Quantitative analysis of Streptococcus pneumoniae TIGR4 response to in vitro iron restriction by 2-D LC ESI MS/MS

Understanding the growth of bacterial pathogens in a micronutrient restricted host environment can identify potential virulence proteins that help overcome this nutritional barrier to productive infection. In this study, we investigated the pneumococcal protein expression response to iron limitation using an in vitro model. We identified S. pneumoniae TIGR4 proteins by 2-D LC ESI MS/MS and determined significant changes in protein expression in response to iron restriction using computer-intensive random resampling methods. Differential protein expression was studied in the context of a S. pneumoniae TIGR4 protein interaction network using Pathway Studio. Our analysis showed that pneumococcal iron restriction response was marked by increased expression of known virulence factors like PsaA. It involved changes in the expression of stress response, and phase variation and biofilm formation proteins. The net effect of changes in all these biological processes could increase the virulence of S. pneumoniae TIGR4 during in vivo infection.     

Structural and conformational changes concomitant with the E1-E2 transition in H(+)K(+)-ATPase: a comparative protein modeling study

Comparative modeling studies on conserved regions of the gastric H(+)K(+)-ATPase reveal that the E1-E2 conformational transition induces significant tertiary structural changes while conserving the secondary structure. The residues 516-530 of the cytoplasmic domain and TM10 within the transmembrane (TM) regions undergo maximum tertiary structural changes. The luminal regions exhibit comparatively lesser tertiary structural deviations. Residues 249-304 show maximum secondary structural deviation in the conformational transition. The Cys-815 and Cys-323 residues involved in inhibitor binding are found to have smaller buried side chain areas in the E1 conformation compared to E2. Retention of activity correlates well with the buried side chain area when selected amino acid residues in TM6 are mutated using modeling techniques with bulkier amino acid residues. Conformational specificity for ion binding is corroborated with the fraction of side chains exposed to polar atoms of the residues E345, D826, V340, A341, V343, and E822.     

Combined effect of alcohol and cigarette smoke on lipid peroxidation and antioxidant status in rats

The effect of long-term administration of alcohol and cigarette smoke independently and both in combination on lipid peroxidation and antioxidant enzymes, such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione-S-transferase (GST) was studied in liver, kidney, heart and lungs of albino rats. The levels of peroxidation products viz., malondialdehyde, hydroperoxides and conjugated dienes were increased in all the tissues of alcohol administered and smoke-exposed rats. Activities of SOD and CAT were decreased in alcohol-treated and alcohol and smoke combination groups, but increased in smoke-exposed group. Activities of GPx and GST have shown an increase, while concentration of reduced glutathione was found decreased in all the three groups.     

Utilization of a nitrobenzoxadiazole (NBD) fluorophore in the design of a Grb2 SH2 domain-binding peptide mimetic

Fluorescence labeling has become a general technique for studying the intracellular accumulation and localization of exogenously administered materials. Reported herein is a low nanomolar affinity Grb2 SH2 domain-binding antagonist that utilizes the environmentally-sensitive nitrobenzoxadiazole (NBD) fluorophore as a naphthyl replacement. This novel agent should serve as a useful tool to visualize the actions of this class of Grb2 SH2 domain-binding antagonists in whole cell systems.     

Conservation of a novel protein associated with an antibiotic efflux operon in Burkholderia cenocepacia

Burkholderia cenocepacia is a significant problem in individuals with cystic fibrosis and is a member of the B. cepacia complex of closely related antibiotic resistant bacteria. A salicylate-regulated antibiotic efflux operon has been identified in B. cenocepacia and one of its four genes, llpE, is without parallel in previously reported efflux operons. PCR amplification and sequencing of llpE from B. cepacia complex isolates demonstrated the highest prevalence in B. cenocepacia with a high degree of sequence conservation. While at least one non-synonymous mutation was identified between isolates from different genomovars, only synonymous differences were identified within the IIIA and IIIB sub-groups of B. cenocepacia. Structural modeling suggests that LlpE is a member of the alpha/beta hydrolase enzyme family. Identification of strong structural homology to hydrolases and a high degree of conservation in B. cenocepacia suggests an enzymatic function for LlpE, benefiting survival in the cystic fibrosis lung.     

What drives adoption of a computerised,multifaceted quality improvement intervention for cardiovascular disease management in primary healthcare settings? A mixed methods analysis using normalisation process theory

Background

A computerised, multifaceted quality improvement (QI) intervention for cardiovascular disease (CVD) management in Australian primary healthcare was evaluated in a cluster randomised controlled trial. The intervention was associated with improved CVD risk factor screening but there was no improvement in prescribing rates of guideline-recommended medicines. The aim of this study was to conduct a process evaluation to identify and explain the underlying mechanisms by which the intervention did and did not have an impact.

Methods/design

Normalisation process theory (NPT) was used to understand factors that supported or constrained normalisation of the intervention into routine practice. A case study design was used in which six of the 30 participating intervention sites were purposively sampled to obtain a mix of size, governance, structure and performance. Multiple data sources were drawn on including trial outcome data, surveys of job satisfaction and team climate (68 staff) and in-depth interviews (19 staff). Data were primarily analysed within cases and compared with quantitative findings in other trial intervention and usual care sites.

Results

We found a complex interaction between implementation processes and several contextual factors affecting uptake of the intervention. There was no clear association between team climate, job satisfaction and intervention outcomes. There were four spheres of influence that appeared to enhance or detract from normalisation of the intervention: organisational mission and history (e.g. strategic investment to promote a QI culture enhanced cognitive participation), leadership (e.g. ability to energise or demotivate others influenced coherence), team environment (e.g. synergistic activities of team members with different skill sets influenced collective action) and technical integrity of the intervention (e.g. tools that slowed computer systems limited reflective action).

Discussion

Use of NPT helped explain how certain contextual factors influence the work that is done by individuals and teams when implementing a novel intervention. Although these factors do not necessarily distil into a recipe for successful uptake, they may assist system planners, intervention developers, and health professionals to better understand the trajectory that primary health care services may take when developing and engaging with QI interventions.

Trial registration

ACTRN 12611000478910. Registered 08 May 2011.

     

Complete genome sequence of uropathogenic <Emphasis Type="Italic">Escherichia coli</Emphasis> isolate UPEC 26-1

Urinary tract infections (UTIs) are among the most common infections in humans, predominantly caused by uropathogenic Escherichia coli (UPEC). The diverse genomes of UPEC strains mostly impede disease prevention and control measures. In this study, we comparatively analyzed the whole genome sequence of a highly virulent UPEC strain, namely UPEC 26-1, which was isolated from urine sample of a patient suffering from UTI in Korea. Whole genome analysis showed that the genome consists of one circular chromosome of 5,329,753 bp, comprising 5064 protein-coding genes, 122 RNA genes (94 tRNA, 22 rRNA and 6 ncRNA genes), and 100 pseudogenes, with an average G+C content of 50.56%. In addition, we identified 8 prophage regions comprising 5 intact, 2 incomplete and 1 questionable ones and 63 genomic islands, suggesting the possibility of horizontal gene transfer in this strain. Comparative genome analysis of UPEC 26-1 with the UPEC strain CFT073 revealed an average nucleotide identity of 99.7%. The genome comparison with CFT073 provides major differences in the genome of UPEC 26-1 that would explain its increased virulence and biofilm formation. Nineteen of the total GIs were unique to UPEC 26-1 compared to CFT073 and nine of them harbored unique genes that are involved in virulence, multidrug resistance, biofilm formation and bacterial pathogenesis. The data from this study will assist in future studies of UPEC strains to develop effective control measures.     

Mitochondrial dysfunction in human skeletal muscle biopsies of lipid storage disorder

Mitochondria regulate the balance between lipid metabolism and storage in the skeletal muscle. Altered lipid transport, metabolism and storage influence the bioenergetics, redox status and insulin signalling, contributing to cardiac and neurological diseases. Lipid storage disorders (LSD s) are neurological disorders which entail intramuscular lipid accumulation and impaired mitochondrial bioenergetics in the skeletal muscle causing progressive myopathy with muscle weakness. However, the mitochondrial changes including molecular events associated with impaired lipid storage have not been completely understood in the human skeletal muscle. We carried out morphological and biochemical analysis of mitochondrial function in muscle biopsies of human subjects with LSD s (n  = 7), compared to controls (n  = 10). Routine histology, enzyme histochemistry and ultrastructural analysis indicated altered muscle cell morphology and mitochondrial structure. Protein profiling of the muscle mitochondria from LSD samples (n  = 5) (vs. control, n  = 5) by high‐throughput mass spectrometric analysis revealed that impaired metabolic processes could contribute to mitochondrial dysfunction and ensuing myopathy in LSD s. We propose that impaired fatty acid and respiratory metabolism along with increased membrane permeability, elevated lipolysis and altered cristae entail mitochondrial dysfunction in LSD s. Some of these mechanisms were unique to LSD apart from others that were common to dystrophic and inflammatory muscle pathologies. Many differentially regulated mitochondrial proteins in LSD are linked with other human diseases, indicating that mitochondrial protection via targeted drugs could be a treatment modality in LSD and related metabolic diseases.

Cover Image for this Issue: doi: 10.1111/jnc.14177 .