Contrasting biogeographical patterns in Margarella (Gastropoda: Calliostomatidae: Margarellinae) across the Antarctic Polar Front.

Members of the trochoidean genus Margarella (Calliostomatidae) are broadly distributed across Antarctic and sub-Antarctic ecosystems. Here we used novel mitochondrial and nuclear gene sequences to clarify species boundaries and phylogenetic relationships among seven nominal species distributed on either side of the Antarctic Polar Front (APF). Molecular reconstructions and species-delimitation analyses recognized only four species: M. antarctica (the Antarctic Peninsula), M. achilles (endemic to South Georgia), M. steineni (South Georgia and Crozet Island) and the morphologically variable M. violacea (=M. expansa, M. porcellana and M. pruinosa), with populations in southern South America, the Falkland/Malvinas, Crozet and Kerguelen Islands. Margarella violacea and M. achilles are sister species, closely related to M. steineni, with M. antarctica sister to all these. This taxonomy reflects contrasting biogeographic patterns on either side of the APF in the Southern Ocean. Populations of Margarella north of the APF (M. violacea) showed significant genetic variation but with many shared haplotypes between geographically distant populations. By contrast, populations south of the APF (M. antarctica, M. steineni and M. achilles) exhibited fewer haplotypes and comprised three distinct species, each occurring across a separate geographical range. We hypothesize that the biogeographical differences may be the consequence of the presence north of the APF of buoyant kelps - potential long-distance dispersal vectors for these vetigastropods with benthic-protected development - and their near-absence to the south. Finally, we suggest that the low levels of genetic diversity within higher-latitude Margarella reflect the impact of Quaternary glacial cycles that exterminated local populations during their maxima.

Application of the ecosystem service concept at a small-scale: The cases of coralligenous habitats in the North-western Mediterranean Sea.

The understanding of ecosystem services is essential to support sustainable use and preservation of ecosystems. Coralligenous habitats, main contributors of the Mediterranean marine biodiversity, are yet understudied in term of services provided. This study presents an original small-scale approach to investigate the services provided by coralligenous habitats of a French study area consisting of two marine sites (Marseille and Port-Cros sites) in order to cover two contrasted anthropogenic pressure despite the small-scale. Our results are based on the opinions of 43 experts who ranked 15 services in terms of existence and level of importance for human well-being: supporting ecological functions were considered the most important, then provisioning and cultural services. Regulating services were considered uncertain due to a lack of knowledge. The small-scale approach highlighted a need for a referential frame to determine the existence of services (e.g. geographical and temporal scales, benefits and beneficiaries levels).

Regional and local environmental conditions do not shape the response to warming of a marine habitat-forming species.

The differential response of marine populations to climate change remains poorly understood. Here, we combine common garden thermotolerance experiments in aquaria and population genetics to disentangle the factors driving the population response to thermal stress in a temperate habitat-forming species: the octocoral Paramuricea clavata. Using eight populations separated from tens of meters to hundreds of kilometers, which were differentially impacted by recent mortality events, we identify 25 °C as a critical thermal threshold. After one week of exposure at this temperature, seven of the eight populations were affected by tissue necrosis and after 30 days of exposure at this temperature, the mean % of affected colonies increased gradually from 3 to 97%. We then demonstrate the weak relation between the observed differential phenotypic responses and the local temperature regimes experienced by each population. A significant correlation was observed between these responses and the extent of genetic drift impacting each population. Local adaptation may thus be hindered by genetic drift, which seems to be the main driver of the differential response. Accordingly, conservation measures should promote connectivity and control density erosion in order to limit the impact of genetic drift on marine populations facing climate change.

Morphological and genetic analyses reveal a cryptic species complex in the echinoid Echinocardium cordatum and rule out a stabilizing selection explanation.

Preliminary analyses revealed the presence of at least five mitochondrial clades within the widespread sea urchin Echinocardium cordatum (Spatangoida). In this study, we analyzed the genetic (two mitochondrial and two nuclear sequence loci) and morphological characteristics (20 indices) from worldwide samples of this taxon to establish the species limits, morphological diversity and differentiation. Co-occurring spatangoid species were also analyzed with mitochondrial DNA. The nuclear sequences confirm that mitochondrial lineages correspond to true genetic entities and reveal that two clades (named A and B1) hybridize in their sympatry area, although a more closely related pair of clades (B1 and B2), whose distributions widely overlap, does not display hybridization. The morphology of all E. cordatum clade pairs was significantly differentiated, but no morphological diagnostic character was evidenced. By contrast, other spatangoid species pairs that diverged more recently than the E. cordatum clades display clear diagnostic characters. Morphological diversity thus appears responsible for the absence of diagnostic characters, ruling out stabilizing selection, a classical explanation for cryptic species. Alternative classical explanations are (i) environmental plasticity or (ii) a high diversity of genes determining morphology, maintained by varying environmental conditions. We suggest a new hypothesis that the observed morphological diversity is selectively neutral and reflects high effective population sizes in the E. cordatum complex. It is supported by the higher abundance of this taxon compared with other taxa, a trend for the genetic and morphological diversity to be correlated in Europe, and the higher genetic and morphological diversities found in clades of E cordatum (except B1) than in other spatangoid samples in Europe. However, the Pacific clades do not confirm these trends.

PCR survey of 50 introns in animals: cross-amplification of homologous EPIC loci in eight non-bilaterian, protostome and deuterostome phyla.

Exon Primed Intron Crossing (EPIC) markers provide molecular tools that are susceptible to be variable within species while remaining amplifiable by PCR using potentially universal primers. In this study we tested the possibility of obtaining PCR products from 50 EPIC markers on 23 species belonging to seven different phyla (Porifera, Cnidaria, Arthropoda, Nematoda, Mollusca, Annelida, Echinodermata) using 70 new primer pairs. A previous study had identified and tested those loci in a dozen species, including another phylum, Urochordata (Chenuil et al., 2010). Results were contrasted among species. The best results were achieved with the oyster (Mollusca) where 28 loci provided amplicons susceptible to contain an intron according to their size. This was however not the case with the other mollusk Crepidula fornicata, which seems to have undergone a reduction in intron number or intron size. In the Porifera, 13 loci appeared susceptible to contain an intron, a surprisingly high number for this phylum considering its phylogenetic distance with genomic data used to design the primers. For two cnidarian species, numerous loci (24) were obtained. Ecdysozoan phyla (arthropods and nematodes) proved less successful than others as expected considering reports of their rapid rate of genome evolution and the worst results were obtained for several arthropods. Some general patterns among phyla arose, and we discuss how the results of this EPIC survey may give new insights into genome evolution of the study species. This work confirms that this set of EPIC loci provides an easy-to-use toolbox to identify genetic markers potentially useful for population genetics, phylogeography or phylogenetic studies for a large panel of metazoan species. We then argue that obtaining diploid sequence genotypes for these loci became simple and affordable owing to Next-Generation Sequencing development. Species surveyed in this study belong to several genera (Acanthaster, Alvinocaris, Aplysina, Aurelia, Crepidula, Eunicella, Hediste, Hemimysis, Litoditis, Lophelia, Mesopodopsis, Mya, Ophiocten, Ophioderma, Ostrea, Pelagia, Platynereis, Rhizostoma, Rimicaris), two of them, belonging to the family Vesicomydae and Eunicidae, could not be determined at the genus level.

Phylogeography of the red coral (Corallium rubrum): inferences on the evolutionary history of a temperate gorgonian.

The red coral Corallium rubrum (Cnidaria, Octocorallia) is an exploited, long-lived sessile species from the Mediterranean Sea and the adjacent coastline in the Atlantic Ocean. Surveys of genetic variation using microsatellites have shown that populations of C. rubrum are characterized by strong differentiation at the local scale but a study of the phylogeography of this species was still lacking. Here, we used seven polymorphic microsatellite loci, together with sequence data from an intron of the elongation factor 1 (EF1) gene, to investigate the genetic structure of C. rubrum across its geographical range in the western Mediterranean Sea and in the Adriatic Sea. The EF1 sequences were also used to analyse the consequences of demographic fluctuations linked with past environmental change. Clustering analysis with microsatellite loci highlighted three to seven genetic groups with the distinction of North African and Adriatic populations; this distinction appeared significant with AMOVA and differentiation tests. Microsatellite and EF1 data extended the isolation by distance pattern previously observed for this species at the western Mediterranean scale. EF1 sequences confirmed the genetic differentiation observed between most samples with microsatellites. A statistical parsimony network of EF1 haplotypes provided no evidence of high sequence divergence among regions, suggesting no long-term isolation. Selective neutrality tests on microsatellites and EF1 were not significant but should be interpreted with caution in the case of EF1 because of the low sample sizes for this locus. Our results suggest that recent Quaternary environmental fluctuations had a limited impact on the genetic structure of C. rubrum.

From global to local genetic structuring in the red gorgonian Paramuricea clavata: the interplay between oceanographic conditions and limited larval dispersal.

Defining the scale of connectivity among marine populations and identifying the barriers to gene flow are tasks of fundamental importance for understanding the genetic structure of populations and for the design of marine reserves. Here, we investigated the population genetic structure at three spatial scales of the red gorgonian Paramuricea clavata (Cnidaria, Octocorallia), a key species dwelling in the coralligenous assemblages of the Mediterranean Sea. Colonies of P. clavata were collected from 39 locations across the Mediterranean Sea from Morocco to Turkey and analysed using microsatellite loci. Within three regions (Medes, Marseille and North Corsica), sampling was obtained from multiple locations and at different depths. Three different approaches (measures of genetic differentiation, Bayesian clustering and spatially explicit maximum-difference algorithm) were used to determine the pattern of genetic structure. We identified genetic breaks in the spatial distribution of genetic diversity, which were concordant with oceanographic conditions in the Mediterranean Sea. We revealed a high level of genetic differentiation among populations and a pattern of isolation by distance across the studied area and within the three regions, underlining short effective larval dispersal in this species. We observed genetic differentiation among populations in the same locality dwelling at different depths, which may be explained by local oceanographic conditions and which may allow a process of local adaptation of the populations to their environment. We discuss the implications of our results for the conservation of the species, which is exposed to various threats.

Fine-scale genetic structure and inferences on population biology in the threatened Mediterranean red coral, Corallium rubrum.

Identifying microevolutionary processes acting in populations of marine species with larval dispersal is a challenging but crucial task because of its conservation implications. In this context, recent improvements in the study of spatial genetic structure (SGS) are particularly promising because they allow accurate insights into the demographic and evolutionary processes at stake. Using an exhaustive sampling and a combination of image processing and population genetics, we highlighted significant SGS between colonies of Corallium rubrum over an area of half a square metre, which sheds light on a number of aspects of its population biology. Based on this SGS, we found the mean dispersal range within sites to be between 22.6 and 32.1 cm, suggesting that the surveyed area approximately corresponded to a breeding unit. We then conducted a kinship analysis, which revealed a complex half-sib family structure and allowed us to quantify the level of self-recruitment and to characterize aspects of the mating system of this species. Furthermore, significant temporal variations in allele frequencies were observed, suggesting low genetic drift. These results have important conservation implications for the red coral and further our understanding of the microevolutionary processes acting within populations of sessile marine species with a larval phase.

Genetic survey of shallow populations of the Mediterranean red coral [Corallium rubrum (Linnaeus, 1758)]: new insights into evolutionary processes shaping nuclear diversity and implications for conservation.

Combined action from over-harvesting and recent mass mortality events potentially linked to ongoing climate changes has led to new concerns for the conservation of shallow populations (5-60 m) of Corallium rubrum, an octocorallian that is mainly found in the Mediterranean Sea. The present study was designed to analyse population structure and relationships at different spatial scales (from 10s of meters to 100s of kilometres) with a focus on dispersal pattern. We also performed the first analysis of the distribution of genetic diversity using a comparative approach between regional-clusters and samples. Forty populations dwelling in four distinct regions between 14 and 60 m in depth were genotyped using 10 microsatellites. Our main results indicate (i) a generalized pair-sample differentiation combined with a weak structure between regional-clusters; (ii) the occurrence of isolation by distance at the global scale, but also within two of the three analysed regional-clusters; (iii) a high level of genetic diversity over the surveyed area with a heterogeneous distribution from regional-cluster to sample levels. The evolutionary consequences of these results are discussed and their management implications are provided.

Defining reproductively isolated units in a cryptic and syntopic species complex using mitochondrial and nuclear markers: the brooding brittle star, Amphipholis squamata (Ophiuroidea).

At a time when biodiversity is threatened, we are still discovering new species, and particularly in the marine realm. Delimiting species boundaries is the first step to get a precise idea of diversity. For sympatric species which are morphologically undistinguishable, using a combination of independent molecular markers is a necessary step to define separate species. Amphipholis squamata, a cosmopolitan brittle star, includes several very divergent mitochondrial lineages. These lineages appear totally intermixed in the field and studies on morphology and colour polymorphism failed to find any diagnostic character. Therefore, these mitochondrial lineages may be totally interbreeding presently. To test this hypothesis, we characterized the genetic structure of the complex in the French Mediterranean coast using sequences of mitochondrial DNA (16S) and for the first time, several nuclear DNA markers (introns and microsatellites). The data revealed six phylogenetic lineages corresponding to at least four biological species. These sibling species seem to live in syntopy. However, they seem to display contrasted levels of genetic diversity, suggesting they have distinct demographic histories and/or life-history traits. Genetic differentiation and isolation-by-distance within the French Mediterranean coasts are revealed in three lineages, as expected for a species without a free larval phase. Finally, although recombinant nuclear genotypes are common within mitochondrial lineages, the data set displays a total lack of heterozygotes, suggesting a very high selfing rate, a feature likely to have favoured the formation of the species complex.

Combined ecological factors permit classification of developmental patterns in benthic marine invertebrates: a discussion note.

Traditional classifications of developmental patterns of marine benthic invertebrates are based on combinations of embryological (direct or indirect development) and ecological (such as nutritional source or habitat) characteristics. Different schemes have been proposed for different reasons, relating to ecology, evolution and/or development. However, these classifications contain interconnected characters that do not efficiently discriminate between developmental patterns and, thus, do not fully apply to either ecological or embryological studies. An ecological multifactor classification based on three independent two-state characters (pelagic/benthic, free/protected, and feeding/non-feeding) is proposed. It discriminates between eight developmental patterns and can encompass any ecological pattern of development among marine benthic invertebrates.

A presumptive developmental role for a sea urchin cyclin B splice variant.

We show that a splice variant-derived cyclin B is produced in sea urchin oocytes and embryos. This splice variant protein lacks highly conserved sequences in the COOH terminus of the protein. It is found strikingly abundant in growing oocytes and cells committed to differentiation during embryogenesis. Cyclin B splice variant (CBsv) protein associates weakly in the cell with Xenopus cdc2 and with budding yeast CDC28p. In contrast to classical cyclin B, CBsv very poorly complements a triple CLN deletion in budding yeast, and its microinjection prevents an initial step in MPF activation, leading to an important delay in oocyte meiosis reinitiation. CBsv microinjection in fertilized eggs induces cell cycle delay and abnormal development. We assume that CBsv is produced in growing oocytes to keep them in prophase, and during embryogenesis to slow down cell cycle in cells that will be committed to differentiation.

Completely Direct Development of Abatus cordatus, a Brooding Schizasterid (Echinodermata: Echinoidea) from Kerguelen, With Description of Perigastrulation, a Hypothetical New Mode of Gastrulation.

Development of the brooding schizasterid Abatus cordatus, a subantarctic echinoid endemic to Kerguelen, is described. Females spawn nonbuoyant eggs 1300 µm in diameter, which are fertilized by elongated sperm (head 1 µm wide and 15 µm long). The main characteristics of this development are (1) incomplete cleavage beginning at the animal pole that becomes holoblastic, giving a filled wrinkled blastula 26 days after fertilization; (2) apparent (fate-mapping studies have not been done) external migration of mesenchyme cells, in the perivitelline space, from the animal to the vegetal pole during gastrulation while the archenteron invaginates; (3) hatching occurring at the end of the gastrulation (65 days after fertilization); (4) differentiation of the vestibule from a thickening of the oral epidermis as soon as the end of gastrulation is attained; and (5) production of a juvenile directly from the gastrula without any larval stage. The juvenile that leaves the brood chamber is 2 mm in diameter and about 250 days old. A. cordatus is a true completely direct developer (no larva and no metamorphosis). We propose to use (1) the term perigastrulation, as a tentative one until more definitive studies are available, to describe the hypothetical peculiar movement of cells during gastrulation and (2) the terms of direct development only for completely direct developing species and abbreviated development for species that have more or less transformed plutei.

The effect of somatic and gonadal size on the rate of oxygen consumption in the subantarctic echinoid Abatus cordatus (Echinodermata) from Kerguelen.

1. The rate of oxygen consumption of this burrowing spatangoid was measured for individuals ranging in size from 3-month old 2.5-mm long juveniles to 39-mm long adults. 2. The decrease in the rate of oxygen consumption/dry weight with increasing body size is greater among mature adults than among juveniles because the increase in aerobic tissue (primarily the test) with body size is less than the increase in anaerobic tissues (mainly the gonads). 3. The rate of oxygen consumption/ash-free dry weight decreased more slowly with increasing body size because of the increase in the level of inorganic material. 4. Replacement of the common fresh weight or dry weight specific oxygen consumption by a more synthetic value calculated from ash-free dry weight specific oxygen consumption measurements, to annulate the body-size effect, is proposed for interspecific comparison over a wide range of body size, taking into account parameters such as temperature.