Fate of microplastic captured in the marine demosponge Halichondria panicea.

Microplastic particles are widespread pollutants in the sea and filter-feeding sponges have recently been suggested as useful monitoring organisms. However, the fate of microplastic particles in sponges is poorly understood, yet crucial for interpreting monitoring data. The present study aims to help develop sponges as more useful monitoring organisms for microplastic in the sea. Here, we describe the fate of inedible (2 and 10 µm) plastic beads compared to that of edible bacteria and algal cells captured in the marine demosponge Halichondria panicea. Small Cyanobium bacillare cells entered the choanocyte chambers and were phagocytized by choanocytes, while larger Rhodomonas salina cells were captured in incurrent canals and phagocytized in the mesohyl. Small 2 µm-beads were captured by choanocytes and subsequently expelled into the excurrent canals after 58 ± 34 min. Larger 10 µm-beads were captured in the incurrent canals and transferred to the mesohyl, where amoeboid cells moved them across the mesohyl before they were expelled into the excurrent canal after 95 ± 36 min. SEM observations further indicated engulfment of plastic beads on the outer sponge surface. This insight provides useful information on how sponges, in general, treat microplastic particles of various sizes. It helps us understand actual measured sizes and concentrations of microplastic particles in sponges in relation to those in the ambient water.

Molecular machineries of ciliogenesis, cell survival, and vasculogenesis are differentially expressed during regeneration in explants of the demosponge Halichondria panicea.

Sponges are interesting animal models for regeneration studies, since even from dissociated cells, they are able to regenerate completely. In particular, explants are model systems that can be applied to many sponge species, since small fragments of sponges can regenerate all elements of the adult, including the oscula and the ability to pump water. The morphological aspects of regeneration in sponges are relatively well known, but the molecular machinery is only now starting to be elucidated for some sponge species. Here, we have used an explant system of the demosponge Halichondria panicea to understand the molecular machinery deployed during regeneration of the aquiferous system. We sequenced the transcriptomes of four replicates of the 5-day explant without an osculum (NOE), four replicates of the 17-18-day explant with a single osculum and pumping activity (PE) and also four replicates of field-collected individuals with regular pumping activity (PA), and performed differential gene expression analysis. We also described the morphology of NOE and PE samples using light and electron microscopy. Our results showed a highly disorganised mesohyl and disarranged aquiferous system in NOE that is coupled with upregulated pathways of ciliogenesis, organisation of the ECM, and cell proliferation and survival. Once the osculum is formed, genes involved in "response to stimulus in other organisms" were upregulated. Interestingly, the main molecular machinery of vasculogenesis described in vertebrates was activated during the regeneration of the aquiferous system. Notably, vasculogenesis markers were upregulated when the tissue was disorganised and about to start forming canals (NOE) and angiogenic stimulators and ECM remodelling machineries were differentially expressed once the aquiferous system was in place (PE and PA). Our results are fundamental to better understanding the molecular mechanisms involved in the formation of the aquiferous system in sponges, and its similarities with the early onset of blood-vessel formation in animal evolution.

The contraction-expansion behaviour in the demosponge Tethya wilhelma is light controlled and follows a diurnal rhythm.

Sponges (phylum Porifera) are metazoans which lack muscles and nerve cells, yet perform coordinated behaviours such as whole-body contractions. Previous studies indicate diurnal variability in both the number of contractions and the expression of circadian clock genes. Here, we show that diurnal patterns are present in the contraction-expansion behaviour of the demosponge Tethya wilhelma, by using infrared videography and a simulated night/day cycle including sunrise and sunset mimics. In addition, we show that this behaviour is at least strongly influenced by ambient light intensity and therefore indicates light-sensing capabilities in this sponge species. This is supported by our finding that T. wilhelma consistently contracts at sunrise, and that this pattern disappears both when the sponge is kept in constant darkness and when it is in constant light.

Growth and single cell kinetics of the loricate choanoflagellate Diaphanoeca grandis.

Choanoflagellates are common members of planktonic communities. Some have complex life histories that involve transitions between multiple cell stages. We have grown the loricate choanoflagellate Diaphanoeca grandis on the bacterium Pantoea sp. and integrated kinetic observations at the culture level and at the single cell level. The life history of D. grandis includes a cell division cycle with a number of recognisable cell stages. Mature, loricate D. grandis were immobile and settled on the bottom substratum. Daughter cells were ejected from the lorica 30 min. after cell division, became motile and glided on the bottom substratum until they assembled a lorica. Single cell kinetics could explain overall growth kinetics in D. grandis cultures. The specific growth rate was 0.72 day-1 during exponential growth while mature D. grandis produced daughter cells at a rate of 0.9 day-1. Daughter cells took about 1.2 h to mature. D. grandis was able to abandon and replace its lorica, an event that delayed daughter cell formation by more than 2 days. The frequency of daughter cell formation varied considerably among individuals and single cell kinetics demonstrated an extensive degree of heterogeneity in D. grandis cultures, also when growth appeared to be balanced.

Helminths in common eiders (Somateria mollissima): Sex, age, and migration have differential effects on parasite loads.

In birds, parasites cause detrimental effects to the individual host, including reduced survival and reproductive output. The level of parasitic infection can vary with a range of factors, including migratory status, body size, sex, and age of hosts, or season. Understanding this baseline variation is important in order to identify the effects of external changes such as climate change on the parasitic load and potential impacts to individuals and populations. In this study, we compared the infection level (prevalence, intensity, and abundance) of gastrointestinal parasites in a total of 457 common eiders (Somateria mollissima) from four different sampling locations (Belcher Islands, Cape Dorset, West Greenland and Newfoundland), and explored the effects of migration, sex and age on levels of parasitism. Across all samples, eiders were infected with one nematode genus, two acanthocephalan genera, three genera of cestodes, and three trematode genera. Migratory phase and status alone did not explain the observed variation in infection levels; the expectation that post-migratory eiders would be more parasitized than pre-migratory eiders, due to the energetic cost of migration, did not fit our results. No effect of age was detected, whereas effects of sex and body size were only detected for certain parasitic taxa and was inconsistent with location. Since gastrointestinal helminths are trophically-transmitted, future studies of the regional and temporal variation in the diet of eiders and the associated variation and infestation level of intermediate hosts might further explain the observed variation of the parasitic load in eiders in different regions.

The house spider genome reveals an ancient whole-genome duplication during arachnid evolution.

BACKGROUND: The duplication of genes can occur through various mechanisms and is thought to make a major contribution to the evolutionary diversification of organisms. There is increasing evidence for a large-scale duplication of genes in some chelicerate lineages including two rounds of whole genome duplication (WGD) in horseshoe crabs. To investigate this further, we sequenced and analyzed the genome of the common house spider Parasteatoda tepidariorum. RESULTS: We found pervasive duplication of both coding and non-coding genes in this spider, including two clusters of Hox genes. Analysis of synteny conservation across the P. tepidariorum genome suggests that there has been an ancient WGD in spiders. Comparison with the genomes of other chelicerates, including that of the newly sequenced bark scorpion Centruroides sculpturatus, suggests that this event occurred in the common ancestor of spiders and scorpions, and is probably independent of the WGDs in horseshoe crabs. Furthermore, characterization of the sequence and expression of the Hox paralogs in P. tepidariorum suggests that many have been subject to neo-functionalization and/or sub-functionalization since their duplication. CONCLUSIONS: Our results reveal that spiders and scorpions are likely the descendants of a polyploid ancestor that lived more than 450 MYA. Given the extensive morphological diversity and ecological adaptations found among these animals, rivaling those of vertebrates, our study of the ancient WGD event in Arachnopulmonata provides a new comparative platform to explore common and divergent evolutionary outcomes of polyploidization events across eukaryotes.

Rotiferan Hox genes give new insights into the evolution of metazoan bodyplans.

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 sister group to the remaining spiralians.Rotifers are microscopic animals with an unusual, nonsegmented body plan consisting of a head, trunk and foot. Here, Fröbius and Funch investigate the role of Hox genes-which are widely used in animal body plan patterning-in rotifer embryogenesis and find non-canonical expression in the nervous system.

Low Oxygen Levels Slow Embryonic Development of Limulus polyphemus.

The American horseshoe crab Limulus polyphemus typically spawns in the upper intertidal zone, where the developing embryos are exposed to large variations in abiotic factors such as temperature, humidity, salinity, and oxygen, which affect the rate of development. It has been shown that embryonic development is slowed at both high and low salinities and temperatures, and that late embryos close to hatching tolerate periodic hypoxia. In this study we investigated the influence of hypoxia on both early and late embryonic development in L. polyphemus under controlled laboratory conditions. Embryos were exposed to four different oxygen levels and their developmental stage was scored every second day. Embryos developed more slowly at both 5% O2 and 10% O2 than at the 21% O2 treatment; late development was arrested when oxygen was reduced to 2%. Our study confims that L. polyphemus not only tolerates pronounced hypoxia in later embryonic developmental stages, but also in earlier, previously unexplored, developmental stages.

Endozoicomonas Are Specific, Facultative Symbionts of Sea Squirts.

Ascidians are marine filter feeders and harbor diverse microbiota that can exhibit a high degree of host-specificity. Pharyngeal samples of Scandinavian and Mediterranean ascidians were screened for consistently associated bacteria by culture-dependent and -independent approaches. Representatives of the Endozoicomonas (Gammaproteobacteria, Hahellaceae) clade were detected in the ascidian species Ascidiella aspersa, Ascidiella scabra, Botryllus schlosseri, Ciona intestinalis, Styela clava, and multiple Ascidia/Ascidiella spp. In total, Endozoicomonas was detected in more than half of all specimens screened, and in 25-100% of the specimens for each species. The retrieved Endozoicomonas 16S rRNA gene sequences formed an ascidian-specific subclade, whose members were detected by fluorescence in situ hybridization (FISH) as extracellular microcolonies in the pharynx. Two strains of the ascidian-specific Endozoicomonas subclade were isolated in pure culture and characterized. Both strains are chemoorganoheterotrophs and grow on mucin (a mucus glycoprotein). The strains tested negative for cytotoxic or antibacterial activity. Based on these observations, we propose ascidian-associated Endozoicomonas to be commensals, living off the mucus continuously secreted into the pharynx. Members of the ascidian-specific Endozoicomonas subclade were also detected in seawater from the Scandinavian sampling site, which suggests acquisition of the symbionts by horizontal transmission. The combined results indicate a host-specific, yet facultative symbiosis between ascidians and Endozoicomonas.

Description of Endozoicomonas ascidiicola sp. nov., isolated from Scandinavian ascidians.

Two gram-negative, facultative anaerobic, chemoorganoheterotrophic, motile and rod-shaped bacteria, strains AVMART05(T) and KASP37, were isolated from ascidians (Tunicata, Ascidiaceae) of the genus Ascidiella collected at Gullmarsfjord, Sweden. The strains are the first cultured representatives of an ascidian-specific lineage within the genus Endozoicomonas (Gammaproteobacteria, Oceanospirillales, Hahellaceae). Both strains feature three distinct 16S rRNA gene paralogs, with identities of 98.9-99.1% (AVMART05(T)) and 97.7-98.8% (KASP37) between paralogs. The strains are closely related to Endozoicomonas atrinae and Endozoicomonas elysicola, with which they share 97.3-98.0% 16S rRNA gene sequence identity. Digital DNA-DNA hybridization, average nucleotide identity, and tetra-nucleotide correlation analysis indicate that both strains belong to a single species distinct from their closest relatives. Both strains feature similar DNA G+C contents of 46.70mol% (AVMART05(T)) and 44.64mol% (KASP37). The fatty acid patterns of AVMART05(T) and KASP37 are most similar to those of Endozoicomonas euniceicola and Endozoicomonas gorgoniicola. Based on the polyphasic approach, we propose the species Endozoicomonas ascidiicola sp. nov. to accommodate the newly isolated strains. E. ascidiicola sp. nov. is represented by the type strain AVMART05(T) (=DSM 100913(T)=LMG 29095(T)) and strain KASP37 (=DSM 100914=LMG 29096).

A Novel Extracellular Gut Symbiont in the Marine Worm Priapulus caudatus (Priapulida) Reveals an Alphaproteobacterial Symbiont Clade of the Ecdysozoa.

Priapulus caudatus (phylum Priapulida) is a benthic marine predatory worm with a cosmopolitan distribution. In its digestive tract we detected symbiotic bacteria that were consistently present in specimens collected over 8 years from three sites at the Swedish west coast. Based on their 16S rRNA gene sequence, these symbionts comprise a novel genus of the order Rickettsiales (Alphaproteobacteria). Electron microscopy and fluorescence in situ hybridization (FISH) identified them as extracellular, elongate bacteria closely associated with the microvilli, for which we propose the name "Candidatus Tenuibacter priapulorum". Within Rickettsiales, they form a phylogenetically well-defined, family-level clade with uncultured symbionts of marine, terrestrial, and freshwater arthropods. Cand. Tenuibacter priapulorum expands the host range of this candidate family from Arthropoda to the entire Ecdysozoa, which may indicate an evolutionary adaptation of this bacterial group to the microvilli-lined guts of the Ecdysozoa.

Spider genomes provide insight into composition and evolution of venom and silk.

Spiders are ecologically important predators with complex venom and extraordinarily tough silk that enables capture of large prey. Here we present the assembled genome of the social velvet spider and a draft assembly of the tarantula genome that represent two major taxonomic groups of spiders. The spider genomes are large with short exons and long introns, reminiscent of mammalian genomes. Phylogenetic analyses place spiders and ticks as sister groups supporting polyphyly of the Acari. Complex sets of venom and silk genes/proteins are identified. We find that venom genes evolved by sequential duplication, and that the toxic effect of venom is most likely activated by proteases present in the venom. The set of silk genes reveals a highly dynamic gene evolution, new types of silk genes and proteins, and a novel use of aciniform silk. These insights create new opportunities for pharmacological applications of venom and biomaterial applications of silk.

Xenoturbella bocki exhibits direct development with similarities to Acoelomorpha.

Xenoturbella bocki, a marine animal with a simple body plan, has recently been suggested to be sister group to the Acoelomorpha, together forming the new phylum Xenacoelomorpha. The phylogenetic position of the phylum is still under debate, either as an early branching bilaterian or as a sister group to the Ambulacraria (hemichordates and echinoderms) within the deuterostomes. Although development has been described for several species of Acoelomorpha, little is known about the life cycle of Xenoturbella. Here we report the embryonic stages of Xenoturbella, and show that it is a direct developer without a feeding larval stage. This mode of development is similar to that of the acoelomorphs, supporting the newly proposed phylum Xenacoelomorpha and suggesting that the last common ancestor of the phylum might have been a direct developer.

Biofilm formation in long-term central venous catheters in children with cancer: a randomized controlled open-labelled trial of taurolidine versus heparin.

Taurolidine has demonstrated inhibition of biofilm formation in vitro. The aim of this study was to compare the effect of catheter locking with taurolidine vs heparin in biofilm formation in central venous catheters. Forty-eight children with cancer were randomized to catheter locking by heparin (n = 22) or taurolidine (n = 26), respectively. After removal, catheters were examined by standardized scanning electron microscopy to assess quantitative biofilm formation. Biofilm was present if morphologically typical structures and bacterial cells were identified. Quantitative and semi-quantitative cultures were also performed. Biofilm was identified in 23 of 26 catheters from the taurolidine group and 21 of 22 catheters from the heparin group. A positive culture was made of six of the catheters locked with taurolidine and heparin, respectively (p = 0.78). The rate of catheter-related bloodstream infections (CRBSI) was 0.1 per 1000 catheter-days using taurolidine and 0.9 per 1000 catheter-days using heparin (p = 0.03). This randomized trial confirmed that the use of taurolidine as catheter-lock compared with heparin reduced the rate of CRBSIs; this reduction was not related to a reduction in the intraluminal biofilm formation and the rate of bacterial colonization detected by scanning electron microscopy in the two groups.

A holomorph approach to xiphosuran evolution--a case study on the ontogeny of Euproops.

Specimens of Euproops sp. (Xiphosura, Chelicerata) from the Carboniferous Piesberg quarry near Osnabrück, Germany, represent a relatively complete growth series of 10 stages. Based on this growth sequence, morphological changes throughout the ontogeny can be identified. The major change affects the shape of the epimera of the opisthosoma. In earlier stages, they appear very spine-like, whereas in later stages the bases of these spine-like structures become broader; the broadened bases are then successively drawn out distally. In the most mature stage known, the epimera are of trapezoidal shape and approach each other closely to form a complete flange around the thoracetron (=fused tergites of the opisthosoma). These ontogenetic changes question the taxonomic status of different species of Euproops, as the latter appear to correspond to different stages of the ontogenetic series reconstructed from the Piesberg specimens. This means that supposed separate species could, in fact, represent different growth stages of a single species. It could alternatively indicate that heterochrony (=evolutionary change of developmental timing) plays an important role in the evolution of Xiphosura. We propose a holomorph approach, i.e., reconstructing ontogenetic sequences for fossil and extant species as a sound basis for a taxonomic, phylogenetic, and evolutionary discussion of Xiphosura.

Ultrastructure and morphology of the cycliophoran female.

Knowledge on the morphology of the cycliophoran female has mostly been based on observations of immature females in brood chambers of feeding stages. With the use of light- and transmission electron microscopy, the morphology and ultrastructure of the free and fully mature female of Cycliophora is described now for the first time. The external morphology is characterized by a ciliation consisting of an anteroventral ciliated field, a posterior ciliated tuft, and four sensory structures extending anteriorly from the anteroventral ciliated field. In addition, a small ciliated structure in the midventral region is interpreted as a round-shaped gonopore. Internally, a bilateral cerebral ganglion is situated in the anterior region and a large oocyte is located medially in the body. Several glands are present anteriorly, while posteriorly a pair of glands is associated with the ciliated tuft. Dorsal and ventral longitudinal muscles, as well as, dorsoventral muscles are identified by electron microscopy. Muscle fibers attach to the endocuticle via the epidermis, by means of attachment fibers. An unknown endosymbiont is present throughout the body of the female. We discuss the functional implications of the morphological and ultrastructural aspects of the cycliophoran female. Finally, we compare this life cycle stage with that fromother phyla, suggested as phylogenetically close.

Molecular phylogeny of extant horseshoe crabs (Xiphosura, Limulidae) indicates Paleogene diversification of Asian species.

Horseshoe crabs are marine invertebrates well known for their exceptionally low rates of diversification during their entire evolutionary history. Despite the low species diversity in the group, the phylogenetic relationships among the extant species, especially among the three Asian species are still unresolved. Here we apply a new set of molecular genetic data in combination with a wide geographic sampling of the intra-specific diversity to reinvestigate the evolutionary history among the four living limulid xiphosurans. Our analysis of the intraspecific diversity reveals low levels of connectivity among Carcinoscorpius rotundicauda lineages, which can be explained by the estuarine-bound ecology of this species. Moreover, a clear genetic break across the Thai-Malay Peninsula suggests the presence of cryptic species in C. rotundicauda. The limulid phylogeny finds strong support for a monophyletic genus Tachypleus and a diversification of the three Asian species during the Paleogene period, with speciation events well separated in time by several million years. The tree topology suggests that the three Asian species originated in central South East Asia from a marine stem group that inhabited the shallow coastal waters between the Andaman Sea, Vietnam, and Borneo. In this region C. rotundicauda probably separated from the Tachypleus stem group by invading estuarine habitats, while Tachypleus tridentatus most likely migrated northeast along the Southern coast of China and towards Japan.

Population dynamics of American horseshoe crabs--historic climatic events and recent anthropogenic pressures.

Populations of the American horseshoe crab, Limulus polyphemus, have declined, but neither the causes nor the magnitude are fully understood. In order to evaluate historic demography, variation at 12 microsatellite DNA loci surveyed in 1218 L. polyphemus sampled from 28 localities was analysed with Bayesian coalescent-based methods. The analysis showed strong declines in population sizes throughout the species' distribution except in the geographically isolated southern-most population in Mexico, where a strong increase in population size was inferred. Analyses suggested that demographic changes in the core of the distribution occurred in association with the recolonization after the Ice Age and also by anthropogenic effects, such as the past overharvest of the species for fertilizer or the current use of the animals as bait for American eel (Anguilla rostrata) and whelk (Busycon spp.) fisheries. This study highlights the importance of considering both climatic changes and anthropogenic effects in efforts to understand population dynamics--a topic which is highly relevant in the ongoing assessments of the effects of climate change and overharvest.

Two types of endosymbiotic bacteria in the enigmatic marine worm Xenoturbella bocki.

Two types of endosymbiotic bacteria were identified in the gastrodermis of the marine invertebrate Xenoturbella bocki (Xenoturbellida, Bilateria). While previously described Chlamydia-like endosymbionts were rare, Gammaproteobacteria distantly related to other endosymbionts and pathogens were abundant. The endosymbionts should be considered when interpreting the poorly understood ecology and evolution of Xenoturbella.

Comparative myoanatomy of cycliophoran life cycle stages.

The metazoan phylum Cycliophora includes small cryptic epibionts that live attached to the mouthparts of clawed lobsters. The life cycle is complex, with alternating sexual and asexual generations, and involves several sessile and free-living stages. So far, the morphological and genetic characterization of cycliophorans has been unable to clarify the phylogenetic position of the phylum. In this study, we add new details on the muscular anatomy of the feeding stage, the attached Prometheus larva, the dwarf male, and the female of one of the two hitherto described species, Symbion pandora. The musculature of the feeding stage is composed of myofibers that run longitudinally in the buccal funnel (two fibers) and in the trunk (variable number of fibers). The mouth opening is lined by a myoepithelial ring musculature. A complex myoepithelial sphincter is situated proximal to the anus. In the attached Prometheus larva, three longitudinal sets of myofilaments run dorsally, laterally, and ventrally along the entire anterior-posterior body axis. The muscular architecture of the dwarf male is complex, especially close to the penis, in the posterior part of the body. An X-shaped muscle structure is found on the dorsal side, whereas on the ventral side, longitudinal muscles and a V-shaped muscle structure are present. These muscles are complemented by additional dorsoventral muscles. The mesodermal muscle fibers attach to the cuticle via the epidermis in all life cycle stages studied herein. The musculature of the female is similar to that of the Pandora larva of Symbion americanus and includes dorsoventral muscles and longitudinal muscles that run in the dorsal and ventral body region. Overall, our results reveal striking similarities in the muscular arrangement of the life cycle stages of both Symbion species.