Temperature induced giant shift of phonon energy in epitaxial boron nitride layers.

The recent progress in the growth of large-area boron nitride epilayers opens up new possibilities for future applications. However, it remains largely unclear how weakly attached two-dimensional BN layers interact with their substrate and how their properties are influenced by defects. In this work, we investigate hBN layers grown by metal organic vapor phase epitaxy using Fourier-transform infrared spectroscopy in the temperature range of 160-540 K. Our measurements reveal strong differences in the character of layer-substrate interaction for as-grown and delaminated epitaxial layers. A much weaker interaction of as-grown layers is explained by wrinkles formation that reduces strain at the layer-substrate interface, which for layers transferred to other substrates occurs only in a limited temperature range. The most striking result is the observation of a giant increase in theE1uphonon energy of up to ∼6 cm-1in a narrow temperature range. We show that the amplitude and temperature range of the anomaly is strongly modified by UV light illumination. The observed giant effect is explained in terms of strain generation resulting from charge redistribution between shallow traps and different defects, which can be interpreted as a result of strong electron-phonon coupling in hBN. The observed narrow temperature range of the anomaly indicates that the effect may be further enhanced for example by electrostrictive effects, expected for sp2boron nitride.

Assessing the potential of nematode metabarcoding for benthic monitoring of offshore oil platforms.

Environmental DNA metabarcoding is gaining momentum as a time and cost-effective tool for biomonitoring and environmental impact assessment. Yet, its use as a replacement for the conventional marine benthic monitoring based on morphological analysis of macrofauna is still challenging. Here we propose to study the meiofauna, which is much better represented in sediment DNA samples. We focus on nematodes, which are the most numerous and diverse group of meiofauna. Our aim is to assess the potential of nematode metabarcoding to monitor impacts associated with offshore oil platform activities. To achieve this goal, we used nematode-optimized marker (18S V1V2-Nema) and universal eukaryotic marker (18S V9) region to analyse 252 sediment DNA samples collected near three offshore oil platforms in the North Sea. For both markers, we analysed changes in alpha and beta diversity in relation to distance from the platforms and environmental variables. We also defined three impact classes based on selected environmental variables that are associated with oil extraction activities and used random forest classifiers to compare the predictive performance of both datasets. Our results show that alpha- and beta-diversity of nematodes varies with the increasing distance from the platforms. The variables directly related to platform activity, such as Ba and THC, strongly influence the nematode community. The nematode metabarcoding data provide more robust predictive models than eukaryotic data. Furthermore, the nematode community appears more stable in time and space, as illustrated by the overlap of nematode datasets obtained from the same platform three years apart. A significative negative correlation between distance and Shannon diversity also advocates for higher performance of the V1V2-Nema over the V9. Overall, these results suggest that the sensitivity of nematodes is higher compared to the eukaryotic community. Hence, nematode metabarcoding has the potential to become an effective tool for benthic monitoring in marine environment.

Multifrequency nonlinear model of magnetic material with artificial intelligence optimization.

Magnetic rings are extensively used in power products where they often operate in high frequency and high current conditions, such as for mitigation of excessive voltages in high-power switchgear equipment. We provide a general model of a magnetic ring that reproduces both frequency and current dependencies with the use of artificial intelligence (AI) optimization methods. The model has a form of a lumped element equivalent circuit that is suitable for power system transient studies. A previously published conventional (non-AI) model, which we take as a starting point, gives a good fit of parameters but uneven characteristics as a function of current, which pose numerical instabilities in transient simulations. We first enforce the Langevin function relationship to obtain smooth characteristics of parameters, which reduces the number of parameters and ensures their even characteristics, however, compromises fit quality. We then use AI metaheuristic optimization methods that give a perfect fit for the model in the whole range of frequency up to 100 MHz and current up to saturation, with smooth characteristics of its parameters. Additionally, for such fitted parameters, we show that it is feasible to find a frequency dependence for the magnetic saturation parameter of the Jiles-Atherton (JA) model, thus enabling frequency-dependent JA.

Environmental DNA metabarcoding for benthic monitoring: A review of sediment sampling and DNA extraction methods.

Environmental DNA (eDNA) metabarcoding (parallel sequencing of DNA/RNA for identification of whole communities within a targeted group) is revolutionizing the field of aquatic biomonitoring. To date, most metabarcoding studies aiming to assess the ecological status of aquatic ecosystems have focused on water eDNA and macroinvertebrate bulk samples. However, the eDNA metabarcoding has also been applied to soft sediment samples, mainly for assessing microbial or meiofaunal biota. Compared to classical methodologies based on manual sorting and morphological identification of benthic taxa, eDNA metabarcoding offers potentially important advantages for assessing the environmental quality of sediments. The methods and protocols utilized for sediment eDNA metabarcoding can vary considerably among studies, and standardization efforts are needed to improve their robustness, comparability and use within regulatory frameworks. Here, we review the available information on eDNA metabarcoding applied to sediment samples, with a focus on sampling, preservation, and DNA extraction steps. We discuss challenges specific to sediment eDNA analysis, including the variety of different sources and states of eDNA and its persistence in the sediment. This paper aims to identify good-practice strategies and facilitate method harmonization for routine use of sediment eDNA in future benthic monitoring.

Assessing the ecological quality status of the highly polluted Bagnoli area (Tyrrhenian Sea, Italy) using foraminiferal eDNA metabarcoding.

Morphology-based benthic foraminifera indices are increasingly used worldwide for biomonitoring the ecological quality of marine sediments. The recent development of foraminiferal eDNA metabarcoding offers a reliable, time-, and cost-effective alternative to morphology-based foraminiferal biomonitoring. However, the practical applications of these new tools are still highly limited. In the present study, we evaluate the response of benthic foraminifera and define the ecological quality status (EcoQS) in the Bagnoli area (Tyrrhenian Sea, Italy) based on a traditional morphology-based approach and eDNA metabarcoding. The geochemical data show that several sites in front of the former industrial plant contain higher concentrations of potentially toxic elements than the effect range median and are characterized by the highest total organic carbon (TOC) content, whereas the distantly located sites can be considered relatively low- to unpolluted. Significant differences (i.e., diversity and assemblage composition) in both morphological and molecular datasets were found between the relatively low- to unpolluted and the most polluted areas. Similarly, the selected ecological indices of both morphological and molecular datasets strikingly and congruently resulted in a clear separation following the environmental stress gradient. The molecular indices (i.e., g-exp(H'bc), g-Foram AMBI, and g-Foram AMBI-MOTUs) reliably identified poor-to-bad EcoQS in the polluted area in front of the former industrial plant. On the other hand, the Foram-AMBI based on morphology well identified an overall trend but seemed to overestimate the EcoQS if the traditional class boundaries were considered. The congruent and complementary trends between morphological and metabarcoding data observed in the case of the Bagnoli site further support the application of foraminiferal metabarcoding in routine biomonitoring to assess the environmental impacts of heavily polluted marine areas.

Expanding ecological assessment by integrating microorganisms into routine freshwater biomonitoring.

Bioindication has become an indispensable part of water quality monitoring in most countries of the world, with the presence and abundance of bioindicator taxa, mostly multicellular eukaryotes, used for biotic indices. In contrast, microbes (bacteria, archaea and protists) are seldom used as bioindicators in routine assessments, although they have been recognized for their importance in environmental processes. Recently, the use of molecular methods has revealed unexpected diversity within known functional groups and novel metabolic pathways that are particularly important in energy and nutrient cycling. In various habitats, microbial communities respond to eutrophication, metals, and natural or anthropogenic organic pollutants through changes in diversity and function. In this review, we evaluated the common trends in these changes, documenting that they have value as bioindicators and can be used not only for monitoring but also for improving our understanding of the major processes in lotic and lentic environments. Current knowledge provides a solid foundation for exploiting microbial taxa, community structures and diversity, as well as functional genes, in novel monitoring programs. These microbial community measures can also be combined into biotic indices, improving the resolution of individual bioindicators. Here, we assess particular molecular approaches complemented by advanced bioinformatic analysis, as these are the most promising with respect to detailed bioindication value. We conclude that microbial community dynamics are a missing link important for our understanding of rapid changes in the structure and function of aquatic ecosystems, and should be addressed in the future environmental monitoring of freshwater ecosystems.

Towards apparent convergence in asymptotically safe quantum gravity.

The asymptotic safety scenario in gravity is accessed within the systematic vertex expansion scheme for functional renormalisation group flows put forward in Christiansen et al. (Phys Lett B 728:114, 2014), Christiansen et al. (Phy Rev D 93:044036, 2016), and implemented in Christiansen et al. (Phys Rev D 92:121501, 2015) for propagators and three-point functions. In the present work this expansion scheme is extended to the dynamical graviton four-point function. For the first time, this provides us with a closed flow equation for the graviton propagator: all vertices and propagators involved are computed from their own flows. In terms of a covariant operator expansion the current approximation gives access to Λ , R, R 2 as well as R µ ν 2 and higher derivative operators. We find a UV fixed point with three attractive and two repulsive directions, thus confirming previous studies on the relevance of the first three operators. In the infrared we find trajectories that correspond to classical general relativity and further show non-classical behaviour in some fluctuation couplings. We also find signatures for the apparent convergence of the systematic vertex expansion. This opens a promising path towards establishing asymptotically safe gravity in terms of apparent convergence.

Stimulation of activin A expression in rat aortic smooth muscle cells by thrombin and angiotensin II correlates with neointimal formation in vivo.

Vasoactive GTP-binding protein-coupled receptor agonists (e.g., angiotensin II [AII] and alpha-thrombin) stimulate the production of mitogenic factors from vascular smooth muscle cells. In experiments to identify mitogens secreted from AII- or alpha-thrombin-stimulated rat aortic smooth muscle (RASM) cells, neutralizing antibodies directed against several growth factors (e.g., PDGF and basic fibroblast growth factor [basic FGF]) failed to inhibit the mitogenic activity of conditioned media samples derived from the cells. In this report, we found that polyclonal neutralizing antibodies directed against purified human placental basic FGF reduced the mitogenic activity of AII-stimulated RASM cell-conditioned media and in immunoblot experiments identified a 26-kD protein (14 kD under reducing conditions) that was distinct from basic FGF. After purification from RASM cell-conditioned medium, amino acid sequence analysis identified the protein as activin A, a member of the TGF-beta superfamily. Increased activin A expression was observed after treatment of the RASM cells with AII, alpha-thrombin, and the protein kinase C agonist PMA. In contrast, PDGF-BB or serum caused only a minor induction of this protein. Although activin A alone only weakly stimulated RASM cell DNA synthesis, it demonstrated a potent comitogenic effect in combination with either EGF or heparin-binding EGF-like growth factor in the RASM cells, increasing DNA synthesis by up to fourfold. Furthermore, in a rat carotid injury model, activin A mRNA was upregulated within 6 h after injury followed by increases in immunoreactive protein detected in the expanding neointima 7 and 14 d later. Taken together, these results indicate that activin A is a vascular smooth muscle cell-derived factor induced by vasoactive agonists that may, either alone or in combination with other vascular derived growth factors, have a role in neointimal formation after arterial injury.

Thrombin receptor activation elicits rapid protein tyrosine phosphorylation and stimulation of the raf-1/MAP kinase pathway preceding delayed mitogenesis in cultured rat aortic smooth muscle cells: evidence for an obligate autocrine mechanism promoting cell proliferation induced by G-protein-coupled receptor agonist.

Treatment of quiescent rat aortic smooth muscle cells with either alpha-thrombin or a thrombin receptor-derived agonist peptide (SFLLRNP) resulted in pronounced increases in [3H]thymidine incorporation that were concentration dependent and reached a maximum of approximately 15-fold above serum-starved controls. However, in contrast to FBS, PDGF-BB, or basic fibroblast growth factor (bFGF), that initiated DNA synthesis promptly after 16-19 h, thymidine incorporation in response to thrombin was delayed by an additional 3-6 h. Delayed mitogenesis correlated with the appearance of a potent mitogenic activity in conditioned media samples obtained from thrombin-stimulated rat aortic smooth muscle cells, as assayed using Swiss 3T3 fibroblasts. This activity was not inhibited by neutralizing antibodies directed against PDGF or bFGF. Furthermore, in the Swiss 3T3 cells, simple addition of either alpha-thrombin or SFLLRNP failed to elicit a significant mitogenic response. In signal transduction studies, both thrombin and SFLLRNP treatment led to rapid tyrosine phosphorylation of proteins with apparent molecular masses of 42, 44, 75, 120, and 190 kD, respectively, as assessed by antiphosphotyrosine immunoblotting. The overall pattern of protein tyrosine phosphorylation was distinct from that observed after PDGF-BB addition. Activation of Raf-1 and the mitogen-activated protein (MAP) kinases p44mapk and p42mapk was also observed. However, the time course and duration of Raf-1/MAP kinase activation after thrombin stimulation were similar to those elicited by PDGF-BB. Taken together, our results indicate that thrombin-stimulated vascular smooth muscle proliferation is delayed and requires the de novo expression of one or more autocrine mitogens. In addition, the rapid induction of discrete intracellular signaling mechanisms by thrombin, including the Raf-1/MAP kinase pathway, appears to be insufficient alone to promote vascular smooth muscle cell mitogenesis.

Rat dihydrodiol dehydrogenase: complexity of gene structure and tissue-specific and sexually dimorphic gene expression.

Dihydrodiol dehydrogenase (DD; EC 1.3.1.20) catalyzes a novel pathway of polycyclic aromatic hydrocarbon (PAH) metabolism in which trans-dihydrodiols (proximate carcinogens) are oxidized to reactive o-quinones which are cytotoxic and genotoxic. In this study, the complementary DNA for rat liver DD was used to examine the structure and regulation of the DD gene. Southern analysis of rat genomic DNA confirmed that DD is a member of the multigene aldo-keto reductase superfamily. Conservative estimates indicate that the rat DD gene is at least 20-25 kilobases in length. Northern analysis showed that the rat liver transcript was 2.4 kilobases whereas the complementary DNA contains an open-reading frame of 966 nucleotides. Primer extension of male and female polyadenylated RNA indicated that the major transcription start sites are only 53 and 54 base pairs upstream from the translation start site, confirming that the RNA has a very long 3'-untranslated region. In male and female tissues, 2.4 kilobase transcripts predominate in liver, small intestine, and lung, which is consistent with a role for the enzyme in PAH metabolism. Transcripts were also detected in male (prostate)- and female (ovary, mammary gland, and uterus)-specific tissues. In the ovary, two transcripts were observed of 2.4 and 1.4 kilobases in length. Using benzenedihydrodiol as a model substrate for PAH trans-dihydrodiols, highest levels of DD activity were observed in the liver and small intestine of both sexes. Enzyme activity is 2.5-fold higher in the female liver versus the male liver. This sexual dimorphism can be explained by increases in the DD mRNA and enzyme protein as measured by dot-blot and immunotitration analyses, respectively. The latter measurements indicate that DD represents 1.0% of the soluble protein in female liver but is only 0.5% of the soluble protein in male liver. Hormonal ablation (ovariectomy and hypophysectomy) abolishes the sexual dimorphism observed in levels of DD mRNA, enzyme protein, and enzyme activity. Administration of estrogens to males is sufficient to establish the female pattern of gene expression. These data indicate that DD gene expression is hormonally regulated, that estrogens exert their effect at the level of the mRNA, and that aldo-keto reductases involved in PAH metabolism may have their expression regulated by female sex hormones.

Chiral fermions in asymptotically safe quantum gravity.

We study the consistency of dynamical fermionic matter with the asymptotic safety scenario of quantum gravity using the functional renormalisation group. Since this scenario suggests strongly coupled quantum gravity in the UV, one expects gravity-induced fermion self-interactions at energies of the Planck scale. These could lead to chiral symmetry breaking at very high energies and thus to large fermion masses in the IR. The present analysis which is based on the previous works (Christiansen et al., Phys Rev D 92:121501, 2015; Meibohm et al., Phys Rev D 93:084035, 2016), concludes that gravity-induced chiral symmetry breaking at the Planck scale is avoided for a general class of NJL-type models. We find strong evidence that this feature is independent of the number of fermion fields. This finding suggests that the phase diagram for these models is topologically stable under the influence of gravitational interactions.

Protist metabarcoding and environmental biomonitoring: Time for change.

High-throughput amplicon sequencing of environmental DNA and/or RNA proved to be a powerful tool to describe protist diversity. This new approach called also the metabarcoding has totally transformed our view of protist diversity, revealing a large number of novel lineages and expanding the range of protist phylogenetic diversity at almost every taxonomic level. However, until now the objectives of the vast majority of metabarcoding studies were purely academic. Practical applications of protist metabarcoding are surprisingly scarce, despite the fact that several groups of protists are commonly used as bioindicators of environmental impacts in freshwater or marine ecosystems. Here, we are reviewing studies that examine the ecological applications of metabarcoding for two groups of well-known protist bioindicators: diatoms and foraminifera. The results of these studies show that despite some biological and technical biases, molecular data quite faithfully reflect the morphology-based biotic indices and provide a similar assessment of ecosystem status. In view of these results, protist metabarcoding appears as a rapid and accurate tool for the evaluation of the quality of aquatic ecosystems. Hence, we plead for integration of protist metabarcoding in future biomonitoring projects as a complement of traditional methods and a source of new biosensors for environmental impact assessment.

Selective zircon accumulation in a new benthic foraminifer, Psammophaga zirconia, sp. nov.

Benthic foraminifera are single-celled eukaryotes that make a protective organic, agglutinated or calcareous test. Some agglutinated, single-chambered taxa, including Psammophaga Arnold, 1982, retain mineral particles in their cytoplasm, but the selective mechanism of accumulation is not clear. Here, we report the ability of a foraminiferal species to select and accumulate zircons and other heavy minerals in their cytoplasm. In particular, the use of Scanning Electron Microscope coupled with an Energy Dispersive X-ray microanalysis system (SEM-EDS) enabled a representative overview of the mineral diversity and showed that the analysed Psammophaga zirconia sp. nov. individuals contained dominantly crystals of zircon (51%), titanium oxides (27%), and ilmenite (11%) along with minor magnetite and other minerals. The studied specimens occur in the shallow central Adriatic Sea where the sediment has a content of zircon below 1% and of other heavy minerals below 4%. For that reason we hypothesize that: (i) P. zirconia may be able to chemically select minerals, specifically zircon and rutile; (ii) the chemical mechanism allowing the selection is based on electrostatic interaction, and it could work also for agglutinated foraminifera (whether for ingestion, like Xenophyophores, or incorporation in the test as in many other described taxa). In particular, this aptitude for high preferential uptake and differential ingestion or retention of zircon is reported here for the first time, together with the selection of other heavy minerals already described in members of the genus Psammophaga. They are generally counted among early foraminifera, constructing a morphologically simple test with a single chamber. Our molecular phylogenetic study confirms that P. zirconia is a new species, genetically distinctive from other Psammophaga, and occurs in the Adriatic as well as in the Black Sea.

Constrictive bronchiolitis obliterans in patient with Castelman's disease.

A 37-year-old woman with hialin- vascular type Castelman's disease (CD) localised in the retroperitoneal region, incompletely resected, developed progressive dyspnoea. The chest radiograph taken 3 months before the operation was normal. The chest CT scan revealed diffused bronchiectases, hyperinflation and air trapping. Pulmonary function tests disclosed severe obstructive impairment with hyperinflation. The bronchoscopic examination of the bronchial tree was normal. Cultures of sputum, bronchial washing and blood were negative. No pemphigus antibodies were found. Mycoplasmal, chlamydial and viral infections were excluded. Histological examination of specimens obtained by open lung biopsy revealed bronchiolar inflammation, submucosal bronchial fibrosis with obliteration of bronchiolar lumen. Constrictive bronchiolitis obliterans (CBO) was diagnosed. Despite slight clinical and spirometric improvements that were achieved due to corticosteroid therapy, one year later she died as a result of respiratory failure. It is widely known that patients with CD develop CBO during the course of paraneoplastic pemphigus. However we present the case of CBO and CD but without any symptoms of this condition.

Accurate assessment of the impact of salmon farming on benthic sediment enrichment using foraminiferal metabarcoding.

Assessing the environmental impact of salmon farms on benthic systems is traditionally undertaken using biotic indices derived from microscopic analyses of macrobenthic infaunal (MI) communities. In this study, we tested the applicability of using foraminiferal-specific high-throughput sequencing (HTS) metabarcoding for monitoring these habitats. Sediment samples and physico-chemical data were collected along an enrichment gradient radiating out from three Chinook salmon (Oncorhynchus tshawytscha) farms in New Zealand. HTS of environmental DNA and RNA (eDNA/eRNA) resulted in 1,875,300 sequences that clustered into 349 Operational Taxonomic Units. Strong correlations were observed among various biotic indices calculated from MI data and normalized fourth-root transformed HTS data. Correlations were stronger using eRNA compared to eDNA data. Quantile regression spline analyses identified 12 key foraminiferal taxa that have potential to be used as bioindicator species. This study demonstrates the huge potential for using this method for biomonitoring of fish-farming and other marine industrial activities.

Novel cardiovascular actions of the activins.

Proliferation and directed migration of vascular cells are key components in vascular diseases such as atherosclerosis and restenosis following percutaneous transluminal coronary angioplasty. However, the precise cellular and molecular mechanisms involved in the control of vascular cell proliferation or migration at the tissue level remain largely undefined. Molecules contributing to these processes are elaborated by distinct cell types and act in both autocrine and paracrine modes. They include two broad classes, polypeptide growth factors and vasoactive G-protein-coupled receptor (GPCR) agonists. Examples of the former, such as platelet-derived growth factor, bind to and activate cell surface receptor tyrosine kinases, initiating intracellular biochemical signaling pathways associated with cell proliferation or migration. In contrast, recent evidence suggests that vasoactive GPCR agonists (e.g. angiotensin II, endothelin-1, alpha-thrombin) elicit cell growth indirectly by inducing the production of autocrine or paracrine factors in vascular cells. Recent studies have identified activin A as a novel component of conditioned medium obtained from GPCR agonist-stimulated vascular smooth muscle cells (SMCs). Although activin A alone only weakly stimulated rat aortic SMC DNA synthesis, it demonstrated a potent co-mitogenic effect in combination with either epidermal growth factor (EGF) or heparin binding EGF-like growth factor in these cells, increasing DNA synthesis by up to 5- and 4-fold respectively. Furthermore, in a rat carotid-injury model, activin A mRNA was upregulated within 6 h after injury, followed by increases in immunoreactive protein detected in the expanding neointima 7 to 14 days later. Taken together, these results indicate that activin A is a common vascular SMC-derived growth factor induced by vasoactive agonists that may, either alone or in combination with other factors, contribute to fibroproliferative vascular diseases.

Morphological and molecular characterization of a new terrestrial allogromiid species: Edaphoallogromia australica gen. et spec. nov. (Foraminifera) from Northern Queensland (Australia).

The foraminiferal family Allogromiidae occurs mainly in marine environments, although some genera are described from brackish and freshwater habitats. We report here the occurrence of a terrestrial allogromiid foraminiferan. Phylogenetic relationships were investigated by sequencing part of the SSU rDNA. DNA sequence analysis confirms a close relationship of the new species to the genus Allogromia. Morphological studies corroborate the affiliation to the family Allogromiidae but the lack of an entosolenian tube and an internal septum as well as the different ecology do not allow a classification into a known genus of this family. Taking the molecular, morphological and ecological differences into account, a new genus Edaphoallogromia with the type species E. australica is erected.

Phylogenetic analysis of the suborder plagiorchiata (Platyhelminthes, Digenea) based on partial lsrDNA sequences.

The phylogenetic relationships and systematic position of the members of the suborder Plagiorchiata, one of the derived and most diverse groups of Digenea, have always been controversial. Here, we present a phylogeny of this group based on the analysis of partial sequences of the lsrDNA in 28 species representing 13 families of Plagiorchiata, as well as four outgroups. Our results show that the Plagiorchiata, as considered by most authors, is not monophyletic, and that the superfamilies Opecoeloidea, and most probably Dicrocoelioidea and Gorgoderoidea, may have to be removed from this suborder. According to our results, the Plagiorchiata includes only parasites of terrestrial vertebrates. We find the Plagiorchiata to be composed of two well-supported clades which can be ranked as superfamilies: (1) Plagiorchioidea, including the Plagiorchiidae, Haematoloechidae, Telorchiidae, Brachycoeliidae and Leptophallidae; and (2) Microphalloidea containing the Microphallidae, Prosthogonimidae, Lecithodendriidae and Pleurogenidae. The genetic analysis also allowed revision of the position of several taxa of Plagiorchiata, including: (1) a confirmation of the position of the Brachycoeliidae within the Plagiorchiata; (2) a close phylogenetic relationships of Macrodera with Paralepoderma, Leptophallus and Metaleptophallus; (3) the grouping of Opisthioglyphe and Telorchis within a distinct and strongly supported clade; and (4) the placement of Allassogonoporus amphoraeformis within the Pleurogenidae, and not close to Lecithodendriidae. Some systematic changes, corresponding to these results, are proposed.

Molecular identification of developmental stages in Opecoelidae (Digenea).

Nuclear ribosomal DNA sequences represent a useful tool for distinction of poorly differentiated developmental stages, such as trematode cercariae or metacercariae. Here, the complete internal transcribed spacer region of the ribosomal DNA (ITS 1 + 5.8S + ITS 2) was sequenced for 29 specimens of the digenean family Opecoelidae, including 16 adult specimens and 13 undescribed larval stages (nine cercariae and four metacercariae) occurring in various marine host organisms. Six cercariae and three metacercariae were found to match their corresponding adult form. This work also revealed that cercariae of the same species are able to infect more than one gastropod host species, suggesting that the specificity for the first intermediate host within the Digenea may be lower than previously thought.