Non-Destructive Evaluation of Thermal Spray Coating Interface Quality by Eddy Current Method

Thermal spray coating is usually applied through directing molten or softened particles at very high velocities onto a substrate. An eddy current non-destructive inspection technique is presented here for thermal spray coating interface quality characterization. Several high-velocity-oxy-fuel (HVOF) coated steel plates were produced with different surface preparation conditions before applying the coating, e.g., grit-blasted surface, wire-brush cleaned surface, and a dirty surface. A quad-frequency eddy current probe was used to manually scan over the coating surface to evaluate the bonding quality. Experimental results show that the three surface preparation conditions can be successfully differentiated by looking into the impedance difference observed from the eddy current probe. The measurement is fairly robust and consistent. More specimens are also prepared with variations of process parameters, such as spray angle, stand-off distance, and application of corrosion protective sealant, etc. They are blindly tested to evaluate the reliability of the eddy current system. Quantitative relations between the coating bond strength and the eddy current response are also established with the support of destructive testing. This non-contact, non-destructive, easy to use technique has the potential for evaluating the coating quality immediately after its application so that any defects can be corrected immediately

Temporal and stochastic control of Staphylococcus aureus biofilm development.

Biofilm communities contain distinct microniches that result in metabolic heterogeneity and variability in gene expression. Previously, these niches were visualized within Staphylococcus aureus biofilms by observing differential expression of the cid and lrg operons during tower formation. In the present study, we examined early biofilm development and identified two new stages (designated multiplication and exodus ) that were associated with changes in matrix composition and a distinct reorganization of the cells as the biofilm matured. The initial attachment and multiplication stages were shown to be protease sensitive but independent of most cell surface-associated proteins. Interestingly, after 6 h of growth, an exodus of the biofilm population that followed the transition of the biofilm to DNase I sensitivity was demonstrated. Furthermore, disruption of the gene encoding staphylococcal nuclease (nuc) abrogated this exodus event, causing hyperproliferation of the biofilm and disrupting normal tower development. Immediately prior to the exodus event, S. aureus cells carrying a nuc::gfp promoter fusion demonstrated Sae-dependent expression but only in an apparently random subpopulation of cells. In contrast to the existing model for tower development in S. aureus, the results of this study suggest the presence of a Sae-controlled nuclease-mediated exodus of biofilm cells that is required for the development of tower structures. Furthermore, these studies indicate that the differential expression of nuc during biofilm development is subject to stochastic regulatory mechanisms that are independent of the formation of metabolic microniches. Importance: In this study, we provide a novel view of four early stages of biofilm formation by the human pathogen Staphylococcus aureus. We identified an initial nucleoprotein matrix during biofilm development that is DNase I insensitive until a critical point when a nuclease-mediated exodus of the population is induced prior to tower formation. Unlike the previously described dispersal of cells that occurs after tower development, we found that the mechanism controlling this exodus event is dependent on the Sae regulatory system and independent of Agr. In addition, we revealed that the gene encoding the secreted staphylococcal nuclease was expressed in only a subpopulation of cells, consistent with a model in which biofilms exhibit multicellular characteristics, including the presence of specialized cells and a division of labor that imparts functional consequences to the remainder of the population

Draft Genome Sequence of a Mycobacterium avium Complex Isolate from a Broadbill Bird

Mycobacterium avium complex (MAC) organisms cause opportunistic infections in humans, yet their epidemiology remains poorly understood. They are slowly growing environmental and animal-associated mycobacteria that have little notoriety except for the strains that cause disseminated infections in HIV- infected humans (1). Most MAC organisms are classified taxonomically as a single species, M. avium, which is divided into at least four subspecies, M. avium subsp. avium, M. avium subsp. hominissuis, M. avium subsp. paratuberculosis, and M. avium subsp. silvaticum (2). The only other species in this group is M. intracellulare. Genotyping of this diverse bacterial group has been achieved using intergenic spacers (3) and rpoB sequence analysis (4, 5)

Effects of Caffeine on the Muscular Endurance, Perceived Pain, and Effort of Resistance Trained Women

Please download pdf version here

Early antibody response against Mycobacterium avium subspecies paratuberculosis antigens in subclinical cattle

<p>Abstract</p> <p>Background</p> <p>Our laboratories have previously reported on the experimental infection of cattle with <it>Mycobacterium avium </it>subsp <it>paratuberculosis </it>(<it>M. paratuberculosis</it>) using an intratonsillar infection model. In addition, we have recently developed a partial protein array representing 92 <it>M. paratuberculosis </it>coding sequences. These combined tools have enabled a unique look at the temporal analysis of <it>M. paratuberculosis </it>antigens within the native host. The primary objective of this study was to identify <it>M. paratuberculosis </it>antigens detected by cattle early during infection. A secondary objective was to evaluate the humoral immune response in cattle during the initial year of infection.</p> <p>Results</p> <p>Sera from two experimentally infected cattle, taken pre-inoculation and at day 70, 194 and 321 post infection, identified dynamic antibody reactivity among antigens with some showing an increased response over time and others showing declining levels of reactivity over the same time period. A <it>M. paratuberculosis </it>specific protein, encoded by MAP0862, was strongly detected initially, but the antibody response became weaker with time. The most reactive protein was a putative surface antigen encoded by MAP1087. A second protein, MAP1204, implicated in virulence, was also strongly detected by day 70 in both cattle. Subsequent experiments showed that these two proteins were detected with sera from 5 of 9 naturally infected cattle in the subclinical stage of Johne's disease.</p> <p>Conclusion</p> <p>Collectively these results demonstrate that <it>M. paratuberculosis </it>proteins are detected by sera from experimentally infected cattle as early as 70 days after exposure. These data further suggest at least two antigens may be useful in the early diagnosis of <it>M. paratuberculosis </it>infections. Finally, the construction and use of a protein array in this pilot study has led to a novel approach for discovery of <it>M. paratuberculosis </it>antigens.</p

Impact of Vancomycin on sarA-Mediated Biofilm Formation: Role in Persistent Endovascular Infections Due to Methicillin-Resistant Staphylococcus aureus

Background. Staphylococcus aureus is the most common cause of endovascular infections. The staphylococcal accessory regulator A locus (sarA) is a major virulence determinant that may potentially impact methicillin-resistant S. aureus (MRSA) persistence in such infections via its influence on biofilm formation. Methods. Two healthcare-associated MRSA isolates from patients with persistent bacteremia and 2 prototypical community-acquired MRSA strains, as well as their respective isogenic sarA mutants, were studied for in vitro biofilm formation, fibronectin-binding capacity, autolysis, and protease and nuclease activities. These assays were done in the presence or absence of sub-minimum inhibitory concentrations (MICs) of vancomycin. In addition, these strain pairs were compared for intrinsic virulence and responses to vancomycin therapy in experimental infective endocarditis, a prototypical biofilm model. Results. All sarA mutants displayed significantly reduced biofilm formation and binding to fibronectin but increased protease production in vitro, compared with their respective parental strains. Interestingly, exposure to sub-MICs of vancomycin significantly promoted biofilm formation and fibronectin-binding in parental strains but not in sarA mutants. In addition, all sarA mutants became exquisitely susceptible to vancomycin therapy, compared with their respective parental strains, in the infective endocarditis model. Conclusions. These observations suggest that sarA activation is important in persistent MRSA endovascular infection, potentially in the setting of biofilm formatio

Mobility Disability in Older Adults: At the Intersection of People and Places

Mobility disability is associated with poor lower body function among older adults. This study examines whether specific types of neighborhood characteristics moderate that association

Low levels of β-lactam antibiotics induce extracellular DNA release and biofilm formation in Staphylococcus aureus.

UNLABELLED: Subminimal inhibitory concentrations of antibiotics have been shown to induce bacterial biofilm formation. Few studies have investigated antibiotic-induced biofilm formation in Staphylococcus aureus, an important human pathogen. Our goal was to measure S. aureus biofilm formation in the presence of low levels of β-lactam antibiotics. Fifteen phylogenetically diverse methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-sensitive S. aureus (MSSA) strains were employed. Methicillin, ampicillin, amoxicillin, and cloxacillin were added to cultures at concentrations ranging from 0× to 1× MIC. Biofilm formation was measured in 96-well microtiter plates using a crystal violet binding assay. Autoaggregation was measured using a visual test tube settling assay. Extracellular DNA was quantitated using agarose gel electrophoresis. All four antibiotics induced biofilm formation in some strains. The amount of biofilm induction was as high as 10-fold and was inversely proportional to the amount of biofilm produced by the strain in the absence of antibiotics. MRSA strains of lineages USA300, USA400, and USA500 exhibited the highest levels of methicillin-induced biofilm induction. Biofilm formation induced by low-level methicillin was inhibited by DNase. Low-level methicillin also induced DNase-sensitive autoaggregation and extracellular DNA release. The biofilm induction phenotype was absent in a strain deficient in autolysin (atl). Our findings demonstrate that subminimal inhibitory concentrations of β-lactam antibiotics significantly induce autolysin-dependent extracellular DNA release and biofilm formation in some strains of S. aureus. IMPORTANCE: The widespread use of antibiotics as growth promoters in agriculture may expose bacteria to low levels of the drugs. The aim of this study was to investigate the effects of low levels of antibiotics on bacterial autoaggregation and biofilm formation, two processes that have been shown to foster genetic exchange and antibiotic resistance. We found that low levels of β-lactam antibiotics, a class commonly used in both clinical and agricultural settings, caused significant autoaggregation and biofilm formation by the important human pathogen Staphylococcus aureus. Both processes were dependent on cell lysis and release of DNA into the environment. The effect was most pronounced among multidrug-resistant strains known as methicillin-resistant S. aureus (MRSA). These results may shed light on the recalcitrance of some bacterial infections to antibiotic treatment in clinical settings and the evolution of antibiotic-resistant bacteria in agricultural settings

Nuclease Modulates Biofilm Formation in Community-Associated Methicillin-Resistant Staphylococcus aureus

Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) is an emerging contributor to biofilm-related infections. We recently reported that strains lacking sigma factor B (sigB) in the USA300 lineage of CA-MRSA are unable to develop a biofilm. Interestingly, when spent media from a USA300 sigB mutant was incubated with other S. aureus strains, biofilm formation was inhibited. Following fractionation and mass spectrometry analysis, the major anti-biofilm factor identified in the spent media was secreted thermonuclease (Nuc). Considering reports that extracellular DNA (eDNA) is an important component of the biofilm matrix, we investigated the regulation and role of Nuc in USA300. The expression of the nuc gene was increased in a sigB mutant, repressed by glucose supplementation, and was unaffected by the agr quorum-sensing system. A FRET assay for Nuc activity was developed and confirmed the regulatory results. A USA300 nuc mutant was constructed and displayed an enhanced biofilm-forming capacity, and the nuc mutant also accumulated more high molecular weight eDNA than the WT and regulatory mutant strains. Inactivation of nuc in the USA300 sigB mutant background partially repaired the sigB biofilm-negative phenotype, suggesting that nuc expression contributes to the inability of the mutant to form biofilm. To test the generality of the nuc mutant biofilm phenotypes, the mutation was introduced into other S. aureus genetic backgrounds and similar increases in biofilm formation were observed. Finally, using multiple S. aureus strains and regulatory mutants, an inverse correlation between Nuc activity and biofilm formation was demonstrated. Altogether, our findings confirm the important role for eDNA in the S. aureus biofilm matrix and indicates Nuc is a regulator of biofilm formation