A Type IV Pilus Mediates DNA Binding during Natural Transformation in Streptococcus pneumoniae

Natural genetic transformation is widely distributed in bacteria and generally occurs during a genetically programmed differentiated state called competence. This process promotes genome plasticity and adaptability in Gram-negative and Gram-positive bacteria. Transformation requires the binding and internalization of exogenous DNA, the mechanisms of which are unclear. Here, we report the discovery of a transformation pilus at the surface of competent Streptococcus pneumoniae cells. This Type IV-like pilus, which is primarily composed of the ComGC pilin, is required for transformation. We provide evidence that it directly binds DNA and propose that the transformation pilus is the primary DNA receptor on the bacterial cell during transformation in S. pneumoniae. Being a central component of the transformation apparatus, the transformation pilus enables S. pneumoniae, a major Gram-positive human pathogen, to acquire resistance to antibiotics and to escape vaccines through the binding and incorporation of new genetic material.     

The Interactomes of Influenza Virus NS1 and NS2 Proteins Identify New Host Factors and Provide Insights for ADAR1 Playing a Supportive Role in Virus Replication

Influenza A NS1 and NS2 proteins are encoded by the RNA segment 8 of the viral genome. NS1 is a multifunctional protein and a virulence factor while NS2 is involved in nuclear export of viral ribonucleoprotein complexes. A yeast two-hybrid screening strategy was used to identify host factors supporting NS1 and NS2 functions. More than 560 interactions between 79 cellular proteins and NS1 and NS2 proteins from 9 different influenza virus strains have been identified. These interacting proteins are potentially involved in each step of the infectious process and their contribution to viral replication was tested by RNA interference. Validation of the relevance of these host cell proteins for the viral replication cycle revealed that 7 of the 79 NS1 and/or NS2-interacting proteins positively or negatively controlled virus replication. One of the main factors targeted by NS1 of all virus strains was double-stranded RNA binding domain protein family. In particular, adenosine deaminase acting on RNA 1 (ADAR1) appeared as a pro-viral host factor whose expression is necessary for optimal viral protein synthesis and replication. Surprisingly, ADAR1 also appeared as a pro-viral host factor for dengue virus replication and directly interacted with the viral NS3 protein. ADAR1 editing activity was enhanced by both viruses through dengue virus NS3 and influenza virus NS1 proteins, suggesting a similar virus-host co-evolution.     

Zinc Piracy as a Mechanism of Neisseria meningitidis for Evasion of Nutritional Immunity

The outer membrane of Gram-negative bacteria functions as a permeability barrier that protects these bacteria against harmful compounds in the environment. Most nutrients pass the outer membrane by passive diffusion via pore-forming proteins known as porins. However, diffusion can only satisfy the growth requirements if the extracellular concentration of the nutrients is high. In the vertebrate host, the sequestration of essential nutrient metals is an important defense mechanism that limits the growth of invading pathogens, a process known as “nutritional immunity.” The acquisition of scarce nutrients from the environment is mediated by receptors in the outer membrane in an energy-requiring process. Most characterized receptors are involved in the acquisition of iron. In this study, we characterized a hitherto unknown receptor from Neisseria meningitidis, a causative agent of sepsis and meningitis. Expression of this receptor, designated CbpA, is induced when the bacteria are grown under zinc limitation. We demonstrate that CbpA functions as a receptor for calprotectin, a protein that is massively produced by neutrophils and other cells and that has been shown to limit bacterial growth by chelating Zn²⁺ and Mn²⁺ ions. Expression of CbpA enables N. meningitidis to survive and propagate in the presence of calprotectin and to use calprotectin as a zinc source. Besides CbpA, also the TonB protein, which couples energy of the proton gradient across the inner membrane to receptor-mediated transport across the outer membrane, is required for the process. CbpA was found to be expressed in all N. meningitidis strains examined, consistent with a vital role for the protein when the bacteria reside in the host. Together, our results demonstrate that N. meningitidis is able to subvert an important defense mechanism of the human host and to utilize calprotectin to promote its growth.     

Interactions between peptidyl tRNA hydrolase homologs and the ribosomal release factor Mrf1 in S. pombe mitochondria

Mitochondrial translation synthesizes key subunits of the respiratory complexes. In Schizosaccharomyces pombe, strains lacking Mrf1, the mitochondrial stop codon recognition factor, are viable, suggesting that other factors can play a role in translation termination. S. pombe contains four predicted peptidyl tRNA hydrolases, two of which (Pth3 and Pth4), have a GGQ motif that is conserved in class I release factors. We show that high dosage of Pth4 can compensate for the absence of Mrf1 and loss of Pth4 exacerbates the lack of Mrf1. Also Pth4 is a component of the mitochondrial ribosome, suggesting that it could help recycling stalled ribosomes.     

Metabolomics identifies a biological response to chronic low-dose natural uranium contamination in urine samples

Because uranium is a natural element present in the earth’s crust, the population may be chronically exposed to low doses of it through drinking water. Additionally, the military and civil uses of uranium can also lead to environmental dispersion that can result in high or low doses of acute or chronic exposure. Recent experimental data suggest this might lead to relatively innocuous biological reactions. The aim of this study was to assess the biological changes in rats caused by ingestion of natural uranium in drinking water with a mean daily intake of 2.7 mg/kg for 9 months and to identify potential biomarkers related to such a contamination. Subsequently, we observed no pathology and standard clinical tests were unable to distinguish between treated and untreated animals. Conversely, LC–MS metabolomics identified urine as an appropriate biofluid for discriminating the experimental groups. Of the 1,376 features detected in urine, the most discriminant were metabolites involved in tryptophan, nicotinate, and nicotinamide metabolic pathways. In particular, N-methylnicotinamide, which was found at a level seven times higher in untreated than in contaminated rats, had the greatest discriminating power. These novel results establish a proof of principle for using metabolomics to address chronic low-dose uranium contamination. They open interesting perspectives for understanding the underlying biological mechanisms and designing a diagnostic test of exposure.     

The Rescue of miR-148a Expression in Pancreatic Cancer: An Inappropriate Therapeutic Tool

MicroRNAs are small non-coding RNAs that physiologically modulate proteins expression, and regulate numerous cellular mechanisms. Alteration of microRNA expression has been described in cancer and is associated to tumor initiation and progression. The microRNA 148a (miR-148a) is frequently down-regulated in cancer. We previously demonstrated that its down-regulation by DNA hypermethylation is an early event in pancreatic ductal adenocarcinoma (PDAC) carcinogenesis, suggesting a tumor suppressive function. Here, we investigate the potential role of miR-148a over-expression in PDAC as a therapeutic tool. We first report the consequences of miR-148a over-expression in PDAC cell lines. We demonstrate that miR-148a over-expression has no dramatic effect on cell proliferation and cell chemo-sensitivity in four well described PDAC cell lines. We also investigate the modulation of protein expression by a global proteomic approach (2D-DIGE). We show that despite its massive over-expression, miR-148a weakly modulates protein expression, thus preventing the identification of protein targets in PDAC cell lines. More importantly, in vivo data demonstrate that modulating miR-148a expression either in the epithelia tumor cells and/or in the tumor microenvironment does not impede tumor growth. Taken together, we demonstrate herein that miR-148a does not impact PDAC proliferation both in vitro and in vivo thus suggesting a weak potential as a therapeutic tool.     

Modulation of Protease Activated Receptor 1 Influences Human Metapneumovirus Disease Severity in a Mouse Model

Human metapneumovirus (hMPV) infection causes acute respiratory tract infections (RTI) which can result in hospitalization of both children and adults. To date, no antiviral or vaccine is available for this common viral infection. Immunomodulators could represent an interesting strategy for the treatment of severe viral infection. Recently, the role of protease-activated receptors (PAR) in inflammation, coagulation and infection processes has been of growing interest. Herein, the effects of a PAR1 agonist and a PAR1 antagonist on hMPV infection were investigated in BALB/c mice. Intranasal administration of the PAR1 agonist resulted in increased weight loss and mortality of infected mice. Conversely, the PAR1 antagonist was beneficial to hMPV infection by decreasing weight loss and clinical signs and by significantly reducing pulmonary inflammation, pro-inflammatory cytokine levels (including IL-6, KC and MCP-1) and recruitment of immune cells to the lungs. In addition, a significant reduction in pulmonary viral titers was also observed in the lungs of PAR1 antagonist-treated mice. Despite no apparent direct effect on virus replication during in vitro experiments, an important role for PAR1 in the regulation of furin expression in the lungs was shown for the first time. Further experiments indicated that the hMPV fusion protein can be cleaved by furin thus suggesting that PAR1 could have an effect on viral infectivity in addition to its immunomodulatory properties. Thus, inhibition of PAR1 by selected antagonists could represent an interesting strategy for decreasing the severity of paramyxovirus infections.     

A Systematic Review of Risk Factors Associated with Surgical Site Infections among Surgical Patients

Importance

Surgical site infection (SSI) complicates 2-5% of surgeries in the United States. Severity of SSI ranges from superficial skin infection to life-threatening conditions such as severe sepsis, and SSIs are responsible for increased morbidity, mortality, and economic burden associated with surgery. Staphylococcus aureus (S. aureus) is a commonly-isolated organism for SSI, and methicillin-resistant S. aureus SSI incidence is increasing globally.

Objective

The objective of this systematic review was to characterize risk factors for SSI within observational studies describing incidence of SSI in a real-world setting.

Evidence Review

An initial search identified 328 titles published in 2002-2012; 57 were identified as relevant for data extraction. Extracted information included study design and methodology, reported cumulative incidence and post-surgical time until onset of SSI, and odds ratios and associated variability for all factors considered in univariate and/or multivariable analyses.

Findings

Median SSI incidence was 3.7%, ranging from 0.1% to 50.4%. Incidence of overall SSI and S. aureus SSI were both highest in tumor-related and transplant surgeries. Median time until SSI onset was 17.0 days, with longer time-to-onset for orthopedic and transplant surgeries. Risk factors consistently identified as associated with SSI included co-morbidities, advanced age, risk indices, patient frailty, and surgery complexity. Thirteen studies considered diabetes as a risk factor in multivariable analysis; 85% found a significant association with SSI, with odds ratios ranging from 1.5-24.3. Longer surgeries were associated with increased SSI risk, with a median odds ratio of 2.3 across 11 studies reporting significant results.

Conclusions and Relevance

In a broad review of published literature, risk factors for SSI were characterized as describing reduced fitness, patient frailty, surgery duration, and complexity. Recognition of risk factors frequently associated with SSI allows for identification of such patients with the greatest need for optimal preventive measures to be identified and pre-treatment prior to surgery.     

Element content and expression of genes of interest in guard cells are connected to spatiotemporal variations in stomatal conductance

Element content and expression of genes of interest on single cell types, such as stomata, provide valuable insights into their specific physiology, improving our understanding of leaf gas exchange regulation. We investigated how far differences in stomatal conductance (g_s) can be ascribed to changes in guard cells functioning in amphistomateous leaves. g_s was measured during the day on both leaf sides, on well-watered and drought-stressed trees (two Populus euramericana Moench and two Populus nigra L. genotypes). In parallel, guard cells were dissected for element content and gene expressions analyses. Both were strongly arranged according to genotype, and drought had the lowest impact overall. Normalizing the data by genotype highlighted a structure on the basis of leaf sides and time of day both for element content and gene expression. Guard cells magnesium, phosphorus, and chlorine were the most abundant on the abaxial side in the morning, where g_s was at the highest. In contrast, genes encoding H⁺-ATPase and aquaporins were usually more abundant in the afternoon, whereas genes encoding Ca²⁺-vacuolar antiporters, K⁺ channels, and ABA-related genes were in general more abundant on the adaxial side. Our work highlights the unique physiology of each leaf side and their analogous rhythmicity through the day.