Canning Processes Reduce the DNA-Based Traceability of Commercial Tropical Tunas

Canned tuna is one of the most widely traded seafood products internationally and is of growing demand. There is an increasing concern over the vulnerability of canned tuna supply chains to species mislabelling and fraud. Extensive processing conditions in canning operations can lead to the degradation and fragmentation of DNA, complicating product traceability. We here employed a forensically validated DNA barcoding tool (cytochrome b partial sequences) to assess the effects of canning processes on DNA degradation and the identification of four tropical tuna species (yellowfin, bigeye, skipjack and longtail tuna) collected on a global scale, along their commercial chains. Each species was studied under five different canning processes i.e., freezing, defrosting, cooking, and canning in oil and brine, in order to investigate how these affect DNA-based species identification and traceability. The highest percentage of nucleotide substitutions were observed after brine-canning operations and were greatest for yellowfin and skipjack tuna. Overall, we found that DNA degradation significantly increased along the tuna canning process for most specimens. Consequently, most of the specimens canned in oil or brine were misidentified due to the high rate of nucleotide substitution in diagnostic sequences

To Be, or Not to Be: That Is the Hamletic Question of Cryptic Evolution in the Eastern Atlantic and Mediterranean Raja miraletus Species Complex

The ability to correctly identify specimens at the species level is crucial for assessing and conserving biodiversity. Despite this, species-specific data are lacking for many of South Africa’s catsharks due to a high level of morphological stasis. As comprehensive and curated DNA reference libraries are required for the reliable identification of specimens from morphologically similar species, this study reviewed and contributed to the availability of cytochrome c oxidase subunit I (COI) and nicotinamide adenine dehydrogenase subunit 2 (NADH2) sequences for South Africa’s catsharks. A molecular taxonomic approach, implementing species delimitation and specimen assignment methods, was used to assess and highlight any taxonomic uncertainties and/or errors in public databases. The investigated species were summarised into 47 molecular operational taxonomic units (MOTUs), with some conflicting specimen assignments. Two Apristurus specimens sampled in this study remained unidentified, revealing the presence of previously undocumented genetic diversity. In contrast, haplotype sharing within Haploblepharus—attributed to nucleotide ambiguities—resulted in the delimitation of three congeners into a single MOTU. This study reveals that molecular taxonomy has the potential to flag undocumented species and/or misidentified specimens, and further highlights the need to implement integrated taxonomic assessments on catsharks that represent an irreplaceable component of biodiversity in the region

Strongly structured populations and reproductive habitat fragmentation increase the vulnerability of the Mediterranean starry ray Raja asterias (Elasmobranchii, Rajidae)

1. The Mediterranean starry ray (Raja asterias) populations within the Mediterranean Sea are susceptible to high rates of bycatch in the multispecies trawl fisheries. Understanding its population structure and identifying critical habitats are crucial for assessing species vulnerability and setting the groundwork for specific management measures to prevent population decline. 2. To assess the population structure of R. asterias in the Mediterranean, the genetic variation in nine population samples at one mitochondrial marker and eight nuclear microsatellite loci was analysed. Moreover, 172 egg cases collected in the Strait of Sicily were identified at species level using integrated molecular and morphological approaches. 3. Genetic analyses revealed that the Mediterranean starry ray comprises three distinct units inhabiting the western, the central-western, and the central-eastern areas of the Mediterranean. An admixture zone occurs in the Strait of Sicily and the Ionian Sea, where individuals of the central-western and central-eastern population units intermingle. 4. The joint morphometric–genetic analyses of rajid egg cases confirmed the presence of more than one species in the admixture area, with a predominance of egg cases laid by R. asterias. DNA barcoding revealed that egg cases and embryos of R. asterias shared several haplotypes with adult individuals from the centralwestern and central-eastern Mediterranean Sea, revealing that females of both populations laid numerous eggs in this area. 5. According to these findings, detailed taxonomic determination of egg cases, when combined with seasonal migration studies, could improve the capability to identify important spawning or nursery areas for the Mediterranean starry ray, particularly in those admixture zones relevant to maintaining genetic diversity. 6. Finally, these new insights should be considered to update the Action Plan for the Conservation of Cartilaginous Fishes in the Mediterranean Sea with effective measures to reduce the impact of skate bycatch in trawling and safeguard egg cases in nursery areas

First record of the West Atlantic Ariopsis felis (Linnaeus, 1766) (Actinopterygii : Siluriformes : Ariidae) in the Mediterranean Sea

A single individual of Ariidae was captured in 2022 in the Palestinian waters off the Gaza Strip, eastern Mediterranean. A comprehensive description of the morphological and meristic characters of the specimen was conducted. Molecular identification was performed through sequence analysis of the Cytochrome Oxidase subunit I (COI). The specimen was identified as Ariopsis felis, both morphologically and genetically. The natural range of this species is the western Atlantic and, to the best of our knowledge, the species has never been reported before from Mediterranean waters. The singular occurrence of this fish in the basin prompts further discussion, raising questions about the possible vectors that may be responsible for this new introduction in the Mediterranean Sea.peer-reviewe

Puzzling over spurdogs : molecular taxonomy assessment of the Squalus species in the Strait of Sicily

The actual occurrence of Squalus megalops in the Mediterranean Sea has recently been questioned. Several research works which sought to assess available morphological and meristic features that differentiate S. megalops from other Squalus species in the Mediterranean Sea, revealed poor discriminatory power and high variability of the assessed characters, especially when comparing S. megalops and S. blainville. The application of molecular tools does not support the presence of S. megalops. In the present study, we screened spurdog species from the Strait of Sicily using a molecular taxonomy approach based on two mitochondrial DNA markers and we report the occurrence of two Squalus lineages characterizing specimens collected from the stretch of sea between Tunisia, southern Sicily, Malta and Libya. The results support the hypothesis that a common species, S. blainville, currently inhabits the Mediterranean Sea, while a second and rare species is probably an occasional visitor with high morphological similarity to the S. megalops and S. blainville but is genetically distinct from both. Within this perspective, the occurrence of S. megalops in the Mediterranean Sea is not confirmed and our study highlights the taxonomic uncertainties in relation to the occurrence and distribution of Squalus species in this region. We encourage the establishment of a coordinated international effort to implement a comprehensive and integrated taxonomic assessment on this genus which represents an irreplaceable component of the biodiversity of the area.peer-reviewe

DNA barcoding of atlantic skates (Chondrichthyes, Rajiformes): taxonomic and phylogeographic inferences and conservation impacts

Anthropogenic impacts are responsible for alarming declines in the abundance and distribution of several marine species. Their future conservation and long-term management plans need to be conceived upon a universally recognised key-feature: species identity. In the past, this important assignment resulted particularly arduous among skates (Rajiformes), in which the phenotypic similarity between some taxa and the individual variability in others, entangled accurate species identification. This research project aimed to confirm the power of DNA barcoding for the discrimination between skate species across the Atlantic Ocean and for its use as effective tool to minimize the risk of species misidentification. In this perspective, this work aimed to compile a new and fully available barcode library, the ELASMO-ATL project, which gathered biological and molecular information of Atlantic rajid fauna. In addition, considering the vast geographical distribution of six species, this study investigated whether the mitochondrial gene Cytochrome Oxidase subunit I (COI) can preliminarily identify potential geographical populations. DNA barcodes were obtained from 398 individuals collected during past scientific surveys across four main FAO divisions (FAO 27, FAO 34, FAO 41 and FAO 47), obtained data successfully resolved many taxonomic ambiguities and demonstrated a highly cohesive monophyletic clustering within the order, as well as a high number of concordant Barcode Index Number (BINs). Further inference of evolutionary patterns suitable for addressing management and conservation issues should be undertaken considering the six species selected for the phylogeographic analysis. In conclusion, a well-curated barcode library was established and this novel resource provides samples, specimens information and corresponding reference sequences of the group in question for future studies

Towards the conservation of vulnerable marine large predators: morphometric, molecular and microchemical variation of historical sawfish rostra from Mediterranean collections

Sawfishes (Chondrichthyes, Pristidae) are considered one of the most endangered families among elasmobranchs. Extensive efforts are required worldwide to gather solid information on historical and recent changes in the composition/range of species. In this study, we have implemented an integrative approach to characterize the species diversity and the abundance of historical rostra of sawfishes from museums and private collections of the Mediterranean area. The identification at the species level of 172 dried rostra was carried out through the integration of both traditional and geometric morphometric techniques with molecular tools, allowing the assessment of a robust methodical approach to discriminate species. In addition, we analysed 35 rostral teeth to clarify the past distribution of sawfish species considering the isotopic composition of oxygen and carbon. The morphometric, molecular, and geographical characterization of samples was accompanied by the preliminary evaluation of growth structures and the inspection of the strontium isotope composition in two teeth to unravel movement patterns of individuals across different salinities of water. Results were integrated with currently available data from public repositories and showed that the historical specimens belonged to four nominal species: Pristis zijsron (81), Anoxypristis cuspidata (39), P. pristis (30), and P. pectinata (22). An identification error of 5.41% emerged in the morphological distinction of rostra between juvenile individuals of P. pectinata and P. zijsron. The new approach of carbon and oxygen isotopes, implemented for the first time in these taxa, permitted the identification of the high-probability habitat preferences of these benthopelagic elasmobranchs in about 50% of the analysed specimens. Using this multidisciplinary approach, we successfully assigned the numerous museum rostra with lacking data to a given species and identified their candidate geographical origin, retrieving novel information and data for understanding the species distribution and ecology of past, sometimes locally/regionally extinct sawfish faunas

Another piece of the evolutionary history of Atlantic skates (Chondrichthyes, Rajiformes ): integrating DNA barcoding approach and phylogenetic inferences

Conservation and long-term management plans of marine species need to be conceived upon a universally recognised key-feature: species identity. This important assignment resulted particularly arduous among skates (order Rajiformes), in which the phenotypic similarity between some taxa and the individual variability in others, entangled accurate species identification. This study confirms the power of DNA barcoding for the discrimination between skate species across the Atlantic Ocean and for its use as effective tool to minimize the risk of species misidentification and to elucidate species boundaries. In this perspective, this work compiles and establish a new fully available and well-curated barcode library, the ELASMO-ATL project, which gathered biological and molecular information of 432 skate specimens and covered coastal waters of four FAO Major Fishing Areas (27, 34, 47, 41) of the Atlantic Ocean. The evolutionary histories of 34 skate species were estimated with two concatenated mitochondrial markers (COI and NADH2) through Bayesian and species level phylogeny analyses. It was possible to discover a new evolutionary lineage within the genus Raja in the southern-most part of its distribution area and to enable deepening the relationship between South-African endemic species of Rajella. Once again, Western South African coasts and oceanographic fronts may play a fundamental role among skates\u2019 speciation events in which the paleoclimatic and paleogeographic history joined to hydrography events could have contributed to the formation of refugial areas, characterised by geographical isolation. Subsequent contact zones in these areas between Senegal and Angola seems to constitute a continuum/cline of genetic change among some Raja species. These data successfully resolved many taxonomic ambiguities and demonstrated a highly cohesive monophyletic clustering among the order laying the foundations for further inference of evolutionary patterns suitable for addressing management and conservation issue

Step by step: the unprecedented evolutionary history of family Rajidae

Among skates, Rajidae represents one of the most enigmatic family of cartilaginous fish whose bio- ecological traits contributed to an extraordinary evolutionary success in terms of species richness and endemism. Past and present taxonomic conflicts and species misidentifications are linked to their extraordinary level of morphological stasis. In recent years, these issues have been overcome by wide- scale molecular taxonomy analyses, but also raised questions about their evolutionary history. Concerted actions as the ELASMOMED and ELASMOATL initiatives encouraged and improved large-scale sampling efforts in the Mediterranean and Eastern Atlantic Ocean over years, building unique voucher repositories of thousands of specimens and enabling the exploration of skates\u2019 biogeography. Here, the maximum taxonomic (51 OTUs) and molecular (47 OTUs) diversity of tribes Rajini and Amblyrajini were measured using concatenated mitochondrial genes. We also estimated their evolutionary divergence using the molecular clock approach. Evidences produced so far showed that, despite the ancient origin of Rajidae (97 MYA), the Eastern Atlantic and Mediterranean faunas originated more recently, after the closure of connection between these areas and the Indo-Pacific Ocean (15 MYA). The endemic Mediterranean species originated with the recolonization of the Basin, after the Messinian salinity crisis (7-5 MYA). At least five independent vicariant events contributed from 7.4 to 3.15 MYA to the formation of allopatric or parapatric sister species, each distributed in the N-E Atlantic and S-E Atlantic respectively. On the whole, the Quaternary tectonic movement of continental masses, paleoclimatic events and present oceanographic discontinuities occurring along the western African continental shelf might explain this series of parallel and independent speciation events related to the maintenance of low or null levels of gene flow between closely related sibling and cryptic species

Molecular Taxonomy and Diversification of Atlantic Skates (Chondrichthyes, Rajiformes): Adding More Pieces to the Puzzle of Their Evolutionary History

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