Species and population genomic differentiation in Pocillopora corals (Cnidaria, Hexacorallia).

Correctly delimiting species and populations is a prerequisite for studies of connectivity, adaptation and conservation. Genomic data are particularly useful to test species differentiation for organisms with few informative morphological characters or low discrimination of cytoplasmic markers, as in Scleractinians. Here we applied Restriction site Associated DNA sequencing (RAD-sequencing) to the study of species differentiation and genetic structure in populations of Pocillopora spp. from Oman and French Polynesia, with the objectives to test species hypotheses, and to study the genetic structure among sampling sites within species. We focused here on coral colonies morphologically similar to P. acuta (damicornis type ß). We tested the impact of different filtering strategies on the stability of the results. The main genetic differentiation was observed between samples from Oman and French Polynesia. These samples corresponded to different previously defined primary species hypotheses (PSH), i.e., PSHs 12 and 13 in Oman, and PSH 5 in French Polynesia. In Oman, we did not observe any clear differentiation between the two putative species PSH 12 and 13, nor between sampling sites. In French Polynesia, where a single species hypothesis was studied, there was no differentiation between sites. Our analyses allowed the identification of clonal lineages in Oman and French Polynesia. The impact of clonality on genetic diversity is discussed in light of individual-based simulations.

From seascape ecology to population genomics and back. Spatial and ecological differentiation among cryptic species of the red algae Lithophyllum stictiforme/L. cabiochiae, main bioconstructors of coralligenous habitats.

Ecosystem engineering species alter the physical structure of their environment and can create or modify habitats, having a massive impact on local biodiversity. Coralligenous reefs are highly diverse habitats endemic to the Mediterranean Sea built by calcareous benthic organisms among which Crustose Coralline Algae are the main engineering species. We analyzed the diversity of Lithophyllum stictiforme or L. cabiochiae in coralligenous habitats combining a multiple barcode and a population genomics approach with seascape features. Population genomics allowed disentangling pure spatial effects from environmental effects. We found that these taxa form a complex of eight highly divergent cryptic species that are easily identifiable using classic barcode markers (psbA, LSU, COI). Three factors have a significant effect on the relative abundances of these cryptic species: the location along the French Mediterranean coast, depth and Photosynthetic Active Radiation (PAR). The analysis of around 5000 SNPs for the most abundant species revealed genetic differentiation among localities in the Bay of Marseille but no differentiation between depths within locality. Thus, the effect of depth and PAR on cryptic species communities is not a consequence of restricted connectivity but rather due to differential settlement or survival among cryptic species. This differential is more likely driven by irradiance levels rather than by pressure or temperature. Both the genetic and species diversity patterns are congruent with the main patterns of currents in the Bay. Ecological differentiation among these engineering cryptic species, sensitive to ocean warming and acidification, could have important consequences on the diversity and structure of the coralligenous communities.

Ship-driven biopollution: How aliens transform the local ecosystem diversity in Pacific islands.

Ships moving species across the oceans mix marine communities throughout latitudes. The introduction of new species may be changing the ecosystems even in remote islands. In tropical Pacific islands where maritime traffic is principally local, eDNA metabarcoding and barcoding revealed 75 introduced species, accounting in average for 28% of the community with a minimum of 13% in the very remote Rangiroa atoll. The majority of non-native species were primary producers -from diatoms to red algae, thus the ecosystem is being transformed from the bottom. Primary producers were more shared among sites than other exotics, confirming ship-mediated dispersal in Pacific marine ecosystems. Limited alien share and an apparent saturation of aliens (similar proportion in ports of very different size) suggests the occurrence of "alien drift" in port communities, or random retention of newly introduced aliens that reminds genetic drift of new mutations in a population.

Nonfatal Bites by a Sicklefin Lemon Shark Negaprion acutidens on a Surfer in Makemo Atoll (French Polynesia).

Identifying the species and size of sharks responsible for biting humans is essential for developing strategies to prevent these incidents. Here, we use bite wound characteristics and genetic analysis of a tooth fragment extracted from the wounds to identify a sicklefin lemon shark Negaprion acutidens as the perpetrator of nonfatal bites on the legs of an adult male surfer at Makemo atoll (French Polynesia) in January 2018. The bite was superficial, and N. acutidens are fish predators not known to feed on large prey; hence, foraging is an unlikely explanation for this incident rather linked to territoriality. Lemon sharks are occasionally aggressive toward humans and are site attached with relatively small home ranges; hence, avoiding surfing in the area of a previous bite incident is recommended to decrease the risk of future injuries.

Response of Downy Oak (Quercus pubescens Willd.) to Climate Change: Transcriptome Assembly, Differential Gene Analysis and Targeted Metabolomics.

Global change scenarios in the Mediterranean basin predict a precipitation reduction within the coming hundred years. Therefore, increased drought will affect forests both in terms of adaptive ecology and ecosystemic services. However, how vegetation might adapt to drought is poorly understood. In this report, four years of climate change was simulated by excluding 35% of precipitation above a downy oak forest. RNASeq data allowed us to assemble a genome-guided transcriptome. This led to the identification of differentially expressed features, which was supported by the characterization of target metabolites using a metabolomics approach. We provided 2.5 Tb of RNASeq data and the assembly of the first genome guided transcriptome of Quercus pubescens. Up to 5724 differentially expressed transcripts were obtained; 42 involved in plant response to drought. Transcript set enrichment analysis showed that drought induces an increase in oxidative pressure that is mitigated by the upregulation of ubiquitin-like protein protease, ferrochelatase, oxaloacetate decarboxylase and oxo-acid-lyase activities. Furthermore, the downregulation of auxin biosynthesis and transport, carbohydrate storage metabolism were observed as well as the concomitant accumulation of metabolites, such as oxalic acid, malate and isocitrate. Our data suggest that early metabolic changes in the resistance of Q. pubescens to drought involve a tricarboxylic acid (TCA) cycle shunt through the glyoxylate pathway, galactose metabolism by reducing carbohydrate storage and increased proteolytic activity.

Evidence for stasis and not genetic piracy in developmental expression patterns of Branchiostoma lanceolatum and Branchiostoma floridae, two amphioxus species that have evolved independently over the course of 200 Myr.

Cephalochordates, the most basal extant group in the phylum Chordata, are represented chiefly by about 20 species of the genus Branchiostoma, commonly called amphioxus or lancelets. In recent years, insights into the evolutionary origin of the vertebrates have been gained from molecular genetic studies during the development of three of these amphioxus species (Branchiostoma floridae in North America, Branchiostoma lanceolatum in Europe, and Branchiostoma belcheri in East Asia). In spite of an estimated divergence time of 100-200 Myr among these species, all three are remarkably similar morphologically, and students of amphioxus have tacitly assumed that such resemblances arise during ontogeny from nearly identical networks of developmental genes. We felt that this assumption needed to be reexamined because instances are known--even in comparisons of closely related species--where characters seeming homologous on the basis of morphology actually develop under the control of conspicuously divergent genetic programs (a phenomenon termed "genetic piracy"). In the present work, we tested the hypothesis that morphological similarities reflect strict conservation of developmentally important genes' expression patterns in order to assess whether the developmental genetics of different amphioxus species show evidence of genetic piracy. To these ends, we cloned 18 genes implicated in different developmental functions in B. lanceolatum and compared their gene expression patterns with the known expression patterns of their orthologous genes in B. floridae. We show that, for the most part, conservation of gene expression parallels that of morphology in these two species. We also identified some differences in gene expression, likely reflecting experimental sensitivity, with the exception of Pax1/9, which may result from true developmental specificities in each amphioxus species. Our results demonstrate that morphological conservation reflects stasis in developmental gene expression patterns and find no evidence for genetic piracy. Thus, different species of amphioxus appear to be very similar, not only morphologically, but also in the genetic programs directing the development of their structural features. Moreover, we provide the first catalogue of gene expression data for the European species, B. lanceolatum.

Advances in genotyping microsatellite markers through sequencing and consequences of scoring methods for Ceratonia siliqua (Leguminosae).

PREMISE OF THE STUDY: Simple sequence repeat (SSR) or microsatellite markers have been used in a broad range of studies mostly scoring alleles on the basis of amplicon size as a proxy for the number of repeat units of an SSR motif. However, additional sources of variation within the SSR or in the flanking regions have largely remained undetected. METHODS: In this study, we implemented a next-generation sequencing-based genotyping approach in a newly characterized set of 18 nuclear SSR markers for the carob tree, Ceratonia siliqua. Our aim was to evaluate the effect of three different methods of scoring molecular variation present within microsatellite markers on the genetic diversity and structure results. RESULTS: The analysis of the sequences of 77 multilocus genotypes from four populations revealed SSR variation and additional sources of polymorphism in 87% of the loci analyzed (42 single-nucleotide polymorphisms and five insertion/deletion polymorphisms), as well as divergent paralog copies in two loci. Ignoring sequence variation under standard amplicon size genotyping resulted in incorrect identification of 69% of the alleles, with important effects on the genetic diversity and structure estimates. DISCUSSION: Next-generation sequencing allows the detection and scoring of SSRs, single-nucleotide polymorphisms, and insertion/deletion polymorphisms to increase the resolution of population genetic studies.

From depth to regional spatial genetic differentiation of Eunicella cavolini in the NW Mediterranean.

Connectivity studies in the marine realm are of great importance to understand the evolutionary potential of populations in a context of growing pressures on the marine environment. Here, we investigated the effect of the local, regional, and depth spatial scale on the population genetic structure of the yellow gorgonian Eunicella cavolini, one of the most common octocoral species of the Mediterranean hard-bottom communities. This species, along with other sessile metazoans typical of coralligenous ecosystems, plays an important role in supporting biodiversity, but is also impacted by direct and indirect consequences of human activities, such as physical destruction or mortality events due to thermal anomalies. Samples were taken from 15 sites located in the Ligurian Sea (NW Mediterranean) in two adjacent regions 100 kilometres apart, i.e. from the areas of Marseille (France) and Portofino (Genoa, Italy), and were analysed using six microsatellite loci. A pattern of isolation by distance was observed at the regional as well as the local scales. Although E. cavolini showed less genetic structure than other Mediterranean octocorallian species, we observed a significant genetic differentiation between populations a few kilometres apart. A low genetic differentiation was also observed between shallow and deep populations. The occurrence of genetically differentiated populations of E. cavolini at the scale of kilometres has important consequences for the management of this species and of the associated communities.

The Conservation of the Germline Multipotency Program, from Sponges to Vertebrates: A Stepping Stone to Understanding the Somatic and Germline Origins.

The germline definition in metazoans was first based on few bilaterian models. As a result, gene function interpretations were often based on phenotypes observed in those models and led to the definition of a set of genes, considered as specific of the germline, named the "germline core". However, some of these genes were shown to also be involved in somatic stem cells, thus leading to the notion of germline multipotency program (GMP). Because Porifera and Ctenophora are currently the best candidates to be the sister-group to all other animals, the comparative analysis of gene contents and functions between these phyla, Cnidaria and Bilateria is expected to provide clues on early animal evolution and on the links between somatic and germ lineages. Our present bioinformatic analyses at the metazoan scale show that a set of 18 GMP genes was already present in the last common ancestor of metazoans and indicate more precisely the evolution of some of them in the animal lineage. The expression patterns and levels of 11 of these genes in the homoscleromorph sponge Oscarella lobularis show that they are expressed throughout their life cycle, in pluri/multipotent progenitors, during gametogenesis, embryogenesis and during wound healing. This new study in a nonbilaterian species reinforces the hypothesis of an ancestral multipotency program.

A new renal mitochondrial carrier, KMCP1, is up-regulated during tubular cell regeneration and induction of antioxidant enzymes.

The mitochondrial carrier family transports a variety of metabolites across the inner mitochondrial membrane. We identified and cloned a new member of this family, KMCP1 (kidney mitochondrial carrier protein-1), that is highly homologous to the previously identified protein BMCP1 (brain mitochondrial carrier protein-1). Western blotting and in situ experiments showed that this carrier is expressed predominantly within the kidney cortex in the proximal and distal tubules. KMCP1 was increased during fasting and during the regenerative phase of glycerol-induced renal failure. We show that both situations are associated with transiently increased expression of superoxide-generating enzymes, followed by increased mitochondrial metabolism and antioxidant defenses. Given that KMCP1 expression occurs simultaneously with these latter events, we propose that KMCP1 is involved in situations in which mitochondrial metabolism is increased, in particular when the cellular redox balance tends toward a pro-oxidant status.

Resistance to cerebral ischemic injury in UCP2 knockout mice: evidence for a role of UCP2 as a regulator of mitochondrial glutathione levels.

Uncoupling protein 2 (UCP2) is suggested to be a regulator of reactive oxygen species production in mitochondria. We performed a detailed study of brain injury, including regional and cellular distribution of UCP2 mRNA, as well as measures of oxidative stress markers following permanent middle cerebral artery occlusion in UCP2 knockout (KO) and wild-type (WT) mice. Three days post ischemia, there was a massive induction of UCP2 mRNA confined to microglia in the peri-infarct area of WT mice. KO mice were less sensitive to ischemia as assessed by reduced brain infarct size, decreased densities of deoxyuridine triphosphate nick end-labelling (TUNEL)-labelled cells in the peri-infact area and lower levels of lipid peroxidation compared with WT mice. This resistance may be related to the substantial increase of basal manganese superoxide dismutase levels in neurons of KO mice. Importantly, we found a specific decrease of mitochondrial glutathione (GSH) levels in UCP2 expressing microglia of WT, but not in KO mice after ischemia. This specific association between UCP2 and mitochondrial GSH levels regulation was further confirmed using lipopolysaccharide models of peripheral inflammation, and in purified peritoneal macrophages. Moreover, our data imply that UCP2 is not directly involved in the regulation of ROS production but acts by regulating mitochondrial GSH levels in microglia.