Comparative evolutionary patterns of Burkholderia cenocepacia and B. multivorans during chronic co-infection of a cystic fibrosis patient lung

During chronic respiratory infections of cystic fibrosis (CF) patients, bacteria adaptively evolve in response to the nutritional and immune environment as well as influence other infecting microbes. The present study was designed to gain insights into the genetic mechanisms underlying adaptation and diversification by the two most prevalent pathogenic species of the Burkholderia cepacia complex (Bcc), B. cenocepacia and B. multivorans. Herein, we study the evolution of both of these species during coinfection of a CF patient for 4.4 years using genome sequences of 9 B. multivorans and 11 B. cenocepacia. This co-infection spanned at least 3 years following initial infection by B. multivorans and ultimately ended in the patient’s death by cepacia syndrome. Both species acquired several mutations with accumulation rates of 2.08 (B. cenocepacia) and 2.27 (B. multivorans) SNPs/year. Many of the mutated genes are associated with oxidative stress response, transition metal metabolism, defense mechanisms against antibiotics, and other metabolic alterations consistent with the idea that positive selection might be driven by the action of the host immune system, antibiotic therapy and low oxygen and iron concentrations. Two orthologous genes shared by B. cenocepacia and B. multivorans were found to be under strong selection and accumulated mutations associated with lineage diversification. One gene encodes a nucleotide sugar dehydratase involved in lipopolysaccharide O-antigen (OAg) biosynthesis (wbiI). The other gene encodes a putative two-component regulatory sensor kinase protein required to sense and adapt to oxidative- and heavy metal- inducing stresses. This study contributes to understanding of shared and species-specific evolutionary patterns of B. cenocepacia and B. multivorans evolving in the same CF lung environment

Adaptation and survival of Burkholderia cepacia and B. contaminans during long-term Incubation in saline solutions containing benzalkonium chloride

TheBurkholderia cepaciacomplex (Bcc) is a group of opportunistic pathogenic bacteria with a remarkable metabolic capacity and broad genotypic/phenotypic plasticity, allowing their adaptation to hostile conditions, including nutrient depleted solutions containing antimicrobial agents. Bcc bacteria are feared contaminants in pharmaceutical industries and cause nosocomial outbreaks, posing health threats to immunocompromised individuals and cystic fibrosis (CF) patients. In this study, the adaptation and survival ofB. cepaciaandB. contaminansisolates was investigated after long-term incubation in nutrient depleted saline solutions supplemented with increasing concentrations of the biocidal preservative benzalkonium chloride (BZK), recreating the storage conditions of pharmaceutical products. These epidemiologically related isolates were recovered from intrinsically contaminated saline solutions for nasal application and from two CF patients. Long-term incubation in saline solutions containing BZK led to the development of bacterial sub-populations that survived for at least 16 months, despite an initial 2-3 log decrease in viability, displaying a progressive dose-dependent decrease of colony and cell size, including the appearance of small colony variants (SCVs). Bacterial colonies lost pigmentation, changed the morphotype from rough to smooth and produced more spherical cells during extended incubation with BZK. The development of macroscopically visible cellular aggregates, rich in polysaccharide and harboring viable cells in their interior was triggered by BZK. The existence of a metabolic pathway for BZK degradation was confirmed through genome analysis. This study reveals mechanisms underlying the prevalence of Bcc bacteria as contaminants of pharmaceutical products containing BZK, which often lead to false-negative results during quality control and routine testing

Buttressing staples with cholecyst-derived extracellular matrix (CEM) reinforces staple lines in an ex vivo peristaltic inflation model

This is the author's accepted manuscript. The final published article is available from the link below. Copyright @ Springer Science + Business Media, LLC 2008Background - Staple line leakage and bleeding are the most common problems associated with the use of surgical staplers for gastrointestinal resection and anastomotic procedures. These complications can be reduced by reinforcing the staple lines with buttressing materials. The current study reports the potential use of cholecyst-derived extracellular matrix (CEM) in non-crosslinked (NCEM) and crosslinked (XCEM) forms, and compares their mechanical performance with clinically available buttress materials [small intestinal submucosa (SIS) and bovine pericardium (BP)] in an ex vivo small intestine model. Methods - Three crosslinked CEM variants (XCEM0005, XCEM001, and XCEM0033) with different degree of crosslinking were produced. An ex vivo peristaltic inflation model was established. Porcine small intestine segments were stapled on one end, using buttressed or non-buttressed surgical staplers. The opened, non-stapled ends were connected to a peristaltic pump and pressure transducer and sealed. The staple lines were then exposed to increased intraluminal pressure in a peristaltic manner. Both the leak and burst pressures of the test specimens were recorded. Results - The leak pressures observed for non-crosslinked NCEM (137.8 ± 22.3 mmHg), crosslinked XCEM0005 (109.1 ± 14.1 mmHg), XCEM001 (150.1 ± 16.0 mmHg), XCEM0033 (98.8 ± 10.5 mmHg) reinforced staple lines were significantly higher when compared to non-buttressed control (28.3 ± 10.8 mmHg) and SIS (one and four layers) (62.6 ± 11.8 and 57.6 ± 12.3 mmHg, respectively) buttressed staple lines. NCEM and XCEM were comparable to that observed for BP buttressed staple lines (138.8 ± 3.6 mmHg). Only specimens with reinforced staple lines were able to achieve high intraluminal pressures (ruptured at the intestinal mesentery), indicating that buttress reinforcements were able to withstand pressure higher than that of natural tissue (physiological failure). Conclusions - These findings suggest that the use of CEM and XCEM as buttressing materials is associated with reinforced staple lines and increased leak pressures when compared to non-buttressed staple lines. CEM and XCEM were found to perform comparably with clinically available buttress materials in this ex vivo model.Enterprise Irelan

Stress-Induced Reinstatement of Drug Seeking: 20 Years of Progress

In human addicts, drug relapse and craving are often provoked by stress. Since 1995, this clinical scenario has been studied using a rat model of stress-induced reinstatement of drug seeking. Here, we first discuss the generality of stress-induced reinstatement to different drugs of abuse, different stressors, and different behavioral procedures. We also discuss neuropharmacological mechanisms, and brain areas and circuits controlling stress-induced reinstatement of drug seeking. We conclude by discussing results from translational human laboratory studies and clinical trials that were inspired by results from rat studies on stress-induced reinstatement. Our main conclusions are (1) The phenomenon of stress-induced reinstatement, first shown with an intermittent footshock stressor in rats trained to self-administer heroin, generalizes to other abused drugs, including cocaine, methamphetamine, nicotine, and alcohol, and is also observed in the conditioned place preference model in rats and mice. This phenomenon, however, is stressor specific and not all stressors induce reinstatement of drug seeking. (2) Neuropharmacological studies indicate the involvement of corticotropin-releasing factor (CRF), noradrenaline, dopamine, glutamate, kappa/dynorphin, and several other peptide and neurotransmitter systems in stress-induced reinstatement. Neuropharmacology and circuitry studies indicate the involvement of CRF and noradrenaline transmission in bed nucleus of stria terminalis and central amygdala, and dopamine, CRF, kappa/dynorphin, and glutamate transmission in other components of the mesocorticolimbic dopamine system (ventral tegmental area, medial prefrontal cortex, orbitofrontal cortex, and nucleus accumbens). (3) Translational human laboratory studies and a recent clinical trial study show the efficacy of alpha-2 adrenoceptor agonists in decreasing stress-induced drug craving and stress-induced initial heroin lapse

Stress corrosion cracking: Characteristics, Mechanisms and Experimental study

Stress corrosion cracking (SCC) is a phenomenon in which the cracking of a metal alloy usually results from the combined action of a corrodent and tensile stress. Stresses that cause cracking can be residual or may be applied during service. A degree of mechanistic understanding of SCC will enable most metallic engineering materials to operate safely though stress corrosion cracking failures still continue to occur unexpectedly in industry. In this paper, the characteristics, mechanisms and methods of SCC prevention are reviewed. The results of experimental studies on alpha brass are also reported of which the failure mode conformed with the film-rupture and anodic dissolution mechanism

Regulators of genetic risk of breast cancer identified by integrative network analysis.

Genetic risk for breast cancer is conferred by a combination of multiple variants of small effect. To better understand how risk loci might combine, we examined whether risk-associated genes share regulatory mechanisms. We created a breast cancer gene regulatory network comprising transcription factors and groups of putative target genes (regulons) and asked whether specific regulons are enriched for genes associated with risk loci via expression quantitative trait loci (eQTLs). We identified 36 overlapping regulons that were enriched for risk loci and formed a distinct cluster within the network, suggesting shared biology. The risk transcription factors driving these regulons are frequently mutated in cancer and lie in two opposing subgroups, which relate to estrogen receptor (ER)(+) luminal A or luminal B and ER(-) basal-like cancers and to different luminal epithelial cell populations in the adult mammary gland. Our network approach provides a foundation for determining the regulatory circuits governing breast cancer, to identify targets for intervention, and is transferable to other disease settings.This work was funded by Cancer Research UK and the Breast Cancer Research Foundation. MAAC is funded by the National Research Council (CNPq) of Brazil. TEH held a fellowship from the US DOD Breast Cancer Research Program (W81XWH-11-1-0592) and is currently supported by an RAH Career Development Fellowship (Australia). TEH and WDT are funded by the NHMRC of Australia (NHMRC) (ID: 1008349 WDT; 1084416 WDT, TEH) and Cancer Australia/National Breast Cancer Foundation (ID 627229; WDT, TEH). BAJP is a Gibb Fellow of Cancer Research UK. We would like to acknowledge the support of The University of Cambridge, Cancer Research UK and Hutchison Whampoa Limited.This is the author accepted manuscript. The final version is available from NPG via http://dx.doi.org/10.1038/ng.345

Skeletal Muscle Differentiation Evokes Endogenous XIAP to Restrict the Apoptotic Pathway

Myotube apoptosis occurs normally during muscle development and aging but it can lead to destruction of skeletal muscle in neuromuscular diseases. Therefore, understanding how myotube apoptosis is regulated is important for developing novel strategies for treatment of muscle loss. We investigated the regulation of apoptosis in skeletal muscle and report a striking increase in resistance to apoptosis following differentiation. We find mitotic C2C12 cells (myoblast-like cells) are sensitive to cytosolic cytochrome c microinjection. However, differentiated C2C12 cells (myotube-like cells) and primary myotubes are markedly resistant. This resistance is due to endogenous X-linked inhibitor of apoptotic protein (XIAP). Importantly, the selective difference in the ability of XIAP to block myotube but not myoblast apoptosis is not due to a change in XIAP but rather a decrease in Apaf-1 expression. This decrease in Apaf-1 links XIAP to caspase activation and death. Our findings suggest that in order for myotubes to die, they may degrade XIAP, functionally inactivate XIAP or upregulate Apaf-1. Importantly, we identify a role for endogenous Smac in overcoming XIAP to allow myotube death. However, in postmitotic cardiomyocytes, where XIAP also restricts apoptosis, endogenous Smac was not capable of overcoming XIAP to cause death. These results show that as skeletal muscle differentiate, they become resistant to apoptosis because of the ability of XIAP to regulate caspase activation. The increased restriction of apoptosis in myotubes is presumably important to ensure the long term survival of these postmitotic cells as they play a vital role in the physiology of organisms

Aerosolized Delivery of Antifungal Agents

Pulmonary infections caused by Aspergillus species are associated with significant morbidity and mortality in immunocompromised patients. Although the treatment of pulmonary fungal infections requires the use of systemic agents, aerosolized delivery is an attractive option in prevention because the drug can concentrate locally at the site of infection with minimal systemic exposure. Current clinical evidence for the use of aerosolized delivery in preventing fungal infections is limited to amphotericin B products, although itraconazole, voriconazole, and caspofungin are under investigation. Based on conflicting results from clinical trials that evaluated various amphotericin B formulations, the routine use of aerosolized delivery cannot be recommended. Further research with well-designed clinical trials is necessary to elucidate the therapeutic role and risks associated with aerosolized delivery of antifungal agents. This article provides an overview of aerosolized delivery systems, the intrapulmonary pharmacokinetic properties of aerosolized antifungal agents, and key findings from clinical studies