Bacteria of the Burkholderia cepacia complex are cyanogenic under biofilm and colonial growth conditions

<p>Abstract</p> <p>Background</p> <p>The <it>Burkholderia cepacia </it>complex (Bcc) is a collection of nine genotypically distinct but phenotypically similar species. They show wide ecological diversity and include species that are used for promoting plant growth and bio-control as well species that are opportunistic pathogens of vulnerable patients. Over recent years the Bcc have emerged as problematic pathogens of the CF lung. <it>Pseudomonas aeruginosa </it>is another important CF pathogen. It is able to synthesise hydrogen cyanide (HCN), a potent inhibitor of cellular respiration. We have recently shown that HCN production by <it>P. aeruginosa </it>may have a role in CF pathogenesis. This paper describes an investigation of the ability of bacteria of the Bcc to make HCN.</p> <p>Results</p> <p>The genome of <it>Burkholderia cenocepacia </it>has 3 putative HCN synthase encoding (<it>hcnABC</it>) gene clusters. <it>B. cenocepacia </it>and all 9 species of the Bcc complex tested were able to make cyanide at comparable levels to <it>P. aeruginosa</it>, but only when grown surface attached as colonies or during biofilm growth on glass beads. In contrast to <it>P. aeruginosa </it>and other cyanogenic bacteria, cyanide was not detected during planktonic growth of Bcc strains.</p> <p>Conclusion</p> <p>All species in the Bcc are cyanogenic when grown as surface attached colonies or as biofilms.</p

Cyclic-di-adenosine monophosphate (c-di-AMP) is required for osmotic regulation in Staphylococcus aureus but dispensable for viability in anaerobic conditions

Cyclic di-adenosine monophosphate (c-di-AMP) is a recently discovered signaling molecule important for the survival of Firmicutes, a large bacterial group that includes notable pathogens such as Staphylococcus aureus. However, the exact role of this molecule has not been identified. dacA, the S. aureus gene encoding the diadenylate cyclase enzyme required for c-di-AMP production, cannot be deleted when bacterial cells are grown in rich medium, indicating that c-di-AMP is required for growth in this condition. Here, we report that an S. aureus dacA mutant can be generated in chemically defined medium. Consistent with previous findings, this mutant had a severe growth defect when cultured in rich medium. Using this growth defect in rich medium, we selected for suppressor strains with improved growth to identify c-di-AMP-requiring pathways. Mutations bypassing the essentiality of dacA were identified in alsT and opuD, encoding a predicted amino acid and osmolyte transporter, the latter of which we show here to be the main glycine betaine-uptake system in S. aureus. Inactivation of these transporters likely prevents the excessive osmolyte and amino acid accumulation in the cell, providing further evidence for a key role of c-di-AMP in osmotic regulation. Suppressor mutations were also obtained in hepS, hemB, ctaA and qoxB, coding for proteins required for respiration. Furthermore, we show that dacA is dispensable for growth in anaerobic conditions. Together, these finding reveal an essential role for the c-di-AMP signaling network in aerobic, but not anaerobic, respiration in S. aureus

DNA replication initiation in Bacillus subtilis: Structural and functional characterization of the essential DnaA-DnaD interaction

© 2018 The Author(s). The homotetrameric DnaD protein is essential in low G+C content gram positive bacteria and is involved in replication initiation at oriC and re-start of collapsed replication forks. It interacts with the ubiquitously conserved bacterial master replication initiation protein DnaA at the oriC but structural and functional details of this interaction are lacking, thus contributing to our incomplete understanding of the molecular details that underpin replication initiation in bacteria. DnaD comprises N-terminal (DDBH1) and C-terminal (DDBH2) domains, with contradicting bacterial two-hybrid and yeast two-hybrid studies suggesting that either the former or the latter interact with DnaA, respectively. Using Nuclear Magnetic Resonance (NMR) we showed that both DDBH1 and DDBH2 interact with the N-terminal domain I of DnaA and studied the DDBH2 interaction in structural detail. We revealed two families of conformations for the DDBH2-DnaA domain I complex and showed that the DnaA-interaction patch of DnaD is distinct from the DNA-interaction patch, suggesting that DnaD can bind simultaneously DNA and DnaA. Using sensitive single-molecule FRET techniques we revealed that DnaD remodels DnaA-DNA filaments consistent with stretching and/or untwisting. Furthermore, the DNA binding activity of DnaD is redundant for this filament remodelling. This in turn suggests that DnaA and DnaD are working collaboratively in the oriC to locally melt the DNA duplex during replication initiation

Effect of sweeteners and carbonation on aroma partitioning and release in beverage systems

The effect of monosaccharides (glucose, fructose and galactose) and disaccharides (sucrose and lactose) at 10, 20 and 30 % w/v on the in-vitro aroma partitioning of C - C aldehydes and ethyl esters, as well as limonene (concentration of aroma compounds at 1 μg mL ), was studied using atmospheric pressure chemical ionisation-mass spectrometry. An increase in sugar concentration from 0 to 30 % w/v resulted in a significant increase in partitioning under static headspace conditions for the majority of the compounds (p  0.05). The complexity of the system was increased to model a soft drink design - comprising water, sucrose (10, 20 and 30 % w/v), acid (0.15 % w/v), carbonation (∼7.2 g/L CO ) and aroma compounds representative of an apple style flavouring, namely ethyl butanoate and hexanal (10 μg mL each). Although the addition of sucrose had no significant in-vivo effect, carbonation significantly decreased breath-by-breath (in-vivo) aroma delivery (p  0.05). [Abstract copyright: Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.

Presence of bacteria and bacteriophages in full-scale trickling filters and an aerated constructed wetland

Aerated Constructed Wetlands are a state-of-the-art design that provides a different physical and chemical environment (compared to traditional passive wetland designs) for the wastewater treatment processes and, thus, may have different pathogen removal characteristics. In order to establish the fate of bacterial and viral indicators, a field study was carried out at a Sewage Treatment Works (STW) in the UK (serving 20,000 pe). The STW consists of primary and secondary sedimentation tanks and trickling filters (TF) as the biological stage. A large (1,160 m2) pilot aerated Vertical Flow Constructed Wetland (AVFCW) was constructed at the STW as tertiary stage receiving ¼ of the total flow rate, i.e., 1250 m3/day. Effluent quality of the AVFCW complied with national and international standards for environmental discharge and reuse. For the first time, two sets of bacterial (Faecal coliforms, E.coli and intestinal enterococci) and viral indicators (Somatic coliphages, F-RNA specific bacteriophages and human-specific B. fragilis GB124 phages) were simultaneously investigated in an AVFCW and TF. High elimination rates were detected (up to 3.7 and 2.2 log reduction for bacteria indicators and phages, respectively) and strong correlations between the two sets were found. The superior efficiency of the aerated Constructed Wetlands in microbiological contamination removal compared to passive wetland systems was established for the first time, which may have implications for process selection for wastewater reuse. This field study therefore provides new evidence on the fate of bacteriophages and a first indication of their potential use for performance evaluation in TF and aerated Constructed Wetlands. It also demonstrates that the combination of TF with aerated constructed wetlands could be a novel and effective treatment scheme for new STW or for the upgrade of existing STW

Hypertonic saline therapy in cystic fibrosis: do population shifts caused by the osmotic sensitivity of infecting bacteria explain the effectiveness of this treatment?

Cystic fibrosis (CF) is caused by a defect in the CF transmembrane regulator that leads to depletion and dehydration of the airway surface liquid (ASL) of the lung epithelium, providing an environment that can be infected by bacteria leading to increased morbidity and mortality. Pseudomonas aeruginosa chronically infects more than 80% of CF patients and one hallmark of infection is the emergence of a mucoid phenotype associated with a worsening prognosis and more rapid decline in lung function. Hypertonic saline (HS) is a clinically proven treatment that improves mucociliary clearance through partial rehydration of the ASL of the lung. Strikingly, while HS therapy does not alter the prevalence of P. aeruginosa in the CF lung it does decrease the frequency of episodes of acute, severe illness known as infective exacerbations among CF patients. In this article, we propose a hypothesis whereby the positive clinical effects of HS treatment are explained by the osmotic sensitivity of the mucoid sub-population of P. aeruginosa in the CF lung leading to selection against this group in favor of the osmotically resistant non-mucoid variants

Increased posterior default mode network activity and structural connectivity in young adult APOE-ε4 carriers: a multi-modal imaging investigation

Young adult APOE-ε4 carriers show increased activity in posterior regions of the default mode network (pDMN), but how this is related to structural connectivity is unknown. Thirty young adults (one half of whom were APOE-ε4 carriers; mean age 20 years) were scanned using both diffusion and functional magnetic resonance imaging. The parahippocampal cingulum bundle (PHCB)—which links the pDMN and the medial temporal lobe—was manually delineated in individual participants using deterministic tractography. Measures of tract microstructure (mean diffusivity and fractional anisotropy) were then extracted from these tract delineations. APOE-ε4 carriers had lower mean diffusivity and higher fractional anisotropy relative to noncarriers in PHCB, but not in a control tract (the inferior longitudinal fasciculus). Furthermore, PHCB microstructure was selectively associated with pDMN (and medial temporal lobe) activity during a scene discrimination task known to be sensitive to Alzheimer's disease. These findings are consistent with a lifespan view of Alzheimer's disease risk, where early-life, connectivity-related changes in specific, vulnerable “hubs” (e.g., pDMN) lead to increased neural activity. Critically, such changes may reflect reduced network efficiency/flexibility in APOE-ε4 carriers, which in itself may portend a faster decline in connectivity over the lifespan and ultimately trigger early amyloid-β deposition in later life

A β-mannanase with a lysozyme-like fold and a novel molecular catalytic mechanism

The enzymatic cleavage of β-1,4-mannans is achieved by endo-β-1,4-mannanases, enzymes involved in germination of seeds and microbial hemicellulose degradation, and which have increasing industrial and consumer product applications. β- Mannanases occur in a range of families of the CAZy sequence-based glycoside hydrolase (GH) classification scheme including families 5, 26, and 113. In this work we reveal that β- mannanases of the newly described GH family 134 differ from other mannanase families in both their mechanism and tertiary structure. A representative GH family 134 endo-β-1,4-mannanase from a Streptomyces sp. displays a fold closely related to that of hen egg white lysozyme but acts with inversion of stereochemistry. A Michaelis complex with mannopentaose, and a product complex with mannotriose, reveal ligands with pyranose rings distorted in an unusual inverted chair conformation. Ab initio quantum mechanics/molecular mechanics metadynamics quantified the energetically accessible ring conformations and provided evidence in support of a 1C4 → 3H4 ‡ → 3S1 conformational itinerary along the reaction coordinate. This work, in concert with that on GH family 124 cellulases, reveals how the lysozyme fold can be co-opted to catalyze the hydrolysis of different polysaccharides in a mechanistically distinct manner

The lipidome and proteome of oil bodies from Helianthus annuus (common sunflower).

In this paper we report the molecular profiling, lipidome and proteome, of the plant organelle known as an oil body (OB). The OB is remarkable in that it is able to perform its biological role (storage of triglycerides) whilst resisting the physical stresses caused by changes during desiccation (dehydration) and germination (rehydration). The molecular profile that confers such extraordinary physical stability on OBs was determined using a combination of (31)P/(1)H nuclear magnetic resonance (NMR), high-resolution mass spectrometry and nominal mass-tandem mass spectrometry for the lipidome, and gel-electrophoresis-chromatography-tandem mass spectrometry for the proteome. The integrity of the procedure for isolating OBs was supported by physical evidence from small-angle neutron-scattering experiments. Suppression of lipase activity was crucial in determining the lipidome. There is conclusive evidence that the latter is dominated by phosphatidylcholine (∼60 %) and phosphatidylinositol (∼20 %), with a variety of other head groups (∼20 %). The fatty acid profile of the surface monolayer comprised palmitic, linoleic and oleic acids (2:1:0.25, (1)H NMR) with only traces of other fatty acids (C24:0, C22:0, C18:0, C18:3, C16:2; by MS). The proteome is rich in oleosins (78 %) with the remainder being made up of caleosins and steroleosins. These data are sufficiently detailed to inform an update of the understood model of this organelle and can be used to inform the use of such components in a range of molecular biological, biotechnological and food industry applications. The techniques used in this study for profiling the lipidome throw a new light on the lipid profile of plant cellular compartments