Extracellular secretion of a recombinant therapeutic peptide by Bacillus halodurans utilizing a modified flagellin type III secretion system

<p>Abstract</p> <p>Background</p> <p>Through modification of the flagellin type III secretion pathway of <it>Bacillus halodurans </it>heterologous peptides could be secreted into the medium as flagellin fusion monomers. The stability of the secreted monomers was significantly enhanced through gene-targeted inactivation of host cell extracellular proteases. In evaluating the biotechnological potential of this extracellular secretion system an anti-viral therapeutic peptide, Enfuvirtide, was chosen. Currently, Enfuvirtide is synthesised utilizing 106 chemical steps. We used Enfuvirtide as a model system in an effort to develop a more cost-effective biological process for therapeutic peptide production.</p> <p>Results</p> <p>An attempt was made to increase the levels of the fusion peptide by two strategies, namely strain improvement through gene-targeted knock-outs, as well as vector and cassette optimization. Both approaches proved to be successful. Through chromosomal inactivation of the <it>spo0A, lytC </it>and <it>lytE </it>genes, giving rise to strain <it>B. halodurans </it>BhFDL05S, the secretion of recombinant peptide fusions was increased 10-fold. Cassette optimization, incorporating an expression vector pNW33N and the N- and C-terminal regions of the flagellin monomer as an in-frame peptide fusion, resulted in a further 3.5-fold increase in the secretion of recombinant peptide fusions.</p> <p>Conclusions</p> <p>The type III flagellar secretion system of <it>B. halodurans </it>has been shown to successfully secrete a therapeutic peptide as a heterologous flagellin fusion. Improvements to both the strain and expression cassette led to increased levels of recombinant peptide, showing promise for a biotechnological application.</p

Candida albicans Hypha Formation and Mannan Masking of β-Glucan Inhibit Macrophage Phagosome Maturation

Received 28 August 2014 Accepted 28 October 2014 Published 2 December 2014 This is an open-access article distributed under the terms of the Creative Commons Attribution 3.0 Unported license. ACKNOWLEDGMENTS We thank Janet Willment, Aberdeen Fungal Group, University of Aberdeen, for kindly providing the soluble Dectin-1-Fc reporter. All microscopy was performed with the assistance of the University of Aberdeen Core Microscopy & Histology Facility, and we thank the IFCC for their assistance with flow cytometry. We thank the Wellcome Trust for funding (080088, 086827, 075470, 099215, 097377, and 101873). E.R.B. and A.J.P.B. are funded by the European Research Council (ERC-2009-AdG-249793), and J.L. is funded by a Medical Research Council Clinical Training Fellowship.Peer reviewedPublisher PD

Response to recharge variation of thin rainwater lenses and their mixing zone with underlying saline groundwater

In coastal zones with saline groundwater, fresh groundwater lenses may form due to infiltration of rain water. The thickness of both the lens and the mixing zone, determines fresh water availability for plant growth. Due to recharge variation, the thickness of the lens and the mixing zone are not constant, which may adversely affect agricultural and natural vegetation if saline water reaches the root zone during the growing season. In this paper, we study the response of thin lenses and their mixing zone to variation of recharge. The recharge is varied using sinusoids with a range of amplitudes and frequencies. We vary lens characteristics by varying the Rayleigh number and Mass flux ratio of saline and fresh water, as these dominantly influence the thickness of thin lenses and their mixing zone. Numerical results show a linear relation between the normalised lens volume and the main lens and recharge characteristics, enabling an empirical approximation of the variation of lens thickness. Increase of the recharge amplitude causes increase and the increase of recharge frequency causes a decrease in the variation of lens thickness. The average lens thickness is not significantly influenced by these variations in recharge, contrary to the mixing zone thickness. The mixing zone thickness is compared to that of a Fickian mixing regime. A simple relation between the travelled distance of the centre of the mixing zone position due to variations in recharge and the mixing zone thickness is shown to be valid for both a sinusoidal recharge variation and actual records of daily recharge data. Starting from a step response function, convolution can be used to determine the effect of variable recharge in time. For a sinusoidal curve, we can determine delay of lens movement compared to the recharge curve as well as the lens amplitude, derived from the convolution integral. Together the proposed equations provide us with a first order approximation of lens characteristics using basic lens and recharge parameters without the use of numerical models. This enables the assessment of the vulnerability of any thin fresh water lens on saline, upward seeping groundwater to salinity stress in the root zone

A collaborative and evolving response to the needs of frontline workers, patients and families during the COVID-19 pandemic at Tygerberg Hospital, Western Cape Province, South Africa

The global devastation caused by the COVID-19 pandemic and its mental health impact is undeniable. The physical and psychological consequences are wide-ranging – affecting patients fighting the disease, frontline workers in the trenches with them, healthcare staff deployed in high-care settings, and families disconnected from their loved ones in their darkest hours. Within 6 weeks of the COVID-19 outbreak in South Africa, the Department of Psychiatry at Stellenbosch University established the TBH/SU COVID Resiliency Clinic to provide psychological support to frontline workers at Tygerberg Hospital. Identified barriers in healthcare workers accessing mental healthcare resulted in moving towards an on-site visibility to try to remove some of these barriers. This greater on-site presence enabled networking and building of relationships with frontline staff that over time highlighted other frontline needs, such as providing psychosocial and spiritual support to patients and their families. We share challenges, lessons learned and recommendations from two initiatives: the TBH/SU COVID-19 Resiliency Clinic, and an embedded COVID Care Team (CCT). We describe the establishment, roll-out and progress of the Clinic and the subsequent CCT

Shallow rainwater lenses in deltaic areas with saline seepage

In deltaic areas with saline seepage, freshwater availability is often limited to shallow rainwater lenses lying on top of saline groundwater. Here we describe the characteristics and spatial variability of such lenses in areas with saline seepage and the mechanisms that control their occurrence and size. Our findings are based on different types of field measurements and detailed numerical groundwater models applied in the south-western delta of the Netherlands. By combining the applied techniques we could extrapolate measurements at point scale (groundwater sampling, temperature and electrical soil conductivity (TEC)-probe measurements, electrical cone penetration tests (ECPT)) to field scale (continuous vertical electrical soundings (CVES), electromagnetic survey with EM31), and even to regional scale using helicopter-borne electromagnetic measurements (HEM). The measurements show a gradual mixing zone between infiltrating fresh rainwater and upward flowing saline groundwater. The mixing zone is best characterized by the depth of the centre of the mixing zone &lt;i&gt;D&lt;/i&gt;&lt;sub&gt;mix&lt;/sub&gt;, where the salinity is half that of seepage water, and the bottom of the mixing zone &lt;i&gt;B&lt;/i&gt;&lt;sub&gt;mix&lt;/sub&gt;, with a salinity equal to that of the seepage water (Cl-conc. 10 to 16 g l&lt;sup&gt;−1&lt;/sup&gt;). &lt;i&gt;D&lt;/i&gt;&lt;sub&gt;mix&lt;/sub&gt; is found at very shallow depth in the confining top layer, on average at 1.7 m below ground level (b.g.l.), while &lt;i&gt;B&lt;/i&gt;&lt;sub&gt;mix&lt;/sub&gt; lies about 2.5 m b.g.l. The model results show that the constantly alternating upward and downward flow at low velocities in the confining layer is the main mechanism of mixing between rainwater and saline seepage and determines the position and extent of the mixing zone (&lt;i&gt;D&lt;/i&gt;&lt;sub&gt;mix&lt;/sub&gt; and &lt;i&gt;B&lt;/i&gt;&lt;sub&gt;mix&lt;/sub&gt;). Recharge, seepage flux, and drainage depth are the controlling factors

Knee movement patterns of injured and uninjured adolescent basketball players when landing from a jump: A case-control study

BACKGROUND: A common knee injury mechanism sustained during basketball is landing badly from a jump. Landing is a complex task and requires good coordination, dynamic muscle control and flexibility. For adolescents whose coordination and motor control has not fully matured, landing badly from a jump can present a significant risk for injury. There is currently limited biomechanical information regarding the lower limb kinetics of adolescents when jumping, specifically regarding jump kinematics comparing injured with uninjured adolescents. This study reports on an investigation of biomechanical differences in landing patterns of uninjured and injured adolescent basketball players. METHODS: A matched case-control study design was employed. Twenty-two basketball players aged 14–16 years participated in the study: eleven previously knee-injured and eleven uninjured players matched with cases for age, gender, weight, height and years of play, and playing for the same club. Six high-speed, three-dimensional Vicon 370 cameras (120 Hz), Vicon biomechanical software and SAS Version 8 software were employed to analyse landing patterns when subjects performed a "jump shot". Linear correlations determined functional relationships between the biomechanical performance of lower limb joints, and paired t-tests determined differences between the normalised peak biomechanical parameters. RESULTS: The average peak vertical ground reaction forces between the cases and controls were similar. The average peak ground reaction forces between the cases and controls were moderately correlated (r = -0.47). The control (uninjured) players had significantly greater hip and knee flexion angles and significantly greater eccentric activity on landing than the uninjured cases (p < 0.01). CONCLUSION: The findings of the study indicate that players with a history of knee injuries had biomechanically compromised landing techniques when compared with uninjured players matched for gender, age and club. Descriptions (norms) of expected levels of knee control, proprioceptive acuity and eccentric strength relative to landing from a jump, at different ages and physical developmental stages, would assist clinicians and coaches to identify players with inappropriate knee performance comparable to their age or developmental stage

A conceptual model for the analysis of multi-stressors in linked groundwater–surface water systems

Groundwater and surface water are often closely coupled and are both under the influence of multiple stressors. Stressed groundwater systems may lead to a poor ecological status of surface waters but to date no conceptual framework to analyse linked multi-stressed groundwater – surface water systems has been developed. In this paper, a framework is proposed showing the effect of groundwater on surface waters in multiple stressed systems. This framework will be illustrated by applying it to four European catchments, the Odense, Denmark, the Regge and Dinkel, Netherlands, and the Thames, UK, and by assessing its utility in analysing the propagation or buffering of multi-stressors through groundwater to surface waters in these catchments. It is shown that groundwater affects surface water flow, nutrients and temperature, and can both propagate stressors towards surface waters and buffer the effect of stressors in space and time. The effect of groundwater on drivers and states depends on catchment characteristics, stressor combinations, scale and management practises. The proposed framework shows how groundwater in lowland catchments acts as a bridge between stressors and their effects within surface waters. It shows water managers how their management areas might be influenced by groundwater, and helps them to include this important, but often overlooked part of the water cycle in their basin management plans. The analysis of the study catchments also revealed a lack of data on the temperature of both groundwater and surface water, while it is an important parameter considering future climate warming

Shaping public Opinion for confrontation : catalan independence claims as represented in spanish, catalan, valencian, and basque editorials

Editorials represent a newspaper's principal means of ideological positioning and serve to identify the attitude of each newspaper and its readership to some of the main actors in any given conflict, especially those that are political in nature. Catalonia's independence movement has experienced a surge in recent years. The turning point came in July 2010, after Spain's constitutional court ruled against some aspects of Catalonia's Statute of Autonomy (the legislation regulating the relationship between the Catalan autonomous community and the Spanish state). The ruling annulled some of the articles of the statute. The demonstration in support of Catalan independence that took place on September 11, 2012, was considered the most important among several mass rallies held for this purpose over successive years on Catalonia's National Day. This article presents the results of a qualitative and quantitative analysis of editorial articles published around these two key dates, with the goal of studying each newspaper's treatment of political actors and their degree of identification with these actors. Their use of arguments aimed at persuading readers and legitimizing a given position during the specified time periods in 2010 and 2012 will also be studied. The study focuses on the editorial articles published by several newspapers in four distinct regional contexts: Catalonia, Valencia, the Basque Country, as well as the whole of Spain. The analyzed articles were published a few days before and after July 10, 2010, and September 11, 2012