Molecular evidence for the hybridity ofIlex × wandoensis and the phylogenetic study of KoreanIlex based on ITS sequence data

In this study, internal transcribed spacer (ITS) regions within the nuclear ribosomal DNA of KoreanIlex were analyzed in order to investigate any molecular evidence thatI. × wandoensis could serve as a putative hybrid betweenI. cornuta andI. integra. We also sought to clearly identify taxonomic relationships and problems caused by consecutive external morphological characters among taxa in the genus. We sequenced 20 clones fromI. × wandoensis and found these individuals displayed intra-genomic polymorphisms within ITS regions. The analysis of the clones clearly demonstrated the presence of discrete sequences from bothI. cornuta andI. integra, thereby confirmingI. × wandoensis is a species that was formed by crossingI. cornuta andI. integra. Lastly, the subgenusIlex, which contains the evergreen species, failed to form a monophyletic group in a strict consensus tree that was prepared based upon ITS regions.I. crenata var.microphylla in the subgenus KoreaIlex, which has presented taxonomic problems previously, formed an independent clade within the consensus tree, thereby showing distinction fromI. crenata.) Genetic discontinuity ofI. macropoda andI. Macropoda for.pseudo-macropoda individuals couldn't be confirmed.     

How good are Bayesian belief networks for environmental management? A test with data from an agricultural river catchment

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Restriction data from chloroplast DNA for phylogenetic reconstruction: Is there only one accurate way of scoring?

Information from the same restriction analysis of chloroplast DNA of 33 taxa ofRubiaceae was scored in four different ways, two of which were based on fragments, and two on restriction sites, and they were subsequently analysed with Wagner parsimony. The methods resulted in different phylogenetic trees. The inherent differences between the methods relate to the amount of non-homologous characters and dependent characters, but none of the methods will systematically bias the resulting cladograms. The fragment analyses are much less time-consuming, but probably less accurate, than the site analyses. The choice of method is dependent on a trade-off between accuracy and resources (time). One important recommendation is made: all phylogenetic analyses of chloroplast DNA data should be accompanied by a data matrix and contain information on how the matrix was compiled.     

SC²ATmd: a tool for integration of the figure of merit with cluster analysis for gene expression data

Standard and Consensus Clustering Analysis Tool for Microarray Data (SC2ATmd) is a MATLAB-implemented application specifically designed for the exploration of microarray gene expression data via clustering. Implementation of two versions of the clustering validation method figure of merit allows for performance comparisons between different clustering algorithms, and tailors the cluster analysis process to the varying characteristics of each dataset. Along with standard clustering algorithms this application also offers a consensus clustering method that can generate reproducible clusters across replicate experiments or different clustering algorithms. This application was designed specifically for the analysis of gene expression data, but may be used with any numerical data as long as it is in the right format. AVAILABILITY: SC2ATmd may be freely downloaded from http://www.compbiosci.wfu.edu/tools.htm.     

Spatial analyses of the phylogenetic diversity of Minaria (Apocynaceae): assessing priority areas for conservation in the Espinhaço Range,Brazil

The protection of areas that shelter high evolutionary diversity represented by geographically and phylogenetically isolated lineages is becoming an important conservation strategy. Nevertheless, the spatial distribution of this component of biodiversity is still unknown for most groups, which limits its application for selecting priority areas to conserve. In the present study, we reconstructed the phylogeny of Minaria (Apocynaceae) based on plastid (trnH-psbA, rps16, trnS-trnG and trnD-trnT) and nuclear (ITS and ETS) DNA markers and 34 morphological characters, and analysed the geographic distribution of the phylogenetic diversity (PD) and endemism (PE) in this genus. Minaria includes 21 species that are highly concentrated in the Espinhaço Range, in eastern Brazil, most of which (～75%) are narrowly distributed. The spatial analyses of PD and PE of Minaria indicate four evolutionary relevant areas in this region. The Serra do Cipó and the Diamantina Plateau contain 10 endemic species and present the highest levels of PD. However, the two other areas also deserve special attention. Rio de Contas has high levels of PE, because of two endemic sister species that represent a phylogenetically isolated lineage and the Southern Espinhaço Range houses the most critically endangered species of the genus. Most endemic species of Minaria occur in vegetation islands on rocky outcrops (campos rupestres). These low-fuel areas are less susceptible to fire, suggesting that the Espinhaço Range has served as a historical refuge for fire-sensitive lineages. Our results suggest that conservation units in the Espinhaço Range cover a great proportion of the evolutionary diversity of Minaria and that fire management is probably an important strategy to preserve this endemic biodiversity.     

Beyond taxonomic diversity patterns: how do α, β and γ components of bird functional and phylogenetic diversity respond to environmental gradients across France?

Aim To test how far can macroecological hypotheses relating diversity to environmental factors be extrapolated to functional and phylogenetic diversities, i.e. to the extent to which functional traits and evolutionary backgrounds vary among species in a community or region. We use a spatial partitioning of diversity where regional or γ‐diversity is calculated by aggregating information on local communities, local or α‐diversity corresponds to diversity in one locality, and turnover or β‐diversity corresponds to the average turnover between localities and the region. Location France. Methods We used the Rao quadratic entropy decomposition of diversity to calculate local, regional and turnover diversity for each of three diversity facets (taxonomic, phylogenetic and functional) in breeding bird communities of France. Spatial autoregressive models and partial regression analyses were used to analyse the relationships between each diversity facet and environmental gradients (climate and land use). Results Changes in γ‐diversity are driven by changes in both α‐ and β‐diversity. Low levels of human impact generally favour all three facets of regional diversity and heterogeneous landscapes usually harbour higher β‐diversity in the three facets of diversity, although functional and phylogenetic turnover show some relationships in the opposite direction. Spatial and environmental factors explain a large percentage of the variation in the three diversity facets (>60%), and this is especially true for phylogenetic diversity. In all cases, spatial structure plays a preponderant role in explaining diversity gradients, suggesting an important role for dispersal limitations in structuring diversity at different spatial scales. Main conclusions Our results generally support the idea that hypotheses that have previously been applied to taxonomic diversity, both at local and regional scales, can be extended to phylogenetic and functional diversity. Specifically, changes in regional diversity are the result of changes in both local and turnover diversity, some environmental conditions such as human development have a great impact on diversity levels, and heterogeneous landscapes tend to have higher diversity levels. Interestingly, differences between diversity facets could potentially provide further insights into how large‐ and small‐scale ecological processes interact at the onset of macroecological patterns.     

Reconstructing long-term ecological data from annual census returns: A test for observer bias in counts of bird populations on Skokholm 1928–2002

Long-term ecological data are essential for conservation and to monitor and evaluate the effects of environmental change. Bird populations have been routinely assessed on islands off the British coast for many years and here long term data for one such island, Skokholm, is evaluated for robustness in the light of some 20 changes in observers (wardens) on the island over nearly eight decades. It was found that the dataset was robust when compared to bootstrap data with no species showing significant changes in abundance in years when wardens changed. It is concluded that the breeding bird populations on Skokholm and other British offshore islands are an important scientific resource and that protocols should be enacted to ensure the archiving of records, the continuance of data collection using standardised protocols into the future, and the recognition of such long-term data for science in terms of an appropriate conservation designation.     

RNA–LIM: A novel procedure for analyzing protein/single-stranded RNA propensity data with concomitant estimation of interface structure

RNA–LIM is a procedure that can analyze various pseudo-potentials describing the affinity between single-stranded RNA (ssRNA) ribonucleotides and surface amino acids to produce a coarse-grained estimate of the structure of the ssRNA at the protein interface. The search algorithm works by evolving an ssRNA chain, of known sequence, as a series of walks between fixed sites on a protein surface. Optimal routes are found by application of a set of minimal “limiting” restraints derived jointly from (i) selective sampling of the ribonucleotide amino acid affinity pseudo-potential data, (ii) limited surface path exploration by prior determination of surface arc lengths, and (iii) RNA structural specification obtained from a statistical potential gathered from a library of experimentally determined ssRNA structures. We describe the general approach using a NAST (Nucleic Acid Simulation Tool)-like approximation of the ssRNA chain and a generalized pseudo-potential reflecting the location of nucleic acid binding residues. Minimum and maximum performance indicators of the methodology are established using both synthetic data, for which the pseudo-potential defining nucleic acid binding affinity is systematically degraded, and a representative real case, where the RNA binding sites are predicted by the amplified antisense RNA (aaRNA) method. Some potential uses and extensions of the routine are discussed. RNA–LIM analysis programs along with detailed instructions for their use are available on request from the authors.     

Single cell variability of L. monocytogenes grown on liver pâté and cooked ham at 7 degrees C: comparing challenge test data to predictive simulations

AIMS: The variability in growth between individual Listeria monocytogenes cells was investigated on liver paté and cooked ham. These results were compared to Monte Carlo simulations based on data collected previously in broths (Francois et al., submitted for publication). METHODS AND RESULTS: Single cells were isolated by a dilution protocol and inoculated on 15 g samples of liver paté and cooked ham, pasteurized in the packaging. Of each product, 250 samples were inoculated, of which 50 samples were analysed for L. monocytogenes on each analysis day. Results were compared to simulations, based on distributions that describe the variability of the individual cell lag phases and generation times of L. monocytogenes cultivated in broths. Based on the same simulation techniques, the variability effect was investigated for different inoculum levels (10, 100, 10,00 and 10,000 cells). It was demonstrated that the expected variability of the outgrowth of L. monocytogenes in a challenge test is very high for low inoculum levels. CONCLUSIONS: The variability in growth characteristics observed between different single L. monocytogenes cells on foods is very large. The simulations based on the previously collected optical density data in broths, could be confirmed by foods inoculated with single L. monocytogenes cells. SIGNIFICANCE AND IMPACT OF THE STUDY: The large variability between different individual L. monocytogenes cells has serious consequences for the experimental design of a challenge test. One thousand cells per portion are necessary in order to reduce the variability to acceptable levels and quantify the behaviour of the pathogen consistently with a reasonable number of challenge tests.     

Complete mitochondrial genome of a new vole Proedromys liangshanensis (Rodentia: Cricetidae) and phylogenetic analysis with related species: are there implications for the validity of the genus Proedromys?

AIM. The complete mitochondrial genome sequence of a newly discovered vole, Proedromys liangshanensis (Rodentia: Cricetidae: Arvicolinae), was determined. RESULTS. The mitogenome of P. liangshanensis is 16,296 bp in length. As with most other mammals, it contains the same gene order and an identical number of genes or regions, including 13 protein-coding genes, two ribosomal RNA genes, 22 transfer RNA genes, and one putative control region. The A+T content of the control region is 58.2%, the lowest proportion detected so far in Myomorpha. To confirm the phylogenetic position of P. liangshanensis, we carried out phylogenetic analyses based on complete mitochondrial genomic data using Bayesian, maximum parsimony, and maximum likelihood methods. CONCLUSION. All results revealed that P. liangshanenis is sister to Microtus. Although the results do not bear light on the validity of the genus Proedromys, based on the morphological characters, we suggest that Proedromys is an independent genus of equal rank to the genus Microtus.     

Development of genotoxicity test procedures with Episkin, a reconstructed human skin model: towards new tools for in vitro risk assessment of dermally applied compounds?

Today reconstructed skin models that simulate human skin, such as Episkin, are widely used for safety or efficacy pre-screening. Moreover, they are of growing interest for regulatory purposes in the framework of alternatives to animal testing. In order to reduce and eventually replace results of in vivo genotoxicity testing with in vitro data, there is a need to develop new complementary biological models and methods with improved ability to predict genotoxic risk. This can be achieved if these new assays do take into account exposure conditions that are more relevant than in the current test systems. In an attempt to meet this challenge, two new applications using a human reconstructed skin model for in vitro genotoxicity assessment are proposed. The skin is the target organ for dermally exposed compounds or environmental stress. Although attempts have been made to develop genotoxicity test procedures in vivo on mouse skin, human reconstructed skin models have not been used for in vitro genotoxicity testing so far, although they present clear advantages over mouse skin for human risk prediction. This paper presents the results of the development of a specific protocol allowing to perform the comet assay, a genotoxicity test procedure, on reconstructed skin. The comet assay was conducted after treatment of Episkin with UV, Lomefloxacin and UV or 4-nitroquinoline-N-oxide (4NQO). Treatment with the sunscreen Mexoryl was able to reduce the extent of comet signal. A second approach to use reconstructed epidermis in genotoxicity assays is also proposed. Indeed, the skin is a biologically active barrier driving the response to exposure to chemical agents and their possible metabolites. A specific co-culture system (Figure 1) using Episkin to perform the regular micronucleus assay is presented. Micronucleus induction in L5178Y cells cultured underneath Episkin was assessed after treatment of the reconstructed epidermis with mitomycin C, cyclophosphamide or apigenin. This second way of using human reconstructed skin for genotoxicity testing aims at improving the relevance of exposure conditions in in vitro genotoxicity assays for dermally applied compounds.     

Biomimetic oxidative treatment of spruce wood studied by pyrolysis–molecular beam mass spectrometry coupled with multivariate analysis and 13C-labeled tetramethylammonium hydroxide thermochemolysis: implications for fungal degradation of wood

In this work, pyrolysis–molecular beam mass spectrometry analysis coupled with principal components analysis and ¹³C-labeled tetramethylammonium hydroxide thermochemolysis were used to study lignin oxidation, depolymerization, and demethylation of spruce wood treated by biomimetic oxidative systems. Neat Fenton and chelator-mediated Fenton reaction (CMFR) systems as well as cellulosic enzyme treatments were used to mimic the nonenzymatic process involved in wood brown-rot biodegradation. The results suggest that compared with enzymatic processes, Fenton-based treatment more readily opens the structure of the lignocellulosic matrix, freeing cellulose fibrils from the matrix. The results demonstrate that, under the current treatment conditions, Fenton and CMFR treatment cause limited demethoxylation of lignin in the insoluble wood residue. However, analysis of a water-extractable fraction revealed considerable soluble lignin residue structures that had undergone side chain oxidation as well as demethoxylation upon CMFR treatment. This research has implications for our understanding of nonenzymatic degradation of wood and the diffusion of CMFR agents in the wood cell wall during fungal degradation processes.     

Abundance, diversity and fidelity of macroinvertebrates sheltering beneath rocks during tidal emersion in an intertidal cobble field: Does the intermediate disturbance hypothesis hold for less exposed shores with smaller rocks?

The community structure of macroinvertebrates occurring beneath cobbles during tidal emersion was investigated in an intertidal cobble field on the east coast of the south island of New Zealand. In order to test whether the intermediate disturbance hypothesis (IDH) holds in less exposed shores comprised exclusively of smaller rocks (cobbles) we measured 1) the size frequency distribution of cobbles (mass and basal surface area), 2) the incidence of barnacle encrustation as an indirect measure of cobble disturbance, 3) the abundances of the dominant macroinvertebrates occurring beneath cobbles (the crab Hemigrapsus crenulatus, the chiton Sypharochiton pelliserpentis and the snail Diloma zealandica), and 4) macroinvertebrate fidelity beneath a range of cobble sizes. Only large cobbles were found to be encrusted with barnacles, indicative of their greater resistance to physical disturbance by waves and currents. Mark and relocate experiments indicated fidelity to a given cobble over two tidal cycles was 11-14% for crabs and did not vary with cobble size. In contrast, fidelity to medium and large cobbles was higher for chitons (26-45%) and snails (17-31%) and generally increased with increasing cobble size. In contrast to the predictions of the IDH, there was a positive correlation between increasing cobble size and increasing species diversity of organisms found under the cobbles. This pattern is not simply attributable to greater space available beneath large cobbles as there was a significantly greater density of macroinvertebrates beneath medium and large, compared to the small cobbles. Increasing species diversity with increasing cobble size appears to be independent of patterns of species recolonization associated with small cobbles being subject to high disturbance or interspecific competitive hierarchies associated with low disturbance beneath large cobbles. Instead, factors such as aggregative behaviors, and reducing vulnerability to predators and desiccation stress are likely to explain the relationship between cobbles and their associated macroinvertebrate communities.     

Gravimetric screening method for fungal decay of paper: inoculation with <Emphasis Type="Italic">Trametes</Emphasis> <Emphasis Type="Italic">versicolor</Emphasis>

The European standard test EN 113 for fungal degradation of solid wood has been adapted for degradation of paper by white rot fungus (Trametes versicolor). Fungal degradation of paper sheets may potentially be used for screening different wood preservatives on paper instead of solid wood. The paper samples showed higher relative mass losses compared to wood, and samples pretreated with boric acid, copper sulfate and polymerized linseed oil were successfully tested for biodegradation using the paper sheet method. The results on paper degradation were compared with wood, both as wood blocks (according to standard test) and wood cut in sections forming layered structures mimicking paper layers.     

‘Can he have the test for bipolar,doctor? His dad's got it’: exploring the potential of general practitioners to work with children and young people presenting in primary care with common mental health problems – a clinical initiative

Background General practitioners (GPs) play a key role in assessing and managing adult mental health problems, but this input is not seen in their management of child and adolescent mental health. Mental health problems in 5–19-year-olds are common, yet detection rates in primary care are low. The symptoms of most adult diagnoses of mental health problems are present by mid-adolescence, yet the typical time from onset to diagnosis is 5–15 years. The role of general practice in this area has been underexplored.Aim This pilot study explores the potential of GPs to respond to common mental health problems in children and adolescents.Design Children and young people who would have ordinarily been referred to Child and Adolescent Mental Health Services (CAMHS) were seen in a GP setting. In a UK general practice surgery serving a disadvantaged population.Method Children and young people were seen for an initial biopsychosocial assessment and formulation of the presenting concerns. GP-based interventions were offered as appropriate or referred to CAMHS.Results Data from the first 50 children (2–19 years) are presented. Twenty younger children (10 years and under) and 30 older children (11 years and above) were seen. Eighteen referrals were made to CAMHS. GP interventions included watchful waiting, brief behavioural interventions, non-directive counselling, brief cognitive– behavioural therapy (CBT) and liaison with colleagues in education, CAMHS and the voluntary sector.Conclusion This clinical pilot demonstrates that with adequate time, access to supervision and practice support, children and young people experiencing emotional and behavioural problems associated with common mental health issues can be helped in primary care.     

The phylogenetic position of <Emphasis Type="Italic">Choerodonicola</Emphasis> Cribb, 2005 (Digenea: Opecoelidae) with a partial life-cycle for a new species from the blue-barred parrotfish <Emphasis Type="Italic">Scarus ghobban</Emphasis> Forsskål (Scaridae) in Moreton Bay,Australia

Choerodonicola Cribb, 2005 is a minor genus of opecoelid trematodes defined for species with exceptionally small eggs but otherwise generalised morphology. Four species are currently recognised, all from fishes collected in Japanese waters but each from different perciform families: a labrid, a scarid, a sparid and pinguipeds. We report on a new species, Choerodonicola arothokoros n. sp., from the blue-barred parrotfish Scarus ghobban Forsskål (Scaridae) collected in subtropical waters of Moreton Bay, south-east Queensland, Australia. Using genetic sequence data for the ITS2 rDNA marker, we matched adult C. arothokoros to intramollsucan stages discovered in an intertidal gastropod Herpetopoma atratum (Gmelin) (Vetigastropoda: Chilodontidae) collected in close proximity to the fish hosts. Notably, the cercariae lack a penetration stylet and are among the smallest known in the Opecoelidae. We provide the first assessment of the phylogenetic position of Choerodonicola based on sequence data generated for the phylogenetically informative 18S and 28S rRNA coding regions, for C. arothokoros and also C. renko Machida, 2014, which we recollected from the yellowback seabream Dentex hypselosomus Bleeker from the fish market in Minabe, Wakayama Prefecture, Japan. In our analyses, species of Choerodonicola resolved to neither of the major marine Plagioporinae (sensu lato) clades, clustering instead with Trilobovarium parvvatis Martin, Cutmore & Cribb, 2017, Podocotyloides parupenei (Manter, 1963) Pritchard, 1966 and Macvicaria magellanica Laskowski, Je?ewski & Zdzitowiecki, 2013. This clade is phylogenetically distinctive such that it has the potential to be recognised as a new opecoelid subfamily, but further investigation is required to establish the bounds for such a grouping and to determine the morphological and/or life-history patterns reflected by the phylogeny. Finally, we propose C. interruptus (Manter 1954) n. comb. for a species previously recognised in Plagioporus Stafford, 1904 and known only from Pseudolabrus miles (Schneider & Forster), a labrid endemic to New Zealand.