bluuebunny/crossref_metadata_2025_split huggingface dataset:
Rapid Identification of Pseudallescheria and Scedosporium Strains by Using Rolling Circle Amplification
Authors: Lackner Michaela , Najafzadeh Mohammad Javad , Sun Jiufeng , Lu Qiaoyun , de Hoog G. Sybren 
Published: November 2011

ABSTRACTThePseudallescheria boydiicomplex, comprising environmental pathogens withScedosporiumanamorphs, has recently been subdivided into five main species:Scedosporium dehoogii,S. aurantiacum,Pseudallescheria minutispora,P. apiosperma, andP. boydii, while the validity of some other taxa is being debated. SeveralPseudallescheriaandScedosporiumspecies are indicator organisms of pollution in soil and water.Scedosporium dehoogiiin particular is enriched in soils contaminated by aliphatic hydrocarbons. In addition, the fungi may cause life-threatening infections involving the central nervous system in severely impaired patients. For screening purposes, rapid and economic tools for species recognition are needed. Our aim is to establish rolling circle amplification (RCA) as a screening tool for species-specific identification ofPseudallescheriaandScedosporium. With this aim, a set of padlock probes was designed on the basis of the internal transcribed spacer (ITS) region, differing by up to 13 fixed mutations. Padlock probes were unique as judged from sequence comparison by BLAST search in GenBank and in dedicated research databases at CBS (Centraalbureau voor Schimmelcultures Fungal Biodiversity Centre). RCA was applied as anin vitrotool, tested with pure DNA amplified from cultures. The species-specific padlock probes designed in this study yielded 100% specificity. The method presented here was found to be an attractive alternative to identification by restriction fragment length polymorphism (RFLP) or sequencing. The rapidity (<1 day), specificity, and low costs make RCA a promising screening tool for environmentally and clinically relevant fungi.
Culture and Molecular Method for Detection of Mycobacterium tuberculosis Complex and Mycobacterium avium subsp. paratuberculosis in Milk and Dairy Products
Authors: Messelhäusser U. , Kämpf P. , Hörmansdorfer S. , Wagner B. , Schalch B. , Busch U. , Höller C. , Wallner P. , Barth G. , Rampp A. 
Published: November 2011

ABSTRACT

A combined molecular and cultural method for the detection of the
Mycobacterium tuberculosis
complex (MTBC) and
Mycobacterium avium
subsp.
paratuberculosis
was developed and tested with artificially contaminated milk and dairy products. Results indicate that the method can be used for a reliable detection as a basis for first risk assessments.
Association of Constitutive Hyperphosphorylation of Hsf1p with a Defective Ethanol Stress Response in Saccharomyces cerevisiae Sake Yeast Strains
Authors: Noguchi Chiemi , Watanabe Daisuke , Zhou Yan , Akao Takeshi , Shimoi Hitoshi 
Published: November 2011

ABSTRACT

Modern sake yeast strains, which produce high concentrations of ethanol, are unexpectedly sensitive to environmental stress during sake brewing. To reveal the underlying mechanism, we investigated a well-characterized yeast stress response mediated by a heat shock element (HSE) and heat shock transcription factor Hsf1p in
Saccharomyces cerevisiae
sake yeast. The HSE-
lacZ
activity of sake yeast during sake fermentation and under acute ethanol stress was severely impaired compared to that of laboratory yeast. Moreover, the Hsf1p of modern sake yeast was highly and constitutively hyperphosphorylated, irrespective of the extracellular stress. Since
HSF1
allele replacement did not significantly affect the HSE-mediated ethanol stress response or Hsf1p phosphorylation patterns in either sake or laboratory yeast, the regulatory machinery of Hsf1p is presumed to function differently between these types of yeast. To identify phosphatases whose loss affected the control of Hsf1p, we screened a series of phosphatase gene deletion mutants in a laboratory strain background. Among the 29 mutants, a Δ
ppt1
mutant exhibited constitutive hyperphosphorylation of Hsf1p, similarly to the modern sake yeast strains, which lack the entire
PPT1
gene locus. We confirmed that the expression of laboratory yeast-derived functional
PPT1
recovered the HSE-mediated stress response of sake yeast. In addition, deletion of
PPT1
in laboratory yeast resulted in enhanced fermentation ability. Taken together, these data demonstrate that hyperphosphorylation of Hsf1p caused by loss of the
PPT1
gene at least partly accounts for the defective stress response and high ethanol productivity of modern sake yeast strains.
Locked Nucleic Acid and Flow Cytometry-Fluorescence
In Situ
Hybridization for the Detection of Bacterial Small Noncoding RNAs
Authors: Robertson Kelly L. , Vora Gary J. 
Published: November 2011

ABSTRACT

We describe the development and testing of a high-throughput method that enables the detection of small noncoding RNAs (ncRNAs) from single bacterial cells using locked nucleic acid probes (LNA) and flow cytometry-fluorescence
in situ
hybridization (flow-FISH). The LNA flow-FISH method and quantitative reverse transcription-PCR (qRT-PCR) were used to monitor the expression of three ncRNAs (6S, CsrB, and TPP-2) in
Vibrio campbellii
ATCC BAA-1116 cultures during lag phase, mid-log phase, and stationary phase. Both LNA flow-FISH and qRT-PCR revealed that CsrB and TPP-2 were highly expressed during lag phase but markedly reduced in mid-log phase and stationary phase, whereas 6S demonstrated no to little expression during lag phase but increased thereafter. Importantly, while LNA flow-FISH and qRT-PCR demonstrated similar overall expression trends, only LNA flow-FISH, which enabled the detection of ncRNAs in individual cells as opposed to the lysate-based ensemble measurements generated by qRT-PCR, was able to capture the cell-to-cell heterogeneity in ncRNA expression. As such, this study demonstrates a new method that simultaneously enables the
in situ
detection of ncRNAs and the determination of gene expression heterogeneity within an isogenic bacterial population.
Disease Dynamics and Persistence of Musca domestica Salivary Gland Hypertrophy Virus Infections in Laboratory House Fly (Musca domestica) Populations
Authors: Lietze Verena-Ulrike , Geden Christopher J. , Doyle Melissa A. , Boucias Drion G. 
Published: November 2011

ABSTRACT

Past surveys of feral house fly populations have shown that
Musca domestica
salivary gland hypertrophy virus (MdSGHV) has a worldwide distribution, with an average prevalence varying between 0.5% and 10%. How this adult-specific virus persists in nature is unknown. In the present study, experiments were conducted to examine short-term transmission efficiency and long-term persistence of symptomatic MdSGHV infections in confined house fly populations. Average rates of disease transmission from virus-infected to healthy flies in small populations of 50 or 100 flies ranged from 3% to 24% and did not vary between three tested geographical strains that originated from different continents. Introduction of an initial proportion of 40% infected flies into fly populations did not result in epizootics. Instead, long-term observations demonstrated that MdSGHV infection levels declined over time, resulting in a 10% infection rate after passing through 10 filial generations. In all experiments, induced disease rates were significantly higher in male flies than in female flies and might be explained by male-specific behaviors that increased contact with viremic flies and/or virus-contaminated surfaces.
Characterization of Streptomyces padanus JAU4234, a Producer of Actinomycin X
2
, Fungichromin, and a New Polyene Macrolide Antibiotic
Authors: Xiong Zhi-Qiang , Zhang Zhi-Ping , Li Jiang-Huai , Wei Sai-Jin , Tu Guo-Quan 
Published: November 2011

ABSTRACT

Strain JAU4234, identified as
Streptomyces padanus
, was isolated from soil collected in Jiangxi Province, China. It produced actinomycin X2, fungichromin, and a new polyene macrolide compound with antifungal activity, antifungalmycin 702. Antifungalmycin 702 had good general antifungal activity and may have potential future agricultural and/or clinical applications.
A Trypanosomal Pentatricopeptide Repeat Protein Stabilizes the Mitochondrial mRNAs of Cytochrome Oxidase Subunits 1 and 2
Authors: Pusnik Mascha , Schneider André 
Published: November 2011

ABSTRACT

The pentatricopeptide repeat (PPR) protein family consists of organellar proteins predicted to bind to specific RNA sequences. Plants have hundreds of distinct PPR proteins, whereas other eukaryotes generally have many fewer. The genome of the parasitic protozoon
Trypanosoma brucei
is predicted to encode more than 30 different PPR proteins, which is an extraordinarily high number for a nonplant organism. Here we report the characterization
T. brucei
PPR9 (TbPPR9). Epitope tagging shows that the protein is exclusively mitochondrially localized. Interestingly, while in induced RNA interference cell lines TbPPR9 is efficiently downregulated, the level of its mRNA is not affected. Ablation of TbPPR9 selectively abolishes oxidative but not mitochondrial substrate-level phosphorylation. The molecular basis of this phenotype is the fact that TbPPR9 is required for the stability of the cytochrome oxidase subunit 1 (COX1) and COX2 mRNAs. This is supported by the observation that ablation of TbPPR9 destabilizes the COX complex but not the cytochrome
bc
1
or the ATP synthase complex. Moreover, it was shown by blue native gel electrophoresis that TbPPR9 is present in a large complex of unknown composition.
Proteomic Analysis of Trichoderma atroviride Reveals Independent Roles for Transcription Factors BLR-1 and BLR-2 in Light and Darkness
Authors: Sánchez-Arreguín Alejandro , Pérez-Martínez Ana Silvia , Herrera-Estrella Alfredo 
Published: November 2011

ABSTRACT

The genus
Trichoderma
is one of the most widely used biological control agents of plant-pathogenic fungi. The main mechanism for survival and dispersal of
Trichoderma
is through the production of asexual spores (conidia). The transition from filamentous growth to conidiation can be triggered by light, nutrient deprivation, and mechanical damage of the mycelium. We conducted proteomic profiling analyses of
Trichoderma atroviride
after a blue light pulse. The use of two-dimensional electrophoresis (2-DE) and mass spectrometry (MS) analysis allowed us to identify 72 proteins whose expression was affected by blue light. Functional category analysis showed that the various proteins are involved in metabolism, cell rescue, and protein synthesis. We determined the relationship between mRNA levels of selected genes 30 min after a light pulse and protein expression levels at different times after the pulse and found this correlation to be very weak. The correlation was highest when protein and mRNA levels were compared for the same time point. The transcription factors BLR-1 and BLR-2 are vital to the photoconidiation process; here we demonstrate that both BLR proteins are active in darkness and affect several elements at both the transcript and protein levels. Unexpectedly, in darkness, downregulation of proteins prevailed in the Δ
blr-1
mutant, while upregulation of proteins predominated in the Δ
blr-2
mutant. Our data demonstrate that the BLR proteins play roles individually and as a complex.
Characterization of DNA Sequences Required for the CcrAB-Mediated Integration of Staphylococcal Cassette Chromosome
mec
, a Staphylococcus aureus Genomic Island
Authors: Wang Lei , Safo Martin , Archer Gordon L. 
Published: November 2011

ABSTRACT

The mobile element staphylococcal cassette chromosome
mec
(SC
Cmec
), which carries
mecA
, the gene responsible for methicillin resistance in staphylococci, inserts into the chromosome at a specific site,
attB
, mediated by serine recombinases, CcrAB and CcrC, encoded on the element. This study sought to determine the sequence specificity for CcrB DNA binding
in vitro
and for CcrAB-mediated SCC
mec
insertion
in vivo
. CcrB DNA binding, as assessed
in vitro
by electrophoretic mobility shift assay (EMSA), revealed that a 14-bp sequence (
CGTATCATAAGTAA
; the terminal sequence of the
orfX
gene) was the minimal requirement for binding, containing an invariant sequence (
TATCATAA
) found in all chromosomal (
attB
) and SCC
mec
(
attS
) integration sites. The sequences flanking the minimal
attB
and
attS
binding sites required for insertion
in vivo
were next determined. A plasmid containing only 37 bp of
attS
and flanking sequences was required for integration into the
attB
site at 92% efficiency. In contrast, at least 200 bp of sequence within
orfX
, 5′ to the
attB
core, and 120 bp of specific sequence 3′ to the
orfX
stop site and
attB
core were required for the highest insertion frequency. Finally, an
attS
-containing plasmid was inserted into wild-type
Staphylococcus aureus
strains without integrated SCC
mec
(methicillin susceptible) at various frequencies which were determined both by sequences flanking the
att
site and by the presence of more than one
att
site on either the chromosome or the integration plasmid. This sequence specificity may play a role in the epidemiology of SCC
mec
acquisition.
Novel Roles of SoxR, a Transcriptional Regulator from
X
anthomonas campestris, in Sensing Redox-Cycling Drugs and Regulating a Protective Gene That Have Overall Implications for Bacterial Stress Physiology and Virulence on a Host Plant
Authors: Mahavihakanont Aekkapol , Charoenlap Nisanart , Namchaiw Poommaree , Eiamphungporn Warawan , Chattrakarn Sorayut , Vattanaviboon Paiboon , Mongkolsuk Skorn 
Published: November 2011

ABSTRACT

In
Xanthomonas campestris
pv. campestris, SoxR likely functions as a sensor of redox-cycling drugs and as a transcriptional regulator. Oxidized SoxR binds directly to its target site and activates the expression of
xcc0300
, a gene that has protective roles against the toxicity of redox-cycling compounds. In addition, SoxR acts as a noninducible repressor of its own expression.
X. campestris
pv. campestris requires SoxR both for protection against redox-cycling drugs and for full virulence on a host plant. The
X. campestris
model of the gene regulation and physiological roles of SoxR represents a novel variant of existing bacterial SoxR models.
Network Identification and Flux Quantification of Glucose Metabolism in Rhodobacter sphaeroides under Photoheterotrophic H
2
-Producing Conditions
Authors: Tao Yongzhen , Liu Deng , Yan Xing , Zhou Zhihua , Lee Jeong K. , Yang Chen 
Published: November 2011

ABSTRACT

The nonsulfur purple bacteria that exhibit unusual metabolic versatility can produce hydrogen gas (H
2
) using the electrons derived from metabolism of organic compounds during photoheterotrophic growth. Here, based on
13
C tracer experiments, we identified the network of glucose metabolism and quantified intracellular carbon fluxes in
Rhodobacter sphaeroides
KD131 grown under H
2
-producing conditions. Moreover, we investigated how the intracellular fluxes in
R. sphaeroides
responded to knockout mutations in hydrogenase and poly-β-hydroxybutyrate synthase genes, which led to increased H
2
yield. The relative contribution of the Entner-Doudoroff pathway and Calvin-Benson-Bassham cycle to glucose metabolism differed significantly in hydrogenase-deficient mutants, and this flux change contributed to the increased formation of the redox equivalent NADH. Disruption of hydrogenase and poly-β-hydroxybutyrate synthase resulted in a significantly increased flux through the phosphoenolpyruvate carboxykinase and a reduced flux through the malic enzyme. A remarkable increase in the flux through the tricarboxylic acid cycle, a major NADH producer, was observed for the mutant strains. The
in vivo
regulation of the tricarboxylic acid cycle flux in photoheterotrophic
R. sphaeroides
was discussed based on the measurements of
in vitro
enzyme activities and intracellular concentrations of NADH and NAD
+
. Overall, our results provide quantitative insights into how photoheterotrophic cells manipulate the metabolic network and redistribute intracellular fluxes to generate more electrons for increased H
2
production.
The Cyanobacterial NAD Kinase Gene
sll1415
Is Required for Photoheterotrophic Growth and Cellular Redox Homeostasis in Synechocystis sp. Strain PCC 6803
Authors: Gao Hong , Xu Xudong 
Published: November 2011

ABSTRACT

NAD kinase (NADK), which phosphorylates NAD to NADP, is one of the key enzymes regulating the cellular NADP(H) level. In
Synechocystis
sp. strain PCC 6803,
slr0400
and
sll1415
were shown to encode NAD kinases. The NADP(H) pool in the cyanobacterium was remarkably reduced by an
sll1415
-null mutation but slightly reduced by an
slr0400
-null mutation. The reduction of the NADP(H) level in the
sll1415
mutant led to a significant accumulation of glucose-6-phosphate and a loss of photoheterotrophic growth. As the primary NADK gene,
sll1415
was found to inhibit the transcription of genes involved in redox homeostasis and to exert stronger effects on methyl viologen tolerance than
slr0040
.
BpsR Modulates Bordetella Biofilm Formation by Negatively Regulating the Expression of the Bps Polysaccharide
Authors: Conover Matt S. , Redfern Crystal J. , Ganguly Tridib , Sukumar Neelima , Sloan Gina , Mishra Meenu , Deora Rajendar 
Published: November 2011

ABSTRACT

Bordetella
bacteria are Gram-negative respiratory pathogens of animals, birds, and humans. A hallmark feature of some
Bordetella
species is their ability to efficiently survive in the respiratory tract even after vaccination.
Bordetella bronchiseptica
and
Bordetella pertussis
form biofilms on abiotic surfaces and in the mouse respiratory tract. The Bps exopolysaccharide is one of the critical determinants for biofilm formation and the survival of
Bordetella
in the murine respiratory tract. In order to gain a better understanding of regulation of biofilm formation, we sought to study the mechanism by which Bps expression is controlled in
Bordetella
. Expression of
bpsABCD
(
bpsA-D
) is elevated in biofilms compared with levels in planktonically grown cells. We found that
bpsA-D
is expressed independently of BvgAS. Subsequently, we identified an open reading frame (ORF), BB1771 (designated here
bpsR
), that is located upstream of and in the opposite orientation to the
bpsA-D
locus. BpsR is homologous to the MarR family of transcriptional regulators. Measurement of
bpsA
and
bpsD
transcripts and the Bps polysaccharide levels from the wild-type and the Δ
bpsR
strains suggested that BpsR functions as a repressor. Consistent with enhanced production of Bps, the
bpsR
mutant displayed considerably more structured biofilms. We mapped the
bpsA-D
promoter region and showed that purified BpsR protein specifically bound to the
bpsA-D
promoter. Our results provide mechanistic insights into the regulatory strategy employed by
Bordetella
for control of the production of the Bps polysaccharide and biofilm formation.
Effects on Growth by Changes of the Balance between GreA, GreB, and DksA Suggest Mutual Competition and Functional Redundancy in Escherichia coli
Authors: Vinella Daniel , Potrykus Katarzyna , Murphy Helen , Cashel Michael 
Published: November 2011

ABSTRACT

It is well known that ppGpp and DksA interact with bacterial RNA polymerase (RNAP) to alter promoter activity. This study suggests that GreA plays a major role and GreB plays a minor role in the ppGpp-DksA regulatory network. We present evidence that DksA and GreA/GreB are redundant and/or share similar functions: (i) on minimal medium GreA overproduction suppresses the growth defects of a
dksA
mutant; (ii) GreA and DksA overexpression partially suppresses the auxotrophy of a ppGpp-deficient strain; (iii) microarrays show that many genes are regulated similarly by GreA and DksA. We also find instances where GreA and DksA seem to act in opposition: (i) complete suppression of auxotrophy occurs by overexpression of GreA or DksA only in the absence of the other protein; (ii) P
gadA
and P
gadE
promoter fusions, along with many other genes, are dramatically affected
in vivo
by GreA overproduction only when DksA is absent; (iii) GreA and DksA show opposite regulation of a subset of genes. Mutations in key acidic residues of GreA and DksA suggest that properties seen here probably are not explained by known biochemical activities of these proteins. Our results indicate that the general pattern of gene expression and, in turn, the ability of
Escherichia coli
to grow under a defined condition are the result of a complex interplay between GreA, GreB, and DksA that also involves mutual control of their gene expression, competition for RNA polymerase binding, and similar or opposite action on RNA polymerase activity.
Voxel-based analysis of apparent diffusion coefficient in perihaematomal oedema: associated factors and outcome predictive value for intracerebral haemorrhage
Authors: Tsai Yuan-Hsiung , Hsu Li-Ming , Weng Hsu-Huei , Lee Ming-Hsueh , Yang Jen-Tsung , Lin Ching-Po 
Published: November 2011

ObjectivesThe pathophysiology of perihaematomal oedema (PO) surrounding a primary intracerebral haemorrhage (ICH) is complicated and incompletely understood. We prospectively investigated the components of PO with voxel-based analysis of the apparent diffusion coefficient (ADC) value and assessed its predictive value for functional outcome.DesignForty-six patients with ICH who were enrolled for clinical evaluation underwent MRI scans within 24 h after ICH. Based on the ADC value of the ipsilateral voxels divided by the mean ADC value of the contralateral mirror region of interest, the voxels with oedema were classified into three categories: cytotoxic, vasogenic and undetermined. The percentages of cytotoxic and vasogenic oedema were then calculated and correlated with clinical outcome according to the modified Rankin Scale (mRS) at 6 months after ICH. The intraobserver and interobserver reliability of this method were examined using intraclass correlation coefficients.ResultsThe intraclass correlation coefficients showed that analysis using the voxel-based method is highly reliable. Among the clinical variables tested, age and serum creatinine levels were positively correlated with percentage of cytotoxic oedema. Age, history of coronary artery disease, National Institutes of Health Stroke Scale score and percentage of cytotoxic oedema were all associated with mRS at 6 months after ICH.ConclusionsThe pathophysiological processes within PO are complicated. Voxel-based analysis of ADC values may help to identify the components of PO and may be beneficial for decision making and predicting outcome.
Management of patient adherence to medications: protocol for an online survey of doctors, pharmacists and nurses in Europe
Authors: Clyne Wendy , Mshelia Comfort , Hall Stephanie , McLachlan Sarah , Jones Peter , Dobbels Fabienne , Ruppar Todd , Siebens Kaat , Morrison Val , De Geest Sabina , Kardas Premyzlaw 
Published: November 2011

IntroductionIt is widely recognised that many patients do not take prescribed medicines as advised. Research in this field has commonly focused on the role of the patient in non-adherence; however, healthcare professionals can also have a major influence on patient behaviour in taking medicines. This study examines the perceptions, beliefs and behaviours of healthcare professionals—doctors, pharmacists and nurses—about patient medication adherence.Methods and analysisThis paper describes the study protocol and online questionnaire used in a cross-sectional survey of healthcare professionals in Europe. The participating countries include Austria, Belgium, France, Greece, The Netherlands, Germany, Poland, Portugal, Switzerland, Hungary, Italy and England. The study population comprises primary care and community-based doctors, pharmacists and nurses involved in the care of adult patients taking prescribed medicines for chronic and acute illnesses.DiscussionKnowledge of the nature, extent and variability of the practices of healthcare professionals to support medication adherence could inform future service design, healthcare professional education, policy and research.