ABSTRACT. The very common and small sabellid polychaete Amphiglena mediterranea (Leyidg, 1854) is known to be the only species of this genus reported in the whole Mediterranean Sea. However, preliminary genetic analyses within the Gulf of Naples, highlighted the occurrence of at least 5 putative sibling species. Here we report the description of 9 new taxa within the genus Amphiglena, whose diversity in the Mediterranean basin has been widely underestimated. Examined material derived from both from new collections along the Italian coast, including four CO2 vents/hydrothermal systems, and from a re-examination of older material previously attributed to A. mediterranea. The morphological analyses revealed the presence of different taxa also consistently with a previous molecular analyses conducted on material from the Gulf of Naples. This led to an increase in the number of species in the genus and to enhance the existence in the Mediterranean of a high diversity within the genus, showing also species found in quite unique environmental conditions, such as hydrothermal vent systems, where 5 of the 9 species have been described. Some taxa, however, remained for the moment undescribed due to the poor preservation of the old material, and the lack of the type material for this taxon. A larger revision of all the previous Mediterranean material previously attributed to A. mediterranea from both morphological and molecular point of views is needed.

ABSTRACT. Aim Taxonomic studies of marine biota can give uncertain results when they are based on sparse morphological differences and molecular delimitation alone without considering biogeographical patterns of the studied region. Recent revisions of the nudibranch family Fionidae s.l. (Gastropoda: Nudibranchia) led to the erection of a number of new taxa on species, generic and family levels. However, in several cases the boundaries between these taxa remain unclear. We aimed to update molecular and morphological data on Zelentia pustulata species complex in Atlantic and Arctic to clarify its identity and provide a new prospect for further revision of this group.

Methods The molecular study included five markers (COI, 16S, H3, ITS1 and ITS2) and implemented Bayesian and Maximum likelihood phylogenetic approaches. Species delimitation and intraspecific divergence were investigated with ABGD, PTP and GMYC methods, and TCS-based haplotype analysis. Ancestral area reconstruction and divergence time estimation were implemented to uncover evolutionary history of this species. Morphological analysis included standard anatomical techniques and SEM.

Results Our integrative analysis and phylogeographic approach showed that Zelentia pustulata and Z. roginskae represent a single species, which is highly heterogeneous at both morphological and molecular levels. Divergence times estimation suggests a complex evolutionary history for this species, following Pleistocene glacial conditions in Arctic and Atlantic region that is reflected by the mtDNA divergence.

Main conclusion We propose to synonymize Zelentia pustulata and Z. roginskae. Forthcoming Arctic biodiversity studies should consider the necessity to account for complex evolutionary history and phylogeographical patterns in delimitation of taxa inhabiting regions undergoing rapid environmental changes.

ABSTRACT. Aim The nudibranch group Flabellinidae s.l. (Gastropoda: Nudibranchia) represents a highly diverse and widely distributed group of marine gastropods. Several independent flabellinid lineages occur in boreal regions with only a few representatives in tropical areas. In addition, most species show overlapping ranges and some of them occur sympatrically in very restricted areas. The main goal of this study was to conduct an updated revision of the family Coryphellidae (an independent lineage within Flabellinidae s.l.) using integrative approach.

Methods The molecular study included five markers (COI, 16S, H3, 28S and 18S) and implemented Bayesian and Maximum likelihood phylogenetic approaches. Species delimitation and intraspecific divergence were investigated with ABGD, PTP and GMYC methods, and TCS-based haplotype analysis. Morphological analysis included standard anatomical techniques and scanning electron microscopy. Ancestral area reconstruction and divergence time estimation were implemented to uncover evolutionary history of this group.

Results Based on integrative analysis, we propose a new classification system for the family Coryphellidae and suggest criteria for further revision of flabellinid taxa. Ancestral area reconstruction (AAR) provides evidence for a tropical Pacific origin of the genus. We conclude that the evolution of Coryphella has been shaped by different processes: initial migration out of the tropics, radiation in the North Pacific influenced by Miocene climate change, and subsequent allopatric speciation resulting from successive closings of the Bering strait and cooling of the Arctic Ocean during the Pliocene-Pleistocene.

Main conclusion This study illustrates a complex evolutionary history, in which multiple, independent processes during different periods of time, produce an unusual biogeographic pattern. This model should be tested in other groups of marine invertebrates with similar diversity patterns to better understand the mechanism of generation of biological diversity in non-tropical regions. This study was supported by Russian Science Foundation grant #20-74-10012.

ABSTRACT. Correct species identification and description are fundamental to understand health status of marine ecosystems. However, the use of a single identification tool for species distinction can lead to species misidentifications, having major consequences on ecological studies. Here, we used an integrative taxonomic approach to identify benthic decapods belonging to the genera Galathea (Fabricius, 1793) and Eualus (Thallwitz, 1891) collected in the Mediterranean Sea. 23 Galathea and 22 Eualus individuals were morphologically analysed and sequenced at the mitochondrial COI gene to confirm their identity using BOLD Identification Engine. Morphological identification revealed the presence of two Galathea and three Eualus species, while species delimitation based on DNA barcoding of COI sequences revealed the presence of three Galathea and four Eualus species. Molecular analyses suggested the potential presence of two still undescribed species: one cryptic to Galathea squamifera and one cryptic to Galathea intermedia. Contrasting results obtained by morphological identification and BOLD Identification Engine impeded the recognition of Eualus specimens and suggested misidentifications among BOLD reference records of Eualus cranchii, Eualus occultus and Eualus pusiolus. These results demonstrated that morphological identification overlooks cryptic species and that misidentifications often occur, highlighting the importance of using an integrative approach to increase the current taxonomic knowledge of benthic invertebrates.

ABSTRACT. Aim During last decades, the integrative taxonomy became a most popular approach to define and describe taxa on different taxonomic levels. However, it is often hard to implement it in taxa with sparse or extremely variable morphological characters. Sponges of the genus Leucosolenia (Calcarea, Calcaronea) encounter more than 40 species distributed worldwide. The systematics history of this group is complicated with different species conceptions proposed by various researches. The main goal of our study is to revise the diversity and taxonomy of the genus Leucosolenia from the White Sea using integrative approach. The additional aims of the study are identifying phylogenetically significant morphological characters and proposing optimal sets of molecular markers for further researches of calcareous sponges. Methods Molecular sampling included more than 250 individuals from the White Sea (Russia), Netherlands and Brittany (France). Molecular study included 18S, 28S and H3 markers and implemented Bayesian and Maximum likelihood phylogenetic approaches. Species delimitation and intraspecific divergence were investigated with ABGD and TCS-based analysis. Morphological analysis included studies of external morphology, general skeletal morphology, spicule morphology and cytology using light and electron microscopy. Results Integrative analysis showed that diversity of the genus Leucosolenia in the White Sea is represented by four species: L. variabilis, L. corallorrhiza and two species new to science. Leucosolenia complicata is restricted to NE Atlantic. Examination of morphological characters revealed a strong correlation between morphology of studied Leucosolenia species and their position on the phylogenetic tree. Species differ in general skeletal organization, spicule set and cellular composition. Main conclusion The integrative taxonomical approach, which included molecular studies with three markers and detailed morphological studies, made it possible to clarify the hidden diversity the genus Leucosolenia in White Sea. The study was supported by RSF n°17-14-01089 (ultrastructural studies), RFBR nos°19-04-00563 and 19-04-00545.

ABSTRACT. Abstract Aim

Prokaryotes play a key role in the coastal environment, regulating numerous biogeochemical processes relevant to ecosystem health. Along coastal regions, flowing rivers represent the first vector of transport of sediments from the land. With them, huge quantities of substances like nutrients, organic matter, and chemical compounds are transferred to the marine ecosystem. Transported nutrients and compounds spread along the marine coastal regions, where under different depositional systems they interact with benthic microbial communities determining their ultimate fate.

Methods To assess the influence of different depositional environments and trophic sources on prokaryotes diversity and distribution, we investigated the variability of benthic prokaryotic communities in the surface sediments of 80 stations of the Middle and Southern Adriatic Sea, along the Italian coast.

Results Investigated organic matter showed typical values of eutrophic environments. This was also confirmed from the prokaryotic diversity found, suggesting a strong role of the prokaryotes in organic matter cycling. To understand which factors affected the prokaryotic community structure, the large-scale microbial diversity dataset was correlated to different environmental data. Since there was a lack of linear relationships between variables, different models were used. A non-linear combination of sediments geochemistry, grain size distribution, and station-coastline distance was identified through General Additive Models analysis as the major factors explaining the variation in the prokaryotic distribution. These variables were associated with Adriatic sediment deposits coming from Alpine and Apennines river inputs, which suggest a strong influence in controlling the characteristics of the sediments, and the distribution of trophic resources and prokaryotic abundances.

Main conclusion These observations suggest a strong role of the sedimentary depositional regime in controlling the benthic microbial community structure, potentially influencing the functions performed at the sediment-water interface for the flux of nutrients to the entire benthic community.

ABSTRACT. Aim In this study we assess the performance of the nuclear ITS2 rRNA marker for species identification and species delimitation in eolid nudibranchs. We also characterized the ITS2 rRNA secondary structure of each taxa that can provide additional taxonomic resolution.

Methods ITS2 rRNA multiple sequence alignments were performed simultaneously considering the secondary structure of each sequence. We compared ITS2 pairwise genetic distance (p-distance and distance corrected by the Kimura two parameters model) between 14 species representing 10 genera within six eolid families with distances calculated with the mitochondrial markers COI and 16S rRNA. We infer the ITS2 secondary structures by contrasting several candidate low free energy folding models and we assess the presence of CBCs (Compensatory Base Changes) and semiCBCs across the ITS2 dataset.

Results Based on the ITS2 pairwise distance it is possible to discriminate species within the same genera. The common derived ITS2 rRNA secondary structure of the eolid nudibranchs is organized in five domains, D1-D5, and in the D5 we identified the molluscan consensus Apical STEM. We also identified CBCs and semiCBCs that are diagnostic at both the species and family level.

Main conclusions In this work, we propose the nuclear ITS2 rRNA marker as a new barcoder for eolid nudibranchs. We characterized for the first time the ITS2 rRNA secondary structure in nudibranchs and we pinpointed the conserved regions. Our results would desirably stimulate the increase of DNA barcoding libraries available for nudibranchs that currently only include mitochondrial DNA sequences.

ABSTRACT. Aim Neuropeptides are small intercellular signalling molecules generally produced by neurons and they regulate animal physiology and behaviour. For example, one of such neuropeptides is the Thyrotropin-Releasing-Hormone (TRH), an hypothalamic tripeptide, that controls metabolism, growth and reproduction acting through the Hypothalamic-Pituitary-Thyroid axis. Neuropeptides evolutionary origin can be tracked back to the common ancestor of bilaterian animals (ubilateria, >550 Mya) and the study of their evolution and function may shed light on the nervous system evolution and our understanding of its structure and function. Aim of our study is to take advantage of the phylogenetic and physiological characteristics of the sea urchin Strongylocentrotus purpuratus to investigate the TRH function in this deuterostomian invertebrate species in order to shed light on the evolution of the TRH pathway. Methods We caracterized the role of a sea urchin TRH neuropeptide (QYPGamide), its precursor (SpTRH) and receptor (SpTRHR) genes in Strongylocentrotus purpuratus using a combination of chromogenic and fluorescent in situ hybridisation, whole-mouth immunostaining, morpholino antisense knock-down and CRISPR/Cas9 knock-out techniques. Results In sea urchins at early pluteus stage, SpTRH is produced by two cells bilaterally distributed at each the sides of the larval apical organ. These cells are also expressing the photoreceptor gene, Go-opsin (opsin 3.2). Our results show that knock-down and knock-out experiments of the SpTRH precursor and SpTRHR inhibit the post embryonic growth of the the skeleton supporting the larval arms, but has no consequences on the skeletal body rods. Main conclusion Our functional analysis of TRH in a non-chordate deuterostomes shows a conserved ancestral role of TRH neuropeptide on regulation of postembryonic growth, validating the hypothesis suggested in 2017 by Van Sinary et al. (Proc Natl Acad Sci U S A. 2017 114: E4065-E4074).

ABSTRACT. Aim Tunicates are the closest relatives of vertebrates able to regenerate a new adult body from adult epithelia and/or mesenchymal cellsvia asexual budding and whole-body regeneration, two forms of non-embryonic development (NED). Interestingly, this regenerative capacity is scattered across the whole sub-phylum, with species able to undergo NED (colonial) closely related to species where this regenerative capabilities are absent or reduced (solitary). This suggests that NED has been acquired or lost several times among the group. A recent phylogeny of the Styelidaefamily suggests that the colonial species Polyandrocarpa zorritensis have acquired the capability of NEDindependently from other budding species. However, adetailed description of the cells and tissues responsible for NED in this species was missing. Methods Histological and ultrastructural analyses were carried in order to identifying the stages of NED under laboratory conditions and characterize the nature of the tissues and cells involved. The participation of cell proliferation has been also tested by EdUassays and anti-PHH3 immunostaining. Results In P. zorritensis, new buds develop through an invagination of vascular epidermis with contribution ofmesenchymal cells. Thistype of NED was undescribed before and we calledit “vasal budding”. During the early stages of vasal budding, undifferentiated mesenchymal cells named hemoblasts cluster and proliferate at the budding point.Hemoblasts are known as putative stem cells responsible for NED in other colonial ascidian and preliminary data on P. zorritensissuggest that hemoblast express the stem cell marker Integrin-alpha-6 at the onset of organogenesis during vasal budding. Main conclusions Overall this work generates basic biological understandings of a new organism, Polyandrocarpa zorritensis,suitable for further comparative studies on the cellular and molecular mechanisms of NED in tunicates and their evolution.

ABSTRACT. Aims The aim of the project consists on the identification of microRNAs and piRNAs involved in sea urchin germ line specification. The species chosen for this project is Paracentrotus lividus and the cell type studied are the small micromeres (sMic). These cells are known to express germ line specific genes and contribute to the left coelomic pouch (LCP). The LCP grows and upon interaction with some cells invaginating from the adjacent ectoderm forms the rudiment that will then become the juvenile sea urchin after metamorphosis. Therefore, sMic descendant cells contribute to the sea urchin adult and contain germ line related genes.

Methods To compile the smallRNA library, sMic enriched RNA is necessary and therefore the sMics are first isolated from the rest of the embryo via life cell labelling with Calcein-AM at the desired developmental stage, to then be single cell dissociated and sorted via fluorescent activated cell sorting (FACS). Subsequently, bulk RNA sequencing and specialised microRNA library-construction sequencing takes place. Bioinformatically, to process the sequencing data, a series of filtering and differential analysis steps are used to find the piRNAs and microRNAs that are believed to be relevant to the germline. The final step involves determining the functional, developmental and morphological importance of smallRNAs for the sea urchin germ line with the characterisation of gene expression via in situ hybridisation.

Results and conclusions We have successfully isolated the sMic cells and the analysis of the sequencing data is undergoing. Previous research has established the importance of smallRNAs (e.g. PIWI-like) for the germ line of the sea urchin, found within gene networks that are crucial for the formation of gonads in the adult. Thus, compiling a library of these smallRNA enriched cells may shed light on the cell specification mechanisms and genetic drivers in these pluripotent and stem-cell-like cells.

ABSTRACT. Bivalves are a diverse mollusc group of economic and ecological importance. Their evolutionary resilience to pollution, parasites and extreme environments make them important model systems for studying adaptation and immunology. Despite substantial progress in sequencing projects of bivalves, this is currently contrasted with a lack of comparative genomics and transcriptomic datasets especially for non-coding and gene-regulatory aspects. We here review the current state of bivalve microRNA complements, the most important and most conserved gene-regulators across Metazoa. We find substantial heterogeneity and incorrect annotations of microRNAs in the five available bivalve species that we corrected by reannotation of conserved microRNA families using recently established criteria for bona fide microRNA genes, and expanded it by adding two novel species, the Mediterranean mussel Mytilus galloprovincialis and the blood clam Scapharca broughtonii. Altogether we found 104 missing microRNA families in the previously published bivalve datasets and 89 families and 147 genes in the two novel species, respectively, that give a very homogeneous and consistent picture of regulatory evolution in bivalves and enable future comparative studies.By re-evaluating and re-annotating conserved miRNAs from previous reports and expanding miRNAome data to two additional species, we showed how incomplete previous annotations were and demonstrated a high conservation of miRNA families, according to the taxonomic position of bivalves. This not unexpected discovery opens up for novel paths to study evolution of bivalves with miRNAs and, vice versa, miRNA evolution in bivalves, as we have identified distinct patterns of miRNA expansion, putative losses and two bivalve-specific miRNA families. The identification of two bivalve specific microRNA families sheds further light into bivalve specific gene-regulatory adaptations and trail-blazes directions of bivalve research in the future.

ABSTRACT. Opsins are light sensitive photopigment proteins responsible for phototransduction, the conversion of light to an electrical potential. Opsins are present in every animal except sea sponges and are crucial in the role of animal vision. Early opsin evolution, focusing on marine life, is critical to our understanding of how nervous systems evolved in the early animals, although no agreement has been reached on the relationships between opsin subfamilies. We resolved opsin family relationships with the reanalysis of a previous dataset that proposed several novel yet enigmatic clades; the relationships between novel clades and the major opsin families is contentious. We also construct a new, high quality dataset that reveals a simple structure of opsin classification, with particular focus on early marine Phyla: Cnidaria and Ctenophora. We show that the use of destabilising outgroups and inadequate models of substitution play a role in the erroneous proposal of additional opsin paralogs in the Bilateria. The new dataset improves the taxonomic sampling of non-bilaterian, marine animals, providing evidence for the diversification of all opsin subfamilies in the eumetazoan and a single origin for the evolution of nervous systems in Metazoa.

ABSTRACT. Aim Theory predicts male strategic allocation of resources to the ejaculate in response to the intensity of sperm competition. However, empirical tests are sparse and focused on sperm, overlooking the strategic allocation on the seminal fluid component of the ejaculate. The black goby, Gobius niger, an external fertilizing fish with guard-sneaker mating tactics, is an ideal model to fill this gap. Territorial males release ejaculates rich in viscous seminal fluid embedding sperm that slowly dissolve during spawning. Sneaker ejaculates contain 10-fold more sperm than those of territorials and 10-fold less seminal fluid, for a rapid and conspicuous sperm release. Sneaker males but not territorials, tailor the number of sperm released to the intensity of sperm competition. Here we test whether territorial males strategically adjust their ejaculate performance by exploiting seminal fluid.

Methods Territorial males were exposed to different levels of sperm competition (absence of competing sneakers, 1 or 3 sneakers). Their ejaculate was manually stripped on a piece of acetate sheet and let to dissolve in sea water. The proportion of sperm released in 30 minutes over the total amount of sperm in the ejaculate was, then, estimated.

Results The total number of sperm per ejaculate did not differ among treatments, but we found that a higher proportion of sperm was released in presence of 1 sneaker as compared to control (sneaker absent). However, when competition was more intense (3 sneakers) the proportion of sperm released was maintained lower, at the same level as in the no-competition treatment.

Main conclusion Black goby territorial males appear to adjust their seminal fluid quality in relation to the perceived level of competition. A rapidly dissolving ejaculate or faster sperm might be advantage to defeat a competitor. By contrast, a slower dissolving ejaculate might allow to better face prolonged intrusions by consecutive sneakers.

ABSTRACT. Aim Which are the genetic bases of the adaptation of penguins to extreme temperature conditions? To address this question, we plan a comparative genomic study between the Emperor penguin (​Aptenodytes forsteri), that breeds only in Antarctica, and its much less cold-adapted sibling species, the King penguin (​A. patagonicus), that reproduces exclusively in ice-free sub-Antarctic islands. We aim to detect signatures of differential selection as well as to reveal genetic traits relevant to adaptation to cold in the Emperor penguin. Methods To identify candidate genes involved in lineage-specific adaptations, we first implemented a selection analysis using the ratio of non-synonymous and synonymous variation (ω) comparing the orthologous genes across several penguin and bird species. Then we ran a haplotype-based test (XP-EHH) using population-level genomic data to discover more recent and less intense selection events that were mostly specific to each species. Finally, to investigate the functions of the candidate genes for positive selection, we performed a GO terms overrepresentation analysis. Results The first analysis revealed strong evidence of differential selection for 68 genes indicating that, for these genes, either the King or the Emperor was indeed evolving at different rates compared to 7 penguin and 13 other bird species in the examined phylogeny. A higher number of candidate SNPs and genes under positive selection was detected in the Emperor penguin by both statistical methods and most of them were related to metabolism, response to stimulus, development and signaling according to GO terms enrichment analysis. Main conclusion Our study showed that the Emperor penguin has been subject to more pervasive selection pressures than the King and that several candidate genes which exhibited signals of positive selection in the Emperor penguin were associated with processes that are relevant for adaptive evolution in the extreme Antarctic conditions. Thematic Areas Adaptation, Populations and Species

ABSTRACT. The shape of the naso-oral region in elasmobranch species changes from elongated and thin to short and flat. That shape may be related to the phylogeny, environmental factors, types of habitat and feeding strategies. In the frame of sensory biology, the shape of the head is related to the surface area for the distribution of the electroreceptors and affects the water flow on the olfactory chamber. Geometric morphometrics is a quantitative analysis useful to compare morphology considering homologous points called landmarks. The data distribution on the multidimensional space through principal component analysis shows the differences responding to environmental or evolutionary pressures. The Triakid is a heterogeneous family, it belongs to the Carcharhiniformes and counts six genera with both benthonic and pelagic deep species, both coastal and oceanic species, both squid-feeder only and generalist species. The preliminary study presented here proposes the analysis of the Triakid’s naso-oral region considering rostrum, mouth, nares, and nasal flaps as landmarks. The naso-oral shape of 42 out of at least 45 Triakids species is plotted on a two dimensions plane, which describes the 53% of shape variance. Moreover, the result is coupled with a phylogenetic tree based on the mitochondrial gene of NADPH2 and shows a different species’ clusters, apparently determined by phylogeny rather than ecological traits. This preliminary work shows that in a numerous sharks’ family, the main morphological differences in naso-oral region are produced by the phylogenetic relationship. More investigations will be necessary to understand the changes in shape all over the taxonomic levels in elasmobranch species.

ABSTRACT. Individuals that forgo their own reproduction in animal societies represent an evolutionary paradox because it is not immediately apparent how natural selection can preserve the genes that underlie non-breeding strategies. Cooperative breeding theory provides a solution to the paradox: non-breeders benefit by helping relatives and/or inheriting breeding positions; non-breeders do not disperse to breed elsewhere because of ecological constraints. However, the question of why non-breeders do not contest to breed within their group has rarely been addressed. Here, we use an experimental approach to investigate the ecological and social drivers for the evolution of cooperative breeding in clownfish (Amphiprion percula). To test the hypothesis that non-breeding individuals do not disperse to breed elsewhere because of strong ecological constraints in the form of risk of mortality during dispersal, we experimentally tested the critical prediction that non-breeding individuals will disperse when the risk of moving between anemones is reduced. To test the hypothesis that non-breeding individuals do not contest for breeding positions because of strong social constraints in the form of evictions of non-cooperative individuals, we experimentally tested the critical prediction that non-breeding individuals will contest for breeding positions when the probability of winning a contest is increased. Our results show it is the combination of ecological and social constraints that promote the evolution of non-breeding strategies: non-breeders will disperse when ecological constraints (risk of mortality during dispersal) are experimentally weakened. In addition, we show non-breeders will contest when social constraints (risk of eviction during contest) are experimentally relaxed. The findings highlight parallels between, and potential for unification of, cooperative breeding theory and economic bargaining theory: individuals will forgo their own reproduction and wait peacefully to inherit breeding positions (engage in cooperative options) when there are harsh ecological constraints (poor outside options) and harsh social constraints (poor inside options).

ABSTRACT. Aim Antarctic fishes adopted a wide set of adaptations that allow them to thrive in the extreme antarctic environment. For example due to the high solubility of gases in cold waters some Channichthydae such as Chionodraco hamatus have completely lost hemoglobin and gasses are simply dissolved in their colorless blood. Although other adaptations, such as an increased activity of carbonic anhydrase activity in gills, have been previously described from a biochemical point of view, these have not been investigated from a genomic perspective. Here we explore cold adaptations of Cryonotothenioidea at a transcriptomic level in a multi-species comparison. Moreover we investigate the transcriptomic response of Trematomus bernacchii to a slight heat stress. Methods Whole transcriptome sequencing data from 14 fish species were collected from NCBI or generated de novo. A bioinformatic approach was used to identify peculiar transcriptomic features of antarctic species compared with species living in temperate environments, and to assess the response of Trematomus bernacchii to a slight heat stress in 3 different tissues (brain, gill, muscle). Results We identified 130 upregulated genes in the gills of Cryonotothenioidea, including two carbonic anhydrases displaying high gill-specificity. Moreover molecular signatures of cobalamin deficiency, which could be linked with the high parasite loads found in these species, were observed. The heat stress experiment revealed the brain as the most sensible tissue after 20 days of exposure, with most changes affecting protein synthesis, vescicular transport, inflammation and cytoskeleton assembly. Comparatively, the gills and muscle tissues displayed an earlier and weaker transcriptomic response. Interestingly, several hsps were regulated in response to stress both from heat and stabling. Main conclusion This study allowed us to identify peculiar transcriptomic markers of cold adaptation in Cryonotothenioidea and to understand how the future sea water warming may affect the response of these organisms in the short term.

ABSTRACT. Abstract

Seagrasses are particularly susceptible to environmental changes such as the intensification of warming and eutrophication events. The occurrence of rapid environmental changes are threating natural populations. Posidonia oceanica is an endemic Mediterranean species and ranks amongst the slowest- growing and longest-lived plants on earth. Recent evidence revealed that P. oceanica plants distributed along bathymetric or latitudinal gradients show different transcriptomic responses to common stress, suggesting a crucial role of pre- adaptation to local environmental conditions in driving response to stress. Here we aimed to analyze the physiological and transcriptional responses of P. oceanica plants with different life histories to multiple stresses. Methods P. oceanica shoots collected from sites with different nutrient conditions (Eu plants = Eutrophic plants; Ol Plants = Oligotrophic plants) were exposed in a common garden experiment to nutrient enrichment, temperature increase and their combination. Leaf tissue was collected after two weeks of exposure to stressed conditions to assess the physiological status of plants and to analyze gene expression profiles, applying the TAG sequencing technique. Results Plants showed different transcriptional profiles against the same stress conditions. Differentially expressed genes varied among plants, with a higher genes activation in plants growing in a more stressed environment, in respect to plants growing in a pristine environment. The effect of temperature at physiological level was lower in comparison to nutrients and the analysis, as confirmed by gene expression results. An interesting expression pattern of methylation processes was observed, including a large number of genes related to histone modifications, supporting their role in modulation and control of gene expression. Conclusion The different life history of P. oceanica plants is crucial to understand future persistence of this species under rapid environmental changes. Local environmental conditions seem to modify plant responses to multiple stresses through transcriptional regulation, which could have an epigenetic basis.

ABSTRACT. Stress granules (SGs) are cytoplasmic foci, composed of non-translating messenger ribonucleoproteins, translational initiation factors and other additional proteins, able to modulate gene expression when cells are subjected to adverse environmental conditions. Very few works have been devoted to study the presence of the molecular components of SGs in invertebrates. In this work, we characterized the transcript sequences for two important mRNA-binding proteins of SGs, TIA-1-related nucleolysin (TIAR) and tristetraprolin (TTP), in the solitary ascidian Ciona robusta. The identified sequences for tiar and ttp transcripts allowed us to start a study on the molecular evolution of these proteins in metazoans and carried out the first studies on their gene expression under stress conditions induced by metals (Cu, Zn, Cd and Fe). For both the proteins, the obtained results do not match the accepted phylogenesis of tunicates as sister group of vertebrates. Data on mRNA expression levels, provided by qRT-PCR, show a generalized decrease at the second day of metal-exposure for both tiar and ttp, suggesting that the induced acute stress promotes the inhibition of the transcription for the two studied proteins and then the disassembly of SGs to release useful mRNA to face the stress. ISH analyses demonstrate that TIAR and TTP antisense riboprobes recognize circulating granular amoebocytes in the hemolymph, visible in both blood lacunae and tunic. The results obtained in this work increase our knowledge on the molecular evolution of anti-stress proteins in metazoans and emphasize the importance of the transcription of tiar and ttp, which represents an efficient physiological response allowing organisms to survive under stress conditions in the environment.

ABSTRACT. Hermodice carunculata (Pallas, 1766) is a large-sized polychaete with an opportunistic scavenger feeding behaviour. Due to its northward spread along the Italian coasts, it is considered as a biological indicator of the tropicalization of the Mediterranean Sea.  To understand its population dynamics and local invasiveness potential, laboratory growth rate estimations were carried out starting from small specimens sampled at shallow waters off Santa Caterina (Ionian Sea, Apulia, Italy). Six small specimens of Hermodice carunculata (mean size 11,4±1,9 cm length; 3,4±1,6 g weight; mean age 13-15 months) were transferred to a 100 l aquarium at T = 22 °C, salinity 35-38‰, with a constant LD 12-12 photoperiod, and fed ad libitum twice a week with a diversified diet. Specimens with different color patterns were chosen for a better identification over the experiment. Individual weight, length, and number of chaetigers were monthly assessed from October 2019 to July 2020. At the end of the experiment, a classical hematoxylin-eosin histological analysis was carried out for determining the state of sexual maturity of the cultured  specimens. Over the course of the 10-months experiment, the mean size of cultured fireworms increased from 11,4±1,9 cm to 22,4±3,5 cm in length, and from 3,4±1,6 g to 17,5±3,5 g in weight, with a daily growth rate of 0,054±0,008 g/day. At the end of the experiment coelomic cavities were filled by sperms or oocytes at different state of maturations. The distinctive colour pattern of each specimen was maintained over the course of the experiment. Under laboratory conditions, fireworms exhibited fast growth rate with large dietary adaptability to different food items, and reached sexual maturity during their second year of life. The occurrence of large specimens in wild populations may be indicative of iteroparous reproduction. These features may be key to understand the high invasive potential of this species across the Mediterranean basin. Thematic Areas Populations and Species, Biodiversity

ABSTRACT. The past five years has seen a rise in the use of the genus Cassiopea as a scyphozoan model organism. Studies have explored behavior and environmental parameter tolerance limits of adults and strobilation of polyps, but there is currently no account of normal Cassiopea development under regulated conditions in a lab setting (35ppt salinity, 7mg/L O2, 25℃). To address this gap in the research, we have taken developmental images and size metrics of 50 Cassiopea from day of strobilation to the 42nd day across a variety of feeling regimes (0 kcal/day, 9.4*10^-6 kcal/day, 18.8*10^-6 kcal/day, 37.5*10-6 kcal/day) to establish a growth curve and mortality in a highly regulated, low PAR environment. All Cassiopea feeding treatment groups developed at consistent rates. While the unfed treatment group had the highest mortality (23%), surviving unfed individuals managed to remain stable in size across all 42 days with no visible transition away from their immediately post-strobila state. All fed groups experienced a significant increase in the mean oral arm length to bell radius ratio, while unfed medusae did not, however within fed medusae this ratio was not correlated with increased feeding amounts. At the end of 42 days, symbiont density was determined by ephyra homogenization and varied dramatically between individuals from 117 cells/g to 1139 cells/g, not well correlated with feeding regime. Unfed medusae retained more ephyra-like behaviors, spending less time settled on the bottom of containers. Together this suggests that unfed ephyrae can maintain size for up to 6 weeks and largely experience a pause in development of adult characteristics without feeding.

ABSTRACT. Abstract Aim Gall crabs live in obligate symbiosis with tropical scleractinian corals and induce skeletal modifications in their hosts that are used as dwellings. Their unique biology has led to remarkable adaptations in morphology and reproductive output. However, due to their diminutive size (< 1 cm) and hidden lifestyle many aspects of their biology are still unknown. The gall crab Hapalocarcinus marsupialis, associated with Pocilloporidae corals, is thought to be a species complex based on genetic data, showing stricter host specificity than previously assumed. Here we use an integrative approach combining morphometrics and reproductive data to study host-adaptive speciation in Hapalocarcinus. Methods To study the diversity in Hapalocarcinus, and possible morphological and reproductive differences between putative species, we collected 163 specimens in the Red Sea from Stylophora and Pocillopora corals. Phylogenetic analyses were conducted using the barcoding gene COI. Carapace, abdomen and chelae measurements were taken using a stereomicroscope equipped with calibrated optical micrometer. Brood mass of ovigerous females was removed, fecundity was determined as number of eggs. Females and their eggs were dried and weighed to calculate reproductive output. Results Phylogenetic analyses revealed five candidate species in the H. marsupialis complex. Species a-b inhabit Stylophora corals, whereas species c-e inhabit Pocillopora. Candidate species differ in carapace, abdomen and chelae morphology. Moreover, crabs collected from Pocillopora are significantly bigger than those collected from Stylophora, and their fecundity is on average 117 % higher. The average reproductive output (69%) of H. marsupialis is high compared to non-symbiotic brachyurans (~20%). Main conclusions Morphometrics and reproductive traits are useful in separating the putative species in the H. marsupialis complex. We hypothesize that host specificity in Hapalocarcinus lead to morphological adaptations in the crabs, likely influenced by the size and morphology of their dwelling. Hence co-evolution is considered a driving factor behind species divergence.

ABSTRACT. The two species of yellow-nosed albatross, Atlantic (Thalassarche chlororhynchos) and Indian (Thalassarche carteri), are morphologically similar, however, they show some differences in behaviour and breeding range. We studied genetic variation within and among the two species using nuclear (microsatellite, Pema7 and Occa9) and mitochondrial (control region) markers. We analysed 354 samples from four breeding islands and two bycatch locations (Atlantic: Nightingale, Inaccessible and Gough; Indian: Amsterdam Island, and bycatch samples from South African and New Zealand). Both sets of markers clearly differentiated the two species. Microsatellite and Occa9 nuclear markers revealed two genetically distinct groups within Atlantic yellow-nosed albatross, separating birds from Nightingale and Inaccessible Islands from those on Gough Island. All of the South African and New Zealand bycatch samples were Indian yellow-nosed albatross, but genetically distinct from the Amsterdam Island birds and from each other. Both species have distinct at-sea distributions. Within both species differences in at-sea distribution might also have resulted in genetic differentiation, but there is currently no evidence of intraspecific differences. The two yellow-nosed albatross species are listed as endangered due to their limited number of breeding sites and threats from introduced diseases, introduced predators and fishing mortality. As such, our results contribute to conservation and management and suggest the need for separate management of genetically distinct groups of Atlantic and Indian yellow-nosed albatrosses breeding at different islands.

ABSTRACT. The pace-of-life syndrome (POLS) hypothesis posits consistent variation in behavior among individuals (personality) to covary with variation in life history traits. Within species, the relationship among these traits is expected to plastically responds to changes in ecological conditions. Trade-off should vary along environmental gradients, but the extent to which relationships between POLS traits vary across environmental conditions remain unclear. Here, we measured consistent among-individual variation in both behavior (personality) and morphology (body shape) within and between groups of juveniles of the big-scale sand smelt (Atherina boyeri) - a fishery-target species in the Venetian Lagoon. We compared individuals caught in environmentally different areas and individuals caught with active or passive fishing techniques. We suggest that among-individual variation in behavior and morphology may emerge in juveniles growing in different conditions. In particular, the chosen areas differ in their distance from the sea harbor (more/less exposed to lagoon-sea water exchange) and in the habitat complexity (seagrass meadows vs sandbanks). Second, some personality, such as boldness or high swimming activity, morphological traits may influence juveniles’ vulnerability to fishing. Such plastic responses in early developmental stages may be critical in a scenario of environmental changes and fishery induced evolution.

ABSTRACT. Aims The fan mussel, Pinna nobilis represents the largest bivalve of the Mediterranean Sea, where it is an endemic flagship species. Unfortunately, since 2016 a dramatic mass mortality has been observed in several Mediterranean areas. Epidemiological surveys indicated as potential cause the co-occurrence of different etiological agents, (e.g. Haplosporidium pinnae, Mycobacterium sp., Vibrio spp. and Perkinsus spp.). Here we performed diagnostic molecular surveys on populations of P. nobilis from Sardinia to provide insights on phylogeny and taxonomic status of the possible etiological agents.

Methods Diagnostic analyses have been performed on 48 samples of P. nobilis and two of the congeneric P. rudis, collected in Sardinia recently, to obtain 18S and 16S ribosomal DNA sequences of the protozoa and bacteria infecting fan mussel tissues.

Results Twenty-seven identical sequences of H. pinnae (18S rDNA) were obtained. All these sequences corresponded exactly to one of those already available on GenBank, therefore no phylogenetic surveys were performed. Four sequences of the 16S rDNA of Mycobacterium sp. were isolated from P. nobilis and P. rudis. Phylogenetic analyses showed that Mycobacterium spp. found in our samples are closely related to common mycobacteria and belong to three never described before taxonomic entities.

Main conclusion Our findings indicate the occurrence of several species of Mycobacterium in the Sardinian marine environment, although they might not play a pathogenic role for fan mussels. Indeed, we isolated sequences of mycobacteria from both dead and healthy individuals of P. nobilis and P. rudis, suggesting that their presence may not be necessarily linked to the causes of the mass mortality. The large number of mycobacteria taxonomic entities found likely infecting the fan mussel, suggests that their overall role in the mass mortality of P. nobilis should be re-evaluated and support the occurrence of a multifactorial disease as a possible explanation of this event.

ABSTRACT. Despite the fact of being considered rare or endangered, many species miss effective monitoring measures or protection. This is the case of Savalia savaglia (Bertoloni, 1819), a parasitic zoanthid listed as “Near Threatened” by IUCN and included in the Annex II of the SPA/BIO Protocol of the Barcelona Convention. S. savaglia is an ecosystem engineer contributing to the biodiversity and the structural complexity of mesophotic benthic assemblages. Owing to its quite deep distribution, records of its presence are increasing only recently, thanks to ROV and technical diving explorations. On the present work we update its distribution and provide a predictive model to address eventual ground truthing surveys. Methods SDMs have been applied on the occurrences of S. savaglia obtained from different sources using bioclimatic variables from Bio-Oracle dataset as covariates. Presence only and presence/absence data were modelled using different algorithms (MaxENT, GAMs, RF, GLM). Model’s predictions have been tested using cross-validation and their performance measured and compared by different evaluation metrics: AUC, TSS and environmental consistency. To finally average the more performant models in an ensemble for both presence only and presence/absence data. Results Considering the overall performances of the SDMs, better predictive powers were obtained using models developed with presence-only data, specially by random forest. Lower performances were obtained using presence-absence data which could be explained by the inconspicuous distributions of the species along with the fact that its parasitic behaviour was not considered by these first pool of SDMs. Conclusion The case study of Savalia savaglia is particularly intriguing owing to its parasitic habitus. SDM is suggested as a promising tool identifying potential areas for future field investigations and helping on the process to protect S. savaglia populations from possible threats. Present results can provide a useful reference for predictive model distribution on rare species at large scale.

ABSTRACT. Abstract Aim Movement is a vital evolutionary and ecological action for life that affects genetic diversity, persistence and connectivity of populations. Species that occupy similar ecological niches, such as seagrasses, can reveal different dispersal extent in the marine environment. In this study we use the seagrass species Halodule wrightii Ascherson, 1868, due to its restricted tendency for self-dispersal, to estimate the genetic connectivity along its eastern Atlantic distributional range, where few known occurrence reports are known, to assess the influence of abiotic (ocean currents) versus biotic (grazers) vectors in the observed genetic patterns. Methods To test the hypothesis, eight genetic markers (microsatellites) were used to genotype Halodule wrightii samples from eleven localities, form Mauritania to Angola including Cape Verde and São Tomé and Príncipe islands. Population genetic structure was analyzed with Bayesian assignment methods and factorial correspondence analysis. Biophysical modelling methods were used to infer the influence of oceanographic currents on Halodule wrightii populations genetic structure. Results Halodule wrightii populations revealed low genetic diversity, a considerable variation of genotypic diversity and a differentiation between north and south populations of the equator. The biophysical models of simulated transportation of seagrass fragments mediated by ocean currents, revealed a very low probability of connectivity between populations along the target area, suggesting that biotic transportation might have an important role in this species dispersal. Main conclusion Our findings provide the first insight into seagrass metapopulation connectivity along the tropical west Africa coast, but future studies are essential to understand the true animal dispersal potential in this region.

ABSTRACT. Seagrasses are a unique group of angiosperms that has evolutionarily recolonized the marine environment some 60-90 mya. Seagrasses exist across thousands of kilometres of the sedimentary shorelines where they provide one of the most valuable sources of food and habitat for marine life, together with being one of Earth’s most efficient carbon sinks with direct effects on mitigating global warming. Nonetheless, seagrasses are negatively being pushed into a rapid evolution driven by both natural and anthropogenic stressors with ocean warming being one of the major players. Recently, alongside the growing recognition of the importance of seagrasses and the recent application of advanced molecular technologies, the number of studies focusing on the interactions of warming and seagrasses has increased rapidly. We have gathered information from published studies to identify potential commonalities in the effects of warming and the responses of seagrasses at four different levels: molecular, biochemical/physiological, morphological/population, and ecosystem/planetary. Warming strongly affect seagrasses at all four levels and alters seagrass distribution by causing massive die-offs in some seagrass populations, whilst also enhancing tropicalization and migration of temperate species. We conclude by discussing the most significant knowledge gaps and future directions for seagrass research. Our comprehensive view about seagrasses in an era of ocean warming not only applies to seagrass scientific and management community but also provides a set of useful information for addressing the response capability of marine organisms in general and to keystone species in particular, such as corals, mangroves, and seaweeds. Furthermore, we hope to encourages local stakeholders and decision-makers on designing appropriate strategies for protecting seagrasses as well as raise public awareness on the importance of seagrass protection and restoration for a sustainable future.

ABSTRACT. Many animal societies have dominance hierarchies in which social rank is correlated with size. In such societies, the growth and size of individuals can be a strategic response to their social environment: in fishes, individuals may decrease their growth rate to remain small and retain a subordinate position; in mammals, individuals may increase their growth rate to become large and attain a dominant position—a strategy called competitive growth. Here, we investigate whether the clown anemonefish, Amphiprion percula, exhibits competitive growth also. We show that juvenile clownfish paired with a size-matched reproductive rival increase their growth rate and size relative to solitary controls. Remarkably, paired individuals achieved this, despite being provided with the same amount of food as solitary controls. Subsequently, we use a suite of experiments manipulating availability of sensory cues to determine which cues must be available for individuals to engage in competitive growth. We show that mechanosensory (pressure and/or touch) cues or unobstructed interactions are necessary for competitive growth to occur. Our results demonstrate that clownfish are able to increase their growth rate in response to social competition, and likely this process is mediated via mechanosensory cues. This study adds to the growing body of evidence that the growth of social vertebrates can be a fine-tuned plastic response to their social environment.

ABSTRACT. Aim The increasing accessibility of next generation sequencing offers new opportunities for the genomic study of non-model organisms, allowing to extend such investigations to multiple individuals and to explore phenomena which have so far remained obscure. The aim of this work was to investigate the impact of the unusual architecture of the Mediterranean mussel (Mytilus galloprovincialis) genome on the high invasiveness, resilience and adaptation potential of this species.

Methods We applied a whole genome resequencing approach to identify genes subject to presence-absence variation (PAV) in different individuals belonging to two independent populations. Resequencing data was mapped to the mussel reference genome assembly, allowing to detect hemizygous genomic regions and associated protein-coding genes subject to PAV. These dispensable genes were subjected to functional enrichment analyses, based on Gene Ontology and Pfam domain annotations, to highlight the terms most frequently associated with the accessory fraction of the genome.

Results The genome of the Mediterranean mussel provides an unprecedented example of widespread gene PAV in Metazoa. The high levels of intraspecific genetic variation can be explained by massive structural variants than include very large hemizygous regions containing protein-coding genes subject to PAV. Overall, the mussel genome comprises 45,000 core genes found in all individuals and 20,000 dispensable genes, which are enriched in functions associated with immune response and survival.

Main conclusion Preliminary analyses carried out on recently published, complete genomes from other species indicate that gene PAV is widespread in mollusks, and particularly prominent in bivalves. The functional enrichment of dispensable genes suggest that these represent useful tools for genomic evolution, allowing the maintenance of additional genetic diversity within the population and providing enhanced chance of survival in diverse environments.

ABSTRACT. Size-selective mortality is a major evolutionary driver of life-history and behavioral traits in human-dominated marine ecosystems. These changes might in turn alter the circadian system and therefore affect daily physiological processes and behavioral outputs. The proximate mechanisms governing evolution are difficult to disentangle in the wild. We overcome this limitation by studying the evolutionary impact of size-selective harvesting on group risk-taking behavior and the circadian system in a model fish species. We exposed zebrafish (Danio rerio) to either large or small size-selective harvesting relative to a control over five generations, followed by eight generations during which harvesting halted. Size-selective mortality affected fine-scale timing of behaviours. In particular, small size-selective mortality, typical of gape-limited predators targeting smaller size classes, increased group risk-taking behavior. Moreover, small size-selective mortality increased early peaks of daily activity as well as extended self-feeding daily activity to the photophase compared to controls. By contrast large size-selective mortality, typical of most wild capture fisheries, only showed an almost significant effect of decreasing group risk-taking behavior and no clear changes in fine-scale timing of daily behavioral rhythms compared to controls. We also found changes in the molecular circadian core clockwork in response to both size selective mortality treatments. These changes disappeared in the clock output pathway because both size-selected lines showed similar transcription profiles suggesting the presence of a molecular switch. To conclude, our experimental harvest left an asymmetrical evolutionary legacy in group risk-taking behavior and in fine-scale daily behavioral rhythms. Yet, the overall timing of activity showed evolutionary resistance probably maintained by a molecular switch. Our experimental findings suggest that human-induced changes in size selective mortality can have consequences for behavior and physiological processes in marine ecosystems.

ABSTRACT. Aim Cytosine methylation is a widespread epigenetic modification found in eukaryotesMethylated CG is the most abundant in plant genomes and generally occurs in repetitive/noncoding regions. Coding regions are also methylated and several studies have found that methylation of gene bodies (gbM) varies according to gene function, where genes with housekeeping functions tend to be more heavily methylated than those with inducible functions. The aim of the present study is to provide a comprehensive evaluation of DNA methylation and gene expression patterns in two marine plants with contrasting life history strategies for understanding the role of epigenetic mechanisms in the adaptation to local environments and in the response to heat stress.

Methods We assessed gbM thorough the in silico transcriptome analysis of CpGO/E ratios in the two Mediterranean seagrass species Posidonia oceanica and Cymodocea nodosa collected along a latitudinal gradient (cold and warm environment) and undergoing a heat stress treatment.

Results We demonstrate that the bulk of Posidonia oceanica and Cymodocea nodosa genes possess high levels of intragenic methylation. We also observed a correlation between low gbM and gene expression flexibility between experimental temperature regimes and between plants from different populations.

Main conclusion The empirical determination of global DNA methylation showed patterns of intra and inter-specific divergence that indicate a relationship between methylation level and the plants’ latitude of origin and life history. We reinforced the hypothesis that low DNA methylation facilitates flexible expression and plasticity under changing conditions, with a lower methylation level in genes with inducible functions.

ABSTRACT. Aim In sessile benthic species, survival of early stages is critical for the resilience of the populations. To increase survival, species have developed different defense strategies. One of the most common is the production of hard structures like skeletons or spicules, as a protection from abrasion or predation. Corallium rubrum is an octocoral species endemic to the Mediterranean Sea which produces two types of biomineral CaCO3 structures: the sclerites and the inner skeleton, composed by a medullar zone (MZ) surrounded by an annular domain (AZ). Skeletogenesis processes are well known for adult colonies but still poorly understood for the early life stages. Methods To understand the dynamics of the skeleton formation during the first 4-years of life, colonies of different ages (17-day-, 1-, 2-, and 4-years- old) have been observed with stereomicroscope and scanning electron microscope and analyzed by synchrotron tomography and Laser Ablation Inductively Coupled Plasma Mass Spectrometry (LA-ICPMS). Results During the first year, colonies did not produce an axial skeleton, but a proto-skeleton made up of sclerites not embedded in the calcitic matrix. During the second year of life, a hard skeleton formed by coalescence of sclerites and calcitic cement is produced, which correspond to the MZ observed in adult colonies. LA-ICPMS analyses showed that the chemical signature of the skeleton of 4-years-old colonies is similar to the MZ adult colonies’one, suggesting that, at that age, annular CaCO3 deposition still not occur. Main conclusion Our findings emphasize the importance of sclerites during the early developmental stages when they could offer mechanical protection from abrasion and predation forming a deformable, fast-growing armor that allows a safe development of the first polyps. Furthermore, sclerites could serve as nuclei for triggering the cementation and foster the formation of the hard axial skeleton.

ABSTRACT. A coiled shell is the most evident feature of the Bauplan of a gastropod mollusc. However, at least 54 families independently evolved an apparently simplified shell morphology: the limpet. Species with this largely uncoiled, depressed shell morphology occur in almost every aquatic habitat and are associated to several different lifestyles and diets. The marine gastropod family Capulidae includes 18 recognised genera, the large majority of which are coiled, but with a number of limpet-like species. Capulid shell plasticity is also associated to a broad range of feeding ecologies, from obligate suspension feeders to kleptoparasites. Aim Given the plasticity of shell shapes and the array of feeding strategies of capulids, this family is an ideal model to investigate (a) the evolution of limpet-like shells, (b) the evolution of kleptoparasitic feeding strategies and (c) the relationship between the two traits. Methods To investigate the evolution of the limpet-like shell in the family Capulidae we performed an ancestral state reconstruction analysis on a time-calibrated phylogenetic tree (COI, 16S, and ITS2) including 16 species representing a good deal of its morphological diversity. Results Our results identified at least three capulid lineages that independently evolved limpet-like shells, suggesting that a recurrent limpetization process characterizes this family. At least one of the limpet-like genera was undescribed and deserves the introduction of a new genus-level taxon. Main conclusions We suggest that capulids evolved from a lineage of coiled suspension feeders and that the shift to kleptoparasitism in the family ancestor may have represented a strategy to save energy through the exploitation of the water current produced by the host. Probably the major drivers of shell evolution in capulids are related to their ecology, most of them being kleptoparasites, include the shape and the kind of host substrate, and lead to the repeated acquisition of a limpet-like shape.

ABSTRACT. Abstract

The vision is a fundamental aspect of our daily life, and its evolution has puzzled scientists since Darwin. At the molecular level, it requires a sophisticated molecular machinery that is encoded by different genes. This process starts with photons of light activating a photosensitive photopigment, typically an opsin gene. This event results in the activation of a phototransduction cascade that transforms the light information into an electric signal. Finally, to function in the visual process, all these genes need to be expressed in a specialized photoreceptor cell.

Methods

We integrated phylogenetics methods and single-cell genomics to understand when the molecular components of vision evolved.

Results and conclusion

The result indicates that vision in metazoan evolved gradually, and that several molecular components largely predate the origin of eyes.