The Bryozoan Cauloramphus magnus (Cheilostomata: Calloporidae) in Northern Japan Includes Multiple, Co-occurring Cryptic Species.

The cheilostome bryozoan Cauloramphus magnus is common in the rocky intertidal habitat from southeastern Alaska to northern Japan. We examined its phylogeography by analyzing 576 bp of the mitochondrial COI (cox1) gene sequenced for 298 colonies from 16 localities in northern Japan. A maximum-likelihood phylogeny detected three main clades (A, B, C). Clades A and B occurred throughout the study area but differed in frequency, haplotype diversity, and haplotype distribution; each resolved into three divergent subclades (AI-III, BI-III). Clade A shared none among 15 haplotypes between the Pacific and Sea of Japan sides of Hokkaido. In contrast, Clade B (29 haplotypes) was thrice as common as Clade A among samples, with haplotype B28 common on both sides. Divergent Clade C (nine haplotypes) was detected only at Rumoi. K2P divergences of 12.3-28.3% among Clades A-C suggest these are distinct biological species, a conclusion supported by different inferred evolutionary histories. A bPTP species delimitation analysis indicated nine phylogenetic species among the sequences included in our phylogeny (AI-III, BI-III, C, and one specimen each from Alaska and the Commander Islands), with K2P divergences of 3.9-6.5% among subclades in A or B. Statistical and principal components analyses suggested weak morphological differentiation between Clades A + B and C, although overlapping ranges of measurements preclude identification to clade; these three clades are morphologically cryptic. For taxonomy, we suggest retaining the name C. magnus for lineages within this species complex across its range, followed by a clade designation, if known.

Phylogenomics reveals deep relationships and diversification within phylactolaemate bryozoans.

Bryozoans are mostly sessile colonial invertebrates that inhabit all kinds of aquatic ecosystems. Extant bryozoan species fall into two clades with one of them, Phylactolaemata, being the only exclusively freshwater clade. Phylogenetic relationships within the class Phylactolaemata have long been controversial owing to their limited distinguishable characteristics that reflect evolutionary relationships. Here, we present the first phylogenomic analysis of Phylactolaemata using transcriptomic data combined with dense taxon sampling of six families to better resolve the interrelationships and to estimate divergence time. Using maximum-likelihood and Bayesian inference approaches, we recovered a robust phylogeny for Phylactolaemata in which the interfamilial relationships are fully resolved. We show Stephanellidae is the sister taxon of all other phylactolaemates and confirm that Lophopodidae represents the second offshoot within the phylactolaemate tree. Plumatella fruticosa clearly falls outside Plumatellidae as previous investigations have suggested, and instead clusters with Pectinatellidae and Cristatellidae as the sister taxon of Fredericellidae. Our results demonstrate that cryptic speciation is very likely in F. sultana and in two species of Plumatella (P. repens and P. casmiana). Divergence time estimates show that Phylactolaemata appeared at the end of the Ediacaran and started to diverge in the Silurian, although confidence intervals were large for most nodes. The radiation of most extant phylactolaemate families occurred mainly in the Palaeogene and Neogene highlighting post-extinction diversification.

Morphospecies and molecular diversity of 'lace corals': the genus Reteporella (Bryozoa: Cheilostomatida) in the central North Atlantic Azores Archipelago.

BACKGROUND: As in most bryozoans, taxonomy and systematics of species in the genus Reteporella Busk, 1884 (family Phidoloporidae) has hitherto almost exclusively been based on morphological characters. From the central North Atlantic Azores Archipelago, nine Reteporella species have historically been reported, none of which have as yet been revised. Aiming to characterise the diversity and biogeographic distribution of Azorean Reteporella species, phylogenetic reconstructions were conducted on a dataset of 103 Azorean Reteporella specimens, based on the markers cytochrome C oxidase subunit 1, small and large ribosomal RNA subunits. Morphological identification was based on scanning electron microscopy and complemented the molecular inferences. RESULTS: Our results reveal two genetically distinct Azorean Reteporella clades, paraphyletic to eastern Atlantic and Mediterranean taxa. Moreover, an overall concordance between morphological and molecular species can be shown, and the actual bryozoan diversity in the Azores is greater than previously acknowledged as the dataset comprises three historically reported species and four putative new taxa, all of which are likely to be endemic. The inclusion of Mediterranean Reteporella specimens also revealed new species in the Adriatic and Ligurian Sea, whilst the inclusion of additional phidoloporid taxa hints at the non-monophyly of the genus Reteporella. CONCLUSION: Being the first detailed genetic study on the genus Reteporella, the high divergence levels inferred within the genus Reteporella and family Phidoloporidae calls for the need of further revision. Nevertheless, the overall concordance between morphospecies and COI data suggest the potential adequacy of a 3% cut-off to distinguish Reteporella species. The discovery of new species in the remote Azores Archipelago as well as in the well-studied Mediterranean Sea indicates a general underestimation of bryozoan diversity. This study constitutes yet another example of the importance of integrative taxonomical approaches on understudied taxa, contributing to cataloguing genetic and morphological diversity.

Developmental Process of a Heterozooid: Avicularium Formation in a Bryozoan, Bugulina californica.

In bryozoans (phylum Bryozoa), representative colonial animals mostly found in marine environments, some species possess different types of individuals (heterozooids) specialized in different functions such as defense or structural support for their colonies. Among them, the best-known heterozooids are the avicularia, known to function as defenders. The differentiation processes of heterozooids, including avicularia, should be important keys to understand the evolutionary significance of bryozoans. However, the developmental process of avicularium formation remains to be fully understood. In this study, therefore, in order to understand the detailed developmental process and timing of avicularium formation, extensive observations were carried out in a bryozoan species, Bugulina californica (Cheilostomata, Bugulidae), that possesses adventitious avicularia, by performing stereomicroscopy on live materials, in addition to scanning electron microscopy and histological observations. The whole process can be divided into seven stages based on developmental events. Especially notably, at the earlier stages, there are three major budding events that produce proliferating cell masses corresponding to primordial tissues: (1) budding of the peduncle cushion at the outer margin of the distal part of a young autozooid, (2) budding of the head-part primordium from the peduncle cushion, and (3) budding of the polypide inside the head part. Experimental control of temperature showed that 20°C would be the best to maintain B. californica colonies.

A broadly resolved molecular phylogeny of New Zealand cheilostome bryozoans as a framework for hypotheses of morphological evolution.

Larger molecular phylogenies based on ever more genes are becoming commonplace with the advent of cheaper and more streamlined sequencing and bioinformatics pipelines. However, many groups of inconspicuous but no less evolutionarily or ecologically important marine invertebrates are still neglected in the quest for understanding species- and higher-level phylogenetic relationships. Here, we alleviate this issue by presenting the molecular sequences of 165 cheilostome bryozoan species from New Zealand waters. New Zealand is our geographic region of choice as its cheilostome fauna is taxonomically, functionally and ecologically diverse, and better characterized than many other such faunas in the world. Using this most taxonomically broadly-sampled and statistically-supported cheilostome phylogeny comprising 214 species, when including previously published sequences, and 17 genes (2 nuclear and 15 mitochondrial) we tested several existing systematic hypotheses based solely on morphological observations. We find that lower taxonomic level hypotheses (species and genera) are robust while our inferred trees did not reflect current higher-level systematics (family and above), illustrating a general need for the rethinking of current hypotheses. To illustrate the utility of our new phylogeny, we reconstruct the evolutionary history of frontal shields (i.e., a calcified body-wall layer in ascus-bearing cheilostomes) and ask if its presence has any bearing on the diversification rates of cheilostomes.

Mitochondrial gene order of the freshwater bryozoan Cristatella mucedo retains ancestral lophotrochozoan features.

Bryozoans are aquatic colonial suspension-feeders abundant in many marine and freshwater benthic communities. At the same time, the phylum is under studied on both morphological and molecular levels, and its position on the metazoan tree of life is still disputed. Bryozoa include the exclusively marine Stenolaemata, predominantly marine Gymnolaemata and exclusively freshwater Phylactolaemata. Here we report the mitochondrial genome of the phylactolaemate bryozoan Cristatella mucedo. This species has the largest (21,008 bp) of all currently known bryozoan mitogenomes, containing a typical metazoan gene compendium as well as a number of non-coding regions, three of which are longer than 1500 bp. The trnS1/trnG/nad3 region is presumably duplicated in this species. Comparative analysis of the gene order in C. mucedo and another phylactolaemate bryozoan, Pectinatella magnifica, confirmed their close relationships, and revealed a stronger similarity to mitogenomes of phoronids and other lophotrochozoan species than to marine bryozoans, indicating the ancestral nature of their gene arrangement. We suggest that the ancestral gene order underwent substantial changes in different bryozoan cladesshowing mosaic distribution of conservative gene blocks regardless of their phylogenetic position. Altogether, our results support the early divergence of Phylactolaemata from the rest of Bryozoa.

Comparative transcriptomics and host-specific parasite gene expression profiles inform on drivers of proliferative kidney disease.

The myxozoan parasite, Tetracapsuloides bryosalmonae has a two-host life cycle alternating between freshwater bryozoans and salmonid fish. Infected fish can develop Proliferative Kidney Disease, characterised by a gross lymphoid-driven kidney pathology in wild and farmed salmonids. To facilitate an in-depth understanding of T. bryosalmonae-host interactions, we have used a two-host parasite transcriptome sequencing approach in generating two parasite transcriptome assemblies; the first derived from parasite spore sacs isolated from infected bryozoans and the second from infected fish kidney tissues. This approach was adopted to minimize host contamination in the absence of a complete T. bryosalmonae genome. Parasite contigs common to both infected hosts (the intersect transcriptome; 7362 contigs) were typically AT-rich (60-75% AT). 5432 contigs within the intersect were annotated. 1930 unannotated contigs encoded for unknown transcripts. We have focused on transcripts encoding proteins involved in; nutrient acquisition, host-parasite interactions, development, cell-to-cell communication and proteins of unknown function, establishing their potential importance in each host by RT-qPCR. Host-specific expression profiles were evident, particularly in transcripts encoding proteases and proteins involved in lipid metabolism, cell adhesion, and development. We confirm for the first time the presence of homeobox proteins and a frizzled homologue in myxozoan parasites. The novel insights into myxozoan biology that this study reveals will help to focus research in developing future disease control strategies.

Artificial habitats induce plasticity in colonies of the marine bryozoan Schizoporella errata.

Modular organization provides flexibility for colonial animals to deal with variable and unpredictable environmental conditions since each module has specific tasks within the colony, such as feeding, defending or reproducing. Depending on the selecting pressures, sessile organisms may phenotypically adjust the morphology of each module or modify their density, increasing individual fitness. Here we used the marine bryozoan Schizoporella errata (Cheilostomata, Schizoporellidae) to test how the divergent conditions between two artificial habitats, the location inside a marina (IM) and the external wall of the breakwater (BW), affect colony size and the density of the distinct modules. The density of avicularia and ovicells, modules related to defense and reproduction, respectively, did not differ between habitats. However, colonies growing in the turbulent waters of BW were, in general, larger and had higher density of feeding autozooids than those at IM. Reciprocal transplants of bryozoan clones indicated that trait variation is genotype-dependent but varies according to the environmental conditions at the assigned location. The occurrence of larger colonies with more zooids in BW is probably linked to the easier feeding opportunity offered by the small diffusive boundary layer around the colony at this location. Since in colonial polymorphic organisms each module (zooid) performs a specific function, the phenotypic response is not uniform across colonies, affecting only those modules that are susceptible to variations in the main selective pressures. Understanding the importance of colony-level plasticity is relevant to predict how modularity will contribute to organisms to deal with human-induced environmental changes in coastal habitats.

Draft genome of Bugula neritina, a colonial animal packing powerful symbionts and potential medicines.

Many animal phyla have no representatives within the catalog of whole metazoan genome sequences. This dataset fills in one gap in the genome knowledge of animal phyla with a draft genome of Bugula neritina (phylum Bryozoa). Interest in this species spans ecology and biomedical sciences because B. neritina is the natural source of bioactive compounds called bryostatins. Here we present a draft assembly of the B. neritina genome obtained from PacBio and Illumina HiSeq data, as well as genes and proteins predicted de novo and verified using transcriptome data, along with the functional annotation. These sequences will permit a better understanding of host-symbiont interactions at the genomic level, and also contribute additional phylogenomic markers to evaluate Lophophorate or Lophotrochozoa phylogenetic relationships. The effort also fits well with plans to ultimately sequence all orders of the Metazoa.

The visual pigment xenopsin is widespread in protostome eyes and impacts the view on eye evolution.

Photoreceptor cells in the eyes of Bilateria are often classified into microvillar cells with rhabdomeric opsin and ciliary cells with ciliary opsin, each type having specialized molecular components and physiology. First data on the recently discovered xenopsin point towards a more complex situation in protostomes. In this study, we provide clear evidence that xenopsin enters cilia in the eye of the larval bryozoan Tricellaria inopinata and triggers phototaxis. As reported from a mollusc, we find xenopsin coexpressed with rhabdomeric-opsin in eye photoreceptor cells bearing both microvilli and cilia in larva of the annelid Malacoceros fuliginosus. This is the first organism known to have both xenopsin and ciliary opsin, showing that these opsins are not necessarily mutually exclusive. Compiling existing data, we propose that xenopsin may play an important role in many protostome eyes and provides new insights into the function, evolution, and possible plasticity of animal eye photoreceptor cells.

Differential Gene Expression Between Polymorphic Zooids of the Marine Bryozoan Bugulina stolonifera.

Bryozoans are a diverse phylum of marine and freshwater colonial invertebrates containing approximately 6,300 described living species. Bryozoans grow by budding new physiologically connected colony members (zooids) from a founding individual that forms from a metamorphosed larva. In some species these zooids come in different shapes and sizes and are specialized to serve different tasks within the colony. A complex interaction of genotype, environment, and developmental pathway shapes zooid fate, however, the specific mechanisms underlying the establishment of this division of labor remain unknown. Here, the first characterization of differential gene expression between polymorphic zooids of a bryozoan colony is presented. The development of different zooid types of lab-cultured Bugulina stolonifera colonies including feeding autozooids, avicularia (derived non-feeding zooids that are homologous to feeding autozooids but shaped like a bird's beak), and rhizoids (a branching network of non-feeding anchoring zooids) was explored using RNA sequencing, de novo transcriptome assembly, and differential gene expression analyses. High throughput sequencing of cDNA libraries yielded an average of 14.9 ± 1.3 (SE) million high-quality paired-end reads per sample. Data for the first de novo transcriptome assemblies of B. stolonifera and the first characterization of genes involved in the formation and maintenance of zooid types within a bryozoan colony are presented. In a comparison between autozooid and avicularium tissues, 1,097 significant differentially expressed genes were uncovered. This work provides a much-needed foundation for understanding the mechanisms involved in the development of polymorphic zooids and the establishment of division of labor in bryozoans.

Transcriptome Analysis Elucidates the Key Responses of Bryozoan Fredericella sultana during the Development of Tetracapsuloides bryosalmonae (Myxozoa).

Bryozoans are sessile, filter-feeding, and colony-building invertebrate organisms. Fredericella sultana is a well known primary host of the myxozoan parasite Tetracapsuloides bryosalmonae. There have been no attempts to identify the cellular responses induced in F. sultana during the T. bryosalmonae development. We therefore performed transcriptome analysis with the aim of identifying candidate genes and biological pathways of F. sultana involved in the response to T. bryosalmonae. A total of 1166 differentially up- and downregulated genes were identified in the infected F. sultana. Gene ontology of biological processes of upregulated genes pointed to the involvement of the innate immune response, establishment of protein localization, and ribosome biogenesis, while the downregulated genes were involved in mitotic spindle assembly, viral entry into the host cell, and response to nitric oxide. Eukaryotic Initiation Factor 2 signaling was identified as a top canonical pathway and MYCN as a top upstream regulator in the differentially expressed genes. Our study provides the first transcriptional profiling data on the F. sultana zooid's response to T. bryosalmonae. Pathways and upstream regulators help us to understand the complex interplay in the infected F. sultana. The results will facilitate the elucidation of innate immune mechanisms of bryozoan and will lay a foundation for further analyses on bryozoan-responsive candidate genes, which will be an important resource for the comparative analysis of gene expression in bryozoans.

Community assembly in a modular organism: the impact of environmental filtering on bryozoan colony form and polymorphism.

Understanding community assembly is a key goal in community ecology. Environmental filtering influences community assembly by excluding ill-adapted species, resulting in communities with similar functional traits. An RLQ (a four-way ordination) analysis incorporating spatial data was run on a data set of 642 species of cheilostomes (Bryozoa) from 779 New Zealand sites, and results were compared to trends in other sessile, epibenthic taxa. This revealed environmental filtering of colony form: encrusting-cemented taxa were predominant in shallow environments with hard substrata (<200 m), while erect-rooted taxa characterized deeper environments with soft substrata (>200 m). Furthermore, erect taxa found in shallow environments with high current speeds were typically jointed. Polymorphism also followed environmental gradients. External ovicells (brood chambers) were more common in deeper, low-oxygen water than immersed and internal ovicells. This may reflect the oxygen needs of the embryo or increased predation intensity in shallow environments. Bryozoans with costae tended to be found in deeper water as well, while bryozoans with calcified frontal shields were found in shallow environments with a higher concentration of CaCO3 . Avicularia did not appear to be related to environmental conditions, and changes in pivot bar structure with depth likely represent a phylogenetic signal. The importance of substratum type as a strict environmental filter suggests that anchoring structures, like rootlets, may be "key innovations" for other sessile, epibenthic taxa like sponges and ascidians.

Global Distribution and Variation of the Invasive Cheilostome Bryozoan Cribrilina mutabilis.

Viable populations of the cheilostome bryozoan Cribrilina mutabilis Ito, Onishi & Dick exist in the NW Pacific (Russian Far East and northern Japan), NE Atlantic (Scandinavia and Scotland), and NW Atlantic (Maine, USA). The first NE and NW Atlantic records are from Norway (2008) and Casco Bay, Maine, USA (2018), respectively, indicating a relatively recent introduction to the region. Mitochondrial COI gene sequences from North Atlantic populations (Sweden, Norway, and Maine) showed two haplotypes differing by one substitution, but differed from two haplotypes from Akkeshi, northern Japan, by 6-8 substitutions. North Atlantic populations differed morphologically from the Akkeshi population in that some zooids formed a suboral projection, and frontal zooids were more common. While C. mutabilis in northern Japan has been found only on natural or artificial eelgrass (Zostera marina), across its range it has been found on several species of algae, plastic panels and strips, several species of Zostera, and mollusc shells. Similar frequencies of heteromorphic zooids with differing degree of frontal wall calcification, i.e., R (rib)-, I (intermediate)-, and S (shield)-type zooids, in colonies on eelgrass at comparable times of the season and across populations suggest an innate response to seasonal environmental fluctuations, although zooid frequencies were different on non-eelgrass substrates. The increase in trans-Arctic shipping along the Northern Sea Route in recent decades, and previous documentation of C. mutabilis on ship hulls in the Sea of Japan, indicate a clear mechanism for anthropogenic introduction from the Far East to Europe in recent decades.

A genome-skimmed phylogeny of a widespread bryozoan family, Adeonidae.

BACKGROUND: Understanding the phylogenetic relationships among species is one of the main goals of systematic biology. Simultaneously, credible phylogenetic hypotheses are often the first requirement for unveiling the evolutionary history of traits and for modelling macroevolutionary processes. However, many non-model taxa have not yet been sequenced to an extent such that statistically well-supported molecular phylogenies can be constructed for these purposes. Here, we use a genome-skimming approach to extract sequence information for 15 mitochondrial and 2 ribosomal operon genes from the cheilostome bryozoan family, Adeonidae, Busk, 1884, whose current systematics is based purely on morphological traits. The members of the Adeonidae are, like all cheilostome bryozoans, benthic, colonial, marine organisms. Adeonids are also geographically widely-distributed, often locally common, and are sometimes important habitat-builders. RESULTS: We successfully genome-skimmed 35 adeonid colonies representing 6 genera (Adeona, Adeonellopsis, Bracebridgia, Adeonella, Laminopora and Cucullipora). We also contributed 16 new, circularised mitochondrial genomes to the eight previously published for cheilostome bryozoans. Using the aforementioned mitochondrial and ribosomal genes, we inferred the relationships among these 35 samples. Contrary to some previous suggestions, the Adeonidae is a robustly supported monophyletic clade. However, the genera Adeonella and Laminopora are in need of revision: Adeonella is polyphyletic and Laminopora paraphyletically forms a clade with some Adeonella species. Additionally, we assign a sequence clustering identity using cox1 barcoding region of 99% at the species and 83% at the genus level. CONCLUSIONS: We provide sequence data, obtained via genome-skimming, that greatly increases the resolution of the phylogenetic relationships within the adeonids. We present a highly-supported topology based on 17 genes and substantially increase availability of circularised cheilostome mitochondrial genomes, and highlight how we can extend our pipeline to other bryozoans.

Modularity is the mother of invention: a review of polymorphism in bryozoans.

Modularity is a fundamental concept in biology. Most taxa within the colonial invertebrate phylum Bryozoa have achieved division of labour through the development of specialized modules (polymorphs), and this group is perhaps the most outstanding exemplar of the phenomenon. We provide a comprehensive description of the diversity, morphology and function of these polymorphs and the significance of modularity to the evolutionary success of the phylum, which has >21000 described fossil and living species. Modular diversity likely arose from heterogeneous microenvironmental conditions, and cormidia (repeated clusters of associated modules) are an emergent property of the cue thresholds governing zooid plasticity. Polymorphs in a colony have, during phylogeny, transitioned into associated non-zooidal structures (appendages), increasing colonial integration. While the level of module compartmentalization is important for the evolution of bryozoan polymorphism, it may be less influential for other colonial invertebrates.

Divergent responses to warming of two common co-occurring Mediterranean bryozoans.

Climate change threatens the structure and function of marine ecosystems, highlighting the importance of understanding the response of species to changing environmental conditions. However, thermal tolerance determining the vulnerability to warming of many abundant marine species is still poorly understood. In this study, we quantified in the field the effects of a temperature anomaly recorded in the Mediterranean Sea during the summer of 2015 on populations of two common sympatric bryozoans, Myriapora truncata and Pentapora fascialis. Then, we experimentally assessed their thermal tolerances in aquaria as well as different sublethal responses to warming. Differences between species were found in survival patterns in natural populations, P. fascialis showing significantly lower survival rates than M. truncata. The thermotolerance experiments supported field observations: P. fascialis started to show signs of necrosis when the temperature was raised to 25-26 °C and completely died between 28-29 °C, coinciding with the temperature when we observed first signs of necrosis in M. truncata. The results from this study reflect different responses to warming between these two co-occurring species, highlighting the importance of combining multiple approaches to assess the vulnerability of benthic species in a changing climate world.

Chemoenzymatic Dissection of Polyketide ß-Branching in the Bryostatin Pathway.

ß-Branching is an expansion upon canonical polyketide synthase extension that allows for the installation of diverse chemical moieties in several natural products. Several of these moieties are unique among natural products, including the two vinyl methylesters found in the core structure of bryostatins. This family of molecules is derived from an obligate bacterial symbiont of a sessile marine bryozoan, Bugula neritina. Within this family, bryostatin 1 has been investigated as an anticancer, neuroprotective, and immunomodulatory compound. We have turned to the biosynthetic gene cluster within the bacterial symbiont to investigate the biosynthesis of bryostatins. Recent sequencing efforts resulted in the annotation of two missing genes: bryT and bryU. Using novel chemoenzymatic techniques, we have validated these as the missing enoyl-CoA hydratase and donor acyl carrier protein, essential components of the ß-branching cassette of the bryostatin pathway. Together, this cassette installs the vinyl methylester moieties essential to the activity of bryostatins.

Phylogenetically Widespread Polyembryony in Cyclostome Bryozoans and the Protracted Asynchronous Release of Clonal Brood-Mates.

Polyembryony-the production of multiple cloned embryos from a single fertilised egg-is a seemingly paradoxical combination of reproductive modes that nevertheless persists in diverse taxa. We document features of polyembryony in the Cyclostomata (Bryozoa)-an ancient order of modular colonial marine invertebrates-that suggest a substantial reduction in the paradoxical nature of this enigmatic reproductive mode. Firstly, we provide molecular evidence for polyembryony in three exemplar species, supporting the widely cited inference that polyembryony characterises the entire order. Secondly, genotyping demonstrates protracted release of cloned offspring from the primary embryo in a given gonozooid (chamber for embryonic incubation), thus exposing the same genotype to changing environmental conditions over time. Finally, we confirm that each gonozooid produces a distinct genotype, with each primary embryo being the result of a separate fertilisation event. We hypothesise that the sustained release of one or a few genotypes against varying environmental conditions achieves levels of risk-spreading similar to those in organisms that release multiple, unique genotypes at a single time. We argue that polyembryony, specifically with the production of a large number of progeny per fertilisation event, has been favoured in the Cyclostomata over long geological periods.

Phylogenomics of Lophotrochozoa with Consideration of Systematic Error.

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 data set. [Annelida; Brachiopoda; Bryozoa; Entoprocta; Mollusca; Nemertea; Phoronida; Platyzoa; Polyzoa; Spiralia; Trochozoa.].