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The potentially toxic genus Pseudo-nitzschia is constantly present in phytoplankton community in the Black Sea often proliferating to bloom concentrations. Production of domoic acid (DA) has been confirmed by local P. calliantha strain and the presence of the neurotoxin has been detected in cultured molluscs and plankton samples from the Bulgarian Black Sea coast. This study aims to provide an overview of the long-term trends of Pseudo-nitzschia blooms in the Black Sea based on inventory of the available information for the period 1959-2019 and assess the relation with key environmental variables. Pseudo-nitzschia species diversity and identification constraints are critically analyzed along with data about the presence of DA in the basin. The results demonstrate that Pseudo-nitzschia spp. may bloom at any time of the year under different environmental regimes. Application of advanced identification techniques, technologically adequate regular monitoring, complemented by toxin analysis is essential to improve our understanding of Pseudo-nitzschia bloom dynamics in order to protect public and ecosystem health hazards.

Background The Black Sea is the largest brackish water body in the world, although it is connected to the Mediterranean Sea and presents an upper water layer similar to some regions of the former, albeit with lower salinity and temperature. Despite its well-known hydrology and physicochemical features, this enormous water mass remains poorly studied at the microbial genomics level. Results We have sampled its different water masses and analyzed the microbiome by shotgun and genome-resolved metagenomics, generating a large number of metagenome-assembled genomes (MAGs) from them. We found various similarities with previously described Black Sea metagenomic datasets, that show remarkable stability in its microbiome. Our datasets are also comparable to other marine anoxic water columns like the Cariaco Basin. The oxic zone resembles to standard marine (e.g. Mediterranean) photic zones, with Cyanobacteria (Synechococcus but a conspicuously absent Prochlorococcus), and photoheterotrophs domination (largely again with marine relatives). The chemocline presents very different characteristics from the oxic surface with many examples of chemolithotrophic metabolism (Thioglobus) and facultatively anaerobic microbes. The euxinic anaerobic zone presents, as expected, features in common with the bottom of meromictic lakes with a massive dominance of sulfate reduction as energy-generating metabolism, a few (but detectable) methanogenesis marker genes, and a large number of “dark matter” streamlined genomes of largely unpredictable ecology. Conclusions The Black Sea oxic zone presents many similarities to the global ocean while the redoxcline and euxinic water masses have similarities to other similar aquatic environments of marine (Cariaco Basin or other Black Sea regions) or freshwater (meromictic monimolimnion strata) origin. The MAG collection represents very well the different types of metabolisms expected in this kind of environment. We are adding critical information about this unique and important ecosystem and its microbiome.

Seas and oceans are a global reservoir of antibiotic resistance genes (ARGs). Only a few studies investigated the dynamics of ARGs along the water column of the Black Sea, a unique environment, with a peculiar geology, biology and history of anthropogenic pollution. In this study, we analyzed metagenomic data from two sampling campaigns (2013 and 2019) collected across three different sites in the Western Black Sea at depths ranging from 5 to 2000 m. The data were processed to annotate ARGs, metal resistance genes (MRGs) and integron integrase genes. The ARG abundance was significantly higher in the deep water layers and depth was the main driver of beta-diversity both for ARGs and MRGs. Moreover, ARG and MRG abundances strongly correlated (r = 0.95). The integron integrase gene abundances and composition were not influenced by the water depth and did not correlate with ARGs. The analysis of the obtained MAGs showed that some of them harbored intI gene together with several ARGs and MRGs, suggesting the presence of multidrug resistant bacteria and that MRGs and integrons could be involved in the selection of ARGs. These results demonstrate that the Black Sea is not only an important reservoir of ARGs, but also that they accumulate in the deep water layers where co-selection with MRGs could be assumed as a relevant mechanism of their persistence.

One of the assets, assigned to the phytoplankton resting stages, is that of serving as the “memory” of the aquatic ecosystems and preserved biodiversity in the course of time. However, an accurate cyst identification proves to be a more difficult and extremely challenging process, even today. In order to gain a better taxonomic coverage of cyst assemblages in the Black Sea, an integrated approach of the classical morphological identification with metabarcoding methods (MySeq sequencing of V7-V9 regions of the 18S rDNA) was applied on thirteen surface sediment samples collected from different sites. A total number of 112 dinoflagellate taxa was detected at the species level and ascribed to 51 genera. In general, it is the molecular analysis that yields a higher number of taxa as compared to those obtained through the morphological taxonomy (66 taxa based on the DNA sequences versus 56 morphologically-identified taxa). Besides, it should be pointed out that the integrated dataset includes 14 potentially toxic dinoflagellate species. Discerned, subsequently, was a good dataset consistency for ten species, followed by some discrepancies as to a number of taxa, identified with one of the methods only, due to specific methodological biases. On the whole, it could be concluded that the combination of morphological and molecular methods is likely to increase the potential for a more reliable taxonomic assessment of phytoplankton diversity in marine sediments which, in turn, proves conclusively the utmost importance of the integrated approach.

The study focuses on the impact of life excretion and mucus released by the "biological pollutants" invasive ctenophore Mnemiopsis leidyi and its predator Beroe ovata on the marine environment and lower trophic levels of the Black Sea ecosystem (bacteria, pico-phytoplankton, nano-autotrophic/heterotrophic flagellates, micro-phytoplankton, chlorophyll a, primary production (PP), micro-zooplankton). The chemical and biological variables were analysed in two sets of lab experiments with natural communities from mesotrophic (Gelendzhik) and eutrophic (Varna) coastal waters. While both species altered the chemical properties of experimental media, exerting structural and functional changes in the low food-web biological compartments, the results showed a stronger effect of B. ovata, most likely related to the measured higher rate of excretion and amount of released mucus. In addition the alterations in the Gelendzhik experiment were more pronounced, indicating that environmental implications on lower food-web are more conspicuous in mesotrophic than in eutrophic coastal waters.

The Black Sea is the largest meromictic sea with a reservoir of anoxic water extending from 100 to 1000 m depth. These deeper layers are characterised by a poorly understood fluorescence signal called "deep red fluorescence", a chlorophyll a-(Chl a) like signal found in deep dark oceanic waters. In two cruises, we repeatedly found up to 10(3) cells ml(-1) of picocyanobacteria at 750 m depth in these waters and isolated two phycoerythrin-rich Synechococcus sp. strains (BS55D and BS56D). Tests on BS56D revealed its high adaptability, involving the accumulation of Chl a in anoxic/dark conditions and its capacity to photosynthesise when re-exposed to light. Whole-genome sequencing of the two strains showed the presence of genes that confirms the putative ability of our strains to survive in harsh mesopelagic environments. This discovery provides new evidence to support early speculations associating the "deep red fluorescence" signal to viable picocyanobacteria populations in the deep oxygen-depleted oceans, suggesting a reconsideration of the ecological role of a viable stock of Synechococcus in dark deep waters.

The Black Sea is an extraordinarily complex water body for ocean color remote sensing, as it belong to Case 2 waters, which are characterized by relatively high absorption by Colored Dissolved Organic Matter (CDOM) while the concentration of non-pigmented particulate matter does not co-vary in a predictable manner with chlorophyll a . The optical complexity of the Black Sea is the reason why the standard bio-optical algorithms developed for Case 1 waters, are the source of large uncertainties (of the order of hundreds of percent) of chlorophyll a concentration in the coastal and shelf regions. In the framework of ESA contract “BIO-OPTICS FOR OCEAN COLOR REMOTE SENSING OF THE BLACK SEA - Black Sea Color” we developed empirical ocean color algorithm for chlorophyll a retrieval from Sentinel 3A/OLCI primary ocean color products using the in situ reference bio-optical datasets collected in the Black Sea in the period 2012-2019. Results obtained from the assessment of operational S3A/OLCI chlorophyll products, highlighted and confirmed that the specific regional algorithm is essential for the Black Sea. The coefficients of the regional algorithm were derived from the regression of log-transformed pigment concentrations and remote sensing reflectance ratio at 490nm and 560 nm with determination coefficient R2 =0.88 and number of samples N=186. The algorithm predicts chlorophyll a values using a cubic polynomial formulation. The result of assessment of the regional chlorophyll a product against independent in situ measurements from the data utilized for algorithm development, showed relatively high accuracy (31.7%), fewer underestimations (MPD=-9.2%) and a good agreement (R2=0.66) between datasets indicating that the regional algorithm is more effective in reproducing the pigment concentration in the Black Sea waters in comparison to the standard Sentinel 3A/OLCI algorithms. Our analysis revealed the importance of providing regional algorithms strictly required to suit the peculiar bio-optical properties featuring this basin. However, this requires collection of accurate in situ measurements in the different parts of the Black Sea. The validity of the reported empirical algorithm obviously depends on the size of the dataset used for its development. The Black Sea waters vary at a basin level due to the sub-regional features, environmental factors and seasonal variability, consequently the presented regional algorithm might have a limited generalization capability. Clearly, more in situ data with improved spatial and temporal coverage are critically needed for further calibration and validation of the ocean color products in the Black Sea.

Background: The Black Sea is the largest brackish water body in the world, although it is connected to the Mediterranean Sea and presents an upper water layer similar to some regions of the former albeit with lower salinity and (mostly) temperature. In spite of its well-known hydrology and physico chemistry, this enormous water mass remains poorly studied at the microbial genomics level. Results: We have sampled its different water masses and analyzed the microbiome by classic and genome-resolved metagenomics generating a large number of metagenome-assembled genomes (MAGs) from them. The oxic zone presents many similarities to the global ocean while the euxinic water mass has similarities to other similar aquatic environments of marine or freshwater (meromictic monimolimnion strata) origin. The MAG collection represents very well the different types of metabolisms expected in this kind of environments and includes Cyanobacteria (Synechococcus), photoheterotrophs (largely with marine relatives), facultative/microaerophilic microbes again largely marine, chemolithotrophs (N and S oxidizers) and a large number of anaerobes, mostly sulfate reducers but also a few methanogens and a large number of “dark matter” streamlined genomes of largely unpredictable ecology. Conclusions: The Black Sea presents a mixture of similarities to other water bodies. The photic zone has many microbes in common with that of the Mediterranean with the relevant exception of the absence of Prochlorococcus. The chemocline already presents very different characteristics with many examples of chemolithotrophic metabolism (Thioglobus) and facultatively anaerobic microbes. Finally the euxinic anaerobic zone presents, as expected, features in common with the bottom of meromictic lakes with a massive dominance of sulfate reduction as energy generating metabolism and a small but detectable methanogenesis.We are adding critical information about this unique and important ecosystem and its microbiome.

BackgroundThe Black Sea is the largest brackish water body in the world, although it is connected to the Mediterranean Sea and presents an upper water layer similar to some regions of the former albeit with lower salinity and (mostly) temperature. In spite of its well-known hydrology and physico chemistry, this enormous water mass remains poorly studied at the microbial genomics level.ResultsWe have sampled its different water masses and analyzed the microbiome by classic and genome-resolved metagenomics generating a large number of metagenome-assembled genomes (MAGs) from them. The oxic zone presents many similarities to the global ocean while the euxinic water mass has similarities to other similar aquatic environments of marine or freshwater (meromictic monimolimnion strata) origin. The MAG collection represents very well the different types of metabolisms expected in this kind of environments and includes Cyanobacteria (Synechococcus), photoheterotrophs (largely with marine relatives), facultative/microaerophilic microbes again largely marine, chemolithotrophs (N and S oxidizers) and a large number of anaerobes, mostly sulfate reducers but also a few methanogens and a large number of “dark matter” streamlined genomes of largely unpredictable ecology.ConclusionsThe Black Sea presents a mixture of similarities to other water bodies. The photic zone has many microbes in common with that of the Mediterranean with the relevant exception of the absence of Prochlorococcus. The chemocline already presents very different characteristics with many examples of chemolithotrophic metabolism (Thioglobus) and facultatively anaerobic microbes. Finally the euxinic anaerobic zone presents, as expected, features in common with the bottom of meromictic lakes with a massive dominance of sulfate reduction as energy generating metabolism and a small but detectable methanogenesis.We are adding critical information about this unique and important ecosystem and its microbiome.

MASRI – Infrastructure for Sustainable Development of Marine Research Including the Participation of Bulgaria in the European Infrastructure Euro-Argo is a project of the National roadmap for scientific Infrastructure (2017-2023) of Bulgaria. The mission of MASRI is to build and utilize a modern research infrastructure which will provide the basis for highly efficient marine and maritime research to expand our knowledge of the marine environment and to support blue growth and implementation of marine policy and maritime spatial planning in order to achieve UN Sustainable Development Goal 14: Conservation and sustainable use of oceans, seas and marine resources for sustainable development.MASRI activities include the modernization of existing unique resources and equipment and the establishment of new facilities. The research infrastructure consists of four main modules: Research fleet; National Operational Marine Observing System – NOMOS; Data and information center and Research laboratory complex, each representing a distinct on functional basis part of the scientific infrastructure, and consists of separate components distributed physically in different scientific organizations, in the city of Varna. Thus, MASRI is intended to be a large-scale, interdisciplinary multifunctional (physics, chemistry, biology, geology, aquacultures, medicine, energy, underwater, and offshore technologies) marine research infrastructure of scientific significance and will provide unique facilities (including databases and computer network) which will be widely accessible on national, regional and international level for multidisciplinary researches.Research vessels are intended to provide access to the investigated medium – the sea and they are providing a working platform for conducting research. NOMOS is a system of systems to measure in situ parameters of the marine environment and the surrounding atmosphere. It is designed to provide information on the state of the marine environment for scientific research, forecasting and marine industry. Data and information center provide a computing environment, communication environment and environment for quality control and reliable storage of data and information within the scientific infrastructure. Research laboratory Complex represents a system of research laboratories for chemical, biological and geological analyzes and for relevant research on marine medicine as well as of laboratories for marine resources and technologies research.As an important module of MASRI, NOMOS includes several components: BulArgo – a system of profiling floats to measure the profiles of the characteristics of the marine environment in the depth up to 2000m; waves and currents monitoring system; national sea level observing system; moorings network; coast research bases and metrological control laboratory.MASRI is also intended to support the participation of Bulgaria in European research infrastructure consortia Euro-Argo ERIC. Al least three floats are provided and launched in the Black sea every year in the frame of the BulArgo project. Thus, BulArgo gives an important contribution to the Argo program in particular in the Black sea, providing a significant volume of very important in-situ data both for climatic research, for assimilation into the models and verification of the forecasts.

Antibiotic resistance genes (ARGs) and antibiotic resistant bacteria (ARB) are worldwide considered as emerging contaminants of large interest, and a primary threat to human health. It is becoming clear that the environment plays a central role in the transmission, spread, and evolution of antibiotic resistance. Although marine systems have been largely investigated, only a few studies have considered the presence of ARGs in meso- and bathypelagic waters. To date, no molecular based studies have yet been made to investigate the occurrence of ARGs in the Black Sea, the largest meromictic basin in the world, receiving water from a number of important European rivers and their residues of anthropogenic activities in permanently stratified mesopelagic water masses. In this study, we determined the presence and the abundance of five ARGs (blaCTXM, ermB, qnrS, sul2, tetA) and of the heavy metal resistance gene (HMRG) czcA, in different sampling sites in the eastern and western Black Sea, at several depths (up to 1000 m) and various distances from the shoreline. Three ARGs (blaCTXM, sul2, and tetA) and czcA were present in at least 43% of the analysed samples, whereas ermB and qnrS were never detected. In particular, sul2 abundances increased significantly in coastal location, whereas tetA increased with sampling depth. These findings point out the Black Sea as a source of ARGs and HMRGs distributed along the whole water column.

International audience; Autonomous Reef Monitoring Structures (ARMS) have been applied worldwide to characterize the critical yet frequently overlooked biodiversity patterns of marine benthic organisms. In order to disentangle the relevance of environmental factors in benthic patterns, here, through standardized metabarcoding protocols, we analyze sessile and mobile (

Bio-optical data, obtained during six cruises in the Black Sea carried out during periods of seasonal stratification in years between 1996 and 2016, have been used to parametrize phytoplankton light absorption ($${a_{ph}}\left({\rm{\lambda }} \right)$$) in the deep chlorophyll maximum (DCM) layer located near the bottom of euphotic zone. Relationships between $${a_{ph}}\left({\rm{\lambda }} \right)$$ and the sum of chlorophyll-a and phaeopigment concentrations (Chl-a) differed from those for the summertime upper mixed layer (UML). Notably, chlorophyll a specific absorption coefficients ($$a_{ph}^\ast\left(\lambda \right)$$) were lower in the DCM and more comparable with $$a_{ph}^\ast\left(\lambda \right)$$ values typical for winter phytoplankton in the Black Sea. The $${a_{ph}}\left({\rm{\lambda }} \right)$$ spectral shapes in the DCM differed markedly from those in winter and in the summer UML, due to a shoulder at ~490 nm and a local maximum at ~550 nm corresponding to the absorption bands of phycourobilin and phycoerythrobilin. Light absorbing properties of phytoplankton in the DCM (amplitude and spectral shape of $$a_{ph}^\ast\left(\lambda \right)$$) reflected physiological acclimation to local conditions on the cellular level and population shifts leading to changes in the biomass-dominant species, with Synechococcus spp. domination in the DCM. The parameterization of phytoplankton absorption in the DCM will enable refined spectral models of the downwelling radiance and primary production in the Black Sea.

In this study the plankton diversity in 13 environmental samples from Varna Bay (in the western Black Sea) was analyzed using massively parallel sequencing (MPS). This preliminary study was undertaken to assess the potential of this technology for future implementation in monitoring programs in the Black Sea. Amplicon sequences of the 18S rRNA gene (V4-5 regions) were obtained using the Illumina MiSeq 250PE platform. A total of 1137 operational taxonomic units (OTUs) were obtained among which 242 OTUs with >0.990 BLAST top hit similarity (21.3% of all detected OTUs) closely related to sequences belonging to −protists. A large portion (175 OTUs = 72.3%) was identified at the species levels, including species typical for the Bulgarian Black Sea plankton community, as well as many that haven’t been reported earlier in the Bulgarian Black Sea coast (124 OTUs = 51.2%). Dinoflagellates were represented by the highest species number (77 OTUs comprising 31.8% of protist species), with dominant genera Gyrodinium and Heterocapsa . The present survey revealed the presence of 12 species listed as harmful, some of which have been previously overlooked, such as Cochlodinium polykrikoides , Karenia bicuneiformis , and Karlodinium veneficum . Species identification was possible for 10.3–36.0% of the detected OTUs in the six major supergroups. The frequency in Rhizaria was significantly lower than that in other major groups ( p 0.05–0.01), implying difficulties in the classification from morphology-based observations. The metagenetic data had an insufficient resolution of the 18S rRNA gene for species identification in many genera. These issues may hamper the implementation of MPS-based surveys for plankton monitoring, especially for detecting harmful algal blooms (HAB). The sequencing technology is steadily improving and it is expected that sequence length and quality issues will be resolved in the near future. The ongoing efforts to register taxonomic information and quality controls in the international nucleotide sequence databases (INSDs) will be essential for improving taxonomic identification power.

Size spectra exhibit common patterns of variation and predictable responses to pressures across ecosystem types, functional guilds and taxonomic groups. Here, we extend the size spectra approach to phytoplankton ecological status assessment in transitional waters by developing, testing and validating a multi-metric index of size spectra sensitivity (ISS-phyto), which integrates size structure metrics with others such as phytoplankton diversity, biomass and sensitivity of size classes to anthropogenic disturbance. The ability of various theoretical models of size spectra sensitivity to discriminate between disturbed and undisturbed ecosystems and levels of anthropogenic stress was evaluated. We used data on phytoplankton samples collected in 14 Mediterranean and Black sea transitional water ecosystems (coastal lagoons) from Italy, Albania, Greece, Bulgaria and Romania, and compared the models’ efficiency by looking at their pressure–impact response along salinity and enrichment gradients, the latter quantified as variations in dissolved inorganic nitrogen (DIN) and dissolved inorganic phosphorus (DIP). Data from a fifteenth Mediterranean lagoon was used for external validation purposes. Right asymmetric models of size class sensitivity, implying higher sensitivity of smaller cell size classes, were found to contribute to the ISS-phyto multimetric tool more effectively than symmetric and left asymmetric models, distinguishing disturbed from undisturbed lagoons and disturbed from undisturbed stations within the same lagoon. When based on right asymmetric sensitivity models, i.e., those that were most efficient in identifying anthropogenic impacts, ISS-phyto also showed the best fit of pressure–response relationships along the salinity and enrichment gradients; at low to high levels of impact ISS response was driven by size class sensitivity and at very high impacts by phytoplankton biomass. A scheme for the classification of Ecological Quality Status based on ISS-phyto is proposed and validated. The validation procedure found that ISS-phyto is an effective and sensitive monitoring tool, robust, easy to apply and to inter-calibrate among laboratories.

To estimate the role of atmospheric deposition on Black Sea biogeochemistry, the chemical composition (particulate matter, carbon, major and trace metals) of both atmospheric deposition and sediment traps material was studied. Lithogenic material was found to be the most abundant constituent in sediment traps (31–34%) followed by carbonates (CC; 21%) and particulate organic carbon (POC; 10%). Particulate matter fluxes in sediment traps were low from January to March with maxima from November to December for both depths (930 m and 1930 m). Particulate organic carbon and lithogenic material present common peaks with particulate matter, indicating that during blooms, phytoplankton produces aggregates and sweep from the surface both lithogenic and anthropogenic elements which are transported rapidly to the deep. Atmospheric deposition was found to supply quantitatively a considerable amount of major and trace metals to the seawater of the Black Sea. More precisely, on average from the surface towards the deeper water layers of the Black Sea 26% and 83% of Al, V, Cr, Mn, Fe, Ni, Cu, Zn, Cd and Pb fluxes were provided by atmospheric deposition, demonstrating its significant role as an external source of major and trace metals in the area.

Patterns of phytoplankton size spectra variation with gradients of environmental stress have been observed in freshwater, transitional waters and marine ecosystems, driving the development of size spectra based assessment tools. In this study, we have tested on transitional and coastal waters a new Index of Size spectra Sensitivity of Phytoplankton (ISS-Phyto), which integrates simple size spectra metrics, size class sensitivity to anthropogenic disturbance, phytoplankton biomass (chlorophyll a) and taxonomic richness thresholds. ISS-Phyto has been tested both among and within ecosystems along pressure gradients based on expert view assessment; the adequacy of symmetric and both left and right asymmetric models of phytoplankton size class sensitivity have been compared. The results showed that ISS-Phyto consistently discriminated between anthropogenic and natural disturbance conditions. Left asymmetric models of size spectra sensitivity, assuming greater disturbance tolerance with respect to eutrophication and organic enrichment of increasingly large size classes, showed the best fit comparing all ecosystems; in three of the four considered ecosystems (Varna, Helsinki, Mompas-Pasaia), they seemed to discriminate best between different levels of disturbance also within ecosystems. Moreover, they demonstrated significant and inverse patterns of variation along the overall pressure gradient as well as along the inorganic phosphorus (DIP), chlorophyll a and trophic index (TRIX) gradients. Therefore, ISS-Phyto, originally developed for transitional waters, seems to be an adequate assessment tool of ecological status also in coastal marine waters; moreover, it seems adequate to describe within ecosystem disturbance gradients. Hence, ISS-Phyto helps to understand the relationships between anthropogenic impact and ecosystem response from the individual point of view, with reference to the simple parameter of body size.

Abundance and carbon biomass of different phytoplankton groups obtained by microscopy were compared with taxonomy derived from pigment measurements and CHEMTAX analysis of samples collected in June 2006 in the NW Black Sea. The diatom Chaetoceros curvisetus was dominant in terms of carbon biomass based on cell volume at inshore stations, while the coccolithophore Emiliania huxleyi was prevalent at offshore. Emiliania huxleyi reached bloom abundance of 3.3 × 106 cells L− 1. The chlorophyll a (chl a) concentration within phytoplankton groups as allocated by CHEMTAX was in agreement with microscopy derived carbon biomasses of the taxonomic groups diatoms, dinoflagellates and cryptophytes only. Carbon biomass of less abundant phytoplankton taxa (cyanophytes, euglenophytes and chlorophytes) did not correlate with group-specific chl a. It was not possible to detect E. huxleyi bloom by CHEMTAX analysis probably due to much higher biomass of other species containing 19’-hexanoyloxyfucoxanthin. Nutrient concentrations were generally high in the waters where diatom and dinoflagellates dominated the community but low in the area of E. huxleyi bloom. A good correlation between total carbon biomass of phytoplankton and chl a was found and the estimated C:chl a ratio of phytoplankton varied between 36 and 256 (in average 124 ± 50).

The aim of this investigation was to examine biochemical differences in the gastropod Rapana venosa (R. venosa) from polluted (RvP) and nonpolluted (RvN) sites of the Black Sea's Bulgarian coast that may serve as bioindicators of environmental quality. Mussels Mytilus galloprovincialis (MMg) were collected from polluted (MMgP) and nonpolluted (MMgN) sites at the same time for comparison. Bioactive compounds and the antioxidant activity of dry matter (DM) methanol extracts from RvP and RvN were determined. Three-dimensional fluorometry (3D-FL) and Fourier transform infrared (FTIR) analyses were used to evaluate polyphenols and proteins. DM methanol extracts from RvP, and to lesser extent RvN, showed high amounts of total phenolics (23.22±2.9 and 15.60±1.8 mg GAE · g−1 DW) and exhibited high levels of antioxidant activity in some radical scavenging assays (μ MTE · g−1 DW): 56.38±5.2 and 33.79±3.3 by 2,2azino-bis (3-ethyl-benzothiazoline-6-sulfonic acid) diamonium salt (ABTS·+); 54.79±5.6 and 33.7±2.6 by cup...

The polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs) and organotins were analyzed in mussels Mytilus galloprovincialis from polluted and unpolluted sites from Mokpo Bay, Korea. The total PAH’s concentrations (10 � 3 mg kg � 1 ) measured by GC–MS were in the range from 31 ± 23 to 1 ± 1. Among the eight PAHs the predominant ones were fluoranthene, phenanthrene and pyrene and accounted approximately 63% of the total PAHs. Among the four detected PCBs the highest content was of PCB 153, which accounted about 47% of the total PCBs. The main organotin compounds were dibutyltindichloride (DBT) and tributyltinchloride (TBT) and their composition was approximately 33% and 24%. PAHs, PCBs and organotins were found only in the mussels from polluted site. The antioxidant activity by ABTS [2,2 0 -azinobis (3-ethylbenzothiazoline-6-sulfonic acid)] test was higher in mussels from polluted than from unpolluted sites (P < 0.05). It was found a correlation between the determined compounds (PAHs, PCBs and organotins) and the antioxidant activity of the mussel tissue from polluted site and the correlation coefficients were 0.96, 0.92 and 0.80, respectively. Such correlation can be explained by the properties of mussels. Since the mussel cell wall and tissues are hydrophobic, they can concentrate a number of hydrophobic pollutants like PAHs and PCBs from the marine environment by solubility rules. On the other hand, proteins are lipophilic compounds having antioxidant properties. Certain amino acid residues and thiol (-SH) groups, contained in proteins, respond to the ABTS antioxidant activity assay. Thus there may be a correlation between the total antioxidant activity of the organism and the PAH–PCB pollutants which were concentrated from its environment. The studied properties of mussels from polluted site can be used as an additional indicator of pollution.

1. Leaf litter decomposition rates in aquatic ecosystems are known to be related to many abiotic and biotic factors. 2. Field experiments were carried out during spring 2005 in 16 ecosystems, each with four sampling sites, using the litter bag technique to investigate the influence of abiotic factors on patterns of reed litter breakdown in different physiographic, hydrological and physico-chemical gradients occurring in transitional water ecosystems in the Eastern Mediterranean and Black Sea. 3. Significant differences in leaf litter decomposition were observed among the studied ecosystems along univariate gradients of tidal range, water temperature, salinity and sinuosity index. 4. Overall, 71% of variance in the litter breakdown rate was explained by the hydrological, physico-chemical and physiographic components. Specifically, tidal range, salinity and sinuosity index are among the key factors in the most commonly used typological schemes for classifying transitional water ecosystems (i.e. Confinement Concept and Venice System), due to their influence on abundance and distribution of benthic macroinvertebrates and other guilds. 5. The patterns observed at the regional scale of the study suggest that certain key abiotic factors are likely to play a major role as drivers of plant detritus decomposition processes, through their influence on the overall metabolism of microorganisms and benthic macroinvertebrates. 6. These observations have implications for the identification of reference conditions for transitional water ecosystems in the studied area, on which all processes of classification and conservation of their ecological status are based.

1. A large number of ecological indicators have been applied to environmental quality assessment, including taxonomic indices based on one or more biotic elements and eventually environmental variables. Several structural parameters of benthic invertebrate assemblages (i.e. diversity, abundance and proportion of disturbance-sensitive taxa) have been included in the European Water Framework Directive (WFD) to rank the ecological status of the transitional waters. 2. Several macrobenthic biotic indices based on species diversity and sensitivities to anthropogenic disturbance, were proposed as effective tools to answer the requirements of the WFD, however, little is known about the responses of these indices under differing ecological conditions. 3. In the present study patterns of variation of five benthic biotic indices (AMBI, M-AMBI, BENTIX, Engle’s B-IBI, and Paul’s B-IBI) have been compared in eight Mediterranean and Black Sea transitional water ecosystems characterized by different habitat typologies and anthropogenic pressures. 4. Overall, biotic indices provided contrasting responses in terms of environmental quality assessments. Both the Engle and Paul multimetric benthic biotic indices of integrity were developed for North American estuaries and therefore require to be recalibrated for the Mediterranean and Black Sea transitional waters. M-AMBI applied to each habitat typology promises to be more effective, but the high heterogeneity of transitional waters and the lack of reference condition along the European coasts limit its application. Further work is needed to develop a biotic index that provides useful and unambiguous information under different anthropogenic pressures and in different transitional waters.

Long-term trends (1960se1990s) in surface winter nutrients and summer oxygen concentration of the euphotic zone, as well as seasonal and interannual variability in surface chlorophyll a (chl a) and Secchi depth (Zd) were investigated for different shelf regions (depth

Crise, A. ... et. al.-- 12 pages, 1 figure, 5 tables, PERSEUS project aims to identify the most relevant pressures exerted on the ecosystems of the Southern European Seas (SES), highlighting knowledge and data gaps that endanger the achievement of SES Good Environmental Status (GES) as mandated by the Marine Strategy Framework Directive (MSFD). A complementary approach has been adopted, by a meta-analysis of existing literature on pressure/impact/knowledge gaps summarized in tables related to the MSFD descriptors, discriminating open waters from coastal areas. A comparative assessment of the Initial Assessments (IAs) for five SES countries has been also independently performed. The comparison between meta-analysis results and IAs shows similarities for coastal areas only. Major knowledge gaps have been detected for the biodiversity, marine food web, marine litter and underwater noise descriptors. The meta-analysis also allowed the identification of additional research themes targeting research topics that are requested to the achievement of GES. © 2015 The Authors, This work has been partially funded by the EC FP7 PERSEUS Project (Grant. Agr. 287600)

Characterisation of phytoplankton communities is important for classification of the ecological status of marine waters. In order to design a monitoring programme, it is important to know what degree of variation in the measurements occur at each level (water body, station and sample), so that resources can be spent in a way that maximise the precision of the measured parameters. Seven European water bodies were sampled to assess the variation in pigment concentrations and population densities attributed to water body, station and sample levels. It was found that the main proportion of the variation between pigment measurements was explained by the variation between stations (12-91% of variation) followed by the variation between water bodies (0-89% of variation). For measurements of population density, the main proportion of the variation between densities of cells recorded was explained by the variation between the taxonomists counting the samples (61%), whilst the main proportion of the variation between numbers of taxa recorded was explained by the variation between water bodies (83%). When the cell density of the nine dominant classes were analysed separately, the main proportion of variation was explained at the water body level for all but two class. © 2012 Springer Science+Business Media Dordrecht.

In the recent years, the consumption of seafood has increased. There are no results on the studies of Rapana venosa (Rv) as a supplementation to the diets. We hypothesized that Rv would increase antioxidant capacity and reduce blood lipids, based on the composition of bioactive compounds and fatty acids. Therefore, the aim of this investigation was to evaluate in vitro and in vivo actions of Rv from contaminated (C) and non-C (NC) regions of collection on lipid profiles, antioxidant capacity, and enzyme activities in serum of rats fed an atherogenic diet. Twenty-four male Wistar rats were divided into 4 groups of 6 each and named control, cholesterol (Chol), Chol/RvC and Chol/RvNC. Rats of all 4 groups were fed the basal diet, which included wheat starch, casein, soybean oil, cellulose, vitamin (American Institute of Nutrition for laboratory animals vitamin mixtures), and mineral mixtures (American Institute of Nutrition for laboratory animals mineral mixtures). During 28 days of the experiment, the rats of the control group received the basal diet only, and the diets of the other 3 groups were supplemented with 1% of Chol, 1% of Chol, and 5% of Rv dry matter from C and NC areas. Dry matter from C and NC areas supplemented diets slightly hindered the rise in serum lipids vs. Chol group: total Chol, 13.18% and 11.63% and low-density lipoprotein Chol, 13.57% and 15.08%, respectively. Cholesterol significantly decreased the value of total antioxidant capacity. The supplementation of Rv to the Chol diet significantly affected the increase of antioxidant capacity in serum of rats, expressed by the 2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) method. The water extracts of Rv exhibited high binding properties with bovine serum albumin in comparison with quercetin. In conclusion, atherogenic diets supplemented with Rv from C and NC areas hindered both the rise in serum lipids levels and the decrease in the antioxidant capacity. Based on fluorescence and electrospray-ionization mass spectrometry profiles and in vivo studies, changes in the intensity of the found peaks were estimated in the serum samples after supplemented diets. These findings indicate that the supplementation of Rv to the atherogenic diets improve the lipid profiles and the antioxidant status in serum of rats.

Kopf, Anna ... et. al.-- 5 pages, 1 figure.-- This manuscript is NOAA-GLERL contribution number 1763, Ocean Sampling Day was initiated by the EU-funded Micro B3 (Marine Microbial Biodiversity, Bioinformatics, Biotechnology) project to obtain a snapshot of the marine microbial biodiversity and function of the world’s oceans. It is a simultaneous global mega-sequencing campaign aiming to generate the largest standardized microbial data set in a single day. This will be achievable only through the coordinated efforts of an Ocean Sampling Day Consortium, supportive partnerships and networks between sites. This commentary outlines the establishment, function and aims of the Consortium and describes our vision for a sustainable study of marine microbial communities and their embedded functional traits, This work was supported by the Micro B3 project, which is funded from the European Union’s Seventh Framework Programme (FP7; Joint Call OCEAN.2011‐2: Marine microbial diversity – new insights into marine ecosystems functioning and its biotechnological potential) under the grant agreement no 287589

Strong, James Asa ... et. al.-- 19 pages, 4 figures, 3 tables, There is an increasing demand for environmental assessments of the marine environment to include ecosystem function. However, existing schemes are predominantly based on taxonomic (i.e. structural) measures of biodiversity. Biodiversity and Ecosystem Function (BEF) relationships are suggested to provide a mechanism for converting taxonomic information into surrogates of ecosystem function. This review assesses the evidence for marine BEF relationships and their potential to be used in practical monitoring applications (i.e. operationalized). Five key requirements were identified for the practical application of BEF relationships: (1) a complete understanding of strength, direction and prevalence of marine BEF relationships, (2) an understanding of which biological components are influential within specific BEF relationships, (3) the biodiversity of the selected biological components can be measured easily, (4) the ecological mechanisms that are the most important for generating marine BEF relationships, i.e. identity effects or complementarity, are known and (5) the proportion of the overall functional variance is explained by biodiversity, and hence BEF relationships, has been established. Numerous positive and some negative BEF relationships were found within the literature, although many reproduced poorly the natural species richness, trophic structures or multiple functions of real ecosystems (requirement 1). Null relationships were also reported. The consistency of the positive and negative relationships was often low that compromised the ability to generalize BEF relationships and confident application of BEF within marine monitoring. Equally, some biological components and functions have received little or no investigation. Expert judgement was used to attribute biological components using spatial extent, presence and functional rate criteria (requirement 2). This approach highlighted the main biological components contributing the most to specific ecosystem functions, and that many of the particularly influential components were found to have received the least amount of research attention. The need for biodiversity to be measureable (requirement 3) is possible for most biological components although difficult within the functionally important microbes. Identity effects underpinned most marine BEF relationships (requirement 4). As such, processes that translated structural biodiversity measures into functional diversity were found to generate better BEF relationships, This review has resulted from the DEVOTES (DEVelopment Of innovative Tools for understanding marine biodiversity and assessing good Environmental Status) project funded by the European Union Seventh Programme for Research, Technological Development and Demonstration, ‘The Ocean of Tomorrow’ Theme (grant agreement no. 308392), www.devotes-project.eu

Ecosystem-based management (EBM) is promoted as the solution for sustainable use. An ecosystem-wide assessment methodology is therefore required. In this paper, we present an approach to assess the risk to ecosystem components from human activities common to marine and coastal ecosystems. We build on: (i) a linkage framework that describes how human activities can impact the ecosystem through pressures, and (ii) a qualitative expert judgement assessment of impact chains describing the exposure and sensitivity of ecological components to those activities. Using case study examples applied at European regional sea scale, we evaluate the risk of an adverse ecological impact from current human activities to a suite of ecological components and, once impacted, the time required for recovery to pre-impact conditions should those activities subside. Grouping impact chains by sectors, pressure type, or ecological components enabled impact risks and recovery times to be identified, supporting resource managers in their efforts to prioritize threats for management, identify most at-risk components, and generate time frames for ecosystem recovery.

In the present report the changes in mussels Mytilus galloprovincialis protein profile, as a reaction of water pollution, is presented. The antioxidant capacity was determined by the efficacy to scavenge the derived radical in mussel samples using the Trolox Equivalent Antioxidant Capacity (TEAC) assay. The highest percentage of inhibition was estimated in protein mussels from the dirty area (DM) in comparison with the clean samples (CM). The amounts of both zinc and copper determined by atomic spectroscopy were significantly higher in the protein mussels from the dirty than from the clean areas (P

The objective of this study was to investigate long-term trends in the subsurface sum- mer phytoplankton community and their underlying mechanisms in relation to nutrient enrichment of the open Black Sea. We analysed parameters describing the summer chlorophyll a (chl a) and annual mean nitrate depth distribution in terms of their maximum concentration and its depth, the mean nitrate gradient for the layer between the 2 maxima, and Secchi depth. Investigating the entire period from 1964 to 1996 did not reveal any direct relationship between the nitrate and phytoplank- ton levels. The nitrate level in the open Black Sea increased until 1984/85, corresponding with gen- eral increases in anthropogenic riverine nutrient inputs to the sea. Thus, the average maximum nitrate concentration and the nitrate gradient for all depth profiles increased between 1969 and 1984 and then remained constant. Despite the early increase in nitrate characteristics, the maximum in the chl a profile remained unaltered from 1964 to 1984. After the mid-1980s, the magnitude of the chl a profile gradually increased to 1991 and peaked with an exceptionally high concentration in 1992. In this same period there were only moderate fluctuations in the nitrate profile characteristics. The change of the summer phytoplankton biomass levels since 1984/85 coincided with an intensification of the Rim Current and the associated transport of nitrate to the euphotic zone. Prior to 1984/85 strong stratification and a deep halocline resulted in nitrate accumulation only, whereas uplifting of the halocline, combined with intensified advective currents, increased the transport of nitrate to the euphotic zone during the summer. This resulted in an associated increase in phytoplankton produc- tion and biomass in the following period. While this study shows that the open Black Sea, like the coastal regions, has been impacted by eutrophication, the response is different from the coastal region due to the different physical characteristics of the open waters.

The aim of this investigation was to evaluate the total antioxidant radical scavenging capacity (TARSC) in tissue extracts for understanding biochemical adaptations involving the antioxidant defense system of a bivalve mussel, Mytilus galloprovincialis, sampled in polluted (Pol) and non-polluted (Npol) sites from Black Sea coasts. Antioxidant-rich polyphenol fractions were extracted from whole dry mussel tissue with methanol and water in different proportions. The extracts were screened for polyphenol content and their potential as antioxidants using various in vitro models, such as 1,1-diphenyl-2-picrylhydrazyl (DPPHc), b-carotenelinoleate (b-carotene) and nitric oxide (NOc). The total polyphenol extract of the whole mussel tissue from polluted site (TPMEPol) showed the highest antioxidant capacity among all of the extracts with the tested methods and was comparable with the antioxidant capacity of butylated hydroxyanisole (BHA). The antioxidant capacities of this extract determined by DPPHc, b-carotene and NOc methods and polyphenol content showed the highest correlation coefficients (R 2 ) such as 0.9985, 0.9915 and 0.9895, respectively. The free polyphenol extracts (FPMEPol and FPMENPol) had lower antioxidant capacity than the two others of total polyphenols (TPMEPol and TPMENPol) with three scavenging methods. Responses were linear in all tested methods and the antioxidant capacity values of soluble antioxidants showed the following relative order: TroloxOBHAOTPMEPolOTPMENPolO FPMEPolOFPMENPol. Nutritional antioxidants, such as polyphenols, were probably the main antioxidant contribution to mussel antioxidants. The mussel extracts from the polluted site exhibited higher antioxidant capacity than from the non-polluted one. The results presented in this report indicate that the antioxidant capacity of the whole tissue of the mussel extract could possibly be a useful biomarker for aquatic environments. � 2003 Elsevier Ltd. All rights reserved.

It was shown that phytoplankton from the Varna Bay, Black Sea, has significantly more suspended carbohydrates, proteins and biomass in July than in April. The dominant species were Bacillariophyceae and Dinophyceae. Electrophoretic and fluorescent spectra have shown the main differences in molecular weight and stability of phytoplankton proteins. Phytoplankton included specific proteins distributed over a limited range of molecular weights between 14 and 72 kilodaltons (kDa). The most abundant protein constituents in phytoplankton samples collected in April were around 45–55 kDa. The seasonal variations of the environment influence the quantitative and qualitative changes in phytoplankton.

The present paper is an attempt to test the applicability of the trophic state index (TRIX) for scaling the eutrophication along the Bulgarian Black Sea coastal zone in concert with a number of chemical and biological descriptors aimed at selection of relevant indicators of marine coastal area ecological quality. The following environmental parameters have been considered: to, salinity, nutrients - inorganic P, N and dissolved Si, dissolved oxygen and oxygen saturation, phytoplankton - taxonomic structure, abundance and biomass, chlorophyll a, zooplankton - taxonomic structure, abundance and biomass. Principal Component Analysis was applied in order to figure out and score the most relevant combination of parameters to discriminate between sites and select representative descriptors (pressure/state) of eutrophication. The following variables are defined as relevant descriptors for classification of the sites: nutrients (N, P, Si) and their molar ratios (N:P and Si:P), the capacity of the system to produce and sustain organic matter (chlorophyll a, phytoplankton biomass), phytoplankton taxonomic dominance (Bacilariophyceae:Dinophyceae biomass ratio), grazing pressure (phytoplankton:zooplankton biomass, Bacilariophyceae:Copepoda), plankton diversity index (Hb and Ha) and the trophic state index (TRIX). The investigated sites under a different anthropogenic impact are classified according to selected descriptors and their water quality state.

Extensive data sets on surface chlorophyll a (chl a), depth-integrated primary production (DIPP) and phosphate (PO4, µM) averaged for the upper 25 m layer in 1964, 1973, 1978 and for a period (1980 to 1996) of regular measurements have been used to evaluate long-term changes in the upper portion of the euphotic layer of the entire open (>1000 m) Black Sea. After preliminary analysis of sea- sonal dynamics, special attention was given to data obtained during those periods of the year with rel- atively stable values, revealing interannual and long-term fluctuations and trends. Of 2 phytoplankton characteristics (chl a and DIPP), long-term trends were obtained only for chl a since only these data covered the entire open sea and all periods investiged. A positive correlation was found between DIPP (using 2 different 14 C methods: actual in situ and simulated in situ) and the more numerous chl a data for different monthly intervals, with significant correlation coefficients (r = 0.51 to 0.82). This means that the observed patterns in long-term variability for chl a may be valid also for DIPP. The results show that interannual fluctuations in chl a are more pronounced during the warm months, from ap- proximately May to September. Chl a levels within this interval were moderate, with a mean of 0.15 ± 0.04 mg m -3 during the first 'quiet' period (1964 to 1986), but increased steadily at a rate of 0.06 mg m -3 yr -1 during 1988 to 1991 and sharply in 1992 (mainly due to high July values) to 0.99 ± 0.7 mg m -3 . In contrast, negative trends were characteristic of the third period (1993 to 1996): an abrupt decrease in chl a to 0.26 ± 0.08 mg m -3 in 1993 and a negative trend (-0.02 mg m -3 yr -1 ) during 1993 to 1996. Low concentrations of PO4 (0.015 to 0.138 µM) in the upper 25 m layer throughout the year and the absence of statistically reliable interannual trends in distribution suggest that this nutrient was limiting the level of primary production during the second period. This means that phosphate concentration in the upper 25 m layer cannot be considered a reliable indicator of the presence or absence of anthro- pogenic eutrophication in the open Black Sea. This long-term variability in phytoplankton character- istics agrees well with the data on interannual changes in other ecological variables of the open Black Sea characterized by a collapse of the Black Sea ecosystem during the second period and its recovery after 1992. Comparison of changes in the open and shelf areas of the Black Sea between these 3 peri- ods, and similar changes in the plankton community and in pelagic fish stocks in the second half of the 1980s to the beginning of the 1990s in other regions of the northern hemisphere, connected with changes in the climatic regime during this period, have led us to conclude that global climatic pro- cesses have played an important role in changing the phytoplankton characteristics of the open Black Sea and affecting the whole pelagic ecosystem, especially over the last 2 decades.

The similarities and differences in phytoplankton response to anthropogenic nutrient enrichment were studied in coastal sites of the Western Black Sea and the Eastern Mediterranean (Aegean Sea). Although by physico-chemical parameters, two different environments have been compared, the two regions exhibited similar features: 14 common bloom species in the list of bloom producing algae, the highest rate of occurrence and variety of microalgae involved in summer, the highest average level of competitors-strategies species during the summer outbursts, the similar range of maximum bloom densities attained, and similar capacity to sustain critical biomass during all seasons. The low taxonomic similarity (Bray-Curtis index lower than 30%) between the different basins and among sites, could be attributed to both natural factors and dissimilarities and to the gradients in nutrient levels and their ratios, as affected by the different nature and severity of the anthropogenic pressures on the coastal ecosystems. The results of the PCA analysis indicate that temperature and salinity are factors contributing to the differences outlined between the Aegean Sea and the Black Sea ecosystems, but the differences among the selected sites give support to the importance of nutrients and their ratios. Although competitive outcomes in phytoplankton species selection and succession cannot yet be predicted, the analysis suggests that anthropogenic nutrient enrichment could play an important role in driving the phytoplankton bloom performance. The comparative studies between different basins could be a step forward to highlight common patterns and modes of ecosystem response to anthropogenic eutrophication and to suggest common indices to scale eutrophication impact.

The unfolding of human serum proteins (HSP) was studied by measuring the intrinsic fluorescence intensity at a wavelength of excitation corresponding to tryptophan's or typosine's fluorescence and surface hydrophobicity. The maxima emission wavelengths (lambdamax) of human serum albumin (HSA) and human serum globulin (HSG) before beer consumption (BC) were 336.0 and 337.0 nm and after BC shifted to 335.0 and 334.0 nm, respectively. The surface hydrophobicity slightly increased after BC. In a solution of 8 M urea the lambdamax of BSA shifted to 346.4 and that of BSG to 342.5 nm. In contrast, in the same solution but after BC the lambdamax positions of HSA and HSG shifted to 355.9 and 357.7 nm, respectively. A decrease in fluorescence intensity, a shift in the maximum of emission, and an increase in surface hydrophobicity which reflected unfolding of proteins were observed. Here we provide evidence that the loosening of the HSP structure takes place primarily in various concentrations of urea before and after beer consumption. Differences in the fluorescence behavior of the proteins are attributed to disruption of the structure of proteins by denaturants as well as by the change in their compactability as a result of ethanol consumption.

The Black Sea northwestern shelf (NWS) ecosystem has been subjected to the strongest anthropogenic pressure of the entire Black Sea as about 80% of the freshwater runoff is discharged there. This paper presents a review of the global environmental changes related mainly to increased eutrophication in the western Black Sea basin. A case study (CoMSBlack 92a cruise) attempts to highlight the interaction of some natural and anthropogenic factors responsible for specific chemical and biological features in the western Black Sea environment. The significance of processes located near the Danube river mouth, which, due to their relatively small space scale, have not been recognized before, is clarified. Hydrological processess of specific importance to the distribution and transformation of antropogenic inputs are river plume dynamics, coastal upwelling and mixing and downwelling over the shelf break and slope in this part of the basin. A layer of low hydrological variability (“conservative layer”) appears to be a natural feature of the area, hypothesized to precondition specific biological and chemical processes there. The results suggest that the interplay between the Danube anthropogenic nutrient load with the natural hydrological fronts and gradients provides opportunities for enhanced biological activity thus contributing to the global environmental changes in the Black Sea NWS.