying and removing such cross contaminants from assembled transcriptomes. Using multiple, recently published sequence datasets, we show that cross contamination is consistently present at varying levels in real data. Using real and simulated data, we demonstrate that CroCo detects contaminants efficiently and correctly. Using a real example from a molecular phylogenetic dataset, we show that contaminants, if not eliminated, can have a decisive, deleterious impact on downstream comparative analyses.
Conclusions:
Cross contamination is pervasive in new and published datasets and, if undetected, can have serious deleterious effects on downstream analyses. CroCo is a database-independent, multi-platform tool, designed for ease of use, that efficiently and accurately detects and removes cross contamination in assembled transcriptomes to avoid these problems. We suggest that the use of CroCo should become a standard cleaning step when processing multiple samples for transcriptome sequencing.
It has been hypothesized that a condensed nervous system with a medial ventral nerve cord is an ancestral character of Bilateria. The presence of similar dorsoventral molecular patterns along the nerve cords of vertebrates, flies, and an annelid has been interpreted as support for this scenario. Whether these similarities are generally found across the diversity of bilaterian neuroanatomies is unclear, and thus the evolutionary history of the nervous system is still contentious. Here we study representatives of Xenacoelomorpha, Rotifera, Nemertea, Brachiopoda, and Annelida to assess the conservation of the dorsoventral nerve cord patterning. None of the studied species show a conserved dorsoventral molecular regionalization of their nerve cords, not even the annelid Owenia fusiformis, whose trunk neuroanatomy parallels that of vertebrates and flies. Our findings restrict the use of molecular patterns to explain nervous system evolution, and suggest that the similarities in dorsoventral patterning and trunk neuroanatomies evolved independently in Bilateria.
The relationships at the root of the animal tree have proven difficult to resolve, with the current debate focusing on whether sponges (phylum Porifera) or comb jellies (phylum Ctenophora) are the sister group of all other animals [1–5]. The choice of evolu- tionary models seems to be at the core of the prob- lem because Porifera tends to emerge as the sister group of all other animals (‘‘Porifera-sister’’) when site-specific amino acid differences are modeled (e.g., [6, 7]), whereas Ctenophora emerges as the sis- ter group of all other animals (‘‘Ctenophora-sister’’) when they are ignored (e.g., [8–11]). We show that two key phylogenomic datasets that previously sup- ported Ctenophora-sister [10, 12] display strong het- erogeneity in amino acid composition across sites and taxa and that no routinely used evolutionary model can adequately describe both forms of het- erogeneity. We show that data-recoding methods [13–15] reduce compositional heterogeneity in these datasets and that models accommodating site-spe- cific amino acid preferences can better describe the recoded datasets. Increased model adequacy is associated with significant topological changes in support of Porifera-sister. Because adequate modeling of the evolutionary process that generated the data is fundamental to recovering an accurate phylogeny [16–20], our results strongly support sponges as the sister group of all other animals and provide further evidence that Ctenophora-sister rep- resents a tree reconstruction artifact.
The phylum Rotifera consists of minuscule, nonsegmented animals with a unique body plan and an unresolved phylogenetic position. The presence of pharyngeal articulated jaws supports an inclusion in Gnathifera nested in the Spiralia. Comparison of Hox genes, involved in animal body plan patterning, can be used to infer phylogenetic relationships. Here, we report the expression of five Hox genes during embryogenesis of the rotifer Brachionus manjavacas and show how these genes define different functional components of the nervous system and not the usual bilaterian staggered expression along the anteroposterior axis. Sequence analysis revealed that the lox5-parapeptide, a key signature in lophotrochozoan and platyhelminthean Hox6/lox5 genes, is absent and replaced by different signatures in Rotifera and Chaetognatha, and that the MedPost gene, until now unique to Chaetognatha, is also present in rotifers. Collectively, our results support an inclusion of chaetognaths in gnathiferans and Gnathifera as sis
The position of Xenacoelomorpha in the tree of life remains a major unresolved question in the study of deep animal relationships. Xenacoelomorpha, comprising Acoela, Nemertodermatida, and Xenoturbella, are bilaterally symmetrical marine worms that lack several features common to most other bilaterians, for example an anus, nephridia, and a circulatory system. Two conflicting hypotheses are under debate: Xenacoelomorpha is the sister group to all remaining Bilateria (= Nephrozoa, namely protostomes and deuterostomes) or is a clade inside Deuterostomia. Thus, determining the phylogenetic position of this clade is pivotal for understanding the early evolution of bilaterian features, or as a case of drastic secondary loss of complexity. Here we show robust phylogenomic support for Xenacoelomorpha as the sister taxon of Nephrozoa. Our phylogenetic analyses, based on 11 novel xenacoelomorph transcriptomes and using different models of evolution under maximum likelihood and Bayesian inference analyses, strongly corroborate this result. Rigorous testing of 25 experimental data sets designed to exclude data partitions and taxa potentially prone to reconstruction biases indicates that long-branch attraction, saturation, and missing data do not influence these results. The sister group relationship between Nephrozoa and Xenacoelomorpha supported by our phylogenomic analyses implies that the last common ancestor of bilaterians was probably a benthic, ciliated acoelomate worm with a single opening into an epithelial gut, and that excretory organs, coelomic cavities, and nerve cords evolved after xenacoelomorphs separated from the stem lineage of Nephrozoa.
Acorn worms, also known as enteropneust (literally, 'gut-breathing') hemichordates, are marine invertebrates that share features with echinoderms and chordates. Together, these three phyla comprise the deuterostomes. Here we report the draft genome sequences of two acorn worms, Saccoglossus kowalevskii and Ptychodera flava. By comparing them with diverse bilaterian genomes, we identify shared traits that were probably inherited from the last common deuterostome ancestor, and then explore evolutionary trajectories leading from this ancestor to hemichordates, echinoderms and chordates. The hemichordate genomes exhibit extensive conserved synteny with amphioxus and other bilaterians, and deeply conserved non-coding sequences that are candidates for conserved gene-regulatory elements. Notably, hemichordates possess a deuterostome-specific genomic cluster of four ordered transcription factor genes, the expression of which is associated with the development of pharyngeal 'gill' slits, the foremost morphological innovation of early deuterostomes, and is probably central to their filter-feeding lifestyle. Comparative analysis reveals numerous deuterostome-specific gene novelties, including genes found in deuterostomes and marine microbes, but not other animals. The putative functions of these genes can be linked to physiological, metabolic and developmental specializations of the filter-feeding ancestor.
Despite rapid advances in the study of metazoan evolutionary history [1], phylogenomic analyses have so far neglected a number of microscopic lineages that possess a unique combination of characters and are thus informative for our understanding of morphological evolution. Chief among these lineages are the recently described animal groups Micrognathozoa and Loricifera, as well as the two interstitial "Problematica" Diurodrilus and Lobatocerebrum [2]. These genera show a certain resemblance to Annelida in their cuticle and gut [3, 4]; however, both lack primary annelid characters such as segmentation and chaetae [5]. Moreover, they show unique features such as an inverted body-wall musculature or a novel pharyngeal organ. This and their ciliated epidermis have led some to propose relationships with other microscopic spiralians, namely Platyhelminthes, Gastrotricha, and in the case of Diurodrilus, with Micrognathozoa [6, 7]-lineages that are grouped by some analyses into "Platyzoa," a clade whose status remains uncertain [1, 8-11]. Here, we assess the interrelationships among the meiofaunal and macrofaunal members of Spiralia using 402 orthologs mined from genome and transcriptome assemblies of 90 taxa. Lobatocerebrum and Diurodrilus are found to be deeply nested members of Annelida, and unequivocal support is found for Micrognathozoa as the sister group of Rotifera. Analyses using site-heterogeneous substitution models further recover a lophophorate clade and position Loricifera + Priapulida as sister group to the remaining Ecdysozoa. Finally, with several meiofaunal lineages branching off early in the diversification of Spiralia, the emerging concept of a microscopic, acoelomate, direct-developing ancestor of Spiralia is reviewed.
Copyright © 2015 Elsevier Ltd. All rights reserved.
Next generation sequencing has made it possible to perform differential gene expression studies in non-model organisms. For these studies, the need for a reference genome is circumvented by performing de novo assembly on the RNA-seq data. However, transcriptome assembly produces a multitude of contigs, which must be clustered into genes prior to differential gene expression detection. Here we present Corset, a method that hierarchically clusters contigs using shared reads and expression, then summarizes read counts to clusters, ready for statistical testing. Using a range of metrics, we demonstrate that Corset out-performs alternative methods. Corset is available from https://code.google.com/p/corset-project/.
Electronic supplementary material
The online version of this article (doi:10.1186/s13059-014-0410-6) contains supplementary material, which is available to authorized users.
Chaetognatha are a phylum of marine carnivorous animals which includes more than 130 extant species. The internal systematics of this group have been intensively debated since it was discovered in the 18(th) century. While they can be traced back to the earlier Cambrian, they are an extraordinarily homogeneous phylum at the morphological level - a fascinating characteristic that puzzled many a scientist who has tried to clarify their taxonomy. Recent studies which have attempted to reconstruct a phylogeny using molecular data have relied on single gene analyses and a somewhat restricted taxon sampling. Here, we present the first large scale phylogenetic study of Chaetognatha based on a combined analysis of nearly the complete ribosomal RNA (rRNA) genes. We use this analysis to infer the evolution of some morphological characters. This work includes 36 extant species, mainly obtained from Tara Oceans Expedition 2009/2012, that represent 16 genera and 6 of the 9 extant families.
Cladistic and phenetic analysis of morphological characters, geometric morphometrics and molecular small subunit (SSU rRNA) and large subunit (LSU rRNA) ribosomal genes phylogenies provided new insights into the relationships and the evolutionary history of Chaetognatha. We propose the following clade structure for the phylum: (((Sagittidae, Krohnittidae), Spadellidae), (Eukrohniidae, Heterokrohniidae)), with the Pterosagittidae included in the Sagittidae. The clade (Sagittidae, Krohnittidae) constitutes the monophyletic order of Aphragmophora. Molecular analyses showed that the Phragmophora are paraphyletic. The Ctenodontina/Flabellodontina and Syngonata/Chorismogonata hypotheses are invalidated on the basis of both morphological and molecular data. This new phylogeny also includes resurrected and modified genera within Sagittidae.
The distribution of some morphological characters traditionally used in systematics and for species diagnosis suggests that the diversity in Chaetognatha was produced through a process of mosaic evolution. Moreover, chaetognaths have mostly evolved by simplification of their body plan and their history shows numerous convergent events of losses and reversions. The main morphological novelty observed is the acquisition of a second pair of lateral fins in Sagittidae, which represents an adaptation to the holoplanktonic niche.
Large phylogenomics data sets require fast tree inference methods, especially for maximum-likelihood (ML) phylogenies. Fast
programs exist, but due to inherent heuristics to find optimal trees, it is not clear whether the best tree is found. Thus,
there is need for additional approaches that employ different search strategies to find ML trees and that are at the same
time as fast as currently available ML programs. We show that a combination of hill-climbing approaches and a stochastic perturbation
method can be time-efficiently implemented. If we allow the same CPU time as RAxML and PhyML, then our software IQ-TREE found
higher likelihoods between 62.2% and 87.1% of the studied alignments, thus efficiently exploring the tree-space. If we use
the IQ-TREE stopping rule, RAxML and PhyML are faster in 75.7% and 47.1% of the DNA alignments and 42.2% and 100% of the protein
alignments, respectively. However, the range of obtaining higher likelihoods with IQ-TREE improves to 73.3–97.1%. IQ-TREE
is freely available at http://www.cibiv.at/software/iqtree.
Based on molecular data three major clades have been recognized within Bilateria: Deuterostomia, Ecdysozoa and Spiralia. Within Spiralia, small-sized and simply organized animals such as flatworms, gastrotrichs and gnathostomulids have recently been grouped together as Platyzoa. However, the representation of putative platyzoans was low in the respective molecular phylogenetic studies, in terms of both, taxon number and sequence data. Furthermore, increased substitution rates in platyzoan taxa raised the possibility that monophyletic Platyzoa represents an artefact due to long-branch attraction. In order to overcome such problems, we employed a phylogenomic approach, thereby substantially increasing i) the number of sampled species within Platyzoa and ii) species-specific sequence coverage in datasets of up to 82,162 amino acid positions. Using established and new measures (long-branch score) we disentangled phylogenetic signal from misleading effects such as long-branch attraction. In doing so, our phylogenomic analyses did not recover a monophyletic origin of platyzoan taxa that, instead, appeared paraphyletic with respect to the other spiralians. Platyhelminthes and Gastrotricha formed a monophylum, which we name Rouphozoa. To the exclusion of Gnathifera, Rouphozoa and all other spiralians represent a monophyletic group, which we name Platytrochozoa. Platyzoan paraphyly suggests that the last common ancestor of Spiralia was a simple-bodied organism lacking coelomic cavities, segmentation and complex brain structures, and that more complex animals such as annelids evolved from such a simply organized ancestor. This conclusion contradicts alternative evolutionary scenarios proposing an annelid-like ancestor of Bilateria and Spiralia and several independent events of secondary reduction.
Phylogenies are increasingly used in all fields of medical and biological research. Moreover, because of the next generation sequencing revolution, datasets used for conducting phylogenetic analyses grow at an unprecedented pace. RAxML (Randomized Axelerated Maximum Likelihood) is a popular program for phylogenetic analyses of large datasets under maximum likelihood. Since the last RAxML paper in 2006, it has been continuously maintained and extended to accommodate the increasingly growing input datasets and to serve the needs of the user community.
I present some of the most notable new features and extensions of RAxML, such as, a substantial extension of substitution models and supported data types, the introduction of SSE3, AVX, and AVX2 vector intrinsics, techniques for reducing the memory requirements of the code and a plethora of operations for conducting post-analyses on sets of trees. In addition, an up-to-date, 50 page user manual covering all new RAxML options is available.
The code is available under GNU GPL at https://github.com/stamatak/standard-RAxML.
Alexandros.Stamatakis@h-its.org.
In recent years, there has been an increasing interest in the potential of codon substitution models for a variety of applications. However, the computational demands of these models have sometimes lead to the adoption of over-simplified assumptions, questionable statistical methods, or a limited focus on small data sets.
Here, we offer a scalable, message-passing-interface-based Bayesian implementation of site-heterogeneous codon models in the mutation-selection framework. Our software jointly infers the global mutational parameters at the nucleotide level, the branch lengths of the tree, and a Dirichlet process governing across-site variation at the amino acid level. We focus on an example estimation of the distribution of selection coefficients from an alignment of several hundred sequences of the influenza PB2 gene, and highlight the site-specific characterization enabled by such a modeling approach. Finally, we discuss future potential applications of the software for conducting evolutionary inferences.
The models are implemented within the PhyloBayes-MPI package, available on our website (phylobayes.org) along with usage details in the accompanying manual.
nicolas.rodrigue@ucalgary.ca.
The discovery of a living coelacanth specimen in 1938 was remarkable, as this lineage of lobe-finned fish was thought to have become extinct 70 million years ago. The modern coelacanth looks remarkably similar to many of its ancient relatives, and its evolutionary proximity to our own fish ancestors provides a glimpse of the fish that first walked on land. Here we report the genome sequence of the African coelacanth, Latimeria chalumnae. Through a phylogenomic analysis, we conclude that the lungfish, and not the coelacanth, is the closest living relative of tetrapods. Coelacanth protein-coding genes are significantly more slowly evolving than those of tetrapods, unlike other genomic features. Analyses of changes in genes and regulatory elements during the vertebrate adaptation to land highlight genes involved in immunity, nitrogen excretion and the development of fins, tail, ear, eye, brain and olfaction. Functional assays of enhancers involved in the fin-to-limb transition and in the emergence of extra-embryonic tissues show the importance of the coelacanth genome as a blueprint for understanding tetrapod evolution.
Current genomic perspectives on animal diversity neglect two prominent phyla, the molluscs and annelids, that together account for nearly one-third of known marine species and are important both ecologically and as experimental systems in classical embryology. Here we describe the draft genomes of the owl limpet (Lottia gigantea), a marine polychaete (Capitella teleta) and a freshwater leech (Helobdella robusta), and compare them with other animal genomes to investigate the origin and diversification of bilaterians from a genomic perspective. We find that the genome organization, gene structure and functional content of these species are more similar to those of some invertebrate deuterostome genomes (for example, amphioxus and sea urchin) than those of other protostomes that have been sequenced to date (flies, nematodes and flatworms). The conservation of these genomic features enables us to expand the inventory of genes present in the last common bilaterian ancestor, establish the tripartite diversification of bilaterians using multiple genomic characteristics and identify ancient conserved long- and short-range genetic linkages across metazoans. Superimposed on this broadly conserved pan-bilaterian background we find examples of lineage-specific genome evolution, including varying rates of rearrangement, intron gain and loss, expansions and contractions of gene families, and the evolution of clade-specific genes that produce the unique content of each genome.
The fate of the blastopore during development in the bilaterian ancestor is currently not well understood. In deuterostomes, the blastopore forms the anus, but its fate in protostome groups is variable [1]. This variability, combined with an absence of information from key taxa, hampers the reconstruction of the ancestral developmental mode of the Protostomia and the Bilateria. The blastopore fate of the bilaterian ancestor plays a crucial role in understanding the transition from radial to bilateral symmetric organisms [2, 3]. Priapulids have a conservative morphology, an abundant Cambrian fossil record, and a phylogenetic position that make them a key group in understanding protostome evolution [4, 5]. Here, we characterize gastrulation and the embryonic expression of genes involved in bilaterian foregut and hindgut patterning in Priapulus caudatus. We show that the blastopore gives rise to the anus at the vegetal pole and that the hindgut markers brachyury and caudal are expressed in the blastopore and anus, whereas the foregut markers foxA and goosecoid are expressed in the mouth in the animal hemisphere. Thereby, gastrulation in the conservatively evolving protostome P. caudatus follows strictly a deuterostomic pattern. These results are more compatible with a deuterostomic rather than protostomic (blastopore forms the mouth) or amphistomic (mouth and anus are formed simultaneously) mode of development in the last common bilaterian ancestor.
The epidermis of both species of Seison is syncytial and has a characteristic internal layer divided into two distinct sublayers. Sublayer I is very thin (0.03 μm)
and bounded to the outer cell membrane of the epidermis. Sublayer II is 0.5 μm thick and separated from sublayer I by a thin
layer of cytoplasm. Intrusions of the outer cell membrane of the epidermis perforate the internal layer, before terminating
within the cytoplasm. The intrusions of the cell membrane of S. annulatus are coated by an electron-dense material, the annulus, when passing through the internal layer. Bundles of filaments are present
in the epidermis of S. nebaliae. A comparison of epidermal structures within the Gnathifera Ahlrichs, 1995, confirms phylogenetic relationships earlier proposed
by the author.
Massively parallel sequencing of cDNA has enabled deep and efficient probing of transcriptomes. Current approaches for transcript reconstruction from such data often rely on aligning reads to a reference genome, and are thus unsuitable for samples with a partial or missing reference genome. Here we present the Trinity method for de novo assembly of full-length transcripts and evaluate it on samples from fission yeast, mouse and whitefly, whose reference genome is not yet available. By efficiently constructing and analyzing sets of de Bruijn graphs, Trinity fully reconstructs a large fraction of transcripts, including alternatively spliced isoforms and transcripts from recently duplicated genes. Compared with other de novo transcriptome assemblers, Trinity recovers more full-length transcripts across a broad range of expression levels, with a sensitivity similar to methods that rely on genome alignments. Our approach provides a unified solution for transcriptome reconstruction in any sample, especially in the absence of a reference genome.
Recent progress in reconstructing animal relationships enables us to draw a better picture of the evolution of important characters such as organ systems and developmental processes. By mapping these characters onto the phylogenetic framework, we can detect changes that have occurred in them during evolution. The spiral mode of development is a complex of characters that is present in many lineages, such as nemerteans, annelids, mollusks, and polyclad platyhelminthes. However, some of these lineages show variations of this general program in which sub-characters are modified without changing the overlying pattern. Recent molecular phylogenies suggest that spiral cleavage was lost, or at least has deviated from its original pattern, in more lineages than was previously thought (e.g., in rotifers, gastrotrichs, bryozoans, brachiopods, and phoronids). Here, I summarize recent progress in reconstructing the spiralian tree of life and discuss its significance for our understanding of the spiral-cleavage character complex. I conclude that more detailed knowledge of the development of spiralian taxa is necessary to understand the mechanisms behind these changes, and to understand the evolutionary changes and adaptations of spiralian embryos.
Xenoturbellida and Acoelomorpha are marine worms with contentious ancestry. Both were originally associated with the flatworms (Platyhelminthes), but molecular data have revised their phylogenetic positions, generally linking Xenoturbellida to the deuterostomes and positioning the Acoelomorpha as the most basally branching bilaterian group(s). Recent phylogenomic data suggested that Xenoturbellida and Acoelomorpha are sister taxa and together constitute an early branch of Bilateria. Here we assemble three independent data sets-mitochondrial genes, a phylogenomic data set of 38,330 amino-acid positions and new microRNA (miRNA) complements-and show that the position of Acoelomorpha is strongly affected by a long-branch attraction (LBA) artefact. When we minimize LBA we find consistent support for a position of both acoelomorphs and Xenoturbella within the deuterostomes. The most likely phylogeny links Xenoturbella and Acoelomorpha in a clade we call Xenacoelomorpha. The Xenacoelomorpha is the sister group of the Ambulacraria (hemichordates and echinoderms). We show that analyses of miRNA complements have been affected by character loss in the acoels and that both groups possess one miRNA and the gene Rsb66 otherwise specific to deuterostomes. In addition, Xenoturbella shares one miRNA with the ambulacrarians, and two with the acoels. This phylogeny makes sense of the shared characteristics of Xenoturbellida and Acoelomorpha, such as ciliary ultrastructure and diffuse nervous system, and implies the loss of various deuterostome characters in the Xenacoelomorpha including coelomic cavities, through gut and gill slits.
The quality of multiple sequence alignments plays an important role in the accuracy of phylogenetic inference. It has been shown that removing ambiguously aligned regions, but also other sources of bias such as highly variable (saturated) characters, can improve the overall performance of many phylogenetic reconstruction methods. A current scientific trend is to build phylogenetic trees from a large number of sequence datasets (semi-)automatically extracted from numerous complete genomes. Because these approaches do not allow a precise manual curation of each dataset, there exists a real need for efficient bioinformatic tools dedicated to this alignment character trimming step.
Here is presented a new software, named BMGE (Block Mapping and Gathering with Entropy), that is designed to select regions in a multiple sequence alignment that are suited for phylogenetic inference. For each character, BMGE computes a score closely related to an entropy value. Calculation of these entropy-like scores is weighted with BLOSUM or PAM similarity matrices in order to distinguish among biologically expected and unexpected variability for each aligned character. Sets of contiguous characters with a score above a given threshold are considered as not suited for phylogenetic inference and then removed. Simulation analyses show that the character trimming performed by BMGE produces datasets leading to accurate trees, especially with alignments including distantly-related sequences. BMGE also implements trimming and recoding methods aimed at minimizing phylogeny reconstruction artefacts due to compositional heterogeneity.
BMGE is able to perform biologically relevant trimming on a multiple alignment of DNA, codon or amino acid sequences. Java source code and executable are freely available at ftp://ftp.pasteur.fr/pub/GenSoft/projects/BMGE/.
Multiple sequence alignment is of central importance to bioinformatics and computational biology. Although a large number of algorithms for computing a multiple sequence alignment have been designed, the efficient computation of highly accurate multiple alignments is still a challenge.
We present MSAProbs, a new and practical multiple alignment algorithm for protein sequences. The design of MSAProbs is based on a combination of pair hidden Markov models and partition functions to calculate posterior probabilities. Furthermore, two critical bioinformatics techniques, namely weighted probabilistic consistency transformation and weighted profile-profile alignment, are incorporated to improve alignment accuracy. Assessed using the popular benchmarks: BAliBASE, PREFAB, SABmark and OXBENCH, MSAProbs achieves statistically significant accuracy improvements over the existing top performing aligners, including ClustalW, MAFFT, MUSCLE, ProbCons and Probalign. Furthermore, MSAProbs is optimized for multi-core CPUs by employing a multi-threaded design, leading to a competitive execution time compared to other aligners.
The source code of MSAProbs, written in C++, is freely and publicly available from http://msaprobs.sourceforge.net.
OMA is a project that aims to identify orthologs within publicly available, complete genomes. With 657 genomes analyzed to date, OMA is one of the largest projects of its kind.
The algorithm of OMA improves upon standard bidirectional best-hit approach in several respects: it uses evolutionary distances instead of scores, considers distance inference uncertainty, includes many-to-many orthologous relations, and accounts for differential gene losses. Herein, we describe in detail the algorithm for inference of orthology and provide the rationale for parameter selection through multiple tests.
OMA contains several novel improvement ideas for orthology inference and provides a unique dataset of large-scale orthology assignments.
The chaetognaths, or arrowworms, constitute a small and enigmatic phylum of marine invertebrates whose phylogenetic affinities have long been uncertain. A popular hypothesis is that the chaetognaths are the sister group of the major deuterostome phyla: chordates, hemichordates, and echinoderms. Here we attempt to determine the affinities of the chaetognaths by using molecular sequence data. We describe the isolation and nucleotide sequence determination of 18S ribosomal DNA from one species of chaetognath and one acanthocephalan. Extensive phylogenetic analyses employing a suite of phylogenetic reconstruction methods (maximum parsimony, maximum likelihood, evolutionary parsimony, and two distance methods) suggest that the hypothesized relationship between chaetognaths and the deuterostomes is incorrect. In contrast, we propose that the lineage leading to the chaetognaths arose prior to the advent of the coelomate metazoa.
We present the isolation of six Hox genes in the chaetognath Spadella cephaloptera. We identified one member of the paralogy group 3, four median genes and a mosaic gene that shares features of both median and posterior classes ( SceMedPost). Several hypotheses may account for the presence of a mosaic Hox gene in this animal. Here we propose that SceMedPost may represent an ancestral gene, which has not diverged totally into a posterior or a median one. This hypothesis has interesting implications for the reconstruction of the evolutionary history of Hox genes and suggests that Chaetognatha lineage divergence could predate the deuterostome/protostome split. Such a phylogenetic position is considered in the light of their embryological and morphological characters.
While the phylogenetic position of Chaetognatha has became central to the question of early bilaterian evolution, the internal systematics of the phylum are still not clear. The phylogenetic relationships of the chaetognaths were investigated using newly obtained small subunit ribosomal RNA nuclear 18S (SSU rRNA) sequences from 16 species together with 3 sequences available in GenBank. As previously shown with the large subunit ribosomal RNA 28S gene, two classes of Chaetognatha SSU rRNA gene can be identified, suggesting a duplication of the whole ribosomal cluster; allowing the rooting of one class of genes by another in phylogenetic analyses. Maximum Parsimony, Maximum Likelihood and Bayesian analyses of the molecular data, and statistical tests showed (1) that there are three main monophyletic groups: Sagittidae/Krohnittidae, Spadellidae/Pterosagittidae, and Eukrohniidae/Heterokrohniidae, (2) that the group of Aphragmophora without Pterosagittidae (Sagittidae/Krohnittidae) is monophyletic, (3) the Spadellidae/Pterosagittidae and Eukrohniidae/Heterokrohniidae families are very likely clustered, (4) the Krohnittidae and Pterosagittidae groups should no longer be considered as families as they are included in other groups designated as families, (5) suborder Ctenodontina is not monophyletic and the Flabellodontina should no longer be considered as a suborder, and (6) the Syngonata/Chorismogonata and the Monophragmophora/Biphragmophora hypotheses are rejected. Such conclusions are considered in the light of morphological characters, several of which are shown to be prone to homoplasy.
Until recently, molecular phylogenies based on a single or few orthologous genes often yielded contradictory results. Using multiple genes in a large concatenation was proposed to end these incongruences. Here we show that single-gene phylogenies often produce incongruences, albeit ones lacking statistically significant support. By contrast, the use of different tree reconstruction methods on different partitions of the concatenated supergene leads to well-resolved, but real (i.e. statistically significant) incongruences. Gathering a large amount of data is not sufficient to produce reliable trees, given the current limitation of tree reconstruction methods, especially when the quality of data is poor. We propose that selecting only data that contain minimal nonphylogenetic signals takes full advantage of phylogenomics and markedly reduces incongruence.
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We investigate the use of Markov models of evolution for reduced amino acid alphabets or bins of amino acids. The use of reduced amino acid alphabets can ameliorate effects of model misspecification and saturation. We present algorithms for 2 different ways of automating the construction of bins: minimizing criteria based on properties of rate matrices and minimizing criteria based on properties of alignments. By simulation, we show that in the absence of model misspecification, the loss of information due to binning is found to be insubstantial, and the use of Markov models at the binned level is found to be almost as effective as the more appropriate missing data approach. By applying these approaches to real data sets where compositional heterogeneity and/or saturation appear to be causing biased tree estimation, we find that binning can improve topological estimation in practice.
EVidenceModeler (EVM) is presented as an automated eukaryotic gene structure annotation tool that reports eukaryotic gene structures as a weighted consensus of all available evidence. EVM, when combined with the Program to Assemble Spliced Alignments (PASA), yields a comprehensive, configurable annotation system that predicts protein-coding genes and alternatively spliced isoforms. Our experiments on both rice and human genome sequences demonstrate that EVM produces automated gene structure annotation approaching the quality of manual curation.
The advent of numerical methods for analysing phylogenetic relationships, along with the study of morphology and molecular data, has driven our understanding of animal relationships for the past three decades. Within the protostome branch of the animal tree of life, these data have sufficed to establish its two main side branches, the moulting Ecdysozoa and the non-moulting Lophotrochozoa. In this review, I explore our current knowledge of protostome relationships and discuss progress and future perspectives and strategies to increase resolution within the main lophotrochozoan clades. Novel approaches to coding morphological characters are needed by scoring real observations on species selected as terminals. Still, methodological issues, for example, how to deal with inapplicable characters or the coding of absences, may require novel algorithmic developments. Taxon sampling is another key issue, as phyla should include enough species so as to represent their span of anatomical disparity. On the molecular side, phylogenomics is playing an increasingly important role in elucidating animal relationships, but genomic sampling is still fairly limited within the lophotrochozoan protostomes, for which only three phyla are represented in currently available phylogenies. Future work should therefore concentrate on generating novel morphological observations and on producing genomic data for the lophotrochozoan side of the animal tree of life.
Chaetognaths (arrow worms) are a separate phylum (Chaetognatha) of small carnivorous animals, dominantly pelagic, and a major component of today's plankton [1, 2]. The position of Chaetognatha among metazoan phyla remains equivocal-neither morphological nor molecular data provide definitive evidence [3]. Originating early in the Cambrian period [4], if not earlier [5], chaetognaths quickly became important members of marine metazoan communities [6]. Chaetognath grasping spines, originally reported as conodonts, occur worldwide in many Cambrian marine sediments [6, 7]. Fossilized chaetognath bodies, in contrast, are very rare: only two unequivocal specimens have been reported, both from the early Cambrian of China [8, 9]. Here we describe Capinatator praetermissus, a new genus and species, based on ∼50 specimens from several middle Cambrian Burgess Shale localities in British Columbia, many of which preserve evidence of soft tissues. Capinatator praetermissus reached body lengths of nearly 10 cm exclusive of fins, a much larger size than that of most living forms. Clusters of specimens preserving the body indicate that they were rapidly buried, providing indirect evidence that they swam near the seabed. The feeding apparatus comprises up to ∼25 spines in each half, almost double the maximum number in living chaetognaths. Early chaetognaths apparently occupied ecological niches associated with predatory euarthropods. The large body size and high number of grasping spines in C. praetermissus may indicate that miniaturization and migration to a planktonic lifestyle were secondary.
Despite the large variation in adult bodyplan phenotypes, a worm-shaped morphology is considered plesiomorphic for both Lophotrochozoa and Bilateria. Although almost all larval and adult lophotrochozoan worms have serially arranged ring muscles in their body wall, a comparison of their ontogeny reveals no less than six different developmental pathways that lead to this homogenous arrangement of ring muscles. However, in all taxa, with the exception of chaetodermomorph molluscs and the segmented annelids, ring muscle development starts with synchronous formation of certain pioneer myocytes, which is thus considered basal for Lophotrochozoa. Recent studies on spiralian neurogenesis revealed remnants of ancestral segmentation in echiurans and sipunculans, thus confirming molecular phylogenetic studies that propose a close relationship of these three taxa. Larval entoprocts exhibit a mosaic of larval and adult molluscan characters and, among other apomorphies, share with polyplacophoran Mollusca a complex larval apical organ and a tetraneurous nervous system, strongly suggesting a monophyletic assemblage of Entoprocta and Mollusca. The term Tetraneuralia is proposed herein for this lophotrochozoan clade. Overall, formation of the lophotrochozoan neuromuscular bodyplan appears as a highly dynamic process on both the ontogenetic and the evolutionary timescales, highlighting the importance of insights into these processes for reconstructing ancestral bodyplan features and phylogenetic relationships.
Phylogenomic studies have improved understanding of deep metazoan phylogeny and show promise for resolving incongruences among analyses based on limited numbers of loci. One region of the animal tree that has been especially difficult to resolve, even with phylogenomic approaches, is relationships within Lophotrochozoa (the animal clade that includes molluscs, annelids, and flatworms among others). Lack of resolution in phylogenomic analyses could be due to insufficient phylogenetic signal, limitations in taxon and/or gene sampling, or systematic error. Here, we investigated why lophotrochozoan phylogeny has been such a difficult question to answer by identifying and reducing sources of systematic error. We supplemented existing data with 32 new transcriptomes spanning the diversity of Lophotrochozoa and constructed a new set of Lophotrochozoa-specific core orthologs. Of these, 638 orthologous groups (OGs) passed strict screening for paralogy using a tree-based approach. In order to reduce possible sources of systematic error, we calculated branch-length heterogeneity, evolutionary rate, percent missing data, compositional bias, and saturation for each OG and analyzed increasingly stricter subsets of only the most stringent (best) OGs for these five variables. Principal component analysis of the values for each factor examined for each OG revealed that compositional heterogeneity and average patristic distance contributed most to the variance observed along the first principal component while branch-length heterogeneity and, to a lesser extent, saturation contributed most to the variance observed along the second. Missing data did not strongly contribute to either. Additional sensitivity analyses examined effects of removing taxa with heterogeneous branch lengths, large amounts of missing data, and compositional heterogeneity. Although our analyses do not unambiguously resolve lophotrochozoan phylogeny, we advance the field by reducing the list of viable hypotheses. Moreover, our systematic approach for dissection of phylogenomic data can be applied to explore sources of incongruence and poor support in any phylogenomic dataset.
We present kallisto, an RNA-seq quantification program that is two orders of magnitude faster than previous approaches and achieves similar accuracy. Kallisto pseudoaligns reads to a reference, producing a list of transcripts that are compatible with each read while avoiding alignment of individual bases. We use kallisto to analyze 30 million unaligned paired-end RNA-seq reads in <10 min on a standard laptop computer. This removes a major computational bottleneck in RNA-seq analysis.
A revised six-kingdom system of life is presented, down to the level of infraphylum. As in my 1983 system Bacteria are treated as a single kingdom, and eukaryotes are divided into only five kingdoms: Protozoa, Animalia, Fungi, Plantae and Chromista. Intermediate high level categories (superkingdom, subkingdom, branch, infrakingdom, superphylum, subphylum and infraphylum) are extensively used to avoid splitting organisms into an excessive number of kingdoms and phyla (60 only being recognized). The two 'zoological' kingdoms, Protozoa and Animalia, are subject to the International Code of Zoological Nomenclature, the kingdom Bacteria to the International Code of Bacteriological Nomenclature, and the three 'botanical' kingdoms (Plantae, Fungi, Chromista) to the International Code of Botanical Nomenclature. Circumscriptions of the kingdoms Bacteria and Plantae remain unchanged since Cavalier-Smith (1981). The kingdom Fungi is expanded by adding Microsporidia, because of protein sequence evidence that these amitochondrial intracellular parasites are related to conventional Fungi, not Protozoa. Fungi are subdivided into four phyla and 20 classes; fungal classification at the rank of subclass and above is comprehensively revised. The kingdoms Protozoa and Animalia are modified in the light of molecular phylogenetic evidence that Myxozoa are actually Animalia, not Protozoa, and that mesozoans are related to bilaterian animals. Animalia are divided into four subkingdoms: Radiata (phyla Porifera, Cnidaria, Placozoa, Ctenophora), Myxozoa, Mesozoa and Bilateria (bilateral animals: all other phyla). Several new higher level groupings are made in the animal kingdom including three new phyla: Acanthognatha (rotifers, acanthocephalans, gastrotrichs, gnathostomulids), Brachiozoa (brachiopods and phoronids) and Lobopoda (onychophorans and tardigrades), so only 23 animal phyla are recognized. Archezoa, here restricted to the phyla Metamonada and Trichozoa, are treated as a subkingdom within Protozoa, as in my 1983 six-kingdom system, not as a separate kingdom. The recently revised phylum Rhizopoda is modified further by adding more flagellates and removing some 'rhizopods' and is therefore renamed Cercozoa. The number of protozoan phyla is reduced by grouping Mycetozoa and Archamoebae (both now infraphyla) as a new subphylum Conosa within the phylum Amoebozoa alongside the subphylum Lobosa, which now includes both the traditional aerobic lobosean amoebae and Multicilia. Haplosporidia and the (formerly microsporidian) metchnikovellids are now both placed within the phylum Sporozoa. These changes make a total of only 13 currently recognized protozoan phyla, which are grouped into two subkingdoms: Archezoa and Neozoa; the latter is modified in circumscription by adding the Discicristata, a new infrakingdom comprising the phyla Percolozoa and Euglenozoa). These changes are discussed in relation to the principles of megasystematics, here defined as systematics that concentrates on the higher levels of classes, phyla, and kingdoms. These principles also make it desirable to rank Archaebacteria as an infrakingdom of the kingdom Bacteria, not as a separate kingdom. Archaebacteria are grouped with the infrakingdom Posibacteria to form a new subkingdom, Unibacteria, comprising all bacteria bounded by a single membrane. The bacterial subkingdom Negibacteria, with separate cytoplasmic and outer membranes, is subdivided into two infrakingdoms: Lipobacteria, which lack lipopolysaccharide and have only phospholipids in the outer membrane, and Glycobacteria, with lipopolysaccharides in the outer leaflet of the outer membrane and phospholipids in its inner leaflet. This primary grouping of the 10 bacterial phyla into subkingdoms is based on the number of cell-envelope membranes, whilst their subdivision into infrakingdoms emphasises their membrane chemistry; definition of the negibacterial phyla, five at least partly photosynthetic, relies chiefly on photosynthetic mechanism and cell-envelope structure and chemistry corroborated by ribosomal RNA phylogeny. The kingdoms Protozoa and Chromista are slightly changed in circumscription by transferring subphylum Opalinata (classes Opalinea, Proteromonadea, Blastocystea cl. nov.) from Protozoa into infrakingdom Heterokonta of the kingdom Chromista. Opalinata are grouped with the subphylum Pseudofungi and the zooflagellate Developayella elegans (in a new subphylum Bigyromonada) to form a new botanical phylum (Bigyra) of heterotrophs with a double ciliary transitional helix, making it necessary to abandon the phylum name Opalozoa, which formerly included Opalinata. The loss of ciliary retronemes in Opalinata is attributed to their evolution of gut commensalism. The nature of the ancestral chromist is discussed in the light of recent phylogenetic evidence.
Animals make up only a small fraction of the eukaryotic tree of life, yet, from our vantage point as members of the animal kingdom, the evolution of the bewildering diversity of animal forms is endlessly fascinating. In the century following the publication of Darwin's Origin of Species, hypotheses regarding the evolution of the major branches of the animal kingdom - their relationships to each other and the evolution of their body plans - was based on a consideration of the morphological and developmental characteristics of the different animal groups. This morphology-based approach had many successes but important aspects of the evolutionary tree remained disputed. In the past three decades, molecular data, most obviously primary sequences of DNA and proteins, have provided an estimate of animal phylogeny largely independent of the morphological evolution we would ultimately like to understand. The molecular tree that has evolved over the past three decades has drastically altered our view of animal phylogeny and many aspects of the tree are no longer contentious. The focus of molecular studies on relationships between animal groups means, however, that the discipline has become somewhat divorced from the underlying biology and from the morphological characteristics whose evolution we aim to understand. Here, we consider what we currently know of animal phylogeny; what aspects we are still uncertain about and what our improved understanding of animal phylogeny can tell us about the evolution of the great diversity of animal life.
The trophi of rotifers resemble the sclerotized jaws of gnathostomulids, but whether trophi are homologous to gnathostomulid jaws, and consequently show phylogenetic relationship between Gnathostomulida and Rotifera, has been unclear. We have found that the trophi of a rotifer in the genus Seison, which is ranked close to the ancestral stock of rotifers, have an ultrastructural feature similar to that reported in the literature for jaws of both scleroperalian and filospermoidean gnathostomulids, as well as for trophi in the more derived bdelloid rotifer genus Philodina. Specifically, these trophi and jaws have arrays of tube-like support rods composed of lucent material surrounding a dense core. Jaw-like structures in other small vermiform animals (certain polychaetes and molluscs) lack this special feature. We propose that jaw substructure shows a homology, and thus a sister-group relationship, between Gnathostomulida and the clade containing Rotifera plus Acanthocephala.
The less than one hundred species of the phylum Chaetognatha constitute an important element in marine Zooplankton communities. The rather uniform organization of these transparent arrow-shaped animals (mm-9 cm) is closely adapted to their mostly planktonic habitats and to their predatory feeding behaviour. Our knowledge of their integument is based on early monographs (e.g. Burfied 1927) and further progress, up to the use of the electron microscope, was summarized in the monographs of Kuhl (1938), Hyman (1959) and Ghirar-delli (1968) and in the reviews of Strenger (1964) and Kirsteuer (1969). Since the contribution of Furnestin (1967), a number of electron microscopic investigations has dealt with aspects of the chaetognath integument. The present review and the illustrations also include the author’s observations from 10 species in 5 genera (Ahnelt 1980). Detailed examinations of ontogeny, cytophysiology and biomechanics of the chaetognath body wall are still awaited. Therefore this report concentrates on morphological features with comments on their possible function.
The teeth and grasping spines of Sagitta are similar in structure, both having a central pulp cavity surrounded by two electron-dense chitinous layers. The cells of the pulp cavity contain microtubules arranged along the long axis. The two chitinous tubes are separated by a less-dense zone crossed by coarse fibrils linking the two. The teeth and spines insert into less electron-dense chitin (presumably flexible) and are moved by processes of anchor cells which pass into the basal chitinous zone. The inner region of the anchor cells is apposed to the connective tissue layer on to which the muscles of the teeth and spines insert. At the base of the pulp cavity, i.e. at the secretory zone where the teeth and spines are formed, the cells of the pulp cavity contain electron-dense granules in which zinc is found; zinc is also present in the inner and outer dense chitin layers at high concentration (0·5–1·0% of the dry wt).
Both spines and teeth are tipped with fibrous cones containing silicon. It is suggested that the zinc associated with the chitin serves to toughen the teeth and spines and render them less liable to fracture, and that the silicon in the tips confers hardness to this vulnerable region.
Despite the large variation in adult bodyplan phenotypes, a worm-shaped morphology is considered plesiomorphic for both Lophotrochozoa and Bilateria. Although almost all larval and adult lophotrochozoan worms have serially arranged ring muscles in their body wall, a comparison of their ontogeny reveals no less than six different developmental pathways that lead to this homogenous arrangement of ring muscles. However, in all taxa, with the exception of chaetodermomorph molluscs and the segmented annelids, ring muscle development starts with synchronous formation of certain pioneer myocytes, which is thus considered basal for Lophotrochozoa. Recent studies on spiralian neurogenesis revealed remnants of ancestral segmentation in echiurans and sipunculans, thus confirming molecular phylogenetic studies that propose a close relationship of these three taxa. Larval entoprocts exhibit a mosaic of larval and adult molluscan characters and, among other apomorphies, share with polyplacophoran Mollusca a complex larval apical organ and a tetraneurous nervous system, strongly suggesting a monophyletic assemblage of Entoprocta and Mollusca. The term Tetraneuralia is proposed herein for this lophotrochozoan clade. Overall, formation of the lophotrochozoan neuromuscular bodyplan appears as a highly dynamic process on both the ontogenetic and the evolutionary timescales, highlighting the importance of insights into these processes for reconstructing ancestral bodyplan features and phylogenetic relationships.
This is the first record of the new phylum from North America.
The jaws of Limnognathia maerski, Micrognathozoa, were investigated with light- and scanning electron microscopy. The study yielded several new structures and sclerites, including the ventral part of main jaw, the pharyngeal lamellae, the manus, the dorsal and ventral fibularium teeth, and a reinterpretation of the fibularium compartmentalization. Furthermore, it was shown that several jaw elements are composed of densely packed rods. Comparison with Rotifera and Gnathostomulida suggested that the micrognathozoan main jaw is homologous with the rotifer incus and the gnathostomulid articularium and that the pseudophalangids (the ventral jaws) and their associated sclerites correspond to the rotifer mallei. These results imply that Micrognathozoa is more closely related to Rotifera than to Gnathostomulida.
Nemerteans have been alleged to belong to a protostome clade called the Trochozoa that includes mollusks, annelids, sipunculids, echiurids, and kamptozoans and is characterized by, among other things, the trochophore larva. The trochophore possesses a prototroch, a preoral belt of specialized ciliary cells, derived from the trochoblast cells. Nemertea is the only trochozoan phylum for which presence of the trochophore larva possessing a prototroch had never been shown. However, so little is known about nemertean larval development that comparing it with development of other trochozoans is difficult. Development in the nemertean clade Pilidiophora is via a highly specialized planktonic larva, the pilidium, and most of the larval body is lost during a drastic metamorphosis. Other nemerteans (hoplonemerteans and palaeonemerteans) lack a pilidium, and their development is direct, forming either an encapsulated or planktonic "planuliform" larva, producing a juvenile without a dramatic change in body plan. We show that early in the development of a member of a basal nemertean assemblage, the palaeonemertean Carinoma tremaphoros, large squamous cells cover the entire larval surface except for the apical and posterior regions. Although apical and posterior cells continue to divide, the large surface cells cleavage arrest and form a contorted preoral belt. Based on its position, cell lineage, and fate, we suggest that this belt corresponds to the prototroch of other trochozoans. Lack of differential ciliation obscures the presence of the prototroch in Carinoma, but differentiation of the trochoblasts is clearly manifested in their permanent cleavage arrest and ultimate degenerative fate. Our results allow a meaningful comparison between the development of nemerteans and other trochozoans. We review previous hypotheses of the evolution of nemertean development and suggest that a trochophore-like larva is plesiomorphic for nemerteans while a pilidium type of development with drastic metamorphosis is derived.
cDNA library construction, EST sequencing and data processing The species Spadella cephaloptera, Busch 1851 was chosen because of its availability related to benthic way of life. Specimens were collected in several stations in the South of France (Brusc lagoon and bay of la Ciotat, near Marseille). To build cDNA library, total RNA was isolated from hatchling to 1-day-old juveniles. Single strand cDNAs were synthesized by using oligo (dT) primers and Superscript RNase H reverse transcriptase (Gibco BRL). Double-stranded cDNAs were cloned into Lambda Triplex 2 vector (Clontech). The title of the primary library was 5.4x10 -6 plaque-forming units/ml. In order to allow efficient sequencing, excised pTriplex 2 were extracted and cut by NotI to select plasmids containing inserts longer than 800 bp. After ligation, BB4 E. coli strain was transformed with this pull of plasmids. This cDNA library was sequenced up to 12664 clones at the Génoscope (Centre National de Séquençage, France) from a 5' primer. These raw sequences were filtered for quality and for vector contamination. After these filters, 11526 validated EST sequences were available for analysis. Then, these ESTs were grouped into contigs using CLOBB (S1) and consensus inferred using PHRAP (P. Green, unpublished data) in order to deal with a quite non-redundant partial transcriptome, appropriate for comparative analyses. Among the 5859 EST contigs, 4303 remained alone and 619 clustered with only two sequences. The average lengths of sequences and contigs are around 530 bp and the distribution displays an enrichment in sequences longer than 500 (data not shown) that allows credible blast gene identification. This high number of original S. cephaloptera transcripts and the large size of sequences both plead for the reliability of our library.
Nemertea and Platyhelminthes have traditionally been grouped together because they possess a so-called acoelomate organization, but lateral vessels and rhynchocoel of nemerteans have been regarded as coelomic cavities. Additionally, both taxa show spiral cleavage patterns prompting the placement of Nemertea as sister to coelomate Protostomia, that is, either to Neotrochozoa (Mollusca and Annelida) or to Teloblastica (Neotrochozoa plus Arthropoda). Some workers maintain a sister group relationship of Nemertea and Platyhelminthes as Parenchymia because of an assumed homology of Götte's and Müller's larvae of polyclad Platyhelminthes and the pilidium larvae of heteronemerteans. So far, molecular data were only able to significantly reject a sister group relationship to Teloblastica. Although phylogenomic data are available for Platyhelminthes, Annelida, Mollusca, and Arthropoda, they are lacking for Nemertea. Herein, we present the first analysis specifically addressing nemertean phylogenetic position using phylogenomic data. More specifically, we collected expressed sequence tag data from Lineus viridis (O.F. Müller, 1774) and combined it with available data to produce a data set of 9,377 amino acid positions from 60 ribosomal proteins. Maximum likelihood analyses and Bayesian inferences place Nemertea in a clade together with Annelida and Mollusca. Furthermore, hypothesis testing significantly rejected a sister group relationship to either Platyhelminthes or Teloblastica. The Coelomata hypothesis, which groups coelomate taxa together to the exclusion of acoelomate and pseudocoelomate taxa, is not congruent with our results. Thus, the supposed acoelomate organization evolved independently in Nemertea and Platyhelminthes. In Nemertea, evolution of acoely is most likely due to a secondary reduction of the coelom as it is found in certain species of Mollusca and Annelida. Though looking very similar, the Götte's and Müller's larvae of polyclad Platyhelminthes are not homologous to the pilidium larvae of heteronemerteans. Finally, the convergent evolution of segmentation in Annelida and Arthropoda is further substantiated.