Microstomidae (Platyhelminthes: Macrostomorpha) diversity has been almost entirely ignored within recent years, likely due to inconsistent and often old taxonomic literature and a general rarity of sexually mature collected specimens. Herein, we reconstruct the phylogenetic relationships of the group using both previously published and new 18S and CO1 gene sequences. We present some taxonomic revisions of Microstomidae and further describe 8 new species of Microstomum based on both molecular and morphological evidence. Finally, we briefly review the morphological taxonomy of each species and provide a key to aid in future research and identification that is not dependent on reproductive morphology. Our goal is to clarify the taxonomy and facilitate future research into an otherwise very understudied group of tiny (but important) flatworms.

The meiofauna is an important part of the marine ecosystem, but its composition and distribution patterns are relatively unexplored. Here we assessed the biodiversity and community structure of meiofauna from five locations on the Swedish western and southern coasts using a high-throughput DNA sequencing (metabarcoding) approach. The mitochondrial cytochrome oxidase 1 (COI) mini-barcode and nuclear 18S small ribosomal subunit (18S) V1-V2 region were amplified and sequenced using Illumina MiSeq technology. Our analyses revealed a higher number of species than previously found in other areas: thirteen samples comprising 6.5 dm3 sediment revealed 708 COI and 1,639 18S metazoan OTUs. Across all sites, the majority of the metazoan biodiversity was assigned to Arthropoda, Nematoda and Platyhelminthes. Alpha and beta diversity measurements showed that community composition differed significantly amongst sites. OTUs initially assigned to Acoela, Gastrotricha and the two Platyhelminthes sub-groups Macrostomorpha and Rhabdocoela were further investigated and assigned to species using a phylogeny-based taxonomy approach. Our results demonstrate that there is great potential for discovery of new meiofauna species even in some of the most extensively studied locations.

Microstomum lineare is a common species of fresh and brackish waters found world-wide. Three genes (18S, CO1 and ITS) were sequenced from specimens of M. lineare collected from four countries, and the levels of cryptic diversity and genetic structuring were assessed. Results showed M. lineare has very wide haplotype distributions suggesting higher than expected dispersal capabilities. In addition, three new species were described on the basis of molecular taxonomy: Microstomum artoisi sp. nov., Microstomum tchaikovskyi sp. nov. and Microstomum zicklerorum sp. nov.

The position of Xenacoelomorpha in the tree of life remains a major unresolved question in the study of deep animal relationships1. 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)2,3 or is a clade inside Deuterostomia4. 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. 

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 soft-bodied 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 Italy.

In July 2019 an international team of 39 senior and junior researchers from nine

countries met at the University of the Azores in Ponta Delgada, São Miguel for a 10-daysworkshop/ summer school to explore the meiofaunal biodiversity in marine sediments of the Azores. In total, we sampled intertidal and subtidal sediments from 54 localities on 14 major sites around São Miguel and additionally explored eight freshwater and terrestrial sites for rotifers. We sorted and investigated more than 2000 living specimens in the field, yielding approximately 180 species of soft-bodied meiofauna, representing most major clades of meiofauna with a focus on nematodes, polychaete annelids, proseriate andrhabdocoel flatworms, gastrotrichs, acoelomorphs, nemerteans, molluscs and cnidarians. Most of the encountered diversity shows similarities to the North-East Atlantic continental meiofauna, but in-depth morphological and molecular analyses are still pending. About 60 of the 180 species could not be assigned a species-level identification in the field, and nearly 15% of the total diversity is expected to be new to science and is awaiting formal description. Herein, we present an overview of the results of the workshop, providing detailed information on the sampling sites, methodology and encountered diversity, and we offer a preliminary discussion on aspects of faunal elements shared with other biogeographic regions. We highlight the most common members of the marine meiofauna of the Azores, provide preliminary diversity estimates and suggest a roadmap for future research towards a better understanding of the meiofauna in this remote archipelago.

Acoels are with few exceptions marine worms and a common component of the interstitial meiofauna. In this study we present new species to science belonging to Isodiametridae and Solenofilomorphidae. The new species, Isodiametra finkei n. sp., Postaphanostoma nilssoni n. sp., Pseudaphanostoma hyalinorhabdoida n. sp. and Solenofilomorpha pellucida n. sp. were all collected in Chile during March 2012. Nucleotide sequences for the ribosomal genes 18S rDNA and 28S rDNA as well as COI mtDNA have been determined for the new species and used in a maximum likelihood analysis to further support their classification.

Gastrotricha is a cosmopolitan group of aquatic invertebrates. To date, approximately 765 species have been described. This study is the first to deal with species delimitation and cryptic species of freshwater Gastrotricha. Three commonly encountered species, Heterolepidoderma ocellatum, Lepidochaetus zelinkai, and Lepidodermella squamata, are investigated for cryptic speciation. Most of the material is based on Swedish specimens but closely related species from other parts of the world are also included. Taxonomic revisions are supported by phylogenies based on 18S rRNA, 28S rRNA, and COI mtDNA of freshwater Chaetonotidae from several genera and inferred from Bayesian and maximum likelihood approaches. Heterolepidoderma ocellatum f. sphagnophilum is raised to species level, under the name H. acidophilum n. sp. Moreover, genetic data based on COI indicate large variation between two morphologically very similar groups of Lepidodermella squamata. The extent of cryptic speciation in L. zelinkai appears low. Based on the phylogenetic hypothesis presented in this article, the new species, Lepidodermella intermedia n. sp., from northern Sweden is also described. The phylogenetic hypothesis generated shows that Chaetonotidae is a nonmonophyletic group.

The freshwater gastrotrich fauna of Sweden is poorly known. Only seven species of freshwater gastrotrichs have been reported so far. This paper is the first in a series of contributions about the Swedish freshwater gastrotrich fauna. Here we describe one new species, Ichthydium skandicum n. sp., from Jämtland, northern Sweden. The new species falls within the boundary of the subgenus Forficulichthys and is morphologically closest to Ichthydium tanytrichum from which it can be differentiated based on the presence of four pairs of dorsal, keeled scales in the posterior trunk region. Moreover, we provide morphometric data for three additional Ichthydium species: I. diacanthum, I. squamigerum and I. tanytrichum, Italian species all of which are reported for the first time outside Italy. Considering the accompanying fauna, a total of thirteen freshwater Gastrotricha are reported for the first time from Sweden. Finally we present a dichotomous key for Ichthydium along with distributional data of the species considered.

Chaetonotidae is the largest family within Gastrotricha with almost 400 nominal species represented in both freshwater and marine habitats. The group is probably non-monophyletic and suffers from a troubled taxonomy. Current classification is to a great extent based on shape and distribution of cuticular structures, characters that are highly variable. We present the most densely sampled molecular study so far where 17 of the 31 genera belonging to Chaetonotida are represented. Bayesian and maximum likelihood approaches based on 18S rDNA, 28S rDNA and COI mtDNA are used to reconstruct relationships within Chaetonotidae. The use of cuticular structures for supra-specific classification within the group is evaluated and the question of dispersal between marine and freshwater habitats is addressed. Moreover, the subgeneric classification of Chaetonotus is tested in a phylogenetic context. Our results show high support for a clade containing Dasydytidae nested within Chaetonotidae. Within this clade, only three genera are monophyletic following current classification. Genera containing both marine and freshwater species never form monophyletic clades and group with other species according to habitat. Marine members of Aspidiophorus appear to be the sister group of all other Chaetonotidae and Dasydytidae, indicating a marine origin of the clade. Halichaetonotus and marine Heterolepidoderma form a monophyletic group in a sister group relationship to freshwater species, pointing towards a secondary invasion of marine environments of these taxa. Our study highlights the problems of current classification based on cuticular structures, characters that show homoplasy for deeper relationships.

Whereas most work to understand impacts of humans on biodiversity on coastal areas has focused on large, conspicuous organisms, we highlight effects of tourist access on the diversity of microscopic marine animals (meiofauna). We used a DNA metabarcoding approach with an iterative and phylogeny-based approach for the taxonomic assignment of meiofauna and relate diversity patterns to the numbers of tourists accessing sandy beaches on an otherwise un-impacted island National Park. Tourist frequentation, independently of differences in sediment granulometry, beach length, and other potential confounding factors, affected meiofaunal diversity in the shallow “swash” zone right at the mean water mark; the impacts declined with water depth (up to 2 m). The indicated negative effect on meiofauna may have a consequence on all the biota including the higher trophic levels. Thus, we claim that it is important to consider restricting access to beaches in touristic areas, in order to preserve biodiversity.

The phylogeny of Nemertodermatida, a group of microscopic marine worms, was analysed using nucleotide sequences from the ribosomal LSU and SSU genes and the protein coding Histone 3 gene. All currently known species except Ascoparia neglecta and A. secunda were included in the study in addition to several yet undescribed species. Ascopariidae and Nemertodermatidae, are retrieved as separate clades, although not in all analyses as sister groups. Non-monophyly of Nemertodermatida was rejected by the Approximately Unbiased test. Nucleotide sequences deposited in Genbank before 2013 as nemertodermatid were validated against our dataset; some of them are shown to be chimeric implying falsification of prior hypotheses about nemertodermatid phylogeny: other sequences should be assigned new names. We also show that the genus Nemertoderma needs revision. 

Nemertodermatida are microscopic marine worms likely to be the sister-group to acoels, forming with them the earliest extant branch of bilaterian animals, although their phylogenetic position is debated. The nervous system of Flagellophora cf. apelti, Sterreria spp. and Nemertoderma cf. westbladi has been investigated by immunohistochemistry and confocal microscopy using anti-tubulin, anti-5-HT and anti-FMRFamide antibodies. The nervous system of Flagellophora cf. apelti is composed of a large neuropile and a loose brain at the level of the statocysts with several nerve fibres surrounding them and innervating the broom organ. Sterreria spp. shows a commissural-like brain and several neurite bundles going frontad and caudad from this. At the level of the statocysts there is also a thicker aggregation of immunoreactive fibres. The nervous system of N. cf. westbladi consists of a nerve ring lying outside the body wall musculature at the level of the statocyst and a pair of ventro-lateral neurite bundles, from which extend thinner fibres innervating the ventral side of the animal. Numerous bottle-shaped glands were observed, innervated by fibres starting both from the brain and the neurite bundles. The nervous system of the nemertodermatids studied to-date displays no common pattern, instead there is considerable plasticity in the general morphology of the nervous system. 

In addition, the musculature of Sterreria spp. has been studied by phalloidin staining. It shows diagonal muscles in the anterior quarter of the body and a simple orthogonal grid in the posterior three quarters, being simpler than that of the other nemertodermatids. High-resolution differential interference contrast microscopy permitted to better visualise some morphological characters of the species studied, such as statocysts, sperm and glands and, in combination with anti-tubulin staining, describe in detail the broom organ in Flagellophora cf. apelti. Finally, we note an apparent absence of innervation of the gut in Nemertodermatida similar to the condition in Xenoturbella.

Within an evolutionary framework of Gastrotricha Marinellina flagellata and Redudasys fornerise bear special interest, as they are the only Macrodasyida that inhabit freshwater ecosystems. Notwithstanding, these rare animals are poorly known; found only once (Austria and Brazil), they are currently systematised as incertae sedis. Here we report on the rediscovery of Redudasys fornerise, provide an account on morphological novelties and present a hypothesis on its phylogenetic relationship based on molecular data. Specimens were surveyed using DIC microscopy and SEM, and used to obtain the 18 S rRNA gene sequence; molecular data was analyzed cladistically in conjunction with data from 42 additional species belonging to the near complete Macrodasyida taxonomic spectrum. Morphological analysis, while providing new information on taxonomically relevant traits (adhesive tubes, protonephridia and sensorial bristles), failed to detect elements of the male system, thus stressing the parthenogenetic nature of the Brazilian species. Phylogenetic analysis, carried out with ML, MP and Bayesian approaches, yielded topologies with strong nodal support and highly congruent with each other. Among the supported groups is the previously undocumented clade showing the alliance between Redudasys fornerise and Dactylopodola agadasys; other strongly sustained clades include the densely sampled families Thaumastodermatidae and Turbanellidae and most genera. A reconsideration of the morphological traits of Dactylopodola agadasys in light of the new information on Redudasys fornerise makes the alliance between these two taxa very likely. As a result, we create Anandrodasys gen. nov. to contain members of the previously described D. agadasys and erect Redudasyidae fam. nov. to reflect this novel relationship between Anandrodasys and Redudasys. From an ecological perspective, the derived position of Redudasys, which is deeply nested within the Macrodasyida clade, unequivocally demonstrates that invasion of freshwater by gastrotrichs has taken place at least twice, in contrast with the single event hypothesis recently put forward.

Background: Phylogenetic relationships within Gastrotricha are poorly known. Attempts to shed light on this subject using morphological traits have led to hypotheses lacking satisfactory statistical support; it seemed therefore that a different approach was needed.

Methodology/Principal Findings: In this paper we attempt to elucidate the relationships within the taxonomically vast family Thaumastodermatidae (Macrodasyida) using molecular sequence data. The study includes representatives of all the extant genera of the family and for the first time uses a multi-gene approach to infer evolutionary liaisons within Gastrotricha. The final data set comprises sequences of three genes (18S, 28S rDNA and COI mtDNA) from 41 species, including 29 thaumastodermatids, 11 non-thaumastodermatid macrodasyidans and a single chaetonotidan. Molecular data was analyzed as a combined set of 3 genes and as individual genes, using Bayesian and maximum likelihood approaches. Two different outgroups were used: Xenotrichula intermedia (Chaetonotida) and members of the putative basal Dactylopodola (Macrodasyida). Thaumastodermatidae and all other sampled macrodasyidan families were found monophyletic except for Cephalodasyidae. Within Thaumastodermatidae Diplodasyinae and Thaumastodermatinae are monophyletic and so are most genera. Oregodasys turns out to be the most basal group within Thaumastodermatinae in analyses of the concatenated data set as well as in analyses of the nuclear genes. Thaumastoderma appears as the sister taxon to the remaining species. Surprisingly, Tetranchyroderma is non-monophyletic in our analyses as one group of species clusters with Ptychostomella while another appears as the sister group of Pseudostomella.

Conclusions/Significance: Results in general agree with the current classification; however, a revision of the more derived thaumastodermatid taxa seems necessary. We also found that the ostensible COI sequences from several species do not conform to the general invertebrate or any other published mitochondrial genetic code; they may be mitochondrially derived nuclear genes (numts), or one or more modifications of the mitochondrial genetic code within Gastrotricha.