Confocal laser scanning microscopy (CLSM) as a new tool for morphological characterisation of both newly collected and museum voucher specimens of the Trypanorhyncha Diesing, 1863 (Platyhelminthes: Cestoda).

Taxonomic issues within Trypanorhyncha, e.g., the inaccurate light microscopic visualisation of the hook patterns, are solvable by confocal laser scanning microscopy (CLSM). We applied CLSM imaging to study Trygonicola macropora (Shipley et Hornell, 1906) and Dollfusiella michiae (Southwell, 1929) from Neotrygon caeruliopunctata Last, White et Séret from Bali, Indonesia. To illustrate the strength and limitations of CLSM, images of Otobothrium cysticum (Mayer, 1842) and Symbothriorhynchus tigaminacantha Palm, 2004, both permanent mounts from a collection, were also processed. The CLSM created image stacks of many layers, and edited with IMARIS Software, these layers resulted in three-dimensional images of the armature patterns and internal organs of both species. BABB (benzylalcohol and benzylbenzuolate) clearing was applied to T. macropora. We conclude that trypanorhynch cestodes stained with Mayer-Schuberg's acetic carmine permanently mounted in Canada balsam are suitable for CLSM, allowing detailed analyses of museum type-material as well as freshly collected and processed worms. BABB resulted in imaging the testes in detail, suggesting other stains to be used for CLSM in trypanorhynch cestode research. Application of CLSM for studies of other cestode groups is highly recommended.

From synonym to valid species: Redescriptions of Ampharete acutifrons (Grube, 1860) and A. cirrata Webster amp; Benedict, 1887, and brief descriptions of A. baltica Eliason, 1955 and A. grubei Malmgren, 1865 (Annelida: Terebellida: Ampharetidae).

This paper aims mainly to provide clarity on the morphological characters of the type species of Ampharete, Ampharete acutifrons (Grube, 1860). Its common occurrence and wide distribution are most likely a result of misidentification of different species. Possible reasons for that are the brief original description of this species that is available solely in Latin and a series of questionable synonymizations, resulting in a confusing history of A. acutifrons. In addition to a detailed redescription of the holotype of A. acutifrons, we describe an Ampharete species from the Baltic Sea that has also been incorrectly identified as A. acutifrons for many decades. The individuals of this species agree in all diagnostic characters with those of the Northwest Atlantic species Ampharete cirrata Webster Benedict, 1887. Since no differences were found, but it is clearly distinct from A. acutifrons, A. cirrata is recognised as a valid species and consequently deleted from the synonym list of A. acutifrons. We additionally examined type material of A. grubei Malmgren, 1865 and those of A. grubei baltica Eliason, 1955, now accepted as A. baltica. According to this, both species are valid, and A. grubei must therefore also be deleted from the list of synonyms of A. acutifrons. Information on five molecular markers (Histone H3, COI, 16S, 18S, 28S) is provided for A. cirrata from the Baltic Sea. DNA sequences (H3, 16S, 28S) were identical to sequences of a specimen found in Iceland that was incorrectly determined as A. acutifrons, supporting the assumption of an amphiatlantic distribution of A. cirrata. By comparing obtained sequences to available sequences in GenBank and BOLD, we found evidence that at least four species were previously misidentified as A. acutifrons. The historical course of the taxonomy of A. acutifrons demonstrates the importance of carefully studying type material and type locality material, respectively. We believe that most previous records and synonyms of A. acutifrons have been identified incorrectly and should be re-evaluated. Additionally, an updated key to all species of Ampharete from the North Atlantic is provided.

Exploring confocal laser scanning microscopy (CLSM) and fluorescence staining as a tool for imaging and quantifying traces of marine microbioerosion and their trace-making microendoliths.

Microscopic organisms that penetrate calcareous structures by actively dissolving the carbonate matrix, namely microendoliths, have an important influence on the breakdown of marine carbonates. The study of these microorganisms and the bioerosion traces they produce is crucial for understanding the impact of their bioeroding activity on the carbonate recycling in environments under global climate change. Traditionally, either the extracted microendoliths were studied by conventional microscopy or their traces were investigated using scanning electron microscopy (SEM) of epoxy resin casts. A visualisation of the microendoliths in situ, that is within their complex microbioerosion structures, was previously limited to the laborious and time-consuming double-inclusion cast-embedding technique. Here, we assess the applicability of various fluorescence staining methods in combination with confocal laser scanning microscopy (CLSM) for the study of fungal microendoliths in situ in partly translucent mollusc shells. Among the tested methods, specific staining with dyes against the DNA (nuclei) of the trace making organisms turned out to be a useful and reproducible approach. Bright and clearly delineated fluorescence signals of microendolithic nuclei allow, for instance, a differentiation between abandoned and still populated microborings. Furthermore, infiltrating the microborings with fluorescently stained resin seems to be of great capability for the visualisation and quantification of microbioerosion structures in their original spatial orientation. Potential fields of application are rapid assessments of endolithic bio- and ichnodiversity and the quantification of the impact of microendoliths on the overall calcium carbonate turnover. The method can be applied after CLSM of the stained microendoliths and retains the opportunity for a subsequent investigation of epoxy casts with SEM. This allows a three-fold approach in studying microendoliths in the context of their microborings, thereby fostering the integration of biological and ichnological aspects of microbial bioerosion.

Bioerosion describes the process of active erosion of hard substrates induced by the activity of living organisms. Beside numerous marine macroscopic bioeroding organisms such as sponges, annelids or bivalves, there is an astonishing 'hidden diversity' of microscopic bioeroding organisms which produce minute tunnels and chambers, for example in calcareous shells and skeletons of other marine organisms. These so-called microendoliths belong to bacteria, microalgae, foraminiferans, or fungi. Due to their lifestyle hidden inside the hard substrate, scientific investigation is often laborious and involves complex preparation techniques, electron microscopy, or even nano-computed tomography. Photo-autotrophic microendoliths (eg cyanobacteria and algae) have been studied with fluorescence microscopy using autofluorescence properties, for example of their chloroplasts. However, microendoliths of aphotic depths, mostly of fungal origin, do not show autofluorescence. With the present study we test different fluorescent dyes staining the microbioeroders 'in situ', that is, inside their microscopic tunnels, and visualise them using three-dimensional confocal laser scanning microscopy (CLSM). Very good results have been obtained with the dye Sybr Green I that stains DNA molecules and thereby the cell nuclei of the microendoliths. This method can be used, for instance, to measure the infestation rate of a given substrate by discriminating between abandoned microborings and those still inhabited by microendoliths. Another approach that was successfully tested in the course of the present study was the infiltration of the cleaned microborings with resin that was previously mixed with the fluorescent dye Safranin-O. The datasets obtained with the CLSM were used to reconstruct 3D-surface models of the microborings of three different microendoliths. Such models can be used to analyse the original spatial arrangement inside the hard substrate and to measure exact volumes. The resulting possibility to make exact quantifications is of high value for future investigations that focus on the role and proportion of microbioerosion in the (re)cycling of marine carbonates.

A new freshwater fabriciid species (Sabellida, Fabriciidae) from the Uruguay River, Argentina, with some remarks on the relationships of Monroika Hartman, 1951, Manayunkia Bourne, 1884 and Brandtika Jones, 1974.

Between the years 2007 and 2009 a hitherto unknown fabriciid species was found in the Lower Uruguay River, Argentina. Initially, it was assumed that this species could represent Manayunkia speciosa, a freshwater species originally described from North America. However, re-examinations have revealed that these specimens are clearly different from M. speciosa but they resemble Monroika africana. However, differences were also found that clearly distinguish this South American species from the African species so that it is described here as M. clarae sp. nov. The characters found in Monroika clarae sp. nov. make it also possible to compare and discuss these characters with those from the West African species Monroika africana, the Southeast Asian species Brandtika asiatica as well as Manayunkia species in terms of their systematic position. This mainly concerns the structure of the radiolar crown, the number of abdominal chaetigers, the presence of transitional chaetae as well as the structure of the thoracic and abdominal uncini. Potential origins of the freshwater Fabriciidae are also discussed.

Generation and properties of the new asphalt binder model using molecular dynamics (MD).

Asphalt binder is the main material for road pavement and building construction. It is a complex mixture composed of a large number of hydrocarbons with different molecular weights. The study of asphalt binders and asphalt concretes from a molecular perspective is an important means to understand the intricate properties of asphalt. Molecular dynamics simulation is based on Newton's law and predicts the microscopic performance of materials by calculating the intra- and intermolecular interactions. The asphalt binder can be divided into four components: saturates, aromatics, resins, and asphaltenes (SARA). A new molecular model of asphalt was proposed and verified in this study. Eight molecules selected from the literature were used to represent the four components of asphalt. The AMBER Cornell Extension Force Field was applied in this study to model building and the calculation of properties. The density of the asphalt model was calculated and compared with experimental results for validity verifications. The results show that the purposed model can be used to calculate the microscopic properties of the asphalt binder because the density of the model is close to the real value in the field. Besides, the proportions of different molecules in the model were adjusted to predict the relationship between the asphalt binder density and the hydrocarbon ratios and heteroatom contents of the molecular model. Moreover, the glass transition temperature of the asphalt binder model is predicted by the simulation of the heating process. The range of the glass transition temperature is determined by calculating the relationship between specific volume and temperature, and the calculated range is close to the experimental value.

Natural history collections recapitulate 200 years of faunal change.

Changing species assemblages represent major challenges to ecosystems around the world. Retracing these changes is limited by our knowledge of past biodiversity. Natural history collections represent archives of biodiversity and are therefore an unparalleled source to study biodiversity changes. In the present study, we tested the value of natural history collections for reconstructing changes in the abundance and presence of species over time. In total, we scrutinized 17 080 quality-checked records for 242 epibenthic invertebrate species from the North and Baltic Seas collected throughout the last 200 years. Our approaches identified eight previously reported species introductions, 10 range expansions, six of which are new to science, as well as the long-term decline of 51 marine invertebrate species. The cross-validation of our results with published accounts of endangered species and neozoa of the area confirmed the results for two of the approaches for 49 to 55% of the identified species, and contradicted our results for 9 to 10%. The results based on relative record trends were less validated. We conclude that, with the proper approaches, natural history collections are an unmatched resource for recovering early species introductions and declines.

Coming and going - Historical distributions of the European oyster Ostrea edulis Linnaeus, 1758 and the introduced slipper limpet Crepidula fornicata Linnaeus, 1758 in the North Sea.

Natural history collections are fundamental for biodiversity research as well as for any applied environment-related research. These collections can be seen as archives of earth´s life providing the basis to address highly relevant scientific questions such as how biodiversity changes in certain environments, either through evolutionary processes in a geological timescale, or by man-made transformation of habitats throughout the last decades and/or centuries. A prominent example is the decline of the European flat oyster Ostrea edulis Linneaus, 1758 in the North Sea and the concomitant invasion of the common limpet slipper Crepidula fornicata, which has been implicated to have negative effects on O. edulis. We used collections to analyse population changes in both species in the North Sea. In order to reconstruct the change in distribution and diversity over the past 200 years, we combined the temporal and spatial information recorded with the collected specimens contained in several European natural history collections. Our data recover the decline of O. edulis in the North Sea from the 19th century to the present and the process of invasion of C. fornicata. Importantly, the decline of O. edulis was nearly completed before C. fornicata appeared in the North Sea, suggesting that the latter had nothing to do with the local extinction of O. edulis in the North Sea.

Molecular Dynamics Simulation of Pure n-Alkanes and Their Mixtures at Elevated Temperatures Using Atomistic and Coarse-Grained Force Fields.

The properties of higher n-alkanes and their mixtures is a topic of significant interest for the oil and chemical industry. However, the experimental data at high temperatures are scarce. The present study focuses on simulating n-dodecane, n-octacosane, their binary mixture at a n-dodecane mole fraction of 0.3, and a model mixture of the commercially available hydrocarbon wax SX-70 to evaluate the performance of several force fields on the reproduction of properties such as liquid densities, surface tension, and viscosities. Molecular dynamics simulations over a broad temperature range from 323.15 to 573.15 K were employed in examining a broad set of atomistic molecular models assessed for the reproduction of experimental data. The well-established united atom TraPPE (TraPPE-UA) was compared against the all atom optimized potentials for liquid simulations (OPLS) reparametrization for long n-alkanes, L-OPLS, as well as Lipid14 and MARTINI force fields. All models qualitatively reproduce the temperature dependence of the aforementioned properties, but TraPPE-UA was found to reproduce liquid densities most accurately and consistently over the entire temperature range. TraPPE-UA and MARTINI were very successful in reproducing surface tensions, and L-OPLS was found to be the most accurate in reproducing the measured viscosities as compared to the other models. Our simulations show that these widely used force fields originating from the world of biomolecular simulations are suitable candidates in the study of n-alkane properties, both in the pure and mixture states.

Description of a new species of Sabellidae (Polychaeta, Annelida) from fresh and brackish waters in Europe, with some remarks on the branchial crown of Laonome.

In 2009, a hitherto unknown Laonome species was found in the Canal Ghent-Terneuzen in the Netherlands and subsequently in other Dutch rivers, canals and estuaries. A few years later, more unknown Laonome specimens were found in the eastern part of the Baltic Sea and in the Don River estuary, Sea of Azov. Initially, it was assumed that these specimens could represent Laonome calida Capa, 2007, originally described from Australia. In the present study we examine all these unknown European Laonome specimens and compare these specimens with the type material of L. calida from Australia. This lead to two main results: First, all specimens from Europe have the same diagnostic characters and therefore belong to one species. This finding was also supported by the results of a correspondence analysis, and genetic analyses using four different DNA sequences (COI, 16S, 28S). Second, it turned out that the type material of L. calida contains two morphologically distinct groups of specimens. The holotype and 7 paratypes are similar to each other but differ significantly from the other also similar 16 paratypes, and from all European specimens. On the basis of these observations, the Laonome specimens from European waters are described here as L. xeprovala sp. nov. We also provide the characters of the branchial crown of three Laonome species for a prospective revision of this genus.

Redescription of P. dorsipapillatus (von Marenzeller, 1893) and additional comments on the other species in the genus Pholoides Pruvot, 1895 (Polychaeta: Sigalionidae).

Pholoides Pruvot, 1895 is a species-poor genus of small scale-bearing polychaetes. Pholoides species are restricted to the continental shelf, living in sandy and muddy substrates, or on hard bottoms. During the DIVA 3 expedition grab samples were taken on four seamounts of the Meteor Seamount complex. One of the most common polychaetes in these samples was P. dorsipapillatus (von Marenzeller, 1893). Based on this material a comparison with the type material is undertaken leading to a re-description of P. dorsipapillatus. Diagnostic characters of all accepted Pholoides species were evaluated using light, scanning electron and confocal laser scanning microscopy. CLSM in particular proved to be an excellent tool for investigating these small species, and especially the type material. Blossom-like sensory buds, found at certain appendages of P. dorsipapillatus, could be an important diagnostic character to distinguish P. dorsipapillatus from other Pholoides species. Sequence information on three different gene fragments, the mitochondrial COI and 16S and the nuclear H3a, were obtained, and could serve for future phylogenetic and phylogeographic studies.

Applications of a bis-urea phenylethynylene self-assembled nanoreactor for [2 + 2] photodimerizations.

Confined environments can be used to alter the selectivity of a reaction by influencing the organization of the reactants, altering the mobility of trapped molecules, facilitating one reaction pathway or selectively stabilizing the products. This manuscript utilizes a series of potentially photoreactive guests to interrogate the utility of the one-dimensional nanochannels of a porous host to absorb and facilitate the reaction of encapsulated guests. The host is a columnar self-assembled phenylethynylene bis-urea macrocycle, which absorbs guests, including coumarin, 6-methyl coumarin, 7-methyl coumarin, 7-methoxy coumarin, acenaphthylene, cis-stilbene, trans-stilbene, and trans-ß-methylstyrene to afford crystalline inclusion complexes. We examine the structure of the host:guest complexes using powder X-ray diffraction, which suggests that they are well-ordered highly crystalline materials. Investigations using solid-state cross-polarized magic angle spinning (13)C{(1)H}CP-MAS NMR spectroscopy indicate that the guests are mobile relative to the host. Upon UV-irradiation, we observed selective photodimerization reactions for coumarin, 6-methyl coumarin, 7-methyl coumarin, and acenaphthylene, while the other substrates were unreactive even under prolonged UV-irradiation. Grand Canonical Monte Carlo simulations suggest that the reactive guests were close paired and preorganized in configurations that facilitate the photodimerization with high selectivity while the unreactive guests did not exhibit similar close pairing. A greater understanding of the factors that control diffusion and reaction in confinement could lead to the development of better catalysts.

Spionidae (Polychaeta: Canalipalpata: Spionida) from seamounts in the NE Atlantic.

Spionidae (Polychaeta) collected from seamounts in the Atlantic Ocean were studied. Altogether six species were found of which two are new to science and one belongs to a new genus. Aonidella cf. dayi Maciolek in López-Jamar, 1989 and Glandulospio orestes gen. et sp. nov. were the most common species and occurred on both the Great and Little Meteor Seamount, the Irving Seamount and the Hyeres Seamount. Laonice norgensis Sikorski, 2003 and Malacoceros jirkovi Sikorski, 1992 have a wider distribution in the North Atlantic, including the Mediterranean Sea in case of L. norgensis. Aonides selvagensis Brito, Núñez and Riera, 2006 is only known from the Macaronesian Region. Dipolydora paracaulleryi sp. nov. has been collected from both the Great and Little Meteor Seamounts. All species are compared with morphological similar species and their taxonomy is discussed. Detailed descriptions are provided for the species new to science and descriptions of the previously known species are amended. Accompanying histological studies revealed the presence of very strong dorsoventral musculature in A. cf. dayi and for G. orestes gen. et sp. nov. the presence of glandular organs in the middle body region. Laonice maciolekae Aguirrezabalaga & Ceberio, 2005 was found to be a junior synonym of L. appellöfi Söderström, 1920 and is formally synonymised. Molecular data suggest gene flow between seamounts and autochthonous as well as allochthonous larval recruitment for different species. The results of previous studies by other authors, that polychaete communities of the North Atlantic Seamounts are characterized by low diversity, low rates of endemism, and the predominance of widely distributed (and cosmopolitan) species is not corroborated by our results. 

Parapodial glandular organs in Spiophanes (Polychaeta: Spionidae)-studies on their functional anatomy and ultrastructure.

Parapodial glandular organs (PGOs) of Spiophanes (Polychaeta: Spionidae) were studied using light and electron microscopy. These organs are found in parapodia of the mid body region, starting on chaetiger 5 and terminating with the appearance of neuropodial hooks (chaetiger 14 or 15 in adult individuals). Large PGOs in anterior chaetigers display different species-specific types of openings whereas small PGOs in posterior parapodia of the mid body region always open in a simple vertical slit. Each PGO is composed of three main complexes: (1) the glandular sac with several distinct epithelia of secretory cells and secretory cell complexes and the reservoir filled with fibrous material, (2) the gland-associated chaetal complex (including the region of chaetoblasts and follicle cells, follicular canals, two chaetal collector canals, the combined conducting canal, the chaetal spreader including the opening of the glandular organ with associated type-1 secretory cells, and the gland-associated chaetae), and (3) a bilayered musculature surrounding the gland. A considerable number of different cell types are involved in the secretory activity, in the guidance of the gland-associated chaetae, and in the final expulsion of the fibrous secretion at the opening slit. Among these different cell types the type-5 secretory cells of the proximal glandular complex with their cup-shaped microvilli emanating thick microfibrils into the lumen of the glandular sac are most conspicuous. Secretory cells with cup-shaped microvilli being involved in the production of ß-chitin microfibrils have so far only been reported from some representatives of the deep-sea inhabiting Siboglinidae (Polychaeta). We suggest that the gland-associated chaetae emerging from inside the PGOs of Spiophanes are typical annelid chaetae formed by chaetoblasts and follicle cells. Functional morphology implies the crucial role of PGOs in tube construction. Furthermore, the PGOs are discussed in consideration of phylogenetic aspects.

Component community of larval trematodes in the mudsnail Hydrobia ventrosa: temporal variations in prevalence in relation to host life history.

Temporal variations in the prevalence of larval trematodes in the short-lived prosobranch mudsnail Hydrobia ventrosa (Montagu) were investigated in relation to host life history and season for 4 successive years in temperate windflats of the southern Baltic Sea. The component community of trematode larvae in H. ventrosa comprises at least 10 species; families (and species) represented include Notocotylidae (1), Echinostomatidae (1 or 2), Heterophyidae (2), Monorchidae (1), Microphallidae (3 or 4), Psilostomatidae (1), and Hemiuridae (1). The notocotylid Paramonostomum alveatum was the most prevalent species, followed by the microphallids Maritrema subdolum and Microphallus sp. Trematode prevalence in H. ventrosa fluctuated seasonally. Prevalence usually peaked in summer between July and September-October and decreased in late winter-early spring. This seasonal change is chiefly explained by the life history patterns of the semelparous snail host. Hydrobia ventrosa has a maximum life span of about 2 yr and reproduces between June and November of its second calendar year. The first trematode infections appeared annually in May when the most abundant cohort of H. ventrosa, the second-calendar-year snails, mature. The prevalence continued to increase until August-September, throughout the reproductive period of the second-calendar-year snails, Prevalence decreased during winter, when most of the second-calendar-year snails died after reproduction. On the basis of longterm laboratory experiments, it has been shown that the late autumn-winter mortality was not the result of trematode infections. Seasonal patterns of prevalence were similar among the trematode species except for the monorchid Asymphylodora demeli, the only one using fish definitive hosts. Species-specific differences in the seasonal occurrence of prepatent infections and the predominance of certain larval stages in winter are interpreted as different strategies of the trematode species to survive the harsh winter conditions, or to survive the death of the first intermediate host in autumn-winter, or both.