Validation of the detection of Pseudo-nitzschia spp. using specific RNA probes tested in a microarray format: Calibration of signal based on variability of RNA content with environmental conditions.

Harmful algal blooms (HAB) occur worldwide and cause health problems and economic damage to fisheries and tourism. Monitoring for toxic algae is therefore essential but is based primarily on light microscopy, which is time consuming and can be limited by insufficient morphological characters such that more time is needed to examine critical features with electron microscopy. Monitoring with molecular tools is done in only a few places world-wide. EU FP7 MIDTAL (Microarray Detection of Toxic Algae) used SSU and LSU rRNA genes as targets on microarrays to identify toxic species. In order to comply with current monitoring requirements to report cell numbers as the relevant threshold measurement to trigger closure of fisheries, it was necessary to calibrate our microarray to convert the hybridisation signal obtained to cell numbers. Calibration curves for two species of Pseudo-nitzschia for use with the MIDTAL microarray are presented to obtain cell numbers following hybridisation. It complements work presented by Barra et al. (2012b. Environ. Sci. Pollut. Res. doi: 10.1007/s11356-012-1330-1v) for two other Pseudo-nitzschia spp., Dittami and Edvardsen (2012a. J. Phycol. 48, 1050) for Pseudochatonella, Blanco et al. (2013. Harmful Algae 24, 80) for Heterosigma, McCoy et al. (2013. FEMS. doi: 10.1111/1574-6941.12277) for Prymnesium spp., Karlodinium veneficum, and cf. Chatonella spp. and Taylor et al. (2014. Harmful Algae, in press) for Alexandrium.

Biodiversity and Biogeography of Chthamalid Barnacles from the North-Eastern Pacific (Crustacea Cirripedia)

The biogeography and ecology of the species of Chthamalus present on the west coast of America are described, using data from 51 localities from Alaska to Panama, together with their zonation on the shore with respect to that of other barnacles. The species present were C. dalli, Pilsbry 1916, C. fissus, Darwin, 1854, C. anisopoma Pilsbry 1916 and four species in the C. panamensis complex. The latter are C. panamensis Pilsbry, 1916, C. hedgecocki, Pitombo & Burton, 2007, C. alani nom. nov. (formerly C. southwardorum Pitombo & Burton, 2007) and C. newmani sp. nov.). These four species were initially separated by enzyme electrophoresis. They could only be partially separated by DNA bar coding but may be separated using morphological characters.

PhytoREF: a reference database of the plastidial 16S rRNA gene of photosynthetic eukaryotes with curated taxonomy

Photosynthetic eukaryotes have a critical role as the main producers in most ecosystems of the biosphere. The ongoing environmental metabarcoding revolution opens the perspective for holistic ecosystems biological studies of these organisms, in particular the unicellular microalgae that often lack distinctive morphological characters and have complex life cycles. To interpret environmental sequences, metabarcoding necessarily relies on taxonomically curated databases containing reference sequences of the targeted gene (or barcode) from identified organisms. To date, no such reference framework exists for photosynthetic eukaryotes. In this study, we built the PhytoREF database that contains 6490 plastidial 16S rDNA reference sequences that originate from a large diversity of eukaryotes representing all known major photosynthetic lineages. We compiled 3333 amplicon sequences available from public databases and 879 sequences extracted from plastidial genomes, and generated 411 novel sequences from cultured marine microalgal strains belonging to different eukaryotic lineages. A total of 1867 environmental Sanger 16S rDNA sequences were also included in the database. Stringent quality filtering and a phylogeny-based taxonomic classification were applied for each 16S rDNA sequence. The database mainly focuses on marine microalgae, but sequences from land plants (representing half of the PhytoREF sequences) and freshwater taxa were also included to broaden the applicability of PhytoREF to different aquatic and terrestrial habitats. PhytoREF, accessible via a web interface (http://phytoref.fr), is a new resource in molecular ecology to foster the discovery, assessment and monitoring of the diversity of photosynthetic eukaryotes using high-throughput sequencing.

PhytoREF: a reference database of the plastidial 16S rRNA gene of photosynthetic eukaryotes with curated taxonomy

Photosynthetic eukaryotes have a critical role as the main producers in most ecosystems of the biosphere. The ongoing environmental metabarcoding revolution opens the perspective for holistic ecosystems biological studies of these organisms, in particular the unicellular microalgae that often lack distinctive morphological characters and have complex life cycles. To interpret environmental sequences, metabarcoding necessarily relies on taxonomically curated databases containing reference sequences of the targeted gene (or barcode) from identified organisms. To date, no such reference framework exists for photosynthetic eukaryotes. In this study, we built the PhytoREF database that contains 6490 plastidial 16S rDNA reference sequences that originate from a large diversity of eukaryotes representing all known major photosynthetic lineages. We compiled 3333 amplicon sequences available from public databases and 879 sequences extracted from plastidial genomes, and generated 411 novel sequences from cultured marine microalgal strains belonging to different eukaryotic lineages. A total of 1867 environmental Sanger 16S rDNA sequences were also included in the database. Stringent quality filtering and a phylogeny-based taxonomic classification were applied for each 16S rDNA sequence. The database mainly focuses on marine microalgae, but sequences from land plants (representing half of the PhytoREF sequences) and freshwater taxa were also included to broaden the applicability of PhytoREF to different aquatic and terrestrial habitats. PhytoREF, accessible via a web interface (http://phytoref.fr), is a new resource in molecular ecology to foster the discovery, assessment and monitoring of the diversity of photosynthetic eukaryotes using high-throughput sequencing.

Are Prorocentrum hoffmannianum and Prorocentrum belizeanum (Dinophyceae, Prorocentrales), the same species? An integration of morphological and molecular data.

The taxonomic assignment of Prorocentrum species is based on morphological characteristics; however, morphological variability has been found for several taxa isolated from different geographical regions. In this study, we evaluated species boundaries of Prorocentrum hoffmannianum and Prorocentrum belizeanum based on morphological and molecular data. A detailed morphological analysis was done, concentrating on the periflagellar architecture. Molecular analyses were performed on partial Small Sub-Unit (SSU) rDNA, partial Large Sub-Unit (LSU) rDNA, complete Internal Transcribed Spacer Regions (ITS1-5.8S-ITS2), and partial cytochrome b (cob) sequences. We concatenated the SSU-ITS-LSU fragments and constructed a phylogenetic tree using Bayesian Inference (BI) and maximum likelihood (ML) methods. Morphological analyses indicated that the main characters, such as cell size and number of depressions per valve, normally used to distinguish P. hoffmannianum from P. belizeanum, overlapped. No clear differences were found in the periflagellar area architecture. Prorocentrum hoffmannianum and P. belizeanum were a highly supported monophyletic clade separated into three subclades, which broadly corresponded to the sample collection regions. Subtle morphological overlaps found in cell shape, size, and ornamentation lead us to conclude that P. hoffmanianum and P. belizeanum might be considered conspecific. The molecular data analyses did not separate P. hoffmannianum and P. belizeanum into two morphospecies, and thus, we considered them to be the P. hoffmannianum species complex because their clades are separated by their geographic origin. These geographic and genetically distinct clades could be referred to as ribotypes: (A) Belize, (B) Florida-Cuba, (C1) India, and (C2) Australia.

Automated analysis of non-mass-enhancing lesions in breast MRI based on morphological, kinetic, and spatio-temporal moments and joint segmentation-motion compensation technique

Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) represents an established method for the detection and diagnosis of breast lesions. While mass-like enhancing lesions can be easily categorized according to the Breast Imaging Reporting and Data System (BI-RADS) MRI lexicon, a majority of diagnostically challenging lesions, the so called non-mass-like enhancing lesions, remain both qualitatively as well as quantitatively difficult to analyze. Thus, the evaluation of kinetic and/or morphological characteristics of non-masses represents a challenging task for an automated analysis and is of crucial importance for advancing current computer-aided diagnosis (CAD) systems. Compared to the well-characterized mass-enhancing lesions, non-masses have no well-defined and blurred tumor borders and a kinetic behavior that is not easily generalizable and thus discriminative for malignant and benign non-masses. To overcome these difficulties and pave the way for novel CAD systems for non-masses, we will evaluate several kinetic and morphological descriptors separately and a novel technique, the Zernike velocity moments, to capture the joint spatio-temporal behavior of these lesions, and additionally consider the impact of non-rigid motion compensation on a correct diagnosis.

Morphological and genetic variation in the north Atlantic copepod, Centropages typicus

This study describes phenotypic and genotypic variations in the planktonic copepod, Centropages typicus (Copepoda: Calanoida) that indicate differentiation between geographical samples. We found consistent differences in the morphology of the chela of the sexually modified fifth pereiopod (P5) of male C. typicus between samples from the Mediterranean, western North Atlantic and eastern North Atlantic. A 560 base pairs (bp) region of the C. typicus mitochondrial cytochrome c oxidase subunit I (COI) and a 462 bp fragment of the nuclear rDNA internal transcribed spacer (ITS) tandem array were analysed to determine whether these morphological variations reflect population genetic differentiation. Mitochondrial haplotype diversity was found to be high with 100 unique COI haplotypes among 116 individuals. Analysis of mtCOI variation suggested differentiation between the Mediterranean and Atlantic populations but no separation was detected within the Atlantic. Intragenomic variation in the ITS array suggested genetic differentiation between samples from the western North Atlantic and those from the eastern North Atlantic and Mediterranean. Breeding experiments would be required to elucidate the extent of genetic isolation between C. typicus from the different population centres.