Cecilia Totti

Trends in Ostreopsis proliferation along the Northern Mediterranean coasts

Harmful benthic microalgae blooms represent an emergent phenomenon in temperate zones, causing health, ecological and economic concern. The main goal of this work was to compile records of Ostreopsis at large temporal and spatial scales, in order to study the relationship between cell abundances, the periodicity and intensity of the blooms and the role of sea water temperature in 14 Spanish, French, Monegasque and Italian sites located along the northern limits of the Mediterranean Sea. General trends were observed in the two considered basins: the north-western Mediterranean Sea, in which higher cell abundances were mostly recorded in mid-summer (end of July), and the northern Adriatic Sea where they occur in early fall (end of September). The sea-water temperature does not seem to be a primary driver, and the maximal abundance periods were site and year specific. Such results represent an important step in the understanding of harmful benthic microalgae blooms in temperate areas, and provide a good base for policy makers and managers in the attempt to monitor and forecast benthic harmful microalgae blooms.

Ostreopsis cf. ovata bloom in the northern Adriatic Sea during summer 2009: Ecology, molecular characterization and toxin profile

Intense blooms of the benthic dinoflagellate Ostreopsis cf. ovata have occurred in the northern Adriatic Sea since 2006. These blooms are associated with noxious effects on human health and with the mortality of benthic organisms because of the production of palytoxin-like compounds. The O. cf. ovata bloom and its relationships with nutrient concentrations at two stations on the Conero Riviera (northern Adriatic Sea) were investigated in the summer of 2009. O. cf. ovata developed from August to November, with the highest abundances in September (1.3×10(6) cells g(-1) fw corresponding to 63.8×10(3) cells cm(-2)). The presence of the single O. cf. ovata genotype was confirmed by a PCR assay. Bloom developed when the seawater temperature was decreasing. Nutrient concentrations did not seem to affect bloom dynamics. Toxin analysis performed by high resolution liquid chromatography-mass spectrometry revealed a high total toxin content (up to 75 pg cell(-1)), including putative palytoxin and all the ovatoxins known so far.

Nutrient, particulate matter and phytoplankton variability in the photic layer of the Otranto strait

The distribution of nutrients, suspended matter and phytoplankton in the photic layer of the Otranto Strait, as observed in four seasonal situations (February, May, August and November 1994), is analysed in relation to hydrography. In winter, two water masses were found: the Adriatic Surface Water (ASW) and the Ionian Surface Water (ISW), located at the western and eastern side of the Strait, respectively. In the fresher and cooler ASW, nutrients, suspended matter and phytoplankton contents were higher than in the warmer and saltier ISW (on average: N–NO3=2.3 μM, POC=3.6 μM, chl.a=0.4 μg dm−3 in the ASW, against N–NO3=1.3 μM, POC=2.5 μM, chl.a=0.3 μg dm−3 in the ISW). In the ASW, the mean N:P ratio (50) revealed an excess of nitrogen with respect to phosphorus; the nitrogen supply, as well as the significant presence of diatoms, made us suppose that new production processes were occurring here. In summer, the main features were: (i) high water column stability, (ii) small horizontal differences in hydrological and biological features, (iii) extremely low concentration of nutrients at the surface and (iv) a deep chlorophyll maximum (DCM) at the nutricline level. Phytoplankton summer communities, mainly consisting of nanoflagellates, coccolithophorids and small naked dinoflagellates, were present in low quantities. The low phytoplankton carbon:chlorophyll a ratio (<20), observed at DCM, indicated a high chlorophyll content per single cell, probably as a physiological response to low irradiance. As to the phytoplankton annual cycle, sediment traps and water column observations were highly correlated, evidencing two abundance peaks in spring and autumn, and a summer minimum. Hydro-chemical and biological data suggest that the winter spread of the ASW was the main factor favouring the enhancement of phytoplankton growth and controlling the species composition in the strait, while the DCM formation represents a more typical characteristic of ISW, with an oligotrophic feature similar to that of other Eastern Mediterranean water masses.

New approach using the real-time PCR method for estimation of the toxic marine dinoflagellate Ostreopsis cf. ovata in marine environment.

Background We describe the development and validation of a new quantitative real time PCR (qrt-PCR) method for the enumeration of the toxic benthic dinoflagellate Ostreopsis cf. ovata in marine environment. The benthic Ostreopsis sp. has a world-wide distribution and is associated during high biomass proliferation with the production of potent palytoxin-like compounds affecting human health and environment. Species-specific identification, which is relevant for the complex of different toxins production, by traditional methods of microscopy is difficult due to the high morphological variability, and thus different morphotypes can be easily misinterpreted. Methodology/Findings The method is based on the SYBR I Green real-time PCR technology and combines the use of a plasmid standard curve with a “gold standard” created with pooled crude extracts from environmental samples collected during a bloom event of Ostreopsis cf. ovata in the Mediterranean Sea. Based on their similar PCR efficiencies (95% and 98%, respectively), the exact rDNA copy number per cell was obtained in cultured and environmental samples. Cell lysates were used as the templates to obtain total recovery of DNA. The analytical sensitivity of the PCR was set at two rDNA copy number and 8.0×10−4 cell per reaction for plasmid and gold standards, respectively; the sensitivity of the assay was of cells g−1 fw or 1−1 in macrophyte and seawater samples, respectively. The reproducibility was determined on the total linear quantification range of both curves confirming the accuracy of the technical set-up in the complete ranges of quantification over time. Conclusions/Significance We developed a qrt-PCR assay specific, robust and high sample throughput for the absolute quantification of the toxic dinoflagellate Ostreopsis cf. ovata in the environmental samples. This molecular approach may be considered alternative to traditional microscopy and applied for the monitoring of benthic toxic microalgal species in the marine ecosystems.

Monitoring toxic microalgae Ostreopsis (dinoflagellate) species in coastal waters of the Mediterranean Sea using molecular PCR-based assay combined with light microscopy.

A molecular PCR-based assay was developed and applied to macrophyte and seawater samples containing mixed microphytobenthic and phytoplanktonic assemblages, respectively, in order to detect toxic Ostreopsis species in Mediterranean Sea. The specificity and sensitivity of the molecular PCR assay were assessed with both plasmidic and genomic DNA of the target genus or species using taxon-specific primers in the presence of background macrophyte DNA. The PCR molecular technique allowed rapid detection of the Ostreopsis cells, even at abundances undetectable within the resolution limit of the microscopy technique. Species-specific identification of Ostreopsis was determined only by PCR-based assay, due to the inherent difficulty of morphological identification in field samples. In the monitoring of the toxic Ostreopsis blooms PCR-based methods proved to be effective tools complementary to microscopy for rapid and specific detection of Ostreopsis and other toxic dinoflagellates in marine coastal environments.

Diatom invasion in the antarctic hexactinellid sponge Scolymastra joubini

Abstract Sponges often host large amounts of symbionts, mainly represented by cyanobacteria and dinoflagellates. Recent findings show a widespread presence of symbiotic diatoms living inside antarctic demosponges. In this paper, the invasion by large populations of the diatom Melosira sp. into specimens of the hexactinellid sponge Scolymastra joubini is reported. SEM analyses support the hypothesis that the embedded living diatoms have a negative impact on sponge tissues, leading to degenerative processes.

Effects of harmful dinoflagellate Ostreopsis cf. ovata exposure on immunological, histological and oxidative responses of mussels Mytilus galloprovincialis

In the last decade massive blooms of the Ostreopsis cf. ovata have occurred in the Mediterranean basin, posing great concern to both environmental and human health. Biotoxicological and chemical studies demonstrated that O. cf. ovata produces palytoxin and ovatoxins; besides direct respiratory effects on humans due to inhalation of marine toxic aerosols, O. cf. ovata blooms can cause adverse effects on benthic invertebrates. The main aim of this study was to highlight the role of immunological, cellular and oxidative mechanisms in modulating the toxicity induced by O. cf. ovata in mussels Mytilus galloprovincialis. Organisms were exposed in laboratory condition to O. cf. ovata and analysed after 7 and 14 days of exposure. Obtained results demonstrated a clear involvement of the immune system with a significant decrement of granulocytes respect to the hyalinocytes type cells, a diminished phagocytosis activity and a reduced lysosomal membrane stability in haemocytes, after both 7 and 14 days of exposure. Histological analyses showed a decrease of the digestive gland wall thickness, dilatation of the tubules, haemocytes infiltration into the digestive gland and a decrement of neutral lipid levels in exposed mussels; similar results suggest a possible inhibition of the feeding activity, with a consequent induction of authophagic phenomena and utilization of stored reserve products such as neutral lipids. Antioxidant parameters revealed a limited role of O. cf. ovata to induce oxidative stress in mussels exposed under laboratory conditions excepting for a certain increase of catalase, glutathione reductase and glutathione peroxidases activities, and a significantly higher capability to neutralize peroxyl radicals in mussels exposed for 14 days. Although the obtained results suggest a non-specific response of mussels to the O. cf. ovata exposure, observed effects on the general health status of exposed mussels should be adequately considered when assessing the ecological relevance of these algal blooms.

Biological effects of palytoxin-like compounds from Ostreopsis cf. ovata: a multibiomarkers approach with mussels Mytilus galloprovincialis.

Massive blooms of the harmful benthic dinoflagellate Ostreopsis cf. ovata are of growing environmental concern in the Mediterranean, having recently caused adverse effects on benthic invertebrates and also some intoxication episodes to humans. The toxicological potential of produced palytoxin-like compounds was investigated in the present study on a typical marine sentinel species, the mussel Mytilus galloprovincialis. Organisms were sampled during various phases of a O. cf. ovata bloom, in two differently impacted sites. The presence of the algal toxins was indirectly assessed in mussels tissues (mouse test and hemolysis neutralization assay), while biological and toxicological effects were evaluated through the measurement of osmoregulatory and neurotoxic alterations (Na(+)/K(+)-ATPase and acetylcholinesterase activities), oxidative stress responses (antioxidant defences and total oxyradical scavenging capacity), lipid peroxidation processes (level of malondialdehyde), peroxisomal proliferation, organelle dysfunctions (lysosomal membrane stability, accumulation of lipofuscin and neutral lipids), immunological impairment (granulocytes percentage). Obtained results demonstrated a significant accumulation of algal toxins in mussels exposed to O. cf. ovata. These organisms exhibited a marked inhibition of the Na(+)/K(+)-ATPase activity and alterations of immunological, lysosomal and neurotoxic responses. Markers of oxidative stress showed more limited variations suggesting that toxicity of the O. cf. ovata toxins is not primarily mediated by an over production of reactive oxygen species. This study provided preliminary results on the usefulness of a multi-biomarker approach to assess biological alterations and toxicological events associated to blooms of O. cf. ovata in marine organisms.

Seasonal variations of epilithic diatoms on different hard substrates, in the northern Adriatic Sea

The colonization of epilithic diatoms on artificial hard substrates (marble, quartzite and slate) was investigated on a seasonal basis in a subtidal site of the northern Adriatic Sea to determine if substrate-dependent differences in colonization occurs and to define the seasonal variations of micro-epilithic communities in terms of abundance, biomass and community structure. Artificial substrates (smoothed small discs mounted on a Plexiglas sheet) were placed at a depth of 8 m in April 2003, July 2003, January 2004 and February 2004. The discs were collected after 6-7 weeks for counting and taxonomic identification of the diatoms. The community structure of epilithic diatoms showed a dominance of motile species over the entire study period, followed by erect, adnate and tube-dwelling diatoms. Diatom density showed a marked seasonal variability, ranging from 365 ±407 (winter 2004) to 557156 ±82602 cells cm -2 (spring 2004). Biomass ranged from 0.02 ±0.01 to 17.53 ±3.20 μg Ccm -2 . Abundance and biomass values did not present any significant differences for the three substrates examined.

PHYTOPLANKTON PHOTOSYNTHETIC ACTIVITY AND GROWTH RATES IN THE NW ADRIATIC SEA

Taxonomic composition, biomass, primary production and growth rates of the phytoplankton community were studied in two stations in the NW Adriatic Sea on a seasonal basis, in areas characterized by differing hydrological and trophic conditions. The main differences between the two stations were quantitative rather than qualitative, most phytoplankton species being common to both stations. The effects of differing nutrient concentrations and plume spreading were evident. Biomass and primary production rates were significantly higher in the coastal station (S1), and the phytoplankton distribution in the water column was markedly stratified in S1 and more even in the offshore station (S3). However, chlorophyll a specific production, potential growth rate and production efficiencies were very similar in both stations, even when phosphorus concentrations were limiting. A discrepancy between potential and actual growth rate was observed: as a feature common to both stations, comparisons between potential and actual growth rates revealed that little carbon produced by phytoplankton accumulated as algal biomass; therefore, very high loss rates were estimated.