bluuebunny/crossref_metadata_2025_split huggingface dataset:
Rapid and Sensitive Quantification of Vibrio cholerae and Vibrio mimicus Cells in Water Samples by Use of Catalyzed Reporter Deposition Fluorescence
In Situ
Hybridization Combined with Solid-Phase Cytometry
Authors: Schauer Sonja , Sommer Regina , Farnleitner Andreas H. , Kirschner Alexander K. T. 
Published: August 2012

ABSTRACT

A new protocol for rapid, specific, and sensitive cell-based quantification of
Vibrio cholerae
/
Vibrio mimicus
in water samples was developed. The protocol is based on catalyzed reporter deposition fluorescence
in situ
hybridization (CARD-FISH) in combination with solid-phase cytometry. For pure cultures, we were able to quantify down to 6
V. cholerae
cells on one membrane with a relative precision of 39% and down to 12 cells with a relative precision of 17% after hybridization with the horseradish peroxidase (HRP)-labeled probe Vchomim1276 (specific for
V. cholerae
and
V. mimicus
) and signal amplification. The corresponding position of the probe on the 16S rRNA is highly accessible even when labeled with HRP. For the first time, we were also able to successfully quantify
V. cholerae
/
V. mimicus
via solid-phase cytometry in extremely turbid environmental water samples collected in Austria. Cell numbers ranged from 4.5 × 10
1
cells ml
−1
in the large saline lake Neusiedler See to 5.6 × 10
4
cells ml
−1
in an extremely turbid shallow soda lake situated nearby. We therefore suggest CARD-FISH in combination with solid-phase cytometry as a powerful tool to quantify
V. cholerae
/
V. mimicus
in ecological studies as well as for risk assessment and monitoring programs.
Farnesol and Cyclic AMP Signaling Effects on the Hypha-to-Yeast Transition in Candida albicans
Authors: Lindsay Allia K. , Deveau Aurélie , Piispanen Amy E. , Hogan Deborah A. 
Published: August 2012

ABSTRACT

Candida albicans
, a fungal pathogen of humans, regulates its morphology in response to many environmental cues and this morphological plasticity contributes to virulence. Farnesol, an autoregulatory molecule produced by
C. albicans
, inhibits the induction of hyphal growth by inhibiting adenylate cyclase (Cyr1). The role of farnesol and Cyr1 in controlling the maintenance of hyphal growth has been less clear. Here, we demonstrate that preformed hyphae transition to growth as yeast in response to farnesol and that strains with increased cyclic AMP (cAMP) signaling exhibit more resistance to farnesol. Exogenous farnesol did not induce the hypha-to-yeast transition in mutants lacking the Tup1 or Nrg1 transcriptional repressors in embedded conditions. Although body temperature is not required for embedded hyphal growth, we found that the effect of farnesol on the hypha-to-yeast transition varies inversely with temperature. Our model of Cyr1 activity being required for filamentation is also supported by our liquid assay data, which show increased yeast formation when preformed filaments are treated with farnesol. Together, these data suggest that farnesol can modulate morphology in preformed hyphal cells and that the repression of hyphal growth maintenance likely occurs through the inhibition of cAMP signaling.
A Candida albicans Temperature-Sensitive
cdc12-6
Mutant Identifies Roles for Septins in Selection of Sites of Germ Tube Formation and Hyphal Morphogenesis
Authors: Li Lifang , Zhang Chengda , Konopka James B. 
Published: August 2012

ABSTRACT

Septins were identified for their role in septation in
Saccharomyces cerevisiae
and were subsequently implicated in other morphogenic processes. To study septins in
Candida albicans
hyphal morphogenesis, a temperature-sensitive mutation was created that altered the C terminus of the essential Cdc12 septin. The
cdc12-6
cells grew well at room temperature, but at 37°C they displayed expected defects in septation, nuclear localization, and bud morphogenesis. Although serum stimulated the
cdc12-6
cells at 37°C to form germ tube outgrowths, the mutant could not maintain polarized hyphal growth and instead formed chains of elongated cell compartments. Serum also stimulated the
cdc12-6
mutant to induce a hyphal reporter gene (
HWP1-GFP
) and a characteristic zone of filipin staining at the leading edge of growth. Interestingly,
cdc12-6
cells shifted to 37°C in the absence of serum gradually displayed enriched filipin staining at the tip, which may be due to the altered cell cycle regulation. A striking difference from the wild type was that the
cdc12-6
cells frequently formed a second germ tube in close proximity to the first. The mutant cells also failed to form the diffuse band of septins at the base of germ tubes and hyphae, indicating that this septin band plays a role in preventing proximal formation of germ tubes in a manner analogous to bud site selection. These studies demonstrate that not only are septins important for cytokinesis, but they also promote polarized morphogenesis and selection of germ tube sites that may help disseminate an infection in host tissues.
Fur-Mediated Global Regulatory Circuits in Pathogenic Neisseria Species
Authors: Yu Chunxiao , Genco Caroline Attardo 
Published: August 2012

ABSTRACT

The
f
erric
u
ptake
r
egulator (Fur) protein has been shown to function as a repressor of transcription in a number of diverse microorganisms. However, recent studies have established that Fur can function at a global level as both an activator and a repressor of transcription through both direct and indirect mechanisms. Fur-mediated indirect activation occurs via the repression of additional repressor proteins, or small regulatory RNAs, thereby activating transcription of a previously silent gene. Fur mediates direct activation through binding of Fur to the promoter regions of genes. Whereas the repressive mechanism of Fur has been thoroughly investigated, emerging studies on direct and indirect Fur-mediated activation mechanisms have revealed novel global regulatory circuits.
The β-Lactam Resistance Protein Blr, a Small Membrane Polypeptide, Is a Component of the Escherichia coli Cell Division Machinery
Authors: Karimova Gouzel , Davi Marilyne , Ladant Daniel 
Published: August 2012

ABSTRACT

In
Escherichia coli
, cell division is performed by a multimolecular machinery called the divisome, made of 10 essential proteins and more than 20 accessory proteins. Through a bacterial two-hybrid library screen, we identified the
E. coli
β-lactam resistance protein Blr, a short membrane polypeptide of 41 residues, as an interacting partner of the essential cell division protein FtsL. In addition to FtsL, Blr was found to associate with several other divisomal proteins, including FtsI, FtsK, FtsN, FtsQ, FtsW, and YmgF. Using fluorescently tagged Blr, we showed that this peptide localizes to the division septum and that its colocalization requires the presence of the late division protein FtsN. Although Blr is not essential, previous studies have shown that the inactivation of the
blr
gene increased the sensitivity of bacteria to β-lactam antibiotics or their resistance to cell envelope stress. Here, we found that Blr, when overproduced, restores the viability of
E. coli ftsQ1
(Ts) cells, carrying a thermosensitive allele of the
ftsQ
gene, during growth under low-osmotic-strength conditions (e.g., in synthetic media or in Luria-Bertani broth without NaCl). In contrast, the inactivation of
blr
increases the osmosensitivity of
ftsQ1
(Ts) cells, and
blr ftsQ1
double mutants exhibit filamentous growth in LB broth even at a moderate salt concentration (0.5% NaCl) compared to parental
ftsQ1
(Ts) cells. Altogether, our results suggest that the small membrane polypeptide Blr is a novel component of the
E. coli
cell division apparatus involved in the stabilization of the divisome under certain stress conditions.
The Accessory Sec Protein Asp2 Modulates GlcNAc Deposition onto the Serine-Rich Repeat Glycoprotein GspB
Authors: Seepersaud Ravin , Bensing Barbara A. , Yen Yihfen T. , Sullam Paul M. 
Published: August 2012

ABSTRACT

The accessory Sec system is a specialized transport system that exports serine-rich repeat (SRR) glycoproteins of Gram-positive bacteria. This system contains two homologues of the general secretory (Sec) pathway (SecA2 and SecY2) and several other essential proteins (Asp1 to Asp5) that share no homology to proteins of known function. In
Streptococcus gordonii
, Asp2 is required for the transport of the SRR adhesin GspB, but its role in export is unknown. Tertiary structure predictions suggest that the carboxyl terminus of Asp2 resembles the catalytic region of numerous enzymes that function through a Ser-Asp-His catalytic triad. Sequence alignment of all Asp2 homologues identified a highly conserved pentapeptide motif (Gly-X-Ser
362
-X-Gly) typical of most Ser-Asp-His catalytic triads, where Ser forms the reactive residue. Site-directed mutagenesis of residues comprising the predicted catalytic triad of Asp2 of
S. gordonii
had no effect upon GspB transport but did result in a marked change in the electrophoretic mobility of the protein. Lectin-binding studies and monosaccharide content analysis of this altered glycoform revealed an increase in glucosamine deposition. Random mutagenesis of the Asp2 region containing this catalytic domain also disrupted GspB transport. Collectively, our findings suggest that Asp2 is a bifunctional protein that is essential for both GspB transport and correct glycosylation. The catalytic domain may be responsible for controlling the glycosylation of GspB, while other surrounding regions are functionally required for glycoprotein transport.
Diversity and Versatility of the Thermotoga maritima Sugar Kinome
Authors: Rodionova Irina A. , Yang Chen , Li Xiaoqing , Kurnasov Oleg V. , Best Aaron A. , Osterman Andrei L. , Rodionov Dmitry A. 
Published: August 2012

ABSTRACT

Sugar phosphorylation is an indispensable committed step in a large variety of sugar catabolic pathways, which are major suppliers of carbon and energy in heterotrophic species. Specialized sugar kinases that are indispensable for most of these pathways can be utilized as signature enzymes for the reconstruction of carbohydrate utilization machinery from microbial genomic and metagenomic data. Sugar kinases occur in several structurally distinct families with various partially overlapping as well as yet unknown substrate specificities that often cannot be accurately assigned by homology-based techniques. A subsystems-based metabolic reconstruction combined with the analysis of genome context and followed by experimental testing of predicted gene functions is a powerful approach of functional gene annotation. Here we applied this integrated approach for functional mapping of all sugar kinases constituting an extensive and diverse sugar kinome in the thermophilic bacterium
Thermotoga maritima
. Substrate preferences of 14 kinases mainly from the FGGY and PfkB families were inferred by bioinformatics analysis and biochemically characterized by screening with a panel of 45 different carbohydrates. Most of the analyzed enzymes displayed narrow substrate preferences corresponding to their predicted physiological roles in their respective catabolic pathways. The observed consistency supports the choice of kinases as signature enzymes for genomics-based identification and reconstruction of sugar utilization pathways. Use of the integrated genomic and experimental approach greatly speeds up the identification of the biochemical function of unknown proteins and improves the quality of reconstructed pathways.
The CRISPR/Cas Adaptive Immune System of Pseudomonas aeruginosa Mediates Resistance to Naturally Occurring and Engineered Phages
Authors: Cady Kyle C. , Bondy-Denomy Joe , Heussler Gary E. , Davidson Alan R. , O'Toole George A. 
Published: August 2012

ABSTRACT

Here we report the isolation of 6 temperate bacteriophages (phages) that are prevented from replicating within the laboratory strain
Pseudomonas aeruginosa
PA14 by the endogenous CRISPR/Cas system of this microbe. These phages are only the second identified group of naturally occurring phages demonstrated to be blocked for replication by a nonengineered CRISPR/Cas system, and our results provide the first evidence that the
P. aeruginosa
type I-F CRISPR/Cas system can function in phage resistance. Previous studies have highlighted the importance of the
p
rotospacer
a
djacent
m
otif (PAM) and a proximal 8-nucleotide seed sequence in mediating CRISPR/Cas-based immunity. Through engineering of a protospacer region of phage DMS3 to make it a target of resistance by the CRISPR/Cas system and screening for mutants that escape CRISPR/Cas-mediated resistance, we show that nucleotides within the PAM and seed sequence and across the non-seed-sequence regions are critical for the functioning of this CRISPR/Cas system. We also demonstrate that
P. aeruginosa
can acquire spacer content in response to lytic phage challenge, illustrating the adaptive nature of this CRISPR/Cas system. Finally, we demonstrate that the
P. aeruginosa
CRISPR/Cas system mediates a gradient of resistance to a phage based on the level of complementarity between CRISPR spacer RNA and phage protospacer target. This work introduces a new
in vivo
system to study CRISPR/Cas-mediated resistance and an additional set of tools for the elucidation of CRISPR/Cas function.
Development of a predictive model to identify inpatients at risk of re-admission within 30 days of discharge (PARR-30)
Authors: Billings John , Blunt Ian , Steventon Adam , Georghiou Theo , Lewis Geraint , Bardsley Martin 
Published: August 2012

ObjectivesTo develop an algorithm for identifying inpatients at high risk of re-admission to a National Health Service (NHS) hospital in England within 30 days of discharge using information that can either be obtained from hospital information systems or from the patient and their notes.DesignMultivariate statistical analysis of routinely collected hospital episode statistics (HES) data using logistic regression to build the predictive model. The model's performance was calculated using bootstrapping.SettingHES data covering all NHS hospital admissions in England.ParticipantsThe NHS patients were admitted to hospital between April 2008 and March 2009 (10% sample of all admissions, n=576 868).Main outcome measuresArea under the receiver operating characteristic curve for the algorithm, together with its positive predictive value and sensitivity for a range of risk score thresholds.ResultsThe algorithm produces a ‘risk score’ ranging (0–1) for each admitted patient, and the percentage of patients with a re-admission within 30 days and the mean re-admission costs of all patients are provided for 20 risk bands. At a risk score threshold of 0.5, the positive predictive value (ie, percentage of inpatients identified as high risk who were subsequently re-admitted within 30 days) was 59.2% (95% CI 58.0% to 60.5%); representing 5.4% (95% CI 5.2% to 5.6%) of all inpatients who would be re-admitted within 30 days (sensitivity). The area under the receiver operating characteristic curve was 0.70 (95% CI 0.69 to 0.70).ConclusionsWe have developed a method of identifying inpatients at high risk of unplanned re-admission to NHS hospitals within 30 days of discharge. Though the models had a low sensitivity, we show how to identify subgroups of patients that contain a high proportion of patients who will be re-admitted within 30 days. Additional work is necessary to validate the model in practice.
Junior doctors and patient safety: evaluating knowledge, attitudes and perception of safety climate
Authors: Durani Piyush , Dias Joseph , Singh Harvinder P , Taub Nicholas 
Published: August 2012

IntroductionEngagement of junior doctors in patient safety initiatives is high on the national agenda, but there is a lack of studies evaluating patient safety attitudes among junior doctors.MethodsThe Junior Doctor–Patient Safety Attitudes and Climate Questionnaire is a multidimensional scale created using items from already-validated scales and inclusion of new items based on further review. It consists of three subscales: ‘knowledge and training’ (10 items), ‘attitudes to patient safety’ (15 items) and ‘perception of workplace safety climate’ (15 items). This was disseminated to foundation trainees, general practice trainees and hospital core and speciality trainees via the Deanery distribution lists and responses were collected anonymously.ResultsA total of 527 complete responses were collected; although self-declared knowledge in patient safety concepts was high, there was less declared understanding of a ‘high reliability organisation’ (74% no/unsure) and the concept of active failures/latent conditions (60% no/unsure). The greatest agreement was demonstrated for the statement ‘Even the most experienced and competent doctors make errors’ (p<0.01). However, more senior trainees and surgical trainees (vs medical trainees) demonstrated greater agreement with ‘Medical error is a sign of incompetence’ (p<0.01). More junior trainees demonstrated greater agreement with ‘Management is more interested in meeting performance targets than focusing on patient safety issues’ (p<0.01).ConclusionsThis study demonstrates subtle differences in attitudes to patient safety among junior doctors of different grades and specialities. These should be taken into account when designing interventions to improve patient safety education and culture among junior doctors.
Prehospital delay in acute stroke and TIA
Authors: Faiz Kashif Waqar , Sundseth Antje , Thommessen Bente , Rønning Ole Morten 
Published: August 2012

BackgroundEarly management improves outcome in acute stroke. This study was designed to assess the prehospital path from symptom onset to arrival in hospital and to identify factors associated with prehospital delay.MethodsA prospective study was conducted including patients with acute ischaemic stroke, intracerebral haemorrhage and transient ischaemic attack admitted to hospital. Time intervals for prehospital delay, background data, severity, type of first medical contact and mode of transport were recorded. Univariate and multivariate analyses were performed to identify factors influencing prehospital delay.ResultsA total of 440 patients were included, with a mean age of 71.4±13.0 years (44.3% female subjects), consisting of 65.9% patients with ischaemic stroke, 11.4% with intracerebral haemorrhage and 22.7% with transient ischaemic attack. The median time from symptom onset to admission was 3.0 h (179 min; IQR 77–542). The median decision delay was 1.5 h (92 min, IQR 25–405) and accounted for 55.1% (median value) of the prehospital delay. 310 (70.5%) patients arrived by ambulance. In the multivariate linear regression analysis, high National Institute of Health Stroke Scale score (p<0.001), transport by ambulance (p<0.001) and lower age (p=0.048) were significantly associated with early admission.ConclusionsSevere strokes, use of ambulance and lower age are associated with reduced prehospital delay. The present study shows that more than half of the delay is caused by the hesitation to contact medical services. Public information campaigns should focus on fast symptom recognition and the importance of immediately contacting the Emergency Medical Services upon symptom onset.
Ultrasound guidance for difficult peripheral venous access: systematic review and meta-analysis
Authors: Egan Grace , Healy Donagh , O'Neill Heidi , Clarke-Moloney Mary , Grace Pierce A , Walsh Stewart R 
Published: August 2012

BackgroundEstablishing intravenous access is often vital in an acute hospital setting but can be difficult. Ultrasound-guided cannulation increases success rates in prospective studies. However, these studies have often lacked a comparative group. This systematic review and meta-analysis aimed to determine the clinical effectiveness of Ultrasound-guided peripheral intravenous cannulation compared with the standard technique in patients known to have difficult access.MethodsElectronic abstract databases, trial registries, article reference lists and internet repositories were searched using the following search terms: ‘peripheral venous cannulation’, ‘peripheral venous access’. Studies meeting the following criteria were included: randomised controlled trial patients of all ages who required peripheral intravenous access; interventions were Ultrasound-guided versus standard cannulation technique; patients were identified as having difficult venous access; inclusion of at least one defined outcome (procedural success time to cannula placement; number of attempts).Results7 trials were identified (289 participants). Ultrasound guidance increases the likelihood of successful cannulation (pooled OR 2.42; 95% CI 1.26 to 4.68; p=0.008). There were no differences in time to successful cannulation, or number of percutaneous skin punctures.ConclusionUltrasound guidance increases the likelihood of successful peripheral cannulation in difficult access patients. We recommend its use in patients who have difficult venous access, and have failed venous cannulation by standard methods. Further randomised controlled trials (RCTs) with larger sample sizes would be of benefit to investigate if Ultrasound has any additional advantages in terms of reducing the procedure time and the number of skin punctures required for successful venous cannulation.
Coeliac disease in Asians in a single centre in southern Derbyshire
Authors: Holmes Geoffrey KT , Moor Fiona 
Published: August 2012

BackgroundCoeliac disease affects adult Asians from north India, Pakistan and Bangladesh in the UK but how commonly this occurs is unknown. An audit of coeliac disease was therefore conducted in a well-defined area in southern Derbyshire.MethodsAll white and Asian patients with biopsy-confirmed coeliac disease diagnosed between 1958 and 2008 were identified. Population data from the Office of National Statistics allowed the calculation of prevalence. Presenting symptoms, adherence to a gluten-free diet and follow-up record were determined for Asians and compared with matched white patients.ResultsAmong 1305 coeliac patients diagnosed between 1958 and 2008, 82 were Asian. Coeliac disease occurred significantly more frequently in Asian than white individuals and this could be attributed to the significantly higher prevalence in women 16 years and older and under 60 years of age. No Asian man over the age of 65 years was diagnosed with coeliac disease. Asians are more likely to present with anaemia and less likely to present with diarrhoea than white individuals. Asians are less likely to adhere to a strict gluten-free diet than white patients.ConclusionsThis baseline audit indicates that increased efforts should be directed towards diagnosing coeliac disease in Asian men over the age of 65 years, in whom at present it is unrepresented. Strategies also need to be developed to help more Asian patients adhere strictly to the gluten-free diet.