An Integrated Geometric and Material Survey for the Conservation of Heritage Masonry Structures

This paper reports the knowledge process and the analyses performed to assess the seismic behavior of a heritage masonry building. The case study is a three-story masonry building that was the house of the Renaissance architect and painter Giorgio Vasari (the Vasari’s House museum). An interdisciplinary approach was adopted, following the Italian “Guidelines for the assessment and mitigation of the seismic risk of the cultural heritage”. This document proposes a methodology of investigation and analysis based on three evaluation levels (EL1, analysis at territorial level; EL2, local analysis and EL3, global analysis), according to an increasing level of knowledge on the building. A comprehensive knowledge process, composed by a 3D survey by Terrestrial Laser Scanning (TLS) and experimental in situ tests, allowed us to identify the basic structural geometry and to assess the value of mechanical parameters subsequently needed to perform a reliable structural assessment. The museum represents a typology of masonry building extremely diffused in the Italian territory, and the assessment of its seismic behavior was performed by investigating its global behavior through the EL1 and the EL3 analyses

Comparison of the action of different proteases on virulence properties related to the staphylococcal surface.

AIMS: The purpose of this study was to evaluate the antimicrobial efficacy of five different proteases belonging to two different families on Staphylococcus aureus and Staphylococcus epidermidis strains. METHODS AND RESULTS: We used three serine proteases and two metalloproteases in single species biofilm formation assays and in human cell invasion processes. Following each protease incubation with bacterial cells, surface protein patterns were analysed by SDS-PAGE and zymography. Some differently expressed proteins were identified by mass spectrometry. CONCLUSIONS: The effect of tested proteases on biofilm formation was not related to the protease category but was strain-dependent and was related to the biofilm formation capacity of each staphylococcal strain. Some proteases showed a nonspecific and indiscriminate effect on surface proteins, while others induced a discrete and reproducible action on protein profiles. SIGNIFICANCE AND IMPACT OF THE STUDY: The inhibition of the surface-related virulence factors is a promising avenue to overcome persistent infections caused by bacterial biofilms. To this end, we show here that proteases, in particular the metalloprotease serratiopeptidase, can interfere with adhesion and invasion of eukaryotic cells and biofilm formation in staphylococci and their use could represent a viable treatment for the development of novel combination therapie

A new anti-infective strategy to reduce the spreading of antibiotic resistance by the action on adhesion-mediated virulence factors in Staphylococcus aureus.

Staphylococcus aureus is a flexible microbial pathogen frequently isolated from community-acquired and nosocomial infections. S. aureus expresses a wide array of secreted and cell surface-associated virulence factors, including proteins that promote adhesion to damaged tissue and to the surface of host cells, and that bind proteins in blood to help evade immune responses. Furthermore, surface proteins have a fundamental role in virulence related properties of S. aureus, including biofilm formation. The present study evaluates the anti-infective capabilities of a secreted protein of Serratia marcescens (serratiopeptidase, SPEP), in impairing some staphylococcal virulence-related properties, such as attachment to inert surfaces and adhesion/invasion on eukaryotic cells. SPEP seems to exert its action by modulating specific proteins. It is not assessed if this action is due to the proteolytic activity of SPEP or to a specific mechanism which triggers an out/inside signal. Proteomic studies performed on surface proteins extracted from SPEP treated S. aureus cultures revealed that a number of proteins are affected by the treatment. Among these we found the adhesin/autolysin Atl, SdrD, Sbi, EF-Tu and EF-G. EF-Tu and EF-G are known to perform a variety of function, depending on their cytoplasmic or surface localization. All these factors can facilitate bacterial colonization, persistence and invasion of host tissues. Our results suggest that SPEP could be developed as a potential "anti-infective agent" capable to hinder the entry of S. aureus into human tissues, and also impairs the ability of this pathogen to adhere to prostheses, catheters and medical device

A simple and reliable methodology to detect egg white in art samples

A protocol for a simple and reliable dot-blot immunoassay was developed and optimized to test work of art samples for the presence of specific proteinaceus material (i.e. ovalbumin-based). The analytical protocol has been extensively set up with respect, among the other, to protein extraction conditions, to densitometric analysis and to the colorimetric reaction conditions. Feasibility evaluation demonstrated that a commercial scanner and a free image analysis software can be used for the data acquisition and elaboration, thus facilitating the application of the proposed protocol to commonly equipped laboratories and to laboratories of museums and conservation centres. The introduction of method of standard additions in the analysis of fresh and artificially aged laboratory-prepared samples, containing egg white and various pigments, allowed us to evaluate the matrix effect and the effect of sample aging and to generate threshold density values useful for the detection of ovalbumin in samples from ancient works of art. The efficacy of the developed dot-blot immunoassay was proved testing microsamples from 13th–16th century mural paintings of Saint Francesco Church in Lodi (Italy). Despite the aging, the altered conditions of conservation, the complex matrix, and the micro-size of samples, the presence of ovalbumin was detected in all those mural painting samples where mass-spectrometry-based proteomic analysis unambiguously detected ovalbumin peptides

Shape oscillations of an oil drop rising in water: effect of surface contamination

Inertial shape oscillations of heptane drops rising in water are investigated experimentally. Diameters from 0.59 to 3.52 mm are considered, corresponding to a regime where the rising motion should not affect shape oscillations for pure immiscible fluids. The interface, however, turns out to be contaminated. The drag coefficient is considerably increased compared to that of a clean drop due to the well-known. Marangoni effect resulting from a gradient of surfactant concentration generated by the fluid motion along the interface. Thanks to the decomposition of the shape into spherical harmonics, the eigenfrequencies and the damping rates of oscillation modes n = 2, 3, 4 and 5 have been measured. Frequencies are not affected by contamination, while damping rates are increased by a considerable amount that depends neither on drop instantaneous velocity nor on diameter. This augmentation, however, depends on the mode number: it is maximum for mode two (multiplied by 2.4) and then relaxes towards the value of a clean drop as n increases. A previous similar investigation of a drop attached to a capillary has not revealed such an increase of the damping rates, indicating that the coupling between rising motion and surface contamination is responsible for this effect

Redox imbalance and morphological changes in skin fibroblasts in typical Rett syndrome.

Evidence of oxidative stress has been reported in the blood of patients with Rett syndrome (RTT), a neurodevelopmental disorder mainly caused by mutations in the gene encoding the Methyl-CpG-binding protein 2. Little is known regarding the redox status in RTT cellular systems and its relationship with the morphological phenotype. In RTT patients (n = 16) we investigated four different oxidative stress markers, F2-Isoprostanes (F2-IsoPs), F4-Neuroprostanes (F4-NeuroPs), nonprotein bound iron (NPBI), and (4-HNE PAs), and glutathione in one of the most accessible cells, that is, skin fibroblasts, and searched for possible changes in cellular/intracellular structure and qualitative modifications of synthesized collagen. Significantly increased F4-NeuroPs (12-folds), F2-IsoPs (7.5-folds) NPBI (2.3-folds), 4-HNE PAs (1.48-folds), and GSSG (1.44-folds) were detected, with significantly decreased GSH (−43.6%) and GSH/GSSG ratio (−3.05 folds). A marked dilation of the rough endoplasmic reticulum cisternae, associated with several cytoplasmic multilamellar bodies, was detectable in RTT fibroblasts. Colocalization of collagen I and collagen III, as well as the percentage of type I collagen as derived by semiquantitative immunofluorescence staining analyses, appears to be significantly reduced in RTT cells. Our findings indicate the presence of a redox imbalance and previously unrecognized morphological skin fibroblast abnormalities in RTT patients

DEVELOPING AN INTEGRATED OMICS TECHNOLOGY TO ELUCIDATE BIOLOGICAL MECHANIISM OF METAL EXPOSURE

About 30% of all annotated proteins reported in the major databases (PDB, NCBInr,and Uniprot) results to be associated with a metal. In some cases the functions of these proteins strongly depend by the interaction with the right metal. Metal ions, associated with macromolecules, are utilized by biological systems in fundamental processes; therefore cytosolic concentrations of metals are tightly controlled. For a complete characterization of cell chemistry, the understanding of mechanisms by which a metal is sensed, stored, or incorporated as a cofactor is required. Some metals, such as Ca, Co, Cr, Cu, Fe, K, Mg, Mn, Na, Ni and Zn, are essential,in fact they serve as micronutrients and are used for redox-processes, to stabilize molecules through electrostatic interactions, as components of various enzymes and for regulation of osmotic pressure. Many other metals, in particular heavy metals, have no biological role (e.g. Ag, Al, Cd, Au, Pb and Hg), and are nonessential and potentially toxic to mammalians and microorganisms. Selenium is a crucial micronutrient for human health. In human Se is a component of several important selenoproteins and enzymes required for different functions, such as antioxidant defense and reduction of inflammation. Several organisms possess the ability to metabolize Se into protein by a specific pathway dedicated to the biosynthesis of proteins which contain the aminoacid selenocysteine (Sec, U), or by a non-specific mechanism. The improvements in high-throughput technology, has led to born of a large number of new “omics” disciplines with the aim to expand the knowledge acquired. Thus “metallomics” was born; this term, coined by Haraguchi, denotes the ensemble of research activities related to metals of biological interest. However detailed knowledge of interaction between metals and proteins within organisms is fundamental to prevent diseases related to heavy metals pollutions, or to use some detoxification process to produce more bio-available forms of selenocontaining compounds. This PhD thesis targeted the characterization of metals involved in relevant biological processes by metallomics and proteomics approaches. Thus the following systems of biotechnological interest have been analyzed in this study: 1) Escherichia coli used as model organism for a study of the effects of heavy-metals contamination. The development of an integrated metallomics-proteomics approach for the investigation of the effects of cadmium exposure on Escherichia coli is reported. 2) HRCE (Human Renal Cortical Epithelial) cells for an investigation of the effects to cadmium and lead exposure. Apoptotic pathway was demonstrated to be induced by either cadmium or lead, with the latter the more toxic for human primary renal cells. 3) Lactobacillus reuteri Lb2 BM-DSM 16143 to study the incorporation of selenium into selenocysteines by an integrated ICP-MS based approach. Characterization of the ability of this probiotic microorganism to recover high concentration of selenium and convert them into selenocysteines residues, within the primary sequence of some proteins, is reported, representing an additional innovative approach to solve human selenium deficiency