Environmental quality improvement of a mariculture plant after its conversion into a multi-trophic system.

Integrated Multitrophic Aquaculture (IMTA) seems to be one of the best solutions for sustainable aquaculture. Within the Remedia LIFE Project, an experimental IMTA plant was put in place in the Mar Grande of Taranto (Mediterranean Sea, Southern Italy). The polyculture of several bioremediating organisms, such as mussels, tubeworms, sponges, and seaweeds, was combined with a coastal cage fish farm, in order to remove organic and inorganic wastes coming from the fish's metabolism. To verify the effectiveness of the system, the ex ante measurement of chemical-physical variables, trophic status, microbial contamination, and zoobenthos community health was compared with the results of the same measurement performed one year and two years after the implementation of the experimental IMTA plant. The results were encouraging, since a reduction in total nitrogen concentration in the seawater (from 43.4 ± 8.9 to 5.6 ± 3.7 µM/l), a reduction in microbial pollution indicators in the seawater (total coliforms: from 280 ± 18 MPN/100 mL to 0; E. coli: from 33 ± 1.3 MPN/100 mL to 0) and in the sediments (total coliforms: from 230 ± 6.2 MPN/100 g to 170 ± 9; E. coli: from 40 ± 9.4 MPN/100 g to 0), an enhancement of the trophic status (TRIX: from 4.45 ± 1.29 to 3.84 ± 0.18), and an increase in the zoobenthic quality indices and biodiversity were recorded (AMBI: from 4.8 to 2.4; M-AMBI: from 0.14 to 0.7). These results prove that the Remedia LIFE project's purpose was achieved. The selected bioremediators worked synergistically, improving water and sediments quality within the fish farm area. Moreover, bioremediating organisms increased their weight as a result of wastes uptake, producing, as co-products, large amounts of additional biomass. This could be commercially exploited, thus being an added value of the IMTA plant. Based on our findings, the promotion of eco-friendly practices to ameliorate ecosystem health should be encouraged.

Microbiological and chemical characteristics of beaches along the Taranto Gulf (Ionian Sea, Southern Italy).

Coastal habitats provide important ecosystem services, such as the maintenance of ecological sustainability, water quality regulation, nutrient recycling, and sandy beaches which are important areas for recreation and tourism. The quality of seawater is generally measured by determining the concentrations of Escherichia coli and intestinal Enterococci, which might be affected by the persistent populations of these bacteria in sand. Sand might thus be a significant source of pathogen exposure to beachgoers. The quality of coastal recreational waters can also be affected by eutrophication, water discoloration, and harmful algal blooms, which pose additional human health risks. Here, we conducted a monitoring of the beaches quality along the Taranto Gulf by determining the concentrations of fecal indicator organisms, as well as other parameters that are not traditionally measured (physicochemical parameters, Pseudomonas aeruginosa, and harmful microalgae), in shallow seawater and sand sampled from three beaches. The concentrations of bacteria were determined using both standard microbiological methods and the IDEXX system. Our results demonstrate the utility of measuring a greater number of parameters in addition to those conventionally measured, as well as the importance of assessing the health risks posed by the sand matrix. Additional work is needed to develop rapid analytical techniques that could be used to monitor the microbiological parameters of solid matrices.

An Integrated Monitoring Approach to the Evaluation of the Environmental Impact of an Inshore Mariculture Plant (Mar Grande of Taranto, Ionian Sea).

The results of an ex-ante survey aiming to assess the impact of a fish farm in the Mar Grande of Taranto (southern Italy, Mediterranean Sea) on the surrounding environment are reported. There, the implementation of an innovative IMTA plant was planned, with the goals of environment bioremediation and commercially exploitable biomass production. Analyses were conducted in February and July 2018. Both seawater and sediments were sampled at the four corners of the fish farm to detect the existing biological and physico-chemical features. The investigation was performed to identify the best area of the farming plant for positioning the bioremediating system, but also to obtain a data baseline, to compare to the environmental status after the bioremediating action. Data were also analyzed by canonical analysis of principal coordinates (CAP). All the measurements, in particular, microbiology and macrobenthic community characterization using AZTI's Marine Biotic Index (AMBI) and the Multivariate-AMBI (M-AMBI) indices, suggest that the effect of fish farm waste was concentrated and limited to a small portion of the investigated area in relation to the direction of the main current. A site named A3, which was found to be the most impacted by the aquaculture activities, especially during the summer season, was chosen to place the bioremediation system.

Microbiological and Chemical Assessment of Wastewater Discharged by Infiltration Trenches in Fractured and Karstified Limestone (SCA.Re.S. Project 2019-2020).

This study investigated the environmental contamination of groundwater as a consequence of the discharge of treated wastewater into the soil. The investigation focused on a wastewater treatment plant located in an area fractured by karst in the Salento peninsula (Apulia, Italy). Water samples were collected at four sites (raw wastewater, treated wastewater, infiltration trench, and monitoring well), monthly from May to December 2019 (with the exception of August), and were tested for (1) panel of bacteria; (2) enteric viruses; and (3) chemical substances. A gradual reduction in the concentration of bacteria, viruses and contaminants of emerging concern was observed across the profile of soil fissured by karst. All monitored bacteria were absent from the monitoring well, except for Pseudomonas aeruginosa. Pepper mild mottle virus and adenovirus were detected at all sampling sites. Personal care products and X-ray contrast media showed the greatest decrease in concentration from infiltration trench to the monitoring well, while the highest residual concentrations in the monitoring well were found for anticonvulsants (78.5%), antimicrobials (41.3%), and antipsychotic drugs (38.6%). Our results show that parameters provided by current law may not always be sufficient to evaluate the sanitary risk relating to the discharge of treated wastewater to the soil.

Screening of Chaetomorpha linum Lipidic Extract as A New Potential Source of Bioactive Compounds.

Recent studies have shown that marine algae represent a great source of natural compounds with several properties. The lipidic extract of the seaweed Chaetomorpha linum (Chlorophyta, Cladophorales), one of the dominant species in the Mar Piccolo of Taranto (Mediterranean, Ionian Sea), revealed an antibacterial activity against Vibrio ordalii and Vibrio vulnificus, common pathogens in aquaculture, suggesting its potential employment to control fish and shellfish diseases due to vibriosis and to reduce the public health hazards related to antibiotic use in aquaculture. This extract showed also an antioxidant activity, corresponding to 170.960 ± 16. mmol Trolox equivalent/g (oxygen radical absorbance capacity assay-ORAC) and to 30.554 ± 2.30 mmol Trolox equivalent/g (Trolox equivalent antioxidant capacity assay-TEAC). The chemical characterization of the extract, performed by 1D and 2D NMR spectroscopy, highlighted the presence of free, saturated (SAFAs), unsaturated (UFAs) and polyunsaturated (PUFAs) fatty acids. The high content of ω-6 and ω-3 PUFAs confirmed also by gas chromatography indicates the potentiality of this algal species in the production of fortified food. The antibacterial activity seems related to the presence of linolenic acid present at high density, while the antioxidant activity could be likely ascribable to molecules such as carotenoids and chlorophylls (characterized also by thin-layer chromatography), known for this property. The presence of polyhydroxybutyrate, a biopolymer with potentiality in the field of biodegradable bioplastics was also detected. The exploitation of C. linum for a future biotechnological application is also encouraged by the results from a first attempt of cultivating this species in an integrated multi-trophic aquaculture (IMTA) system.

Screening of Three Echinoderm Species as New Opportunity for Drug Discovery: Their Bioactivities and Antimicrobial Properties.

Echinoderms are a renewable resource with an economic value due to their increasing demand as food and/or source of bioactive molecules exerting antitumor, antiviral, anticoagulant, antioxidant, and antimicrobial activities. In this framework, the present study is aimed at investigating the antibacterial, antioxidant, and hemolytic activities in the three Echinoderm species Echinaster sepositus, Arbacia lixula, and Sphaerechinus granularis. The sea star E. sepositus showed lysozyme-like activity (mean diameter of lysis of 13.4 ± 0.2 mm), an antimicrobial activity against the human emerging pathogens Staphylococcus aureus, Pseudomonas aeruginosa, and Candida famata, and a strong lytic activity (100 ± 0.05%) towards the human red blood cells. Furthermore A. lixula and E. sepositus had the highest antioxidant activity (1792.75 ± 233.7 and 1765.65 ± 484.58 nmolTE/mL, resp.). From toxicological assays, it was shown that E. sepositus was not toxic towards HeLa cells and Vibrio fischeri, encouraging the exploitation of this species in the pharmaceutical field. Therefore, our findings have implications due to the ongoing explosion of antibiotic-resistant infections because of the new opportunistic pathogens and the need to discover antibacterial agents with new modes of action. Also the recorded antioxidant activity taking into account the need to find natural antioxidants useful for human health is intriguing.

The Potential Exploitation of the Mediterranean Invasive Alga Caulerpa cylindracea: Can the Invasion Be Transformed into a Gain?

Recently, there is a growing interest towards the development of strategies for invasive seaweed control and exploitation as source of secondary metabolites. Here, we investigated the potential of exploitation in biotechnology and recycling options in eradication programs of the lipidic extract of the Mediterranean invasive seaweed Caulerpa cylindracea (Chlorophyta). The chemical characterization was carried out by means of multinuclear and multidimensional NMR spectroscopy. The fatty acid profile of C. cylindracea assessed the presence of several types of molecules known for antioxidant activity such as carotenoids, chlorophylls, pheophytins, and sterols. The NMR spectroscopy showed also the characteristic signals of saturated, unsaturated, and free fatty acids as well as other metabolites including the biopolymer polyhydroxybutyrate. The lipidic extract exerted an antioxidant activity corresponding to 552.14 ± 69.13 mmol Trolox equivalent/g (ORAC) and to 70.3 ± 2.67 mmol Trolox equivalent/g (TEAC). The extract showed an antibacterial activity against several Vibrio species, suggesting its potential use in the control of diseases in mariculture. Our results show that C. cylindracea, representing a critical hazard in coastal areas, could be transformed into a gain supporting specific management actions to reduce the effects of human pressures.

Microbial assemblages for environmental quality assessment: Knowledge, gaps and usefulness in the European Marine Strategy Framework Directive.

The EU Marine Strategy Framework Directive 2008/56/EC (MSFD) defines a framework for Community actions in the field of marine environmental policy in order to achieve and/or maintain the Good Environmental Status (GES) of the European seas by 2020. Microbial assemblages (from viruses to microbial-sized metazoa) provide a major contribution to global biodiversity and play a crucial role in the functioning of marine ecosystems, but are largely ignored by the MSFD. Prokaryotes are only seen as "microbial pathogens," without defining their role in GES indicators. However, structural or functional prokaryotic variables (abundance, biodiversity and metabolism) can be easily incorporated into several MSFD descriptors (i.e. D1. biodiversity, D4. food webs, D5. eutrophication, D8. contaminants and D9. contaminants in seafood) with beneficial effects. This review provides a critical analysis of the current MSFD descriptors and illustrates the reliability and advantages of the potential incorporation of some prokaryotic variables within the set of indicators of marine environmental quality. Following a cost/benefit analysis against scientific and economic criteria, we conclude that marine microbial components, and particularly prokaryotes, are highly effective for detecting the effects of anthropogenic pressures on marine environments and for assessing changes in the environmental health status. Thus, we recommend the inclusion of these components in future implementations of the MSFD.

Genotoxic damage of benzo[a]pyrene in cultured sea bream (Sparus aurata L.) hepatocytes: harmful effects of chronic exposure.

The large majority of studies on the genotoxic hazard of PAHs polluted water widely applied the ENA assay as versatile tool in large number of wild and farmed aquatic species. Nuclear abnormalities are commonly considered to be a direct consequence of genotoxic lesions in DNA macromolecule, and such evaluation might be helpful in identifying the genotoxic damage induced by the most harmful PAHs such as B[a]P. Regarding at the fish species subjected to aquaculture, most of the toxicological data come from wild fish and mainly focus on freshwater fish, but very little is known for other marine major aquacultured species. The gilthead sea bream (Sparus aurata L.) is the most economically important sparid species cultured along the Mediterranean costs, and it has been proved a very sensitive species to acute B[a]P exposure. However, further investigation is needed on several other types of genotoxic assessments, especially for chronic effects. This work was totally based on an in vitro model for chronic toxicity, using long-term S. aurata hepatocytes in primary culture, continuously exposed to low levels of BaP, over a prolonged period of time, to provide evidences for latent toxicity response. We aimed to investigate the kind of nuclear damage in gilthead sea bream hepatocytes continuously exposed to B[a]P sublethal doses. Cells were exposed to several B[a]P concentrations (10 µg/mL, 1 µg/mL, 1 ng/mL, 1 pg/mL) for two exposure times (24 and 72 h), and then tested both for apoptosis induction and for nuclear abnormalities by immunofluorescence analysis. The presence of severe nuclear damage, revealed cells progressing towards abnormal genotypes, due to a series of aberrant mitosis followed by unequal distribution of chromosomal content. The nuclear atypia (NA) more frequently observed were: a) micronuclei (MN); b) nuclear buds or blebs (NBUDs); c) notched nuclei; d) lobed nuclei; e) nuclei with nucleoplasmic bridge (NPBs); f) nuclei squashed, with a residual nuclear membrane; g) open nuclei, with membrane tape unrolled; and h) apoptotic bodies. Our results showed at medium-low doses a sustained genotoxic response, whose potency increased with the exposure time, becoming apparent as apoptosis induction, both by cell surface and nuclear changes. At the lowest doses, the longer was B[a]P exposure, greater was the involvement on masses of replicating cells, establishing the connection between the escape from apoptosis and the selection of tumoral cell evolution. In view of these results, there is no evidence of a threshold dose below which B[a]P was found not to be genotoxic in sea bream cultured hepatocytes.

Sewage-exposed marine invertebrates: survival rates and microbiological accumulation.

A large number of bacteria, including agents responsible for diseases, characterise sewage-polluted seawaters. Apart from standards for bathing waters and bivalve aquaculture waters, there are no general microbiological standards applicable to seawaters to help decide if bacterial pollution is within acceptable ranges. This study represents an attempt towards the issue of comparing the susceptibility of different marine invertebrates subjected to polluted seawater with a high microbial contamination. We explored the survival rates and the microbiological accumulation of mollusc bivalves, echinoderms and crustaceans species exposed to sewage-polluted seawaters. Microbiological analyses were performed on the polluted seawater and on the homogenates of exposed and unexposed specimens. Culturable bacteria (22 °C and 37 °C) and microbial pollution indicators (total coliforms, Escherichia coli and intestinal enterococci) were measured. When exposed to the sewage-polluted seawater, the examined invertebrates showed different survival rates. In the filter feeders, bacterial densities at 22 °C and 37 °C rose after 96 h of exposure to sewage. The highest concentrations of total coliforms and intestinal enterococci were found in exposed bivalve Mytilus galloprovincialis. The concentrations of bacteria growing at 37 °C were lower in the exposed deposit feeders compared to the polluted seawater. Some yeasts were absent in several exposed species although these yeasts were present in the polluted seawater. Our data suggest that the examined filter feeders, given their capability to survive and accumulate bacteria, may counteract the effects of sewage and restore seawater quality.

Filter-feeder macroinvertebrates as key players in culturable bacteria biodiversity control: a case of study with Sabella spallanzanii (Polychaeta: Sabellidae).

The present study investigates the effect of the filtering activity of Sabella spallanzanii on the culturable heterotrophic bacterial community through the comparison of the bacterial diversity in transplanted polychaetes and the surrounding seawater. For isolation of culturable heterotrophic bacteria, seawater samples as well as polychaete homogenates were plated in triplicates onto Bacto Marine Agar 2216 (Difco). All the colonies grown were isolated, subcultured and identified by several morphological, biochemical and cultural methods. Some bacterial genera showed higher average abundances in polychaetes than in seawater (i.e. Lucibacterium and Photobacterium). Aeromonas represented a conspicuous component of the bacterial community both in S. spallanzanii and seawater. The presence of Cytophaga and Pseudomonas was also relevant in the examined seawater samples. The selective concentrations of some bacterial genera inside S. spallanzanii either by grazing on bacteria or their capability as bacterial reservoirs, provides evidence for the role of macrobenthic invertebrates as key determinants for microbial diversity.

Bacterial accumulation by Branchiomma luctuosum (Annelida: Polychaeta): a tool for biomonitoring marine systems and restoring polluted waters.

In this study, we examined the bacterial accumulation in the filter feeder polychaete Branchiomma luctuosumGrube (Sabellidae). Analyses were performed on worm homogenates from 'unstarved' and 'starved' individuals, and seawater from the same sampling site (Gulf of Taranto, Western Mediterranean, Italy). Densities of culturable heterotrophic bacteria (22 degrees C), total culturable bacteria at 37 degrees C and halophilic vibrios at 22 and 35 degrees C were measured on Marine Agar 2216, Plate Count Agar and thiosulphate-citrate-bile-salt-agar (TCBS) plus 2% NaCl, respectively. Total and faecal coliforms as well as faecal streptococci were determined by the Most Probable Number method. Results showed that B. luctuosum is able to accumulate all the considered six microbiological groups with a higher efficiency for autochthonous bacteria. The analysis also indicated that bacterial groups differ in their resistance to digestion by B. luctuosum. Our data suggest the potential role of B. luctuosum as an useful bioindicator to assess low levels of microbiological water pollution as well as bioremediator of microbial polluted waters.

Monitoring of a coastal Mediterranean area: culturable bacteria, phytoplankton, environmental factors and their relationships in the Southern Adriatic Sea.

Culturable heterotrophic bacterial and phytoplanktonic densities were investigated at four sites in the Southern Adriatic Sea (Brindisi, S. Cataldo, Otranto and S. M. di Leuca) over an annual cycle. The main phytoplankton groups, the bacterial biodiversity, as well as the faecal contamination indicators were determined. Culturable bacterial numbers averaged 4.8 +/- 0.2 log CFU ml(-1) whereas phytoplankton numbers averaged 2.1 +/- 0.4 log cells ml(-1). Relationships between culturable bacteria, phytoplankton and the environmental factors were established. Bacterial and phytoplankton densities usually depended significantly on temperature, dissolved oxygen, phosphate and nitrite only in the S. Cataldo transect. In all the examined transects phytoplankton showed a bloom during the January-February period followed by a bacterial peak during the February-March period. Thus we can suppose that the phytoplankton winter bloom is responsible for the availabily of organic matter for bacterial populations in the following months in this oligotrophic ecosystem.

Sabella spallanzanii filter-feeding on bacterial community: ecological implications and applications.

The filtration process of Sabella spallanzanii Gmelin on bacterial community was studied in a coastal area of the Northern Ionian Sea (Mediterranean Sea) at three sites, S. Vito, Lido Gandoli and Lido Silvana, where some specimens of S. spallanzanii were transplanted. Analyses were performed both on water and worm samples. A total of six microbial groups were examined: culturable heterotrophic bacteria, total culturable bacteria at 37 degrees C, culturable vibrios, total and fecal coliforms and fecal streptococci. The bacterial densities were usually orders of magnitude higher in the worm homogenates than in the corresponding seawater and the highest values were observed in August. The ability of S. spallanzanii to accumulate the microbial pollution indicators suggests this species can be employed as a bioindicator for monitoring water quality. Moreover, the accumulation capability of S. spallanzanii for specific micro-organisms provides a potential role in sewage bioremediation.

Mytilus galloprovincialis filter feeding on the bacterial community in a Mediterranean coastal area (Northern Ionian Sea, Italy).

This study was carried out seasonally, throughout a year, to evaluate the filtering activity on bacteria of Mytilus galloprovincialis. Six microbiological parameters were researched in the water and mussels samples collected along the coastal area of the Northern Ionian Sea in three stations, S. Vito, Lido Gandoli and Lido Silvana. We detected the densities of culturable heterotrophic bacteria by spread plate on Marine Agar, total culturable bacteria at 37 degrees C on Plate Count Agar and vibrios abundance on thiosulphate-citrate-bile-sucrose-salt (TCBS) agar. Total and fecal coliforms as well as fecal streptococci were determined by the Most Probable Number. Bacterial concentrations at 20 and 37 degrees C as well as vibrios concentrations were higher in the mussel samples compared to the corresponding seawater throughout the year. The results obtained could contribute to improve the information relatively either to the natural processes existing between bacteria and mussels or to the risk of human infections related to the consumption of mussels.

Presence of vibrios in seawater and Mytilus galloprovincialis (Lam.) from the Mar Piccolo of Taranto (Ionian Sea).

During the spring-summer period, vibrios were detected in water and mussels (Mytilus galloprovincialis) collected in 30 sampling sites located in the Mar Piccolo of Taranto (Ionian Sea, Italy). In order to evaluate the degree of microbial pollution of the investigated area, fecal coliforms and Escherichia coli densities were also determined. Vibrio alginolyticus constituted the predominant component of the total culturable vibrios. Some Vibrio species such as V. mediterranei, V. parahaemolyticus, V. diazotrophicus, V. nereis, and V. splendidus were present in water as well as in mussel samples; selective retention in mussels, however, was demonstrated for other vibrios (V. vulnificus, V. cincinnatiensis, V. orientalis, V. anguillarum, V. marinus, V. hollisae). The isolation of some potential pathogenic vibrio species shows the importance of Vibrio research to estimate water quality and to avoid transmission of infection to man and to other marine organisms.