The blue crab (BC) Portunus segnis is considered an invasive species colonizing Tunisian coasts since 2014. This work aims to explore its associated bacteria potential to produce anionic exopolysaccharides (EPSs) in order to open up new ways of valorization. In this study, different BC samples were collected from the coastal area of Sfax, Tunisia. First, bacterial DNA was extracted from seven different fractions (flesh, gills, viscera, carapace scraping water, and three wastewaters from the production plant) and then sequenced using the metabarcoding approach targeting the V3-V4 region of the 16S rDNA to describe their microbiota composition. Metabarcoding data showed that the dominant bacterial genera were mainly Psychrobacter, Vagococcus, and Vibrio. In parallel, plate counting assays were performed on different culture media, and about 250 bacterial strains were isolated and identified by sequencing the 16S rDNA. EPS production by this new bacterial diversity was assessed to identify new compounds of biotechnological interest. The identification of the bacterial strains in the collection confirmed the dominance of Psychrobacter spp. strains. Among them, 43 were identified as EPS producers, as revealed by Stains-all dye in agarose gel electrophoresis. A Buttiauxella strain produced an EPS rich in both neutral sugars including rare sugars such as rhamnose and fucose and uronic acids. This original composition allows us to assume its potential for biotechnological applications and, more particularly, for developing innovative therapeutics. This study highlights bacterial strains associated with BC; they are a new untapped source for discovering innovative bioactive compounds for health and cosmetic applications, such as anionic EPS.

Serratia sp. cause food losses and waste due to spoilage; it is noteworthy that they represent a dominant population in seafood. The main spoilage associated species comprise S. liquefaciens, S. grimesii, S. proteamaculans and S. quinivorans, also known as S. liquefaciens-like strains. These species are difficult to discriminate since classical 16S rRNA gene-based sequences do not possess sufficient resolution. In this study, a phylogeny based on the short-length luxS gene was able to speciate 47 Serratia isolates from seafood, with S. proteamaculans being the main species from fresh salmon and tuna, cold-smoked salmon, and cooked shrimp while S. liquefaciens was only found in cold-smoked salmon. The genome of the first S. proteamaculans strain isolated from the seafood matrix (CD3406 strain) was sequenced. Pangenome analyses of S. proteamaculans and S. liquefaciens indicated high adaptation potential. Biosynthetic pathways involved in antimicrobial compounds production and in the main seafood spoilage compounds were also identified. The genetic equipment highlighted in this study contributed to gain further insights into the predominance of Serratia in seafood products and their capacity to spoil.

Quantification of lactic acid bacteria (LAB) is essential to control quality of seafood products like cold-smoked salmon (CSS). In the present study, we report the design and optimization of a dual-labelled TaqMan ™ probe targeting the V7 region of 16S rRNA gene for the detection of LAB in CSS. This quantitative PCR (qPCR) assays is useful for the simultaneous detection of the ten LAB genera communally encountered in CSS as Aerococcus, Carnobacterium, Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Macrococcus, Streptococcus, Vagococcus and Weissella. The specificity of this method was demonstrated against 14 genera (44 isolates, 35 species) of Gram-positive bacteria and 19 genera of Gram-negative (40 isolates, 34 species). Calibration of the method was performed in CSS matrix using a mix of equimolar cultured solution of five LAB. Quantification with the qPCR method range from 3.5 to 8.5 Log CFU/g in CSS matrix, covering 5 orders of magnitude. On these artificially contaminated CSS slices, PCR method results correlated successfully (R2 = 0.9945) with the conventional enumeration on Elliker medium. In addition, the new method was successful on commercial CSS from five different origins with a quantification range from 3.7 Log CFU/g to 8.0 Log CFU/g. This one-step quantitative methodology is proposed as a rapid and complementary tool of the cultural methods to investigate the LAB microbiota and biodiversity of CSS.

Tuna fisheries and processing represent economic activities of paramount importance around the world. Most of these products are traded for human consumption and in general are highly demanded commodities. However, not all tuna products achieve the same market price, some consumers are willing to pay a huge amount of money for certain species (i.e. Japanese market for Bluefin tuna) while other species are rather affordable (i.e. Skipjack tuna), therefore mislabelling has been observed frequently. We collected and analysed 545 tuna samples in six European countries, including fresh, frozen and canned products, and we have investigated whether or not these products were correctly labelled under European and national legislations. We found an overall mislabelling rate of 6.79%; in particular, 6.70% of the fresh and frozen tuna products and 7.84% of canned tuna were mislabelled, and only in the case of fresh and frozen tuna samples significant differences among countries were found. Mislabelling rates for Atlantic Bluefin tuna labelled products were very high, ranging from 50 up to 100%. In general, mislabelling was higher when specific names were included in the labels. The "tuna" umbrella term is a very popular one with consumers, but also one that remains vulnerable to ambiguity, hampering efforts towards market transparency and with potential negative consequences to the adequate management of tuna species stocks.

An accurate amplified fragment length polymorphism (AFLP) method, including three primer sets for the selective amplification step, was developed to display the phylogenetic position of Photobacterium isolates collected from salmon products. This method was efficient for discriminating the three species Photobacterium phosphoreum, Photobacterium iliopiscarium and Photobacterium kishitanii, until now indistinctly gathered in the P. phosphoreum species group known to be strongly responsible for seafood spoilage. The AFLP fingerprints enabled the isolates to be separated into two main clusters that, according to the type strains, were assigned to the two species P. phosphoreum and P. iliopiscarium. P. kishitanii was not found in the collection. The accuracy of the method was validated by using gyrB-gene sequencing and luxA-gene PCR amplification, which confirmed the species delineation. Most of the isolates of each species were clonally distinct and even those that were isolated from the same source showed some diversity. Moreover, this AFLP method may be an excellent tool for genotyping isolates in bacterial communities and for clarifying our knowledge of the role of the different members of the Photobacterium species group in seafood spoilage.

In this study, we confirmed that DNA control region can be used to identify potential spawning area from pelagic juveniles species. We used the mitochondrial DNA control region to identify tuna juveniles predated by common dolphinfish and sampled in surface waters around Reunion Island. These were skipjack juveniles at an early stage (small length and young age). The biological results confirmed the species identification based on DNA analysis. Catch data by longline reveal a higher number of skipjack from October to December around Reunion Island corresponding to the period of our skipjack juvenile sampling.

The sea lamprey Petromyzon marinus, which is among the most phylogenetically ancient vertebrates, is a hematophagous ectoparasite that feeds on vertebrates and is considered vulnerable in Europe but is a pest in the North American Great Lakes. We conducted a literature review of helminth parasites of P. marinus and investigated postmetamorphic lampreys sampled in rivers and northeast Atlantic coastal waters (western France) during spawning migration. Based on the literature review, 16 helminth taxa have been recorded in P. marinus, among them 14 in North America but only 2 in Europe, with no species in common between these areas. Specific parasites are lacking, and helminth parasites recorded in P. marinus are mostly opportunistic and are trophically transmitted to fish hosts with both extremely low prevalence and mean intensity. Thus, P. marinus seems an unusual host that is probably infected through accidental ingestion of parasites by microphagous larvae (ammocoetes) and/or hematophagous postmetamorphs. Our field study supports this hypothesis, since only a single third-stage larva of Anisakis simplex sensu stricto was found in 2 postmetamorphic P. marinus among the 115 individuals dissected. This opportunistic, trophically transmitted, and cosmopolitan nematode species has never been recorded in North American sea lampreys and only once in Galician rivers (southern Europe). Infestation pathways of P. marinus by A. simplex are proposed vis-à-vis the feeding strategy of postmetamorphs and fish host species which potentially harbor anisakid larvae in their musculature. More generally, the complexity of biotic interactions is discussed considering P. marinus both as a host for helminth parasites and as a parasite for hosts such as fish and mammals, which are also potential predators of sea lamprey.

Over the span of a decade, genetic identification methods have progressively exposed the inadequacies of the seafood supply chain, revealing previously unrecognized levels of seafood fraud, raising awareness among the public, and serving as a warning to industry that malpractice will be detected. Here we present the outcome of the latest and largest multi-species, transnational survey of fish labeling accuracy to date, which demonstrates an apparent sudden reduction of seafood mislabeling in Europe. We argue that recent efforts in legislation, governance, and outreach have had a positive impact on industry regulation. Coordinated, technology-based, policy-oriented actions can play a pivotal role in shaping a transparent, sustainable global seafood market and in bolstering healthier oceans.

This study describes a polymerase chain reaction using restriction fragment length polymorphism (PCR-RFLP) assay based on the 16S rRNA mitochondrial gene to identify commercial food products of wide range of Penaeidae and Pandalidae shrimp species commercialised in the Tunisian market.Phylogenetic analyses on 16S rRNA mitochondrial gene were used to study the relationships among the considered species.Penaeidae shrimp species was easily differentiated and confirmed by direct sequencing, showing a genetic distance of 0.34 with respect to Pandalidae species.A rapid and reliable PCR method using restriction fragment length polymorphism (RFLP) with three restriction enzymes (HpyCH4III/ MboI / AluI) was optimized for unambiguous differentiation of shrimp from 19 commercial market samples (raw and processed products).Results showed that the restriction fragment length polymorphism technique can be used to identify Tunisian shrimp species and thus to control not only commercial fraud but also efficient restocking program.

To investigate the ability of microalgae to develop stable, long-term resistance to herbicides, the marine microalga Tetraselmis suecica was exposed to the herbicide diuron (5 μg/L) for a 43-generation exposure period followed by a 12-generation depuration phase. During the first 25 generations, diuron-exposed cultures showed doubling times ranging from 1.95 to 2.6 days, which was 2 to 2.5-fold longer than control cultures. Between generations 25 and 38, during diuron exposure, two out of the three exposed cultures exhibited a spontaneous drop in doubling time. These results provided evidence of culture adaptation to diuron. To assess persistence of the diuron adaptation observed on growth performance, one of the adapted cultures (D3) was maintained for 12 months in unexposed conditions and then tested by a second, short-term exposure to diuron 5 μg/L, in parallel with a control culture (C1) for six generations. Flow cytometry analyses were used to monitor cell density, viability, morphology, relative chlorophyll content and intracellular reactive oxygen species (ROS) level. Under these conditions, diuron induced a strong increase of doubling time in exposed-C1 cultures (2.5-fold longer than unexposed-C1 cultures), but no significant increase occurred in exposed D3-cultures compared with unexposed D3- and unexposed C1-cultures, showing the persistence of adaptation in the previously-exposed strain D3. Intracellular ROS level showed the same trend. Significant differences were observed between these strains, with weaker effects of diuron on strain D3 compared with strain C1: forward scatter (FSC), representing relative cell size, decreased in exposed cultures (67.8% and 95% of the controls for C1 and D3, respectively), whereas FL3 as relative chlorophyll content increased in exposed cultures (115.6% and 108.6% of the controls for C1 and D3, respectively). Results of second exposure to diuron revealed that the adaptation of strain D3 had persisted after 12 months of depuration, as no growth impairment was observed. This study demonstrates the possible appearance of stable diuron resistance in microalgae in cases of strong, multigenerational chronic exposure to this herbicide in polluted environments.

Overlapping external morphometric characters easily confound the flatfishes Solea aegyptiaca and Solea solea (Soleidae) in areas of the Mediterranean Sea where both species live in sympatry. This leads to uncertainties in the fisheries and marketing of the species, in addition to misinterpretations in biogeography and conservation studies. This paper describes a simple restriction fragment length-based diagnostic test that differentiates S. solea from S. aegyptiaca, as well as from other species of the Soleidae family. Furthermore, the two species living in sympatry in the Gulf of Kavala (North Aegean Sea, Greece) present significant qualitative differences in muscle fatty acid composition, a property that can also be used to distinguish the two cryptic species.

Maps of the spawning grounds of commercially important fishes are necessary when assessing the level of connectivity between life stages of fishes and for identifying ecologically valuable marine areas. A first step toward mapping the spawning grounds is a reliable and rapid species identification of pelagic fish eggs to assess the spatio-temporal distribution of spawning aggregations. As many species have similar egg sizes and morphology, the molecular validation of visually identified eggs is often essential for the use of such data in fisheries management. In the present study, we developed a rapid 16S rRNA PCR-restriction fragment length polymorphism (RFLP) assay to distinguish between formalin-fixed fish eggs of dab Limanda limanda, flounder Platichthys flesus and pout Trisopterus spp., which were collected during the 2008 International Bottom Trawl Survey in the Eastern English Channel and southern North Sea. A comparison of the rapid 16S rRNA PCR-RFLP method with initial visual identification revealed 93% of correct identifications for dab, 90% for pout, but only 64% for flounder, representing an overall error rate of 17%. Visual misidentification occurred mainly between dab and flounder and between flounder and pout. Egg abundance and the relative proportions of each species were subsequently analysed geostatistically. Molecular identifications were incorporated to obtain corrected interpolated distribution maps, taking into account the results from molecular identifications as a correction factor. This highlighted the distinct spawning grounds for the 3 studied taxa and facilitated the identification of regions of high conservation value for these species.

A better knowledge on spawning grounds of principal commercial fishes in the North Sea is necessary for their monitoring. Fish egg taxonomic identification based on morphological characters cannot always be carried out effectively. This is particularly the case for the eggs of a number of species of the Gadidae (i.e. cod, haddock and whiting and three rockling species) which have the same range of size and for which molecular-based techniques can help improve the accuracy of taxonomic identification of eggs, and hence the reliability of studies on spawning areas. In this study, formalin fixation, egg conservation and DNA extraction were optimized in order to develop a specific polumerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Based on the sequence of the cytochrome b gene of reference specimens, a PCR-RFLP method was developed to distinguish eggs of cod, haddock and whiting on the one hand, and between eggs of three species of rocklings on the other. Among the 404 eggs tested, 80% of positive amplification was obtained for rocklings eggs and 86.36% for Gadidae eggs. Eggs of Enchelyopus cimbrius, Ciliata mustela and Ciliata septentrionalis, and Gadus morhua, Merlangius merlangus and Melanogrammus aeglefinus were distinguished with this method. Based on morphological characters, the percentage of misidentification was less for cod than for whiting.

Traceability in the fish food sector plays an increasingly important role for consumer protection and confidence building. This is reflected by the introduction of legislation and rules covering traceability on national and international levels. Although traceability through labeling is well established and supported by respective regulations, monitoring and enforcement of these rules are still hampered by the lack of efficient diagnostic tools. We describe protocols using a direct sequencing method based on 212-274-bp diagnostic sequences derived from species-specific mitochondria DNA cytochrome b, 16S rRNA, and cytochrome oxidase subunit I sequences which can efficiently be applied to unambiguously determine even closely related fish species in processed food products labeled "anchovy". Traceability of anchovy-labeled products is supported by the public online database AnchovyID (http://anchovyid.jrc.ec.europa.eu), which provided data obtained during our study and tools for analytical purposes.

This report describes a set of 21 polymerase chain reaction primers and amplification conditions developed to barcode practically any teleost fish species according to their mitochondrial cytochrome b and nuclear rhodopsin gene sequences. The method was successfully tested in more than 200 marine fish species comprising the main Actinopterygii family groups. When used in phylogenetic analyses, its combination of two genes with different evolutionary rates serves to identify fish at the species level. We provide a flow diagram indicating our validated polymerase chain reaction amplification conditions for barcoding and species identification applications as well as population structure or haplotyping analyses, adaptable to high-throughput analyses.

The identification, by SDS-PAGE, of four red algae used as sea vegetables or ingredients by the food industry, was performed. They were Palmaria palmata (Dulse), Chondrus crispus (Pioca), Porphyra umbilicalis (Nori), Gracilaria verrucosa (Ogo-nori). For each species, variations in protein patterns were observed, according to the season. However, for all species, some protein bands were always present during the yearly cycle of the plant. The reference pattern of P. palmata was composed of six protein bands with apparent molecular weights between 59.6 and 15.2 kDa. The G. verrucosa pattern was constituted of eight permanent bands. Two pattern bands, with apparent molecular weights of 49.1 and 45.9 kDa,differentiated the G. verrucosa profile from other seaweed patterns. C. crispus could be identified by a reference pattern composed of seven bands; three, with close molecular weights (49.3; 46.2 and 43.2 kDa), were characteristic of this species. Finally, the P. umbilicalis pattern showed seven bands with molecular weights between 73.1 and 15.9 kDa. The presence of a band with a molecular weight above 70 kDa appeared to be specific to the Porphyra pattern. So the SDS-PAGE seemed able to identify the four red species used by the food industry, but this analytical method appeared to be applicable only to raw material dried in mild conditions.

SDS PAGE was tested as an analytical tool for the identification of fourgreen algae (Ulva rigida, U. rotundata, Enteromorphaintestinalis, E. compressa) used as food ingredients. A referencepattern composed of the bands present all year long was performed foreach species. The pattern for Ulva rotundata consists of 7 bandslocated between 69.9 and 15.5 kDa with the presence of triplicate bandsat 29.5, 26.3 and 22.9 kDa. The pattern for Ulva rigida is consistsof three bands with apparent molecular weights of 68.5, 56.4 and 44.7kDa. The Enteromorpha compressa pattern is characterised by sixbands located between 65.8 and 19.8 kDa. A double band withmolecular weights of 23.1 and 23.9 distinguished this pattern from theothers. Six bands situated between 66.4 and 19.4 kDa with a specifictriplicate band of 25.9, 23.9 and 22.5 kDa constituted the specific patternof Enteromorpha intestinalis. SDS PAGE appears to be suitable forthe identification of green seaweed foods.

Three electrophoretic techniques, isoelectric focusing (IEF), sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and urea-IEF were used for species identification of "baby-clams" from Vietnam (Meretrix lyrata), grooved carpet shell (Ruditapes decussatus) and the common edible cockle (Cerastoderma edule). Analysis were performed on proteins extracted from different parts of the mollusks, adductor muscle. mantle, foot, of raw and cooked samples. The electrophoretic patterns of adductor muscle were the most discriminative. Although all three methods tested could be used for bivalve species identification, the electrophoretic profiles with IEF were modified after cooking, whereas those with SDS-PAGE or urea-IEF were identical in raw and cooked samples and thus more suitable for sample authentication.