bluuebunny/crossref_metadata_2025_split huggingface dataset:
Transient Effects of Anesthetics on Dendritic Spines and Filopodia in the Living Mouse Cortex
Authors: Yang Guang , Chang Paul C. , Bekker Alex , Blanck Thomas J.J. , Gan Wen-Biao 
Published: July 2011

Background
Anesthetics are widely used to induce unconsciousness, pain relief, and immobility during surgery. It remains unclear whether the use of anesthetics has significant and long-lasting effects on synapse development and plasticity in the brain. To address this question, the authors examined the formation and elimination of dendritic spines, postsynaptic sites of excitatory synapses, in the developing mouse cortex during and after anesthetics exposure.

Methods
Transgenic mice expressing yellow fluorescence protein in layer 5 pyramidal neurons were used in this study. Mice at 1 month of age underwent ketamine-xylazine and isoflurane anesthesia over a period of hours. The elimination and formation rates of dendritic spines and filopodia, the precursors of spines, were followed over hours to days in the primary somatosensory cortex using transcranial two-photon microscopy. Four to five animals were examined under each experimental condition. Student t test and Mann-Whitney U test were used to analyze the data.

Results
Administration of either ketamine-xylazine or isoflurane rapidly altered dendritic filopodial dynamics but had no significant effects on spine dynamics. Ketamine-xylazine increased filopodial formation whereas isoflurane decreased filopodial elimination during 4 h of anesthesia. Both effects were transient and disappeared within a day after the animals woke up.

Conclusion
Studies suggest that exposure to anesthetics transiently affects the dynamics of dendritic filopodia but has no significant effect on dendritic spine development and plasticity in the cortex of 1-month-old mice.
High Oxygen Partial Pressure Decreases Anemia-induced Heart Rate Increase Equivalent to Transfusion
Authors: Feiner John R. , Finlay-Morreale Heather E. , Toy Pearl , Lieberman Jeremy A. , Viele Maurene K. , Hopf Harriet W. , Weiskopf Richard B. 
Published: July 2011

Background
Anemia is associated with morbidity and mortality and frequently leads to transfusion of erythrocytes. The authors sought to directly compare the effect of high inspired oxygen fraction versus transfusion of erythrocytes on the anemia-induced increased heart rate (HR) in humans undergoing experimental acute isovolemic anemia.

Methods
The authors combined HR data from healthy subjects undergoing experimental isovolemic anemia in seven studies performed by the group. HR changes associated with breathing 100% oxygen by nonrebreathing facemask versus transfusion of erythrocytes at their nadir hemoglobin concentration of 5 g/dl were examined. Data were analyzed using a mixed-effects model.

Results
HR had an inverse linear relationship to hemoglobin concentration with a mean increase of 3.9 beats per min per gram of hemoglobin (beats/min/g hemoglobin) decrease (95% CI, 3.7-4.1 beats/min/g hemoglobin), P < 0.0001. Return of autologous erythrocytes significantly decreased HR by 5.3 beats/min/g hemoglobin (95% CI, 3.8-6.8 beats/min/g hemoglobin) increase, P < 0.0001. HR at nadir hemoglobin of 5.6 g/dl (95% CI, 5.5-5.7 g/dl) when breathing air (91.4 beats/min; 95% CI, 87.6-95.2 beats/min) was reduced by breathing 100% oxygen (83.0 beats/min; 95% CI, 79.0-87.0 beats/min), P < 0.0001. The HR at hemoglobin 5.6 g/dl when breathing oxygen was equivalent to the HR at hemoglobin 8.9 g/dl when breathing air.

Conclusions
High arterial oxygen partial pressure reverses the heart rate response to anemia, probably because of its usability rather than its effect on total oxygen content. The benefit of high arterial oxygen partial pressure has significant potential clinical implications for the acute treatment of anemia and results of transfusion trials.
Multiparameter Viability Assay for Stress Profiling Applied to the Food Pathogen Listeria monocytogenes F2365
Authors: Nocker Andreas , Caspers Martien , Esveld-Amanatidou Athina , van der Vossen Jos , Schuren Frank , Montijn Roy , Kort Remco 
Published: July 2011

ABSTRACT

A novel generic approach for stress profiling was applied to
Listeria monocytogenes
strain F2365. This food-borne pathogen was exposed to gradients of five different stresses of increasing intensity, typically ranging from moderate to lethal conditions. The stress factors included heat, acidic pH, a detergent disinfectant, an oxidant, and hyperosmotic conditions. In addition to CFU counts and lag time, five different molecular viability parameters were measured by fluorescence-based assays, including membrane integrity, membrane potential, esterase activity, redox activity, and intracellular pH stability. The last was measured by our recently invented real-time viability assay. Exposure to all stresses resulted in clear dose-response relationships for all viability parameters with the exception of hyperosmotic conditions. A statistical analysis showed strong correlations for (i) the growth parameters plate counts and lag times, (ii) the enzyme-associated functions redox and esterase activity, and (iii) the membrane-associated pH stability and membrane integrity. Results indicated a pronounced difference in the susceptibilities of the measured parameters depending on the stress factor applied. However, at relatively high stress intensities, all of the viability parameters became affected independent of the stress factor. Applications of the approach presented here include studies on the mechanism of action of unknown compounds with biocidal activity and a comparative analysis of the severities of the impact of stress conditions of interest. It appears that a meaningful evaluation of the impact of mild stress conditions can be obtained only through measurement of multiple viability parameters.
A Limited Microbial Consortium Is Responsible for Extended Bioreduction of Uranium in a Contaminated Aquifer
Authors: Gihring Thomas M. , Zhang Gengxin , Brandt Craig C. , Brooks Scott C. , Campbell James H. , Carroll Susan , Criddle Craig S. , Green Stefan J. , Jardine Phil , Kostka Joel E. , Lowe Kenneth , Mehlhorn Tonia L. , Overholt Will , Watson David B. , Yang Zamin , Wu Wei-Min , Schadt Christopher W. 
Published: July 2011

ABSTRACT

Subsurface amendments of slow-release substrates (e.g., emulsified vegetable oil [EVO]) are thought to be a pragmatic alternative to using short-lived, labile substrates for sustained uranium bioimmobilization within contaminated groundwater systems. Spatial and temporal dynamics of subsurface microbial communities during EVO amendment are unknown and likely differ significantly from those of populations stimulated by soluble substrates, such as ethanol and acetate. In this study, a one-time EVO injection resulted in decreased groundwater U concentrations that remained below initial levels for approximately 4 months. Pyrosequencing and quantitative PCR of 16S rRNA from monitoring well samples revealed a rapid decline in groundwater bacterial community richness and diversity after EVO injection, concurrent with increased 16S rRNA copy levels, indicating the selection of a narrow group of taxa rather than a broad community stimulation. Members of the
Firmicutes
family
Veillonellaceae
dominated after injection and most likely catalyzed the initial oil decomposition. Sulfate-reducing bacteria from the genus
Desulforegula
, known for long-chain fatty acid oxidation to acetate, also dominated after EVO amendment. Acetate and H
2
production during EVO degradation appeared to stimulate NO
3
−
, Fe(III), U(VI), and SO
4
2−
reduction by members of the
Comamonadaceae
,
Geobacteriaceae
, and
Desulfobacterales
. Methanogenic archaea flourished late to comprise over 25% of the total microbial community. Bacterial diversity rebounded after 9 months, although community compositions remained distinct from the preamendment conditions. These results demonstrated that a one-time EVO amendment served as an effective electron donor source for
in situ
U(VI) bioreduction and that subsurface EVO degradation and metal reduction were likely mediated by successive identifiable guilds of organisms.
Real-Time Detection and Identification of Chlamydophila Species in Veterinary Specimens by Using SYBR Green-Based PCR Assays
Authors: Nordentoft Steen , Kabell Susanne , Pedersen Karl 
Published: July 2011

ABSTRACT

Infections caused by members of the
Chlamydiaceae
family have long been underestimated due to the requirement of special laboratory facilities for the detection of this group of intracellular pathogens. Furthermore, new studies of this group of intracellular pathogens have revealed that host specificity of different species is not as clear as recently believed. As most members of the genus
Chlamydophila
have shown to be transmissible from animals to humans, sensitive and fast detection methods are required. In this study, SYBR green-based real-time assays were developed that detect all members of
Chlamydiaceae
and differentiate the most prevalent veterinary
Chlamydophila
species:
Cp. psittaci
,
Cp. abortus
,
Cp. felis
, and
Cp. caviae
. By adding bovine serum albumin to the master mixes, target DNA could be detected directly in crude lysates of enzymatically digested conjunctival or pharyngeal swabs or tissue specimens from heart, liver, and spleen without further purification. The assays were evaluated on veterinary specimens where all samples were screened using a family-specific PCR, and positive samples were further tested using species-specific PCRs.
Cp. psittaci
was detected in 47 birds,
Cp. felis
was found in 10 cats,
Cp. caviae
was found in one guinea pig, and
Cp. abortus
was detected in one sheep. The screening assay appeared more sensitive than traditional microscopical examination of stained tissue smears. By combining a fast, robust, and cost-effective method for sample preparation with a highly sensitive family-specific PCR, we were able to screen for
Chlamydiaceae
in veterinary specimens and confirm the species in positive samples with additional PCR assays.
Changes in Diversity and Functional Gene Abundances of Microbial Communities Involved in Nitrogen Fixation, Nitrification, and Denitrification in a Tidal Wetland versus Paddy Soils Cultivated for Different Time Periods
Authors: Bannert Andrea , Kleineidam Kristina , Wissing Livia , Mueller-Niggemann Cornelia , Vogelsang Vanessa , Welzl Gerhard , Cao Zhihong , Schloter Michael 
Published: July 2011

ABSTRACT

In many areas of China, tidal wetlands have been converted into agricultural land for rice cultivation. However, the consequences of land use changes for soil microbial communities are poorly understood. Therefore, we investigated bacterial and archaeal communities involved in inorganic nitrogen turnover (nitrogen fixation, nitrification, and denitrification) based on abundances and relative species richness of the corresponding functional genes along a soil chronosequence ranging between 50 and 2,000 years of paddy soil management compared to findings for a tidal wetland. Changes in abundance and diversity of the functional groups could be observed, reflecting the different chemical and physical properties of the soils, which changed in terms of soil development. The tidal wetland was characterized by a low microbial biomass and relatively high abundances of ammonia-oxidizing microbes. Conversion of the tidal wetlands into paddy soils was followed by a significant increase in microbial biomass. Fifty years of paddy management resulted in a higher abundance of nitrogen-fixing microbes than was found in the tidal wetland, whereas dominant genes of nitrification and denitrification in the paddy soils showed no differences. With ongoing rice cultivation, copy numbers of archaeal ammonia oxidizers did not change, while that of their bacterial counterparts declined. The
nirK
gene, coding for nitrite reductase, increased with rice cultivation time and dominated its functionally redundant counterpart,
nirS
, at all sites under investigation. Relative species richness showed significant differences between all soils with the exception of the archaeal ammonia oxidizers in the paddy soils cultivated for 100 and 300 years. In general, changes in diversity patterns were more pronounced than those in functional gene abundances.
Novel Virulent and Broad-Host-Range Erwinia amylovora Bacteriophages Reveal a High Degree of Mosaicism and a Relationship to Enterobacteriaceae Phages
Authors: Born Yannick , Fieseler Lars , Marazzi Janine , Lurz Rudi , Duffy Brion , Loessner Martin J. 
Published: July 2011

ABSTRACTA diverse set of 24 novel phages infecting the fire blight pathogenErwinia amylovorawas isolated from fruit production environments in Switzerland. Based on initial screening, four phages (L1, M7, S6, and Y2) with broad host ranges were selected for detailed characterization and genome sequencing. Phage L1 is a member of thePodoviridae, with a 39.3-kbp genome featuring invariable genome ends with direct terminal repeats. Phage S6, another podovirus, was also found to possess direct terminal repeats but has a larger genome (74.7 kbp), and the virus particle exhibits a complex tail fiber structure. Phages M7 and Y2 both belong to theMyoviridaefamily and feature long, contractile tails and genomes of 84.7 kbp (M7) and 56.6 kbp (Y2), respectively, with direct terminal repeats. The architecture of all four phage genomes is typical for tailed phages, i.e., organized into function-specific gene clusters. All four phages completely lack genes or functions associated with lysogeny control, which correlates well with their broad host ranges and indicates strictly lytic (virulent) lifestyles without the possibility for host lysogenization. Comparative genomics revealed that M7 is similar toE. amylovoravirus ΦEa21-4, whereas L1, S6, and Y2 are unrelated to any otherE. amylovoraphage. Instead, they feature similarities to enterobacterial viruses T7, N4, and ΦEcoM-GJ1. In a series of laboratory experiments, we provide proof of concept that specific two-phage cocktails offer the potential for biocontrol of the pathogen.
Determination of the Catalytic Base in Family 48 Glycosyl Hydrolases
Authors: Kostylev Maxim , Wilson David B. 
Published: July 2011

ABSTRACT

The catalytic base in family 48 glycosyl hydrolases has not been previously established experimentally. Based on structural and modeling data published to date, we used site-directed mutagenesis and azide rescue activity assays to show definitively that the catalytic base in
Thermobifida fusca
Cel48A is aspartic acid 225. Of the tested mutants, only Cel48A with the D225E mutation retained partial activity on soluble and insoluble substrates. In azide rescue experiments, only the D225G mutation, in the smallest residue tested, showed an increase in activity with added azide.
Action at a Distance: Amino Acid Substitutions That Affect Binding of the Phosphorylated CheY Response Regulator and Catalysis of Dephosphorylation Can Be Far from the CheZ Phosphatase Active Site
Authors: Freeman Ashalla M. , Mole Beth M. , Silversmith Ruth E. , Bourret Robert B. 
Published: July 2011

ABSTRACT

Two-component regulatory systems, in which phosphorylation controls the activity of a response regulator protein, provide signal transduction in bacteria. For example, the phosphorylated CheY response regulator (CheYp) controls swimming behavior. In
Escherichia coli
, the chemotaxis phosphatase CheZ stimulates the dephosphorylation of CheYp. CheYp apparently binds first to the C terminus of CheZ and then binds to the active site where dephosphorylation occurs. The phosphatase activity of the CheZ
2
dimer exhibits a positively cooperative dependence on CheYp concentration, apparently because the binding of the first CheYp to CheZ
2
is inhibited compared to the binding of the second CheYp. Thus, CheZ phosphatase activity is reduced at low CheYp concentrations. The CheZ21IT gain-of-function substitution, located far from either the CheZ active site or C-terminal CheY binding site, enhances CheYp binding and abolishes cooperativity. To further explore mechanisms regulating CheZ activity, we isolated 10 intragenic suppressor mutations of
cheZ21IT
that restored chemotaxis. The suppressor substitutions were located along the central portion of CheZ and were not allele specific. Five suppressor mutants tested biochemically diminished the binding of CheYp and/or the catalysis of dephosphorylation, even when the suppressor substitutions were distant from the active site. One suppressor mutant also restored cooperativity to CheZ21IT. Consideration of results from this and previous studies suggests that the binding of CheYp to the CheZ active site (not to the C terminus) is rate limiting and leads to cooperative phosphatase activity. Furthermore, amino acid substitutions distant from the active site can affect CheZ catalytic activity and CheYp binding, perhaps via the propagation of structural or dynamic perturbations through a helical bundle.
The Rickettsia prowazekii ExoU Homologue Possesses Phospholipase A
1
(PLA
1
), PLA
2
, and Lyso-PLA
2
Activities and Can Function in the Absence of Any Eukaryotic Cofactors
In Vitro
Authors: Housley Nicole A. , Winkler Herbert H. , Audia Jonathon P. 
Published: July 2011

ABSTRACT

Here we have characterized the
Rickettsia prowazekii
RP534 protein, a homologue of the
Pseudomonas aeruginosa
ExoU phospholipase A (PLA) secreted cytotoxin. Our studies showed that purified recombinant RP534 PLA possessed the predicted PLA
2
and lyso-PLA
2
activities based on what has been published for
P. aeruginosa
ExoU. RP534 also displayed PLA
1
activity under the conditions tested, whereas ExoU did not. In addition, recombinant RP534 displayed a basal PLA activity that could hydrolyze phosphatidylcholine in the absence of any eukaryotic cofactors. Interestingly, the addition of bovine liver superoxide dismutase 1 (SOD1), a known activator of
P. aeruginosa
ExoU, resulted in an increased rate of RP534-catalyzed phospholipid hydrolysis, indicating that mechanisms of activation of the ExoU family of PLAs may be evolutionarily conserved. The mechanism of SOD1-dependent stimulation of RP534 was further examined using active site mutants and a fluorogenic phospholipid substrate whose hydrolysis by RP534 over a short time course is measureable only in the presence of SOD1. These studies suggest a mechanism by which SOD1 stimulates RP534 activity once it has bound to the substrate. We also show that antibody raised against RP534 was useful for immunoprecipitating active RP534 from
R. prowazekii
lysed cell extracts, thus verifying that this protein is expressed and active in rickettsiae isolated from embryonated hen egg yolk sacs.
The Heat-Resistant Agglutinin Family Includes a Novel Adhesin from Enteroaggregative Escherichia coli Strain 60A
Authors: Mancini Justin , Weckselblatt Brooke , Chung Yoonjie K. , Durante Julia C. , Andelman Steven , Glaubman Jessica , Dorff Justin D. , Bhargava Samhita , Lijek Rebeccah S. , Unger Katherine P. , Okeke Iruka N. 
Published: July 2011

ABSTRACT

Heat-resistant agglutinin 1 (Hra1) is an accessory colonization factor of enteroaggregative
Escherichia coli
(EAEC) strain 042. Tia, a close homolog of Hra1, is an invasin and adhesin that has been described in enterotoxigenic
E. coli
. We devised a PCR-restriction fragment length polymorphism screen for the associated genes and found that they occur among 55 (36.7%) of the enteroaggregative
E. coli
isolates screened, as well as lower proportions of enterotoxigenic, enteropathogenic, enterohemorrhagic, and commensal
E. coli
isolates. Overall, 25%, 8%, and 3% of 150 EAEC strains harbored
hra1
alone,
tia
alone, or both genes, respectively. One EAEC isolate, 60A, produced an amplicon with a unique restriction profile, distinct from those of
hra1
and
tia
. We cloned and sequenced the full-length agglutinin gene from strain 60A and have designated it
hra2
. The
hra2
gene was not detected in any of 257 diarrheagenic
E. coli
isolates in our collection but is present in the genome of
Salmonella enterica
serovar Heidelberg strain SL476. The cloned
hra2
gene from strain 60A, which encodes a predicted amino acid sequence that is 64% identical to that of Hra1 and 68% identical to that of Tia, was sufficient to confer adherence on
E. coli
K-12. We constructed an
hra2
deletion mutant of EAEC strain 60A. The mutant was deficient in adherence but not autoaggregation or invasion, pointing to a functional distinction from the autoagglutinin Hra1 and the Tia invasin. Hra1, Tia, and the novel accessory adhesin Hra2 are members of a family of integral outer membrane proteins that confer different colonization-associated phenotypes.
Roles of the Redox-Active Disulfide and Histidine Residues Forming a Catalytic Dyad in Reactions Catalyzed by 2-Ketopropyl Coenzyme M Oxidoreductase/Carboxylase
Authors: Kofoed Melissa A. , Wampler David A. , Pandey Arti S. , Peters John W. , Ensign Scott A. 
Published: July 2011

ABSTRACT

NADPH:2-ketopropyl-coenzyme M oxidoreductase/carboxylase (2-KPCC), an atypical member of the disulfide oxidoreductase (DSOR) family of enzymes, catalyzes the reductive cleavage and carboxylation of 2-ketopropyl-coenzyme M [2-(2-ketopropylthio)ethanesulfonate; 2-KPC] to form acetoacetate and coenzyme M (CoM) in the bacterial pathway of propylene metabolism. Structural studies of 2-KPCC from
Xanthobacter autotrophicus
strain Py2 have revealed a distinctive active-site architecture that includes a putative catalytic triad consisting of two histidine residues that are hydrogen bonded to an ordered water molecule proposed to stabilize enolacetone formed from dithiol-mediated 2-KPC thioether bond cleavage. Site-directed mutants of 2-KPCC were constructed to test the tenets of the mechanism proposed from studies of the native enzyme. Mutagenesis of the interchange thiol of 2-KPCC (C82A) abolished all redox-dependent reactions of 2-KPCC (2-KPC carboxylation or protonation). The air-oxidized C82A mutant, as well as wild-type 2-KPCC, exhibited the characteristic charge transfer absorbance seen in site-directed variants of other DSOR enzymes but with a pK
a
value for C87 (8.8) four units higher (i.e., four orders of magnitude less acidic) than that for the flavin thiol of canonical DSOR enzymes. The same higher pK
a
value was observed in native 2-KPCC when the interchange thiol was alkylated by the CoM analog 2-bromoethanesulfonate. Mutagenesis of the flavin thiol (C87A) also resulted in an inactive enzyme for steady-state redox-dependent reactions, but this variant catalyzed a single-turnover reaction producing a 0.8:1 ratio of product to enzyme. Mutagenesis of the histidine proximal to the ordered water (H137A) led to nearly complete loss of redox-dependent 2-KPCC reactions, while mutagenesis of the distal histidine (H84A) reduced these activities by 58 to 76%. A redox-independent reaction of 2-KPCC (acetoacetate decarboxylation) was not decreased for any of the aforementioned site-directed mutants. We interpreted and rationalized these results in terms of a mechanism of catalysis for 2-KPCC employing a unique hydrophobic active-site architecture promoting thioether bond cleavage and enolacetone formation not seen for other DSOR enzymes.