A Morphometric Approach to Understand Prokaryoplankton: A Study in the Sicily Channel (Central Mediterranean Sea).

A new understanding of plankton ecology has been obtained by studying the phenotypic traits of free-living prokaryotes in the Sicily Channel (Central Mediterranean Sea), an area characterised by oligotrophic conditions. During three cruises carried out in July 2012, January 2013 and July 2013, the volume and morphology of prokaryotic cells were assessed microscopically using image analysis in relation to environmental conditions. The study found significant differences in cell morphologies among cruises. The largest cell volumes were observed in the July 2012 cruise (0.170 ± 0.156 µm3), and the smallest in the January 2013 cruise (0.060 ± 0.052 µm3). Cell volume was negatively limited by nutrients and positively by salinity. Seven cellular morphotypes were observed among which cocci, rods and coccobacilli were the most abundant. Cocci, although they prevailed numerically, always showed the smallest volumes. Elongated shapes were positively related to temperature. Relationships between cell morphologies and environmental drivers indicated a bottom-up control of the prokaryotic community. The morphology/morphometry-based approach is a useful tool for studying the prokaryotic community in microbial ecology and should be widely applied to marine microbial populations in nature.

Thermo-mechanical, antimicrobial and biocompatible properties of PVC blends based on imidazolium ionic liquids.

The aim of this study was the development of antimicrobial polyvinylchloride (PVC) blends loaded with 0.1-10% (w/w) of the ILs 1-hexadecyl-3-methylimidazolium 1,3-dimethyl 5-sulfoisophthalate (HdmimDMSIP) and 1-octyloximethyl-3-methylimidazolium hexafluorophosphate (OOMmimPF6). The synthetized ILs were characterized by 1HNMR, MALDI-TOF, DSC and TGA. PVC/ILs films were obtained by solvent casting.Thermal and mechanical properties (tensile stress TS and elongation at break EB), morphology by SEM, surface wettability, antimicrobial activity, cytotoxicity and ILs release in sterile water from PVC/ILs film blends were determined. Results demonstrated that the presence of both ILs in PVC formulation slightly affected thermal and mechanical properties of blends. The loading of both ILs into PVC matrix made PVC/ILs films hydrophilic, especially at the highest concentration of HdmimDMSIP. The PVC/ILs blends displayed antibacterial activity up to ILs lowest concentrations (0.1-0.5%). The inhibition of Escherichia coli growth was lower than that showed toward Staphylococcus epidermidis. The addition of 10% ILs concentration resulted excessive as demonstrated by accumulation of ILs on film surfaces (SEM) and ILs high release from PVC/ILs blends during the first day of water immersion. Biocompatibility studies highlighted that the addition of low amounts of both ILs into PVC matrix is not cytotoxic for mouse fibroblast cells (L929), supporting their potential use for biomedical porposes.

Regulation of Microbial Activity Rates by Organic Matter in the Ross Sea during the Austral 2017.

The active prokaryotic communities proliferate in the ecosystems of the Antarctic Ocean, participating in biogeochemical cycles and supporting higher trophic levels. They are regulated by several environmental and ecological forcing, such as the characteristics of the water masses subjected to global warming and particulate organic matter (POM). During summer 2017, two polynyas in the Ross Sea were studied to evaluate key-microbiological parameters (the proteasic, glucosidasic, and phosphatasic activities, the microbial respiratory rates, the prokaryotic abundance and biomass) in relation to quantitative and qualitative characteristics of POM. Results showed significant differences in the epipelagic layer between two macro-areas (Terra Nova Bay and Ross Sea offshore area). Proteins and carbohydrates were metabolized rapidly in the offshore area (as shown by turnover times), due to high enzymatic activities in this zone, indicating fresh and labile organic compounds. The lower quality of POM in Terra Nova Bay, as shown by the higher refractory fraction, led to an increase in the turnover times of proteins and carbohydrates. Salinity was the physical constraint that played a major role in the distribution of POM and microbial activities in both areas.

Microbial enzymes in the Mediterranean Sea: relationship with climate changes.

In most of the aquatic ecosystems, microorganisms are major players in the biogeochemical and nutrients cycles (Carbon Nitrogen, Phosphorus), through their enzymatic activities (leucine aminopeptidase, alkaline phosphatase and beta-glucosidase) on organic polymers such as polypeptides, organophosphate esters and polysaccharides, respectively. The small monomers released by decomposition are metabolised by microbes, supporting their growth. Most of the extracellular enzymes are adaptative and their synthesis and activity is strongly affected by environmental factors, consequently the relative importance of leucine aminopeptidase, alkaline phosphatase and beta-glucosidase reflects differences in the composition of organic matter and assume a different meaning. Since more than two decades, at the CNR the influence of climate changes, seasonal variability, depth and coastal input on the patterns of enzymatic activities in the Mediterranean Sea have been studied. Its particular characteristics of a semi-closed basin, high summer evaporation and the occurrence of important water dynamics, make this ecosystem particularly suitable as a model site for climate changes-related observations. The present paper reviews the current information of environmental changes on extracellular enzymatic activity obtained in the Mediterranean areas with the aim of evaluating the effects of environmental changes on the microbial activities. The obtained results revealed significant variations in the rates of hydrolytic activities in relation to space and time, with the highest levels generally found in the epipelagic layer (0-100m) and in coastal zones during warm periods. In the Central Mediterranean Sea their relationship with temperature changes was demonstrated. Spatial variations in the relative enzyme activities also suggested a modulation in the metabolic profiles of the prokaryotic communities, with biogeochemical implications in nutrient regeneration. Long term studies on microbial activity and abundances in relation with rising temperatures can have a predictive value to describe the evolutionary scenario of microbial processes and the response of microbial metabolism to climate changes in the Mediterranean Sea.

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.

Isolation of a novel gene from Photobacterium damselae subsp. piscicida and analysis of the recombinant antigen as promising vaccine candidate.

Photobacterium damselae subsp. piscicida (PDP) is the causative agent of fish pasteurellosis, a bacterial disease causing important losses in marine aquaculture. Vaccines against the pathogen can be a way to control the infection and avoid antibiotic treatments. However, a satisfactory protective vaccine against fish pasteurellosis is not commercially available. In this study, a biotechnogical approach based on reverse vaccinology has been used to identify potential vaccine candidates for the development of a recombinant subunit vaccine. Genome sequencing of clones from a genomic cosmid library of PDP and in silico selection of the surface exposed proteins were the initial steps in vaccine candidate identification. From 370 open reading frames (ORF) eight potential antigens were selected, expressed as recombinant proteins and purified. These vaccine candidates were used to generate specific polyclonal antibodies in mice. Each antibody was then screened in vitro by inhibition adherence assay of live PDP on chinook salmon embryo cells (CHSE-214). A lipoprotein, found to be involved in the adherence of the bacterium to epithelial cells and annotated as PDP_0080, was then selected. The recombinant protein was further investigated in fish vaccination and challenge experiments to assess its ability to protect sea bass, Dicentrarchus labrax, against PDP infection. Immunisation with PDP_0080 recombinant protein elicited high specific antibody titres. Furthermore, the survival rate of fish immunized with the 25 µg dose of protein was significantly higher compared to the control group. The results of the study suggest that the PDP_0080 protein could be a promising candidate for the design of a recombinant vaccine against pasteurellosis.

Susceptibility to antibiotics of Vibrio spp. and Photobacterium damsela ssp. piscicida strains isolated from Italian aquaculture farms.

The antibiotic resistance patterns of aetiological agents responsible for vibriosis and pasteurellosis were studied to contribute to control the spread of these two bacterial diseases in Mediterranean fish farming. Strains of Photobacterium damsela ssp. piscicida, Vibrio fluvialis, Vibrio alginolyticus, Vibrio parahaemolyticus, Vibrio metschnikovii, isolated from Italian aquaculture (fish, shellfish and crustaceans) sites, were assayed for their susceptibility to some antibacterial agents currently used in farming practices. Kirby Bauer and Minimum Inhibitory Concentration (M.I.C.) tests were performed. The bacterial strains showed resistance to ampicillin, carbenicillin, kanamycin, cefalothin, while they were sensitive to chloramphenicol, nitrofurantoin and tobramycin; the sulfadiazine-trimethoprim association was completely ineffective. Conversely, flumequine showed the lowest M.I.C. value (0.97 �g mL-1), suggesting its marked antibiotic effect. Considering that quinolone resistance can be transmitted only by selection of mutations and not by other genetic mechanisms, this study stresses the importance of a more responsible use of this antibacterial drug in aquaculture.

First report on antibody response of Seriola dumerilii (Risso 1810) challenged with Listonella anguillarum.

The greater amberjack, Seriola dumerilii (Risso 1810) is a semi-pelagic fish and a worldwide species; it is considered a promising candidate for the diversification of Mediterranean aquaculture. In this paper an experimental injection with Listonella (Vibrio) anguillarum was performed to study the immune response of S. dumerilii. Antibody titres to L. anguillarum O1 were determined with indirect-ELISA at different times over a period of 42 days. Results showed that the antibody levels against L. anguillarum were significantly higher in the challenged fish compared to the control. They started developing since the 5th day reaching the highest peak on day 20 after injection, indicating a fast response of the immune system. The observed antibody titre was very high versus L. anguillarum if compared to other fish species.