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Marine biodiversity monitoring in the Mediterranean’s increasingly threatened ecosystems is crucial for effective ecosystem conservation and management. Here, we leveraged the Mediterranean International Trawl Survey program (MEDITS) to implement eDNA sampling through the recently tested ‘metaprobe’ procedure and characterize fish assemblages in three separate areas off the Italian coasts: Northern Adriatic Sea (NoAS), Ligurian and Northern Tyrrhenian Sea (LNTS), and Sardinian Sea (SaS). By combining the information from two homologous mitochondrial 12S metabarcodes––i.e., Elas02 and Tele02 targeting elasmobranchs and teleosts, respectively––we identified 108 species, over 60% of which overlapped with those caught by the trawl net. We produced an accurate reconstruction of fish community composition of the examined sites, reflecting differences in species assemblages linked with both geographic area and depth range. Metaprobe eDNA data consistently returned a biodiversity ‘bonus’ mostly consisting of pelagic taxa not captured through bottom trawl surveys, including rare and endangered taxa (e.g., elasmobranchs). Overall, the spatial characterisation of the assemblages across the surveyed areas was better delineated and more robust using eDNA metabarcoding than trawl data. Our results support the operationalisation of the metaprobe as a simple, inexpensive, versatile sampling tool, in association with pre-existing ship surveys, to overcome many of the limitations of marine data collection and strengthen marine management.

Implant-prosthetic characteristics jeopardize accurate diagnosis, professional and domiciliary plaque control around dental implants. Accurate prosthetic design planning and prosthetic features modifications are fundamental in peri-implant diseases' primordial prevention and active treatment. To evaluate the impact of prosthetic emergence angles (EAs) and mucosal tunnel depths (MTDs) on the efficacy of ultrasonic debridement in removing ink stain simulating artificial biofilm in an in vitro model. An in vitro model simulating biofilm around implant abutment, incorporating a 4 mm implant analog replicating a missing single tooth was designed. Titanium abutments with three MTDs (2, 4, and 6 mm) were associated with individualized crowns with different EAs (15°, 30°, and 45°), resulting in nine experimental groups. Abutments were stained with artificial biofilm and subsequently instrumented through ultrasonic debridement. The proportion of residual biofilm (ResB) was quantified and evaluated for the four surfaces. A total of 360 images of 90 instrumented abutments was evaluated. The overall means described a consistent increase of ResB in relation to the progressive increment of both MTD and EA. Mesial and distal surfaces presented more biofilm across all EA-MTD combinations (   < 0.05). Logistic regression models pinpointed MTD and EA as significant predictors. The 6 mm MTD and 45° EA combination demonstrated as the strongest predictor (odds ratio [OR] = 134,33). The combination of a progressively wider prosthetic EA and a deeper mucosal tunnel significantly reduced the efficacy of submucosal instrumentation. Narrower EA (<30°) and shallower MTD (<4 mm) yielded significantly better results in terms of ResB.

Cartilaginous fish are particularly vulnerable to anthropogenic stressors and environmental change because of their K-selected reproductive strategy. Accurate data from scientific surveys and landings are essential to assess conservation status and to develop robust protection and management plans. Currently available data are often incomplete or incorrect as a result of inaccurate species identifications, due to a high level of morphological stasis, especially among closely related taxa. Moreover, several diagnostic characters clearly visible in adult specimens are less evident in juveniles. Here we present results generated by the ELASMOMED Consortium, a regional network aiming to sample and DNA-barcode the Mediterranean Chondrichthyans with the ultimate goal to provide a comprehensive DNA barcode reference library. This library will support and improve the molecular taxonomy of this group and the effectiveness of management and conservation measures. We successfully barcoded 882 individuals belonging to 42 species (17 sharks, 24 batoids and one chimaera), including four endemic and several threatened ones. Morphological misidentifications were found across most orders, further confirming the need for a comprehensive DNA barcoding library as a valuable tool for the reliable identification of specimens in support of taxonomist who are reviewing current identification keys. Despite low intraspecific variation among their barcode sequences and reduced samples size, five species showed preliminary evidence of phylogeographic structure. Overall, the ELASMOMED initiative further emphasizes the key role accurate DNA barcoding libraries play in establishing reliable diagnostic species specific features in otherwise taxonomically problematic groups for biodiversity management and conservation actions.

Over-fishing is one of the most complex and argued issue among the human impacts affecting the marine environment. The development of integrated policies for fish stock conservation and its responsible use are essential. Nowadays, genomics is considered an ultimate tool to define stock boundaries and population structure as it expands the focus on the entire organism’s genetic makeup and its interaction with the environment. An example of effort and capitalization on genomics is shown by the Italian RITMARE Flagship Project, which aims to create a genomic mapping for stocks of two commercially important species, the red mullet (Mullus barbatus L.) and the deep-water rose shrimp (Parapenaeus longirostris Lucas, 1847), through the use of 2b-RAD technology, based on sequencing the uniform fragments produced by type IIB restriction endonucleases. A sampling design covering 8 Geographical Sub-Areas was defined on the basis of nursery areas, abundance and importance in local fishery and it was completed during the MEDITS 2013 survey. After the extraction of high quality genomic DNA from 7 individuals for each species, the 2b-RAD protocol was tested for the first time. Thereafter, a high-throughput sequencing of libraries was accomplished through Illumina® platform and the software STACKS was used to analyze the sequences. Several millions of high quality reads per individual were obtained and approximately from 1.0 to 2.0x10e3 candidate SNP markers were identified. These preliminary results confirm the successful application of 2b-RAD technology on both non model species and, above all, its effective extendibility to larger number of individuals and population samples, necessary to assess stock boundaries and stock units in the Italian seas.

The present study focused on the three species of electric rays known to occur in the Mediterranean Sea: Torpedo torpedo, Torpedo marmorata and Tetronarce nobiliana. Correct identification of specimens is needed to properly assess the impact of fisheries on populations and species. Unfortunately, torpedoes share high morphological similarities, boosting episodes of field misidentification. In this context, genetic data was used (1) to identify specimens caught during fishing operations, (2) to measure the diversity among and within these species, and (3) to shed light on the possible occurrence of additional hidden species in the investigated area. New and already published sequences of COI and NADH2 mitochondrial genes were analyzed, both at a small scale along the Sardinian coasts (Western Mediterranean) and at a large scale in the whole Mediterranean Sea. High levels of genetic diversity were found in Sardinian populations, being significantly different from other areas of the Eastern Mediterranean Sea due to the biotic and abiotic factors here discussed. Sardinian torpedoes can hence be indicated as priority populations/areas to be protected within the Mediterranean Sea. Moreover, sequence data confirmed that only the three species occur in the investigated area. The application of several ‘species-delimitation’ methods found evidence of cryptic species in the three species outside the Mediterranean Sea, as well as in other genera/families, suggesting the urgent need for future studies and a comprehensive revision of the order Torpediniformes for its effective conservation.

Investigating marine species population structure in a multidisciplinary framework can reveal signatures of potential local adaptation and the consequences for management and conservation. In this study we delineate the population structure of common sole (Solea solea) in the Mediterranean Sea using genomic and otolith data, based on single nucleotide polymorphism (SNPs) markers, otolith shape and otolith trace element composition data. We correlated SNPs with environmental and spatial variables to evaluate the impact of the selected features on the actual population structure. Specifically, we used a seascape genetics approach with redundancy (RDA) and genetic-environmental association (GEA) analysis to identify loci potentially involved in local adaptation. Finally, putative functional annotation was investigated to detect genes associated with the detected patterns of neutral and adaptive genetic variation. Results from both genetic and otolith data suggested significant divergence among putative populations of common sole, confirming a clear separation between the Western and Eastern Mediterranean Sea, as well as a distinct genetic cluster corresponding to the Adriatic Sea. Evidence of fine-scale population structure in the Western Mediterranean Sea was observed at outlier loci level and further differentiation in the Adriatic. Longitude and salinity variation accounted for most of the wide and fine spatial structure. The GEA detected significant associated outlier loci potentially involved in local adaptation processes under highly structured differentiation. In the RDA both spatial distribution and environmental features could partially explain the genetic structure. Our study not only indicates that separation among Mediterranean sole population is led primarily by neutral processes because of low connectivity due to spatial segregation and limited dispersal, but it also suggests the presence of local adaptation. These results should be taken into account to support and optimize the assessment of stock units, including a review and possible redefinition of fishery management units.

The Blackspot Seabream, Pagellus bogaraveo, is a commercially valuable species widely distributed in the northeastern Atlantic and Mediterranean. Its biology makes it vulnerable to overfishing, but its population structure and ontogenetic migration strategy remain unclear. Building on previous work based on microsatellite markers, we expanded the investigation by analysing the mitochondrial Control Region (CR) to complement nuclear data. We analysed 199 specimens from 13 sites and combined the new CR sequences with 129 published records to achieve the broadest coverage in terms of biogeographic and genetic data. We calculated genetic diversity and performed AMOVA, pairwise ΦST comparisons, and multivariate analyses. Eighty-eight haplotypes were identified, showing high haplotype diversity (Hd = 0.767–0.945) and moderate nucleotide diversity (π = 0.0026–0.0054). Most genetic variation occurred within populations, and overall analyses indicated genetic homogeneity. However, pairwise analysis and AMOVA confirmed significant differentiation of the Azores population. These results confirm extensive genetic connectivity throughout the Atlantic–Mediterranean range of P. bogaraveo, likely due to a combination of large larval dispersal and a common spawning migration strategy, but identify the Azores as a genetically distinct unit. This highlights the need to consider both large-scale connectivity and local divergence in fisheries management.

Conservation and long-term management plans of marine species need to be based upon the universally recognized key-feature of species identity. This important assignment is particularly challenging in skates (Rajiformes) in which the phenotypic similarity between some taxa and the individual variability in others, hampers accurate species identification. Here, 432 individual skate samples collected from four major ocean areas of the Atlantic were barcoded and taxonomically analysed. A BOLD project ELASMO ATL was implemented with the aim of establishing a new fully available and well curated barcode library containing both biological and molecular information. The evolutionary histories of the 38 skate taxa were estimated with two concatenated mitochondrial markers (COI and NADH2) through Maximum Likelihood and Bayesian inference. New evolutionary lineages within the genus Raja were discovered off Angola, where paleogeographic history coupled with oceanographic discontinuities could have contributed to the establishment of isolated refugia, playing a fundamental role among skates’ speciation events. These data successfully resolved many taxonomic ambiguities, identified cryptic diversity within valid species and demonstrated a highly cohesive monophyletic clustering among the order, laying the background for further inference of evolutionary patterns suitable for addressing management and conservation issues.

The present study, based on microsatellite markers, describes a population genetic analysis of the small-spotted catshark Scyliorhinus canicula (Linnaeus, 1758), representing one of the most abundant and commonly caught cartilaginous fishes in the Mediterranean Sea and adjacent areas. The analyses were performed to unravel the genetic features (variability, connectivity, sex-biased dispersal) of their relative geographic populations, both at the small (around the coast of Sardinia, Western Mediterranean Sea) and at a larger spatial scale (pan-Mediterranean level and between the Atlantic Ocean and the Mediterranean Sea). Individual clustering, multivariate and variance analyses rejected the hypothesis of genetic homogeneity, with significant genetic differences mainly within the Mediterranean between the Western and Eastern basins, as well as between the Mediterranean and the NE Atlantic Ocean. In detail, our results seem to confirm that the Strait of Gibraltar could not represent a complete barrier to the exchange of individuals of small-spotted catshark between the Atlantic Ocean and the Mediterranean Sea. In the latter area, a complex genetic structuring for S. canicula was found. Apart from differences among the Western, Eastern and Adriatic sites, within the Western basin the small-spotted catsharks around Sardinian waters are strongly differentiated from all others (both from the eastern Tyrrhenian Sea and southernmost part of the Algerian basin) and are demographically stable. Several possible mechanisms, both biological and abiotic (e.g., migratory behavior, waterfronts, and oceanographic discontinuities), are discussed here to explain their peculiar characteristics. Overall, the genetic data presented, both at the local and regional level, could represent a baseline information, useful for the temporal monitoring of populations, and to assess the effects of present or future fishing/management/conservation measures.

The deep sea, the largest biome on Earth, is the least explored and understood. This lack of knowledge hampers our ability to understand and protect this important environment. In this study, water and sediment samples were collected at different depths in the central Mediterranean (224–780 m), specifically, within the Dohrn Canyon and the Palinuro Seamount, to investigate the diversity of cephalopods and establish a baseline knowledge of their distribution in these sites to preserve their habitats and estimate the impacts of human-driven environmental changes. Key taxa identified included Heteroteuthis sp., Loligo sp., and Histioteuthis sp., which were the most abundant across all sampling stations. A low overlap in species detection was observed between water and sediment samples, confirming previous findings that the typology of environmental matrices used in eDNA metabarcoding has a significant impact on the organisms detected and, therefore, the integrated use of different matrices to better represent local biodiversity is recommended. Furthermore, this study highlights the limitations posed by gaps in reference databases, particularly for deep-sea organisms, and addresses these by emphasising the need for improved multi-marker approaches and expanded reference databases to enhance the accuracy of eDNA-based biodiversity assessment.