Subdivision of the bacterioferritin comigratory protein family of bacterial peroxiredoxins based on catalytic activity.

© American Chemical Society,2010. Post-print version of article deposited in accordance with SHERPA RoMEO guidelinesPeroxiredoxins are ubiquitous proteins that catalyze the reduction of hydroperoxides, thus conferring resistance to oxidative stress. Using high-resolution mass spectrometry, we recently reclassified one such peroxiredoxin, bacterioferritin comigratory protein (BCP) of Escherichia coli, as an atypical 2-Cys peroxiredoxin that functions through the formation of an intramolecular disulfide bond between the active and resolving cysteine. An engineered E. coli BCP, which lacked the resolving cysteine, retained enzyme activity through a novel catalytic pathway. Unlike the active cysteine, the resolving cysteine of BCP peroxiredoxins is not conserved across all members of the family. To clarify the catalytic mechanism of native BCP enzymes that lack the resolving cysteine, we have investigated the BCP homologue of Burkholderia cenocepacia. We demonstrate that the B. cenocepacia BCP (BcBCP) homologue functions through a 1-Cys catalytic pathway. During catalysis, BcBCP can utilize thioredoxin as a reductant for the sulfenic acid intermediate. However, significantly higher peroxidase activity is observed utilizing glutathione as a resolving cysteine and glutaredoxin as a redox partner. Introduction of a resolving cysteine into BcBCP changes the activity from a 1-Cys pathway to an atypical 2-Cys pathway, analogous to the E. coli enzyme. In contrast to the native B. cenocepacia enzyme, thioredoxin is the preferred redox partner for this atypical 2-Cys variant. BCP-deficient B. cenocepacia exhibit a growth-phase-dependent hypersensitivity to oxidative killing. On the basis of sequence alignments, we believe that BcBCP described herein is representative of the major class of bacterial BCP peroxiredoxins. To our knowledge, this is the first detailed characterization of their catalytic activity. These studies support the subdivision of the BCP family of peroxiredoxins into two classes based on their catalytic activity

How conservation initiatives go to scale

Although a major portion of the planet’s land and sea is managed to conserve biodiversity, little is known about the extent, speed and patterns of adoption of conservation initiatives. We undertook a quantitative exploration of how area-based conservation initiatives go to scale by analysing the adoption of 22 widely recognized and diverse initiatives from across the globe. We use a standardized approach to compare the potential of different initiatives to reach scale. While our study is not exhaustive, our analyses reveal consistent patterns across a variety of initiatives: adoption of most initiatives (82% of our case studies) started slowly before rapidly going to scale. Consistent with diffusion of innovation theory, most initiatives exhibit slow–fast–slow (that is, sigmoidal) dynamics driven by interactions between existing and potential adopters. However, uptake rates and saturation points vary among the initiatives and across localities. Our models suggest that the uptake of most of our case studies is limited; over half of the initiatives will be taken up by <30% of their potential adopters. We also provide a methodology for quantitatively understanding the process of scaling. Our findings inform us how initiatives scale up to widespread adoption, which will facilitate forecasts of the future level of adoption of initiatives, and benchmark their extent and speed of adoption against those of our case studies

Extracellular DNA Chelates Cations and Induces Antibiotic Resistance in Pseudomonas aeruginosa Biofilms

Biofilms are surface-adhered bacterial communities encased in an extracellular matrix composed of DNA, bacterial polysaccharides and proteins, which are up to 1000-fold more antibiotic resistant than planktonic cultures. To date, extracellular DNA has been shown to function as a structural support to maintain Pseudomonas aeruginosa biofilm architecture. Here we show that DNA is a multifaceted component of P. aeruginosa biofilms. At physiologically relevant concentrations, extracellular DNA has antimicrobial activity, causing cell lysis by chelating cations that stabilize lipopolysaccharide (LPS) and the outer membrane (OM). DNA-mediated killing occurred within minutes, as a result of perturbation of both the outer and inner membrane (IM) and the release of cytoplasmic contents, including genomic DNA. Sub-inhibitory concentrations of DNA created a cation-limited environment that resulted in induction of the PhoPQ- and PmrAB-regulated cationic antimicrobial peptide resistance operon PA3552–PA3559 in P. aeruginosa. Furthermore, DNA-induced expression of this operon resulted in up to 2560-fold increased resistance to cationic antimicrobial peptides and 640-fold increased resistance to aminoglycosides, but had no effect on β-lactam and fluoroquinolone resistance. Thus, the presence of extracellular DNA in the biofilm matrix contributes to cation gradients, genomic DNA release and inducible antibiotic resistance. DNA-rich environments, including biofilms and other infection sites like the CF lung, are likely the in vivo environments where extracellular pathogens such as P. aeruginosa encounter cation limitation

Survival of stroke patients after introduction of the 'Dutch Transmural Protocol TIA/CVA'

<p>Background: Earlier research showed that healthcare in stroke could be better organized, aiming for improved survival and less comorbidity. Therefore, in 2004 the Dutch College of General Practitioners (NHG) and the Dutch Association of Neurology (NVN) introduced the 'Dutch Transmural Protocol TIA/CVA' (the LTA) to improve survival, minimize the risk of stroke recurrence, and increase quality of life after stroke. This study examines whether survival improved after implementation of the new protocol, and whether there was an increase in contacts with the general practitioner (GP)/nurse practitioner, registration of comorbidity and prescription of medication.</p><p>Methods: From the primary care database of the Registration Network Groningen (RNG) two cohorts were composed: one cohort compiled before and one after introduction of the LTA. Cohort 1 (n = 131, first stroke 2001-2002) was compared with cohort 2 (n = 132, first stroke 2005-2006) with regard to survival and the secondary outcomes.</p><p>Results: Comparison of the two cohorts showed no significant improvement in survival. In cohort 2, the number of contacts with the GP was significantly lower and with the nurse practitioner significantly higher, compared with cohort 1. All risk factors for stroke were more prevalent in cohort 2, but were only significant for hypercholesterolemia. In both cohorts more medication was prescribed after stroke, whereas ACE inhibitors were prescribed more frequently only in cohort 2.</p><p>Conclusion: No major changes in survival and secondary outcomes were apparent after introduction of the LTA. Although, there was a small improvement in secondary prevention, this study shows that optimal treatment after introduction of the LTA has not yet been achieved.</p>