Nathan V. Whelan

Error, signal, and the placement of Ctenophora sister to all other animals

Significance Traditional interpretation of animal phylogeny suggests traits, such as mesoderm, muscles, and neurons, evolved only once given the assumed placement of sponges as sister to all other animals. In contrast, placement of ctenophores as the first branching animal lineage raises the possibility of multiple origins of many complex traits considered important for animal diversification and success. We consider sources of potential error and increase taxon sampling to find a single, statistically robust placement of ctenophores as our most distant animal relatives, contrary to the traditional understanding of animal phylogeny. Furthermore, ribosomal protein genes are identified as creating conflict in signal that caused some past studies to recover a sister relationship between ctenophores and cnidarians. Elucidating relationships among early animal lineages has been difficult, and recent phylogenomic analyses place Ctenophora sister to all other extant animals, contrary to the traditional view of Porifera as the earliest-branching animal lineage. To date, phylogenetic support for either ctenophores or sponges as sister to other animals has been limited and inconsistent among studies. Lack of agreement among phylogenomic analyses using different data and methods obscures how complex traits, such as epithelia, neurons, and muscles evolved. A consensus view of animal evolution will not be accepted until datasets and methods converge on a single hypothesis of early metazoan relationships and putative sources of systematic error (e.g., long-branch attraction, compositional bias, poor model choice) are assessed. Here, we investigate possible causes of systematic error by expanding taxon sampling with eight novel transcriptomes, strictly enforcing orthology inference criteria, and progressively examining potential causes of systematic error while using both maximum-likelihood with robust data partitioning and Bayesian inference with a site-heterogeneous model. We identified ribosomal protein genes as possessing a conflicting signal compared with other genes, which caused some past studies to infer ctenophores and cnidarians as sister. Importantly, biases resulting from elevated compositional heterogeneity or elevated substitution rates are ruled out. Placement of ctenophores as sister to all other animals, and sponge monophyly, are strongly supported under multiple analyses, herein.

Employing Phylogenomics to Resolve the Relationships among Cnidarians, Ctenophores, Sponges, Placozoans, and Bilaterians

Despite an explosion in the amount of sequence data, phylogenomics has failed to settle controversy regarding some critical nodes on the animal tree of life. Understanding relationships among Bilateria, Ctenophora, Cnidaria, Placozoa, and Porifera is essential for studying how complex traits such as neurons, muscles, and gastrulation have evolved. Recent studies have cast doubt on the historical viewpoint that sponges are sister to all other animal lineages with recent studies recovering ctenophores as sister. However, the ctenophore-sister hypothesis has been criticized as unrealistic and caused by systematic error. We review past phylogenomic studies and potential causes of systematic error in an effort to identify areas that can be improved in future studies. Increased sampling of taxa, less missing data, and a priori removal of sequences and taxa that may cause systematic error in phylogenomic inference will likely be the most fruitful areas of focus when assembling future datasets. Ultimately, we foresee metazoan relationships being resolved with higher support in the near future, and we caution against dismissing novel hypotheses merely because they conflict with historical viewpoints of animal evolution.

Ctenophore relationships and their placement as the sister group to all other animals

Ctenophora, comprising approximately 200 described species, is an important lineage for understanding metazoan evolution and is of great ecological and economic importance. Ctenophore diversity includes species with unique colloblasts used for prey capture, smooth and striated muscles, benthic and pelagic lifestyles, and locomotion with ciliated paddles or muscular propulsion. However, the ancestral states of traits are debated and relationships among many lineages are unresolved. Here, using 27 newly sequenced ctenophore transcriptomes, publicly available data and methods to control systematic error, we establish the placement of Ctenophora as the sister group to all other animals and refine the phylogenetic relationships within ctenophores. Molecular clock analyses suggest modern ctenophore diversity originated approximately 350 million years ago ± 88 million years, conflicting with previous hypotheses, which suggest it originated approximately 65 million years ago. We recover Euplokamis dunlapae—a species with striated muscles—as the sister lineage to other sampled ctenophores. Ancestral state reconstruction shows that the most recent common ancestor of extant ctenophores was pelagic, possessed tentacles, was bioluminescent and did not have separate sexes. Our results imply at least two transitions from a pelagic to benthic lifestyle within Ctenophora, suggesting that such transitions were more common in animal diversification than previously thought.Newly sequenced transcriptomes are combined with existing data to establish Ctenophora as the sister group to all other animals and suggest a radiation around 350 Ma as well as multiple transitions from a pelagic to a benthic lifestyle.

Who Let the CAT Out of the Bag? Accurately Dealing with Substitutional Heterogeneity in Phylogenomic Analyses

&NA; As phylogenetic datasets have increased in size, site‐heterogeneous substitution models such as CAT‐F81 and CAT‐GTR have been advocated in favor of other models because they purportedly suppress long‐branch attraction (LBA). These models are two of the most commonly used models in phylogenomics, and they have been applied to a variety of taxa, ranging from Drosophila to land plants. However, many arguments in favor of CAT models have been based on tenuous assumptions about the true phylogeny, rather than rigorous testing with known trees via simulation. Moreover, CAT models have not been compared to other approaches for handling substitutional heterogeneity such as data partitioning with site‐homogeneous substitution models. We simulated amino acid sequence datasets with substitutional heterogeneity on a variety of tree shapes including those susceptible to LBA. Data were analyzed with both CAT models and partitioning to explore model performance; in total over 670,000 CPU hours were used, of which over 97% was spent running analyses with CAT models. In many cases, all models recovered branching patterns that were identical to the known tree. However, CAT‐F81 consistently performed worse than other models in inferring the correct branching patterns, and both CAT models often overestimated substitutional heterogeneity. Additionally, reanalysis of two empirical metazoan datasets supports the notion that CAT‐F81 tends to recover less accurate trees than data partitioning and CAT‐GTR. Given these results, we conclude that partitioning and CAT‐GTR perform similarly in recovering accurate branching patterns. However, computation time can be orders of magnitude less for data partitioning, with commonly used implementations of CAT‐GTR often failing to reach completion in a reasonable time frame (i.e., for Bayesian analyses to converge). Practices such as removing constant sites and parsimony uninformative characters, or using CAT‐F81 when CAT‐GTR is deemed too computationally expensive, cannot be logically justified. Given clear problems with CAT‐F81, phylogenies previously inferred with this model should be reassessed.

Nemertean toxin genes revealed through transcriptome sequencing

Nemerteans are one of few animal groups that have evolved the ability to utilize toxins for both defense and subduing prey, but little is known about specific nemertean toxins. In particular, no study has identified specific toxin genes even though peptide toxins are known from some nemertean species. Information about toxin genes is needed to better understand evolution of toxins across animals and possibly provide novel targets for pharmaceutical and industrial applications. We sequenced and annotated transcriptomes of two free-living and one commensal nemertean and annotated an additional six publicly available nemertean transcriptomes to identify putative toxin genes. Approximately 63–74% of predicted open reading frames in each transcriptome were annotated with gene names, and all species had similar percentages of transcripts annotated with each higher-level GO term. Every nemertean analyzed possessed genes with high sequence similarities to known animal toxins including those from stonefish, cephalopods, and sea anemones. One toxin-like gene found in all nemerteans analyzed had high sequence similarity to Plancitoxin-1, a DNase II hepatotoxin that may function well at low pH, which suggests that the acidic body walls of some nemerteans could work to enhance the efficacy of protein toxins. The highest number of toxin-like genes found in any one species was seven and the lowest was three. The diversity of toxin-like nemertean genes found here is greater than previously documented, and these animals are likely an ideal system for exploring toxin evolution and industrial applications of toxins.

Miscues misplace sponges.

In contrast to several recent publications (1⇓⇓⇓–5), Pisani et al. (6) claim that ( i ) genomic data do not support ctenophores as the sister group to other animals and ( ii ) independent evolution of complex features (e.g., neurons, muscles) in ctenophores is not supported. These claims are based on selective interpretation, subjective criteria, and improper assumptions about original analyses. Pisani et al.’s (6) conclusions about evolution improperly treat complex traits as single characters. Features such as neuromusclular systems possess several independently evolving cell lineages and mechanisms. Given that underlying components of these systems use remarkably different molecular machinery (1), whether Porifera or Ctenophora is sister to other animals is irrelevant to … [↵][1]2To whom correspondence may be addressed. Email: ken{at}auburn.edu or nwhelan{at}auburn.edu. [1]: #xref-corresp-1-1

Phylogenomics of tubeworms (Siboglinidae, Annelida) and comparative performance of different reconstruction methods

Deep‐sea tubeworms (Annelida, Siboglinidae) represent dominant species in deep‐sea chemosynthetic communities (e.g. hydrothermal vents and cold methane seeps) and occur in muddy sediments and organic falls. Siboglinids lack a functional digestive tract as adults, and they rely on endosymbiotic bacteria for energy, making them of evolutionary and physiological interest. Despite their importance, inferred evolutionary history of this group has been inconsistent among studies based on different molecular markers. In particular, placement of bone‐eating Osedax worms has been unclear in part because of their distinctive biology, including harbouring heterotrophic bacteria as endosymbionts, displaying extreme sexual dimorphism and exhibiting a distinct body plan. Here, we reconstructed siboglinid evolutionary history using 12 newly sequenced transcriptomes. We parsed data into three data sets that accommodated varying levels of missing data, and we evaluate effects of missing data on phylogenomic inference. Additionally, several multispecies‐coalescent approaches and Bayesian concordance analysis (BCA) were employed to allow for a comparison of results to a supermatrix approach. Every analysis conducted herein strongly supported Osedax being most closely related to the Vestimentifera and Sclerolinum clade, rather than Frenulata, as previously reported. Importantly, unlike previous studies, the alternative hypothesis that frenulates and Osedax are sister groups to one another was explicitly rejected by an approximately unbiased (AU) test. Furthermore, although different methods showed largely congruent results, we found that a supermatrix method using data partitioning with site‐homogenous models potentially outperformed a supermatrix method using the CAT‐GTR model and multispecies‐coalescent approaches when the amount of missing data varies in a data set and when taxa susceptible to LBA are included in the analyses.

Discovery and evolution of novel hemerythrin genes in annelid worms

BackgroundDespite extensive study on hemoglobins and hemocyanins, little is known about hemerythrin (Hr) evolutionary history. Four subgroups of Hrs have been documented, including: circulating Hr (cHr), myohemerythrin (myoHr), ovohemerythrin (ovoHr), and neurohemerythrin (nHr). Annelids have the greatest diversity of oxygen carrying proteins among animals and are the only phylum in which all Hr subgroups have been documented. To examine Hr diversity in annelids and to further understand evolution of Hrs, we employed approaches to survey annelid transcriptomes in silico.ResultsSequences of 214 putative Hr genes were identified from 44 annelid species in 40 different families and Bayesian inference revealed two major clades with strong statistical support. Notably, the topology of the Hr gene tree did not mirror the phylogeny of Annelida as presently understood, and we found evidence of extensive Hr gene duplication and loss in annelids. Gene tree topology supported monophyly of cHrs and a myoHr clade that included nHrs sequences, indicating these designations are functional rather than evolutionary.ConclusionsThe presence of several cHrs in early branching taxa suggests that a variety of Hrs were present in the common ancestor of extant annelids. Although our analysis was limited to expressed-coding regions, our findings demonstrate a greater diversity of Hrs among annelids than previously reported.

Toll-like receptor pathway evolution in deuterostomes

Significance Innate immunity provides critical defense against pathogen invasion, and mutations in its cellular mechanisms have been implicated in autoimmunity, immune suppression, and other disease-producing conditions. However, knowledge of innate immunity pathways is largely biased toward model species. Thus, evolutionary interpretations suffer from large taxonomic gaps that ultimately weaken the strength of evolutionary inference. Our phylogenetic approach shows that the molecular machinery of the canonical TLR pathway was present in the last deuterostome ancestor before the rise of chordate lineages. Thus, TLR pathways with multiple gene–gene interactions have been conserved for more than 500 million years within vertebrates. Moreover, we provide evidence suggesting TLR3 may represent an ancient, evolutionarily conserved molecular interface for viral immune stimulation present across Deuterostomia. Animals have evolved an array of pattern-recognition receptor families essential for recognizing conserved molecular motifs characteristic of pathogenic microbes. One such family is the Toll-like receptors (TLRs). On pathogen binding, TLRs initiate specialized cytokine signaling catered to the class of invading pathogen. This signaling is pivotal for activating adaptive immunity in vertebrates, suggesting a close evolutionary relationship between innate and adaptive immune systems. Despite significant advances toward understanding TLR-facilitated immunity in vertebrates, knowledge of TLR pathway evolution in other deuterostomes is limited. By analyzing genomes and transcriptomes across 37 deuterostome taxa, we shed light on the evolution and diversity of TLR pathway signaling elements. Here, we show that the deuterostome ancestor possessed a molecular toolkit homologous to that which drives canonical MYD88-dependent TLR signaling in contemporary mammalian lineages. We also provide evidence that TLR3-facilitated antiviral signaling predates the origin of its TCAM1 dependence recognized in the vertebrates. SARM1, a negative regulator of TCAM1-dependent pathways in vertebrates, was also found to be present across all major deuterostome lineages despite the apparent absence of TCAM1 in invertebrate deuterostomes. Whether the presence of SARM1 is the result of its role in immunity regulation, neuron physiology, or a function of both is unclear. Additionally, Bayesian phylogenetic analyses corroborate several lineage-specific TLR gene expansions in urchins and cephalochordates. Importantly, our results underscore the need to sample across taxonomic groups to understand evolutionary patterns of the innate immunity foundation on which complex immunological novelties arose.

Radular Morphology of Extinct Pleurocerids (Gastropoda: Cerithioidea: Pleuroceridae)

Abstract: The freshwater gastropod family Pleuroceridae suffers from a disproportionately high number of imperiled species and recent extinctions. As pleurocerid diversity has been lost, so too has our ability to study the biology of these animals. However, many extinct species were deposited in natural history collections before their demise. I extracted radulae from dried tissue left in shells of seven extinct species in three genera (Leptoxis Rafinesque, 1819, Gyrotoma Shuttleworth, 1845, Lithasia Haldeman, 1840) to gain insights into the morphological differences separating species and provide data for future researchers. There were notable intergeneric and interspecific differences in radular morphology such as shape of cusps (e.g., dagger-like vs. blunt) and number of denticles on the rachidian, lateral, and marginal teeth. Interestingly, the degree of radular differences among Leptoxis spp. likely corroborates previous hypotheses that the genus is not a natural group. These data are a resource for future studies and should aid in determining the feeding habits and relationships of extinct pleurocerids.