Prokaryotic abundance and heterotrophic metabolism in the deep Mediterranean Sea

A synthesis of field data carried out in the Mediterranean Sea are presented, aimed at contributing to the knowledge of three prokaryotic-mediated processes and their implications on the Carbon cycle. The distribution of exoenzymatic activities, secondary production and respiration rates was studied together with the prokaryotic abundances. Particular attention was paid to the meso- and bathypelagic layers which play an important role in the Mediterranean carbon cycle. The study is noteworthy because of its large spatial scale spanning the entire Mediterranean Sea over 4 years. In addition, two Atlantic stations in front of the Gibraltar Strait were investigated. The longitudinal distribution of prokaryotic activities and abundance along the MED showed different trends along the depthlayers. In particular, higher exoenzymatic rates were detected in the Eastern basin compared to the Western one; carbon respiration rate showed patterns variable with the sampling periods in the epipelagic and bathypelagic layers, while a consistent Westwards decreasing trend at the mesopelagic layers occurred. Specific enzyme activities per cell showed high values in the deepest layers for leucine aminopeptidase. Comparison with Carbon respiration rate data collected before the 2000s showed changing patterns of microbial heterotrophic processes in the Mediterranean Sea

Modelling approach to the assessment of biogenic fluxes at a selected Ross Sea site, Antarctica

Several biogeochemical data have been collected in the last 10 years of Italian activity in Antarctica (ABIOCLEAR, ROSSMIZE, BIOSESO-I/II). A comprehensive 1-D biogeochemical model was implemented as a tool to link observations with processes and to investigate the mechanisms that regulate the flux of biogenic material through the water column. The model is ideally located at station B (175° E–74° S) and was set up to reproduce the seasonal cycle of phytoplankton and organic matter fluxes as forced by the dominant water column physics over the period 1990–2001. Austral spring-summer bloom conditions are assessed by comparing simulated nutrient drawdown, primary production rates, bacterial respiration and biomass with the available observations. The simulated biogenic fluxes of carbon, nitrogen and silica have been compared with the fluxes derived from sediment traps data. The model reproduces the observed magnitude of the biogenic fluxes, especially those found in the bottom sediment trap, but the peaks are markedly delayed in time. Sensitivity experiments have shown that the characterization of detritus, the choice of the sinking velocity and the degradation rates are crucial for the timing and magnitude of the vertical fluxes. An increase of velocity leads to a shift towards observation but also to an overestimation of the deposition flux which can be counteracted by higher bacterial remineralization rates. Model results suggest that the timing of the observed fluxes depends first and foremost on the timing of surface production and on a combination of size-distribution and quality of the autochtonous biogenic material. It is hypothesized that the bottom sediment trap collects material originated from the rapid sinking of freshly-produced particles and also from the previous year's production period

Modelling approach to the assessment of biogenic fluxes at a selected Ross Sea site, Antarctica

Abstract Several biogeochemical data have been collected in the last 10 years of Italian activity in Antarctica (ABIOCLEAR, ROSSMIZE, BIOSESO-I/II). A comprehensive 1-D biogeochemical model was implemented as a tool to link observations with processes and to investigate the mechanisms that regulate the flux of biogenic material through the water column. The model is ideally located at station B (175^{o}E - 74^{o}S) and was set up to reproduce the seasonal cycle of phytoplankton and organic matter fluxes as forced by the dominant water column physics over the period 1990-2001. Austral spring-summer bloom conditions are assessed by comparing simulated nutrient drawdown, primary production rates, bacterial respiration and biomass with the available observations. The simulated biogenic fluxes of carbon, nitrogen and silica have been compared with the fluxes derived from sediment traps data. The model reproduces quite well the magnitude of the biogenic fluxes, expecially those observed in the bottom sediment trap, but the peaks are delayed in time. Sensitivity experiments have shown that the characterization of detritus, the choice of the sinking velocity and the degradation rates are crucial for the timing and magnitude of the vertical fluxes. An increase of velocity leads to a shift towards observation but also to an overestimation of the deposition flux which can be counteracted by higher bacterial remineralization rates. Model results suggest that observed fluxes could be explained by the size-distribution and quality of the locally-produced biogenic material. It is hypothesized that the bottom sediment trap collects material originated from rapid sinking of particles and also from previous years production periods, likely modulated by advective and aggregation mechanisms which are still not resolved by the model

Microbial assemblages in pressurized antarctic brine pockets (Tarn flat, northern Victoria land): A hotspot of biodiversity and activity

Two distinct pressurized hypersaline brine pockets (named TF4 and TF5), separated by a thin ice layer, were detected below an ice-sealed Antarctic lake. Prokaryotic (bacterial and archaeal) diversity, abundances (including virus-like particles) and metabolic profiles were investigated by an integrated approach, including traditional and new-generation methods. Although similar diversity indices were computed for both Bacteria and Archaea, distinct bacterial and archaeal assemblages were observed. Bacteroidetes and Gammaproteobacteria were more abundant in the shallowest brine pocket, TF4, and Deltaproteobacteria, mainly represented by versatile sulphate-reducing bacteria, dominated in the deepest, TF5. The detection of sulphate-reducing bacteria and methanogenic Archaea likely reflects the presence of a distinct synthrophic consortium in TF5. Surprisingly, members assigned to hyperthermophilic Crenarchaeota and Euryarchaeota were common to both brines, indicating that these cold habitats host the most thermally tolerant Archaea. The patterns of microbial communities were different, coherently with the observed microbiological diversity between TF4 and TF5 brines. Both the influence exerted by upward movement of saline brines from a sub-surface anoxic system and the possible occurrence of an ancient ice remnant from the Ross Ice Shelf were the likely main factors shaping the microbial communities

Molecular analysis of TP53, Ki-Ras and P16 methylation status in tissue and plasma of subjects affected by gastrointestinal cancer (GIC)

BACKGROUND: Despite the improvement in detection and surgical therapy in the last years, the outcome of patients affected by colorectal carcinoma (CRC) remains limited by metastatic relapse. The aim of this study was to investigate the presence of free tumor DNA in the plasma of CRC patients in order to understand its possible prognostic role. PATIENTS AND METHODS: Ki-Ras, TP53 mutations and p16(INK4A) methylation status were prospectively evaluated in tumor tissues and plasma of 66 CRC patients. RESULTS: In 50 of the 66 primitive tumor cases (76%) at least one significant alteration was identified in Ki-Ras and/or TP53 and/or p16(INK4A) genes. Eighteen of the 50 patients presented the same alteration both in the plasma and in the tumor tissue. At univariate analysis, Ki-Ras mutations proved to be significantly related to quicker relapse (P <0.01), whereas only a trend towards statistical significance (P = 0.083) was observed for the TP53 mutations CONCLUSIONS: Detection of Ki-Ras and TP53 mutation in plasma should be significantly related to disease recurrence. These data suggest that patients with a high risk of recurrence can be identified by means of the analysis of tumor-derived plasma DNA with the use of fairly non-invasive techniques

Interaction of Chandipura Virus N and P Proteins: Identification of Two Mutually Exclusive Domains of N Involved in Interaction with P

The nucleocapsid protein (N) and the phosphoprotein (P) of nonsegmented negative-strand (NNS) RNA viruses interact with each other to accomplish two crucial events necessary for the viral replication cycle. First, the P protein binds to the aggregation prone nascent N molecules maintaining them in a soluble monomeric (N0) form (N0-P complex). It is this form that is competent for specific encapsidation of the viral genome. Second, the P protein binds to oligomeric N in the nucleoprotein complex (N-RNA-P complex), and thereby facilitates the recruitment of the viral polymerase (L) onto its template. All previous attempts to study these complexes relied on co-expression of the two proteins in diverse systems. In this study, we have characterised these different modes of N-P interaction in detail and for the first time have been able to reconstitute these complexes individually in vitro in the chandipura virus (CHPV), a human pathogenic NNS RNA virus. Using a battery of truncated mutants of the N protein, we have been able to identify two mutually exclusive domains of N involved in differential interaction with the P protein. An unique N-terminal binding site, comprising of amino acids (aa) 1–180 form the N0-P interacting region, whereas, C-terminal residues spanning aa 320–390 is instrumental in N-RNA-P interactions. Significantly, the ex-vivo data also supports these observations. Based on these results, we suggest that the P protein acts as N-specific chaperone and thereby partially masking the N-N self-association region, which leads to the specific recognition of viral genome RNA by N0

NEDDylation is essential for Kaposi's sarcoma-associated herpesvirus latency and lytic reactivation and represents a novel anti-KSHV target.

Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of Kaposi's sarcoma (KS) and primary effusion lymphoma (PEL), which are aggressive malignancies associated with immunocompromised patients. For many non-viral malignancies, therapeutically targeting the ubiquitin proteasome system (UPS) has been successful. Likewise, laboratory studies have demonstrated that inhibition of the UPS might provide a promising avenue for the treatment of KSHV-associated diseases. The largest class of E3 ubiquitin ligases are the cullin-RING ligases (CRLs) that are activated by an additional ubiquitin-like protein, NEDD8. We show that pharmacological inhibition of NEDDylation (using the small molecule inhibitor MLN4924) is cytotoxic to PEL cells by inhibiting NF-κB. We also show that CRL4B is a novel regulator of latency as its inhibition reactivated lytic gene expression. Furthermore, we uncovered a requirement for NEDDylation during the reactivation of the KSHV lytic cycle. Intriguingly, inhibition prevented viral DNA replication but not lytic cycle-associated gene expression, highlighting a novel mechanism that uncouples these two features of KSHV biology. Mechanistically, we show that MLN4924 treatment precluded the recruitment of the viral pre-replication complex to the origin of lytic DNA replication (OriLyt). These new findings have revealed novel mechanisms that regulate KSHV latency and reactivation. Moreover, they demonstrate that inhibition of NEDDylation represents a novel approach for the treatment of KSHV-associated malignancies

Inhibition of nuclear factor kappa-B signaling reduces growth in medulloblastoma in vivo

Abstract Background Medulloblastoma is a highly malignant pediatric brain tumor that requires surgery, whole brain and spine irradiation, and intense chemotherapy for treatment. A more sophisticated understanding of the pathophysiology of medulloblastoma is needed to successfully reduce the intensity of treatment and improve outcomes. Nuclear factor kappa-B (NFκB) is a signaling pathway that controls transcriptional activation of genes important for tight regulation of many cellular processes and is aberrantly expressed in many types of cancer. Methods To test the importance of NFκB to medulloblastoma cell growth, the effects of multiple drugs that inhibit NFκB, pyrrolidine dithiocarbamate, diethyldithiocarbamate, sulfasalazine, curcumin and bortezomib, were studied in medulloblastoma cell lines compared to a malignant glioma cell line and normal neurons. Expression of endogenous NFκB was investigated in cultured cells, xenograft flank tumors, and primary human tumor samples. A dominant negative construct for the endogenous inhibitor of NFκB, IκB, was prepared from medulloblastoma cell lines and flank tumors were established to allow specific pathway inhibition. Results We report high constitutive activity of the canonical NFκB pathway, as seen by Western analysis of the NFκB subunit p65, in medulloblastoma tumors compared to normal brain. The p65 subunit of NFκB is extremely highly expressed in xenograft tumors from human medulloblastoma cell lines; though, conversely, the same cells in culture have minimal expression without specific stimulation. We demonstrate that pharmacological inhibition of NFκB in cell lines halts proliferation and leads to apoptosis. We show by immunohistochemical stain that phosphorylated p65 is found in the majority of primary tumor cells examined. Finally, expression of a dominant negative form of the endogenous inhibitor of NFκB, dnIκB, resulted in poor xenograft tumor growth, with average tumor volumes 40% smaller than controls. Conclusions These data collectively demonstrate that NFκB signaling is important for medulloblastoma tumor growth, and that inhibition can reduce tumor size and viability in vivo. We discuss the implications of NFκB signaling on the approach to managing patients with medulloblastoma in order to improve clinical outcomes.</p

Complete Genome Sequence of Crohn's Disease-Associated Adherent-Invasive E. coli Strain LF82

International audienceBACKGROUND: Ileal lesions of Crohn's disease (CD) patients are abnormally colonized by pathogenic adherent-invasive Escherichia coli (AIEC) able to invade and to replicate within intestinal epithelial cells and macrophages. PRINCIPAL FINDINGS: We report here the complete genome sequence of E. coli LF82, the reference strain of adherent-invasive E. coli associated with ileal Crohn's disease. The LF82 genome of 4,881,487 bp total size contains a circular chromosome with a size of 4,773,108 bp and a plasmid of 108,379 bp. The analysis of predicted coding sequences (CDSs) within the LF82 flexible genome indicated that this genome is close to the avian pathogenic strain APEC_01, meningitis-associated strain S88 and urinary-isolated strain UTI89 with regards to flexible genome and single nucleotide polymorphisms in various virulence factors. Interestingly, we observed that strains LF82 and UTI89 adhered at a similar level to Intestine-407 cells and that like LF82, APEC_01 and UTI89 were highly invasive. However, A1EC strain LF82 had an intermediate killer phenotype compared to APEC-01 and UTI89 and the LF82 genome does not harbour most of specific virulence genes from ExPEC. LF82 genome has evolved from those of ExPEC B2 strains by the acquisition of Salmonella and Yersinia isolated or clustered genes or CDSs located on pLF82 plasmid and at various loci on the chromosome. CONCLUSION: LF82 genome analysis indicated that a number of genes, gene clusters and pathoadaptative mutations which have been acquired may play a role in virulence of AIEC strain LF82

A MSFD complementary approach for the assessment of pressures, knowledge and data gaps in Southern European Seas : the PERSEUS experience

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. Published by Elsevier Ltd. This is an open access article under the CC BY license.peer-reviewe