Evolution of the lactic acid bacterial biofilms on different wood typologies applied for the production of vats intended for cheese making

The present work was carried out to evaluate the lactic acid bacterial biofilms formation on wooden vat produced with seven different woods, derived from trees species grown in Sicily. The biofilm formation on the wooden surfaces was performed as reported by Gaglio et al. Microbiological and scanning electron microscopy analyses did not show differences in terms of microbial levels and composition within the neoformed biofilms (Fig.1). The specific investigation of Salmonella spp., Listeria monocytogenes, Escherichia coli and positive-coagulase staphylococci did not generate any colony for all vats before and after microbial activation. Lactic acid bacteria (LAB) populations dominated the surfaces of all vats and the highest concentration of approximately 7.63 Log CFU/cm2 was registered for thermophilic cocci. All colonies of different morphologies were isolated and characterized for their physiological and biochemical characteristics before being investigated genetically at strain level by RAPD-PCR and species level by a polyphasic approach consisting of 16S rRNA gene sequencing. As reported in a preview investigation conducted on wooden vat surfaces. we found species within the genera Enterococcus, Lactobacillus, Lactococcus, and Streptococcus. The species most frequently present were Lactobacillus fermentum and Lactococcus lactis. The technological characterization of the LAB found at high numbers on to the surfaces of the wooden vats showed interesting dairy properties. In order to deeply investigate on the safety of the wooden vat, a test of artificial contamination on new Calabrian chestnut vats was carried out showing that the acidity and the bacteriocin generated by LAB represent efficient barriers to their adhesion. This study highlights the importance to use the wooden vats for traditional cheese production and provides evidences for their safety in terms of bacterial community evolution

Two distinct conformational states define the interaction of human RAD51-ATP with single-stranded DNA.

An essential mechanism for repairing DNA double-strand breaks is homologous recombination (HR). One of its core catalysts is human RAD51 (hRAD51), which assembles as a helical nucleoprotein filament on single-stranded DNA, promoting DNA-strand exchange. Here, we study the interaction of hRAD51 with single-stranded DNA using a single-molecule approach. We show that ATP-bound hRAD51 filaments can exist in two different states with different contour lengths and with a free-energy difference of ~4 kBT per hRAD51 monomer. Upon ATP hydrolysis, the filaments convert into a disassembly-competent ADP-bound configuration. In agreement with the single-molecule analysis, we demonstrate the presence of two distinct protomer interfaces in the crystal structure of a hRAD51-ATP filament, providing a structural basis for the two conformational states of the filament. Together, our findings provide evidence that hRAD51-ATP filaments can exist in two interconvertible conformational states, which might be functionally relevant for DNA homology recognition and strand exchange

Formation and Characterization of Early Bacterial Biofilms on Different Wood Typologies Applied in Dairy Production

The main hypothesis of this work was that Sicilian forestry resources are suitable for the production of equipment to be used in cheese making and indigenous milk lactic acid bacteria (LAB) are able to develop stable biofilms providing starter and nonstarter cultures necessary for curd fermentation and cheese ripening, respectively. Hence, the present work was carried out with deproteinized whey to evaluate LAB biofilm formation on different woods derived from tree species grown in Sicily. Microbiological and scanning electron microscopy analyses showed minimal differences in microbial levels and compositions for the neoformed biofilms. The specific investigation of Salmonella spp., Listeria monocytogenes, Escherichia coli, coagulase-positive staphylococci (CPS), and sulfite-reducing anaerobes did not generate any colony for all vats before and after bacterial adhesion. LAB populations dominated all vat surfaces. The highest levels (7.63 log CFU/cm2) were registered for thermophilic cocci. Different colonies were characterized physiologically, biochemically, and genetically (at strain and species levels). Six species within the genera Enterococcus, Lactobacillus, Lactococcus, and Streptococcus were identified. The species most frequently present were Lactobacillus fermentum and Lactococcus lactis. LAB found on the surfaces of the wooden vats in this study showed interesting characteristics important for dairy manufacture. To thoroughly investigate the safety of the wooden vat, a test of artificial contamination on new Calabrian chestnut (control wood) vats was carried out. The results showed that LAB represent efficient barriers to the adhesion of the main dairy pathogens, probably due to their acidity and bacteriocin generation. IMPORTANCE This study highlights the importance of using wooden vats for traditional cheese production and provides evidence for the valorization the Sicilian forest wood resources via the production of dairy equipment

Covid-19 And Rheumatic Autoimmune Systemic Diseases: Role of Pre-Existing Lung Involvement and Ongoing Treatments

The Covid-19 pandemic may have a deleterious impact on patients with autoimmune systemic diseases (ASD) due to their deep immune-system alterations

Is neuroglobin a signal transducer?

[No abstract available

Neuroglobin: Enzymatic reduction and oxygen affinity

Neuroglobin (Ngb) is a hexacoordinate globin expressed in the nervous system of vertebrates, involved in neuroprotection. O-2 equilibrium measurements on mouse Ngb yielded significantly different P-50 values, ranging from similar to 2 torr to similar to 10 torr. By a kinetic approach minimizing the effects of protein autoxidation, we measured P-50 = 2.2 torr at 20 degrees C. As predicted from the structure, O-2 binds to the Y44D Ngb mutant more quickly (k = 2.2 s(-1) vs 0.15 s(-1)) and with slightly higher affinity (P-50 = 1.3 torr) than wild-type. In addition, we introduced a novel reduction protocol for metNgb based on NADH:flavorubredoxin oxidoreductase (FlRd-red) from Escherichia. coli, a candidate for the Ngb reducing activity recently identified in E coli extracts. Interestingly, E coli FlRd-red shares sequence similarity with the FAD-binding domain of the human apoptosis-inducing factor, a finding which may have unexpected significance with reference to the mechanism of neuroprotection by Ngb. (c) 2008 Elsevier Inc. All rights reserved

Failure of apoptosis-inducing factor to act as neuroglobin reductase

International audienceNeuroglobin (Ngb) is a hexacoordinate globin expressed in the nervous system of vertebrates, where it protects neurons against hypoxia. Ferrous Ngb has been proposed to favor cell survival by scavenging NO and/or reducing cytochrome c released into the cytosol during hypoxic stress. Both catalytic functions require an as yet unidentified Ngb-reductase activity. Such an activity was detected both in tissue homogenates of human brain and liver and in Escherichia coli extracts. Since NADH:flavorubredoxin oxidoreductase from E. coli, that was shown to reduce ferric Ngb, shares sequence similarity with the human apoptosis-inducing factor (AIF), AIF has been proposed by us as a candidate Ngb reductase. In this study, we tested this hypothesis and show that the Ngb-reductase activity of recombinant human AIF is negligible and hence incompatible with such a physiological function

An X-ray diffraction and X-ray absorption spectroscopy joint study of neuroglobin.

Neuroglobin (Ngb) is a member of the globin family expressed in the vertebrate brain, involved in neuroprotection. A combined approach of X-ray diffraction (XRD) on single crystal and X-ray absorption spectroscopy (XAS) in solution, allows to determine the oxidation state and the structure of the Fe-heme both in the bis-histidine and the CO-bound (NgbCO) states. The overall data demonstrate that under Xray the iron is photoreduced fairly rapidly, and that the previously reported X-ray structure of ferric Ngb [B. Vallone, K. Nienhaus, M. Brunori, G.U. Nienhaus, Proteins 56 (2004) 85-92] very likely refers to a photoreduced species indistinguishable from the dithionite reduced protein. Results from the XAS analysis of NgbCO in solution are in good agreement with XRD data on the crystal. However prolonged X-ray exposure at 15 K determines CO release. This preliminary result paves the way to experiments aimed at the characterization of pentacoordinate ferrous Ngb, the only species competent in binding external ligands such as O-2, CO or N

Engineering the internal cavity of neuroglobin demonstrates the role of the haem-sliding mechanism

Neuroglobin is a member of the globin family involved in neuroprotection; it is primarily expressed in the brain and retina of vertebrates. Neuroglobin belongs to the heterogeneous group of hexacoordinate globins that have evolved in animals, plants and bacteria that are endowed with the capability of reversible intramolecular coordination, allowing the binding of small gaseous ligands (O2, NO and CO). In a unique fashion among haemoproteins, ligand-binding events in neuroglobin are dependent on the sliding of the haem itself within a preformed internal cavity, as revealed by the crystal structure of its CO-bound derivative. Point mutants of the neuroglobin internal cavity have been engineered and their functional and structural characterization shows that hindering the haem displacement leads to a decrease in CO affinity, whereas reducing the cavity volume without interfering with haem sliding has negligible functional effects.Neuroglobin is a member of the globin family involved in neuroprotection; it is primarily expressed in the brain and retina of vertebrates. Neuroglobin belongs to the heterogeneous group of hexacoordinate globins that have evolved in animals, plants and bacteria, endowed with the capability of reversible intramolecular coordination, allowing the binding of small gaseous ligands (O2, NO and CO). In a unique fashion among haemoproteins, ligand-binding events in neuroglobin are dependent on the sliding of the haem itself within a preformed internal cavity, as revealed by the crystal structure of its CO-bound derivative. Point mutants of the neuroglobin internal cavity have been engineered and their functional and structural characterization shows that hindering the haem displacement leads to a decrease in CO affinity, whereas reducing the cavity volume without interfering with haem sliding has negligible functional effects