A comprehensive molecular phylogeny of Dalytyphloplanida (Platyhelminthes: Rhabdocoela) reveals multiple escapes from the marine environment and origins of symbiotic relationships

In this study we elaborate the phylogeny of Dalytyphloplanida based on complete 18S rDNA (156 sequences) and partial 28S rDNA (125 sequences), using a Maximum Likelihood and a Bayesian Inference approach, in order to investigate the origin of a limnic or limnoterrestrial and of a symbiotic lifestyle in this large group of rhabditophoran flatworms. The results of our phylogenetic analyses and ancestral state reconstructions indicate that dalytyphloplanids have their origin in the marine environment and that there was one highly successful invasion of the freshwater environment, leading to a large radiation of limnic and limnoterrestrial dalytyphloplanids. This monophyletic freshwater clade, Limnotyphloplanida, comprises the taxa Dalyelliidae, Temnocephalida, and most Typhloplanidae. Temnocephalida can be considered ectosymbiotic Dalyelliidae as they are embedded within this group. Secondary returns to brackish water and marine environments occurred relatively frequently in several dalyeliid and typhloplanid taxa. Our phylogenies also show that, apart from the Limnotyphloplanida, there have been only few independent invasions of the limnic environment, and apparently these were not followed by spectacular speciation events. The distinct phylogenetic positions of the symbiotic taxa also suggest multiple origins of commensal and parasitic life strategies within Dalytyphloplanida. The previously established higher-level dalytyphloplanid clades are confirmed in our topologies, but many of the traditional families are not monophyletic. Alternative hypothesis testing constraining the monophyly of these families in the topologies and using the approximately unbiased test, also statistically rejects their monophyly

Problematic barcoding in flatworms: A case-study on monogeneans and rhabdocoels (Platyhelminthes)

Some taxonomic groups are less amenable to mitochondrial DNA barcoding than others. Due to the paucity of molecular information of understudied groups and the huge molecular diversity within flatworms, primer design has been hampered. Indeed, all attempts to develop universal flatworm-specific COI markers have failed so far. We demonstrate how high molecular variability and contamination problems limit the possibilities for barcoding using standard COI-based protocols in flatworms. As a consequence, molecular identification methods often rely on other widely applicable markers. In the case of Monogenea, a very diverse group of platyhelminth parasites, and Rhabdocoela, representing one-fourth of all free-living flatworm taxa, this has led to a relatively high availability of nuclear ITS and 18S/28S rDNA sequences on GenBank. In a comparison of the effectiveness in species assignment we conclude that mitochondrial and nuclear ribosomal markers perform equally well. In case intraspecific information is needed, rDNA sequences can guide the selection of the appropriate (i.e. taxon-specific) COI primers if available.This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The attached file is the published pdf.NHM Repositor

Contribution of soft-bodied meiofaunal taxa to Italian marine biodiversity

Meiofauna includes an astonishing diversity of organisms, whose census is far from being complete. Most classic ecological studies have focused on hard-bodied Ecdysozoan taxa (notably Copepoda and Nematoda), whose cuticle allows determination at species-level after fixation, rather than soft-bodied, Spiralian taxa, which most often lose any diagnostic feature in fixed samples. Yet, metabarcoding studies have recently revealed a species-richness of softbodied taxa comparable, and in cases superior, to that of Copepoda and Nematoda together. However, given objective difficulties inherent to their study, which necessarily has to be performed on living individuals, and their limited utilisation for ecological and applicative research, taxonomic expertise on soft-bodied organisms has declined over the years, and diversity of these phyla in most areas of the world is presently completely unknown. Here we present an expert-based survey of current knowledge on the composition and distribution of soft-bodied meiofaunal taxa in Italy, with special references to the predominantly or exclusively meiobenthic phyla Gastrotricha, Gnathostomulida, Platyhelminthes, Rotifera, Xenacoelomorpha, and macrofaunal taxa with conspicuous meiofaunal representatives (Annelida, Mollusca and Nemertea). A total of 638 described species have been reported from Italian coasts; furthermore, the existence of a large number of undescribed species is mentioned. Knowledge of Annelida, Gastrotricha, and Rotifera appears particularly detailed, placing Italy among the best-known country worldwide. In contrast, knowledge of Platyhelminthes and Xenacoelomorpha appears patchy, and limited to few areas. Sampling effort has been uneven, with most species recorded from the Tyrrhenian Sea, while large sections of the Adriatic and Ionian seas have been poorly explored. Results highlight the role that Marine Biological Stations, notably the Zoological Station “Anton Dohrn” in Naples, have had in promoting the study of soft-bodied taxa in Ital

Acoel Flatworms Are Not Platyhelminthes: Evidence from Phylogenomics

Acoel flatworms are small marine worms traditionally considered to belong to the phylum Platyhelminthes. However, molecular phylogenetic analyses suggest that acoels are not members of Platyhelminthes, but are rather extant members of the earliest diverging Bilateria. This result has been called into question, under suspicions of a long branch attraction (LBA) artefact. Here we re-examine this problem through a phylogenomic approach using 68 different protein-coding genes from the acoel Convoluta pulchra and 51 metazoan species belonging to 15 different phyla. We employ a mixture model, named CAT, previously found to overcome LBA artefacts where classical models fail. Our results unequivocally show that acoels are not part of the classically defined Platyhelminthes, making the latter polyphyletic. Moreover, they indicate a deuterostome affinity for acoels, potentially as a sister group to all deuterostomes, to Xenoturbellida, to Ambulacraria, or even to chordates. However, the weak support found for most deuterostome nodes, together with the very fast evolutionary rate of the acoel Convoluta pulchra, call for more data from slowly evolving acoels (or from its sister-group, the Nemertodermatida) to solve this challenging phylogenetic problem

Acoelomorpha: earliest branching bilaterians or deuterostomes?

The Acoelomorpha is an animal group comprised by nearly 400 species of misleadingly inconspicuous flatworms. Despite this, acoelomorphs have been at the centre of a heated debate about the origin of bilaterian animals for 150 years. The animal tree of life has undergone major changes during the last decades, thanks largely to the advent of molecular data together with the development of more rigorous phylogenetic methods. There is now a relatively robust backbone of the animal tree of life. However, some crucial nodes remain contentious, especially the node defining the root of Bilateria. Some studies situate Acoelomorpha (and Xenoturbellida) as the sister group of all other bilaterians, while other analyses group them within the deuterostomes which instead suggests that the last common bilaterian ancestor directly gave rise to deuterostomes and protostomes. The resolution of this node will have a profound impact on our understanding of animal/bilaterian evolution. In particular, if acoelomorphs are the sister group to Bilateria, it will point to a simple nature for the first bilaterian. Alternatively, if acoelomorphs are deuterostomes, this will imply that they are the result of secondary simplification. Here, we review the state of this question and provide potential ways to solve this long-standing issue. Specifically, we argue for the benefits of (1) obtaining additional genomic data from acoelomorphs, in particular from taxa with slower evolutionary rates; (2) the development of new tools to analyse the data; and (3) the use of metagenomics or metatranscriptomics data. We believe the combination of these three approaches will provide a definitive answer as to the position of the acoelomorphs in the animal tree of life

Patterns of Diversity in Soft-Bodied Meiofauna: Dispersal Ability and Body Size Matter

Background: Biogeographical and macroecological principles are derived from patterns of distribution in large organisms, whereas microscopic ones have often been considered uninteresting, because of their supposed wide distribution. Here, after reporting the results of an intensive faunistic survey of marine microscopic animals (meiofauna) in Northern Sardinia, we test for the effect of body size, dispersal ability, and habitat features on the patterns of distribution of several groups.Methodology/Principal Findings: As a dataset we use the results of a workshop held at La Maddalena (Sardinia, Italy) in September 2010, aimed at studying selected taxa of soft-bodied meiofauna (Acoela, Annelida, Gastrotricha, Nemertodermatida, Platyhelminthes and Rotifera), in conjunction with data on the same taxa obtained during a previous workshop hosted at Tjärnö (Western Sweden) in September 2007. Using linear mixed effects models and model averaging while accounting for sampling bias and potential pseudoreplication, we found evidence that: (1) meiofaunal groups with more restricted distribution are the ones with low dispersal potential; (2) meiofaunal groups with higher probability of finding new species for science are the ones with low dispersal potential; (3) the proportion of the global species pool of each meiofaunal group present in each area at the regional scale is negatively related to body size, and positively related to their occurrence in the endobenthic habitat.Conclusion/Significance: Our macroecological analysis of meiofauna, in the framework of the ubiquity hypothesis for microscopic organisms, indicates that not only body size but mostly dispersal ability and also occurrence in the endobenthic habitat are important correlates of diversity for these understudied animals, with different importance at different spatial scales. Furthermore, since the Western Mediterranean is one of the best-studied areas in the world, the large number of undescribed species (37%) highlights that the census of marine meiofauna is still very far from being complete

Coordinated spatial and temporal expression of Hox genes during embryogenesis in the acoel Convolutriloba longifissura

Background: Hox genes are critical for patterning the bilaterian anterior-posterior axis. The evolution of their clustered genomic arrangement and ancestral function has been debated since their discovery. As acoels appear to represent the sister group to the remaining Bilateria (Nephrozoa), investigating Hox gene expression will provide an insight into the ancestral features of the Hox genes in metazoan evolution. Results: We describe the expression of anterior, central and posterior class Hox genes and the ParaHox ortholog Cdx in the acoel Convolutriloba longifissura. Expression of all three Hox genes begins contemporaneously after gastrulation and then resolves into staggered domains along the anterior-posterior axis, suggesting that the spatial coordination of Hox gene expression was present in the bilaterian ancestor. After early surface ectodermal expression, the anterior and central class genes are expressed in small domains of putative neural precursor cells co-expressing ClSoxB1, suggesting an evolutionary early function of Hox genes in patterning parts of the nervous system. In contrast, the expression of the posterior Hox gene is found in all three germ layers in a much broader posterior region of the embryo. Conclusion: Our results suggest that the ancestral set of Hox genes was involved in the anteriorposterior patterning of the nervous system of the last common bilaterian ancestor and were later co-opted for patterning in diverse tissues in the bilaterian radiation. The lack of temporal colinearity of Hox expression in acoels may be due to a loss of genomic clustering in this clade or, alternatively, temporal colinearity may have arisen in conjunction with the expansion of the Hox cluster in the Nephrozoa

A phylogenetic approach to species delimitation in freshwater Gastrotricha from Sweden

Gastrotricha is a cosmopolitan group ofaquatic invertebrates. To date, approximately 765species have been described. This study is the first todeal with species delimitation and cryptic species offreshwater Gastrotricha. Three commonly encounteredspecies, Heterolepidoderma ocellatum, Lepidochaetuszelinkai, and Lepidodermella squamata, areinvestigated for cryptic speciation. Most of thematerial is based on Swedish specimens but closelyrelated species from other parts of the world are alsoincluded. Taxonomic revisions are supported byphylogenies based on 18S rRNA, 28S rRNA, andCOI mtDNA of freshwater Chaetonotidae from severalgenera and inferred from Bayesian and maximumlikelihood approaches. Heterolepidoderma ocellatumf. sphagnophilum is raised to species level, under thename H. acidophilum n. sp. Moreover, genetic databased on COI indicate large variation between twomorphologically very similar groups of Lepidodermellasquamata. The extent of cryptic speciation inL. zelinkai appears low. Based on the phylogenetichypothesis presented in this article, the new species,Lepidodermella intermedia n. sp., from northernSweden is also described. The phylogenetic hypothesisgenerated shows that Chaetonotidae is a nonmonophyleticgroup