bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evaluation of Two DNA Template Preparation Methods for Post-Immunomagnetic Separation Detection of
Cryptosporidium parvum
in Foods and Beverages by PCR
Authors: Frazar Christian D. , Orlandi Palmer A. 
Published: September 2007

ABSTRACT

Cryptosporidium parvum
oocysts were recovered by immunomagnetic separation from six artificially contaminated foods. Two DNA isolation methods were subsequently evaluated by PCR. The FTA Concentrator-PS filter provided rapid and reproducible detection, although variability increased at lower inoculum levels (88% and 15% detection in high- and low-inoculum-level samples, respectively). Total DNA extraction generated consistent results at all oocyst levels but resulted in longer analysis time (100% and 59% detection in high- and low-inoculum-level samples, respectively). Also reflected in this study was that the matrix played an important role in the ability to recover oocysts, as sample turbidity, pH, and PCR inhibitors all influenced detection.
Cyclic AMP-Protein Kinase A and Snf1 Signaling Mechanisms Underlie the Superior Potency of Sucrose for Induction of Filamentation in
Saccharomyces cerevisiae
Authors: Van de Velde Sam , Thevelein Johan M. 
Published: September 2007

ABSTRACT

Under specific environmental conditions, the yeast
Saccharomyces cerevisiae
can undergo a morphological switch to a pseudohyphal growth pattern. Pseudohyphal differentiation is generally studied upon induction by nitrogen limitation in the presence of glucose. It is known to be controlled by several signaling pathways, including mitogen-activated protein kinase, cyclic AMP-protein kinase A (cAMP-PKA), and Snf1 kinase pathways. We show that the alpha-glucoside sugars maltose and maltotriose, and especially sucrose, are more potent inducers of filamentation than glucose. Sucrose even induces filamentation in nitrogen-rich media and in the
mep2
Δ
/mep2
Δ ammonium permease mutant on ammonium-limiting medium. We demonstrate that glucose also inhibits filamentation by means of a pathway parallel to the cAMP-PKA pathway. Deletion of
HXK2
shifted the pseudohyphal growth pattern on glucose to that of sucrose, while deletion of
SNF4
abrogated filamentation on both sugars, indicating a negative role of glucose repression and a positive role for Snf1 activity in the control of filamentation. In all strains and in all media, sucrose induction of filamentation is greatly diminished by deletion of the sucrose/glucose-sensing G-protein-coupled receptor Gpr1, whereas it has no effect on induction by maltose and maltotriose. The competence of alpha-glucoside sugars to induce filamentation is reflected in the increased expression of the cell surface flocculin gene
FLO11
. In addition, sucrose is the only alpha-glucoside sugar capable of rapidly inducing
FLO11
expression in a Gpr1-dependent manner, reflecting the sensitivity of Gpr1 for this sugar and its involvement in rapid sucrose signaling. Our study identifies sucrose as the most potent nutrient inducer of pseudohyphal growth and shows that glucose inactivation of Snf1 kinase signaling is responsible for the lower potency of glucose.
Compromised Outer Membrane Integrity in
Vibrio cholerae
Type II Secretion Mutants
Authors: Sikora Aleksandra E. , Lybarger Suzanne R. , Sandkvist Maria 
Published: September 2007

ABSTRACT

The type II secretion (T2S) system of
Vibrio cholerae
is a multiprotein complex that spans the cell envelope and secretes proteins important for pathogenesis as well as survival in different environments. Here we report that, in addition to the loss of extracellular secretion, removal or inhibition of expression of the T2S genes,
epsC-N
, results in growth defects and a broad range of alterations in the outer membrane that interfere with its barrier function. Specifically, the sensitivity to membrane-perturbing agents such as bile salts and the antimicrobial peptide polymyxin B is increased, and periplasmic constituents leak out into the culture medium. As a consequence, the σ
E
stress response is induced. Furthermore, due to the defects caused by inactivation of the T2S system, the Δ
eps
deletion mutant of
V. cholerae
strain N16961 is incapable of surviving the passage through the infant mouse gastrointestinal tract. The growth defect and leaky outer membrane phenotypes are suppressed when the culture medium is supplemented with 5% glucose or sucrose, although the
eps
mutants remain sensitive to membrane-damaging agents. This suggests that the sugars do not restore the integrity of the outer membrane in the
eps
mutant strains per se but may provide osmoprotective functions.
Chemotactic Control of the Two Flagellar Systems of
Rhodobacter sphaeroides
Is Mediated by Different Sets of CheY and FliM Proteins
Authors: del Campo Ana Martínez , Ballado Teresa , de la Mora Javier , Poggio Sebastian , Camarena Laura , Dreyfus Georges 
Published: September 2007

ABSTRACT

Rhodobacter sphaeroides
expresses two different flagellar systems, a subpolar flagellum (fla1) and multiple polar flagella (fla2). These structures are encoded by different sets of flagellar genes. The chemotactic control of the subpolar flagellum (fla1) is mediated by three of the six different CheY proteins (CheY6, CheY4, or CheY3). We show evidence that CheY1, CheY2, and CheY5 control the chemotactic behavior mediated by fla2 flagella and that RSP6099 encodes the fla2 FliM protein.
Probing the Antiprotease Activity of λCIII, an Inhibitor of theEscherichia coliMetalloprotease HflB (FtsH)
Authors: Halder Sabyasachi , Datta Ajit Bikram , Parrack Pradeep 
Published: September 2007

ABSTRACTThe CIII protein encoded by the temperate coliphage lambda acts as an inhibitor of the ubiquitousEscherichia colimetalloprotease HflB (FtsH). This inhibition results in the stabilization of transcription factor λCII, thereby helping the phage to lysogenize the host bacterium. λCIII, a small (54-residue) protein of unknown structure, also protects σ32, another specific substrate of HflB. In order to understand the details of the inhibitory mechanism of CIII, we cloned and expressed the protein with an N-terminal six-histidine tag. We also synthesized and studied a 28-amino-acid peptide, CIIIC, encompassing the central 14 to 41 residues of CIII that exhibited antiproteolytic activity. Our studies show that CIII exists as a dimer under native conditions, aided by an intersubunit disulfide bond, which is dispensable for dimerization. Unlike CIII, CIIIC resists digestion by HflB. While CIII binds to HflB, it does not bind to CII. On the basis of these results, we discuss various mechanisms for the antiproteolytic activity of CIII.
l-Rhamnose Transport Is Sugar Kinase (RhaK) Dependent inRhizobium leguminosarumbv. trifolii
Authors: Richardson Jason S. , Oresnik Ivan J. 
Published: September 2007

ABSTRACTStrains ofRhizobium leguminosarumwhich are unable to catabolizel-rhamnose, a methyl-pentose sugar, are compromised in the ability to compete for nodule occupancy versus wild-type strains. Previous characterization of the 11-kb region necessary for the utilization of rhamnose identified a locus carrying catabolic genes and genes encoding the components of an ABC transporter. Genetic evidence suggested that the putative kinase RhaK carried out the first step in the catabolism of rhamnose. Characterization of this kinase led to the observation that strains carrying rhamnose kinase mutations were unable to transport rhamnose into the cell. The absence of a functional rhamnose kinase did not stop the transcription and translation of the ABC transporter components. By developing an in vitro assay for RhaK activity, we have been able to show that (i) RhaK activity is consistent with RhaK phosphorylating rhamnose and (ii) biochemical activity of RhaK is necessary for rhamnose transport.
A Wall of Funnels Concentrates Swimming Bacteria
Authors: Galajda Peter , Keymer Juan , Chaikin Paul , Austin Robert 
Published: September 2007

ABSTRACT

Randomly moving but self-propelled agents, such as
Escherichia coli
bacteria, are expected to fill a volume homogeneously. However, we show that when a population of bacteria is exposed to a microfabricated wall of funnel-shaped openings, the random motion of bacteria through the openings is rectified by tracking (trapping) of the swimming bacteria along the funnel wall. This leads to a buildup of the concentration of swimming cells on the narrow opening side of the funnel wall but no concentration of nonswimming cells. Similarly, we show that a series of such funnel walls functions as a multistage pump and can increase the concentration of motile bacteria exponentially with the number of walls. The funnel wall can be arranged along arbitrary shapes and cause the bacteria to form well-defined patterns. The funnel effect may also have implications on the transport and distribution of motile microorganisms in irregular confined environments, such as porous media, wet soil, or biological tissue, or act as a selection pressure in evolution experiments.
Amino Acid Toxicities of
Escherichia coli
That Are Prevented by Leucyl-tRNA Synthetase Amino Acid Editing
Authors: Karkhanis Vrajesh A. , Mascarenhas Anjali P. , Martinis Susan A. 
Published: September 2007

ABSTRACT

Leucyl-tRNA synthetase (LeuRS) has evolved an editing function to clear misactivated amino acids. An
Escherichia coli
-based assay was established to identify amino acids that compromise the fidelity of LeuRS and translation. Multiple nonstandard as well as standard amino acids were toxic to the cell when LeuRS editing was inactivated.
How Moist Heat Kills Spores of
Bacillus subtilis
Authors: Coleman William H. , Chen De , Li Yong-qing , Cowan Ann E. , Setlow Peter 
Published: September 2007

ABSTRACT

Populations of
Bacillus subtilis
spores in which 90 to 99.9% of the spores had been killed by moist heat gave only two fractions on equilibrium density gradient centrifugation: a fraction comprised of less dense spores that had lost their dipicolinic acid (DPA), undergone significant protein denaturation, and were all dead and a fraction with the same higher density as that of unheated spores. The latter fraction from heat-killed spore populations retained all of its DPA, but ≥98% of the spores could be dead. The dead spores that retained DPA germinated relatively normally with nutrient and nonnutrient germinants, but the outgrowth of these germinated spores was significantly compromised, perhaps because they had suffered damage to some proteins such that metabolic activity during outgrowth was greatly decreased. These results indicate that DPA release takes place well after spore killing by moist heat and that DPA release during moist-heat treatment is an all-or-nothing phenomenon; these findings also suggest that damage to one or more key spore proteins causes spore killing by moist heat.
Dissection of 16S rRNA Methyltransferase (KsgA) Function in
Escherichia coli
Authors: Inoue Koichi , Basu Soumit , Inouye Masayori 
Published: September 2007

ABSTRACT

A 16S rRNA methyltransferase, KsgA, identified originally in
Escherichia coli
is highly conserved in all living cells, from bacteria to humans. KsgA orthologs in eukaryotes possess functions in addition to their rRNA methyltransferase activity.
E. coli
Era is an essential GTP-binding protein. We recently observed that KsgA functions as a multicopy suppressor for the cold-sensitive cell growth of an
era
mutant [Era(E200K)] strain (Q. Lu and M. Inouye, J. Bacteriol.
180
:5243-5246, 1998). Here we observed that although KsgA(E43A), KsgA(G47A), and KsgA(E66A) mutations located in the
S
-adenosylmethionine-binding motifs severely reduced its methyltransferase activity, these mutations retained the ability to suppress the growth defect of the Era(E200K) strain at a low temperature. On the other hand, a KsgA(R248A) mutation at the C-terminal domain that does not affect the methyltransferase activity failed to suppress the growth defect. Surprisingly,
E. coli
cells overexpressing wild-type KsgA, but not KsgA(R248A), were found to be highly sensitive to acetate even at neutral pH. Such growth inhibition also was observed in the presence of other weak organic acids, such as propionate and benzoate. These chemicals are known to be highly toxic at acidic pH by lowering the intracellular pH. We found that KsgA-induced cells had increased sensitivity to extreme acid conditions (pH 3.0) compared to that of noninduced cells. These results suggest that
E. coli
KsgA, in addition to its methyltransferase activity, has another unidentified function that plays a role in the suppression of the cold-sensitive phenotype of the Era(E200K) strain and that the additional function may be involved in the acid shock response. We discuss a possible mechanism of the KsgA-induced acid-sensitive phenotype.
One Perturbation of the Mother Cell Gene Regulatory Network Suppresses the Effects of Another during Sporulation of
Bacillus subtilis
Authors: Wang Lijuan , Perpich John , Driks Adam , Kroos Lee 
Published: September 2007

ABSTRACT

In the mother cell of sporulating
Bacillus subtilis
, a regulatory network functions to control gene expression. Four transcription factors act sequentially in the order σ
E
, SpoIIID, σ
K
, GerE. σ
E
and σ
K
direct RNA polymerase to transcribe different regulons. SpoIIID and GerE are DNA-binding proteins that activate or repress transcription of many genes. Several negative regulatory loops add complexity to the network. First, transcriptionally active σ
K
RNA polymerase inhibits early sporulation gene expression, resulting in reduced accumulation of σ
E
and SpoIIID late during sporulation. Second, GerE represses
sigK
transcription, reducing σ
K
accumulation about twofold. Third, SpoIIID represses
cotC
, which encodes a spore coat protein, delaying its transcription by σ
K
RNA polymerase. Partially circumventing the first feedback loop, by engineering cells to maintain the SpoIIID level late during sporulation, causes spore defects. Here, the effects of circumventing the second feedback loop, by mutating the GerE binding sites in the
sigK
promoter region, are reported. Accumulation of pro-σ
K
and σ
K
was increased, but no spore defects were detected. Expression of σ
K
-dependent reporter fusions was altered, increasing the expression of
gerE-lacZ
and
cotC-lacZ
and decreasing the expression of
cotD-lacZ
. Because these effects on gene expression were opposite those observed when the SpoIIID level was maintained late during sporulation, cells were engineered to both maintain the SpoIIID level and have elevated
sigK
expression late during sporulation. This restored the expression of σ
K
-dependent reporters to wild-type levels, and no spore defects were observed. Hence, circumventing the second feedback loop suppressed the effects of perturbing the first feedback loop. By feeding information back into the network, these two loops appear to optimize target gene expression and increase network robustness. Circumventing the third regulatory loop, by engineering cells to express
cotC
about 2 h earlier than normal, did not cause a detectable spore defect.
Functional Characterization of Bacterial Oligosaccharyltransferases Involved in O-Linked Protein Glycosylation
Authors: Faridmoayer Amirreza , Fentabil Messele A. , Mills Dominic C. , Klassen John S. , Feldman Mario F. 
Published: September 2007

ABSTRACTProtein glycosylation is an important posttranslational modification that occurs in all domains of life. Pilins, the structural components of type IV pili, are O glycosylated inNeisseria meningitidis,Neisseria gonorrhoeae, and some strains ofPseudomonas aeruginosa. In this work, we characterized theP. aeruginosa1244 andN. meningitidisMC58 O glycosylation systems inEscherichia coli. In both cases, sugars are transferred en bloc by an oligosaccharyltransferase (OTase) named PglL inN. meningitidisand PilO inP. aeruginosa. We show that, like PilO, PglL has relaxed glycan specificity. Both OTases are sufficient for glycosylation, but they require translocation of the undecaprenol-pyrophosphate-linked oligosaccharide substrates into the periplasm for activity. Whereas PilO activity is restricted to short oligosaccharides, PglL is able to transfer diverse oligo- and polysaccharides. This functional characterization supports the concept that despite their low sequence similarity, PilO and PglL belong to a new family of “O-OTases” that transfer oligosaccharides from lipid carriers to hydroxylated amino acids in proteins. To date, such activity has not been identified for eukaryotes. To our knowledge, this is the first report describing recombinant O glycoproteins synthesized inE. coli.
Local and Global Regulators Linking Anaerobiosis tocupAFimbrial Gene Expression inPseudomonas aeruginosa
Authors: Vallet-Gely Isabelle , Sharp Josh S. , Dove Simon L. 
Published: September 2007

ABSTRACTThecupAgene cluster ofPseudomonas aeruginosaencodes components and assembly factors of a putative fimbrial structure that enable this opportunistic pathogen to form biofilms on abiotic surfaces. InP. aeruginosathe control ofcupAgene expression is complex, with the H-NS-like MvaT protein functioning to repress phase-variable (on/off) expression of the operon. Here we identify four positive regulators ofcupAgene expression, including three unusual regulators encoded by thecgrABCgenes and Anr, a global regulator of anaerobic gene expression. We show that thecupAgenes are expressed in a phase-variable manner under anaerobic conditions and that thecgrgenes are essential for this expression. We show further thatcgrgene expression is negatively controlled by MvaT and positively controlled by Anr and anaerobiosis. Expression of thecupAgenes therefore appears to involve a regulatory cascade in which anaerobiosis, signaled through Anr, stimulates expression of thecgrgenes, resulting in a concomitant increase incupAgene expression. Our findings thus provide mechanistic insight into the regulation ofcupAgene expression and identify anaerobiosis as an inducer of phase-variablecupAgene expression, raising the possibility that phase-variable expression of fimbrial genes important for biofilm formation may occur inP. aeruginosapersisting in the largely anaerobic environment of the cystic fibrosis host lung.
Flavin-Dependent Thymidylate Synthase ThyX Activity: Implications for the Folate Cycle in Bacteria
Authors: Leduc Damien , Escartin Frédéric , Nijhout H. Frederik , Reed Michael C. , Liebl Ursula , Skouloubris Stéphane , Myllykallio Hannu 
Published: September 2007

ABSTRACT

Although flavin-dependent ThyX proteins show thymidylate synthase activity in vitro and functionally complement
thyA
defects in heterologous systems, direct proof of their cellular functions is missing. Using insertional mutagenesis of
Rhodobacter capsulatus thyX
, we constructed the first defined
thyX
inactivation mutant. Phenotypic analyses of the obtained mutant strain confirmed that
R. capsulatus
ThyX is required for de novo thymidylate synthesis. Full complementation of the
R. capsulatus thyX
::
spec
strain to thymidine prototrophy required not only the canonical thymidylate synthase ThyA but also the dihydrofolate reductase FolA. Strikingly, we also found that addition of exogenous methylenetetrahydrofolate transiently inhibited the growth of the different
Rhodobacter
strains used in this work. To rationalize these experimental results, we used a mathematical model of bacterial folate metabolism. This model suggests that a very low dihydrofolate reductase activity is enough to rescue significant thymidylate synthesis in the presence of ThyX proteins and is in agreement with the notion that intracellular accumulation of folates results in growth inhibition. In addition, our observations suggest that the presence of flavin-dependent thymidylate synthase X provides growth benefits under conditions in which the level of reduced folate derivatives is compromised.
Poisonous Effects of Asteroid Impacts or Nuclear Explosions in the Western Regions of the Black Sea
Authors: Badescu Viorel 
Published: September 2007

Abstract
The impact of an asteroid or a very large nuclear explosion in the Black Sea may cause a poisonous hydrogen sulfide gas release into the atmosphere. Some effects of this phenomenon on the western Black Sea coasts are evaluated in this paper. Two simple models are proposed to describe the generation of the H2S cloud. The wind speed plays an important role in H2S cloud dynamics. The land surface area covered by the H2S cloud generated by a 1000-m-sized asteroid during the run-in ranges between about 4480 and 9600 km2. This may affect up to 230 000 people. In the case of a 70-m-sized asteroid, the cloud covers between 70 and 210 km2 of land. This may affect between about 1600 and 5100 people. The evaluations do not include the population of the towns on the seashore and may be a few times underestimated for some particular wind directions. These effects are briefly compared with more usual effects associated with asteroids impacting the sea, such as shock waves and tsunamis. Nuclear explosions of 1 and 50 Mton TNT may be assimilated to the impact by asteroids of about 33- and 120-m diameter, respectively.
The Turbulent Oscillator: A Mechanism of Low-Frequency Variability of the Wind-Driven Ocean Gyres
Authors: Berloff P. , Hogg A. Mc C. , Dewar W. 
Published: September 2007

Abstract
Intrinsic low-frequency variability is studied in the idealized, quasigeostrophic, midlatitude, wind-driven ocean gyres operating at large Reynolds number. A robust decadal variability mode driven by the transient mesoscale eddies is found and analyzed. The variability is a turbulent phenomenon, which is driven by the competition between the eddy rectification process and the potential vorticity anomalies induced by changes of the intergyre transport.
Three-Dimensional Tidal Flow in an Elongated, Rotating Basin
Authors: Winant Clinton D. 
Published: September 2007

Abstract
The three-dimensional tidal circulation in an elongated basin of arbitrary depth is described with a linear, constant-density model on the f plane. Rotation fundamentally alters the lateral flow, introducing a lateral recirculation comparable in magnitude to the axial flow, as long as friction is not too large. This circulation is due to the imbalance between the cross-channel sea level gradient, which is in near-geostrophic balance with the Coriolis acceleration associated with the vertically averaged axial flow, and the Coriolis acceleration associated with the vertically sheared axial flow. During flood condition, for example, the lateral Coriolis acceleration near the surface exceeds the pressure gradient, tending to accelerate the lateral flow, while the converse is true near the bottom. As a result, with rotation, fluid parcels tend to corkscrew into and out of the basin in a tidal period. The axial flow is only weakly modified by rotation. When friction is small, the axial velocity is uniform in each section, except in a narrow bottom boundary layer where it decreases to zero. The boundary layer thickness increases with friction, so that with moderate or large friction, axial velocities are sheared from bottom to surface. When friction is large, the local and Coriolis accelerations are both small and the dynamics are governed by a balance between friction and the pressure gradient.