bluuebunny/crossref_metadata_2025_split huggingface dataset:
Induction of Persistent Double Strand Breaks Following Multiphoton Irradiation of Cycling and G1‐arrested Mammalian Cells—Replication‐induced Double Strand Breaks
Authors: Harper Jane V. , Reynolds Pamela , Leatherbarrow Emma L. , Botchway Stanley W. , Parker Anthony W. , O’Neill Peter 
Published: June 2008

AbstractDNA double strand breaks (DSBs) are amongst the most deleterious lesions induced within the cell following exposure to ionizing radiation. Mammalian cells repair these breaks predominantly via the nonhomologous end joining pathway which is active throughout the cell cycle and is error prone. The alternative pathway for repair of DSBs is homologous recombination (HR) which is error free and active during S‐ and G2/M‐phases of the cell cycle. We have utilized near‐infrared laser radiation to induce DNA damage in individual mammalian cells through multiphoton excitation processes to investigate the dynamics of single cell DNA damage processing. We have used immunofluorescent imaging of γ‐H2AX (a marker for DSBs) in mammalian cells and investigated the colocalization of this protein with ATM, p53 binding protein 1 and RAD51, an integral protein of the HR DNA repair pathway. We have observed persistent DSBs at later times postlaser irradiation which are indicative of DSBs arising at replication, presumably from UV photoproducts or clustered damage containing single strand breaks. Cell cycle studies have shown that in G1 cells, a significant fraction of multiphoton laser‐induced prompt DSBs persists for >4 h in addition to those induced at replication.
Monitoring ALA‐induced PpIX Photodynamic Therapy in the Rat Esophagus Using Fluorescence and Reflectance Spectroscopy
Authors: Kruijt Bastiaan , De Bruijn Henriette S. , Van Der Ploeg‐van den Heuvel Angelique , De Bruin Ron W. F. , Sterenborg Henricus J. C. M. , Amelink Arjen , Robinson Dominic J. 
Published: June 2008

AbstractThe presence of phased protoporphyrin IX (PpIX) bleach kinetics has been shown to correlate with esophageal response to 5‐aminolevulinic acid‐based photodynamic therapy (ALA‐PDT) in animal models. Here we confirm the existence of phased PpIX photobleaching by increasing the temporal resolution of the fluorescence measurements using the therapeutic illumination and long wavelength fluorescence detection. Furthermore fluorescence differential pathlength spectroscopy (FDPS) was incorporated to provide information on the effects of PpIX and tissue oxygenation distribution on the PpIX bleach kinetics during illumination. ALA at a dose of 200 mg kg−1 was orally administered to 15 rats, five rats served as control animals. PDT was performed at an in situ measured fluence rate of 75 mW cm−2 using a total fluence of 54 J cm−2. Forty‐eight hours after PDT the esophagus was excised and histologically examined for PDT‐induced damage. Fluence rate and PpIX photobleaching at 705 nm were monitored during therapeutic illumination with the same isotropic probe. A new method, FDPS, was used for superficial measurement on saturation, blood volume, scattering characteristics and PpIX fluorescence. Results showed two‐phased PpIX photobleaching that was not related to a (systematic) change in esophageal oxygenation but was associated with an increase in average blood volume. PpIX fluorescence photobleaching measured using FDPS, in which fluorescence signals are only acquired from the superficial layers of the esophagus, showed lower rates of photobleaching and no distinct phases. No clear correlation between two‐phased photobleaching and histologic tissue response was found. This study demonstrates the feasibility of measuring fluence rate, PpIX fluorescence and FDPS during PDT in the esophagus. We conclude that the spatial distribution of PpIX significantly influences the kinetics of photobleaching and that there is a complex interrelationship between the distribution of PpIX and the supply of oxygen to the illuminated tissue volume.
Evolutionary origins of invasive populations
Authors: Lee Carol Eunmi , Gelembiuk Gregory William 
Published: June 2008

AbstractWhat factors shape the evolution of invasive populations? Recent theoretical and empirical studies suggest that an evolutionary history of disturbance might be an important factor. This perspective presents hypotheses regarding the impact of disturbance on the evolution of invasive populations, based on a synthesis of the existing literature. Disturbance might select for life‐history traits that are favorable for colonizing novel habitats, such as rapid population growth and persistence. Theoretical results suggest that disturbance in the form of fluctuating environments might select for organismal flexibility, or alternatively, the evolution of evolvability. Rapidly fluctuating environments might favor organismal flexibility, such as broad tolerance or plasticity. Alternatively, longer fluctuations or environmental stress might lead to the evolution of evolvability by acting on features of the mutation matrix. Once genetic variance is generated via mutations, temporally fluctuating selection across generations might promote the accumulation and maintenance of genetic variation. Deeper insights into how disturbance in native habitats affects evolutionary and physiological responses of populations would give us greater capacity to predict the populations that are most likely to tolerate or adapt to novel environments during habitat invasions. Moreover, we would gain fundamental insights into the evolutionary origins of invasive populations.
Environmental and Growth Phase Regulation of the
Streptococcus gordonii
Arginine Deiminase Genes
Authors: Liu Yaling , Dong Yiqian , Chen Yi-Ywan M. , Burne Robert A. 
Published: June 2008

ABSTRACT

A 1,026-bp open reading frame sharing significant similarity with
queA
, which encodes a predicted
S
-adenosylmethionine:tRNA ribosyltransferase-isomerase responsible for queosine modification of tRNAs, was found immediately 5′ of the gene for the transcriptional activator (ArcR) of the arginine deiminase system (ADS) operon of
Streptococcus gordonii
. The role of QueA in bacterial physiology is enigmatic, but loss of QueA has been shown to compromise stationary-phase survival or virulence in certain enteric bacteria. Interestingly,
S. gordonii
appears to be unique among ADS-positive bacteria in the linkage of
queA
with the ADS genes. A putative σ
70
promoter (p

queA

; TTGCCA-N
21
-TATAAT) was mapped 5′ of
queA
by primer extension, and
queA
and
arcR
were shown to be cotranscribed. The expression from p

queA

was found to be constitutive under all conditions tested, but the expression of p

arcA

, which drives the expression of the
arc
structural genes, was enhanced in stationary phase and could be induced by low pH and arginine. QueA and CcpA acted repressively on
arc
transcription, but neither QueA-deficient strains nor CcpA-deficient strains showed significant differences in arginine deiminase enzyme activities compared with the wild-type strain. The growth rate of a QueA-deficient strain did not differ significantly from that of the parental strain, but the QueA-deficient strain did not compete well with the wild-type during serial passage. In addition to the finding that ADS expression can be regulated separately by growth phase and pH, a significant linkage between the ADS, translational efficiency modulated by QueA, and post-exponential-phase survival of
S. gordonii
was found.
Previously Unrecognized Vaccine Candidates ControlTrypanosoma cruziInfection and Immunopathology in Mice
Authors: Bhatia Vandanajay , Garg Nisha Jain 
Published: June 2008

ABSTRACTTrypanosoma cruziis the etiologic agent of Chagas' disease, a major health problem in Latin America and an emerging infectious disease in the United States. Previously, we screened aT. cruzisequence database by a computational-bioinformatic approach and identified antigens that exhibited the characteristics of good vaccine candidates. In this study, we tested the vaccine efficacy of three of the putative candidate antigens againstT. cruziinfection and disease in a mouse model. C57BL/6 mice vaccinated withT. cruzi G1(TcG1)-, TcG2-, or TcG4-encoding plasmids and cytokine (interleukin-12 and granulocyte-macrophage colony-stimulating factor) expression plasmids elicited a strong Th1-type antibody response dominated by immunoglobulin G2b (IgG2b)/IgG1 isotypes. The dominant IgG2b/IgG1 antibody response was maintained after a challenge infection and was associated with 50 to 90% control of the acute-phase tissue parasite burden and an almost undetectable level of tissue parasites during the chronic phase, as determined by a sensitiveT. cruzi18S rRNA gene-specific real-time PCR approach. Splenocytes from vaccinated-and-infected mice, compared to unvaccinated-and-infected mice, exhibited decreased (∼50% lower) proliferation and gamma interferon (IFN-γ) production when stimulated in vitro withT. cruziantigens, thus suggesting that protection from challenge infection was not provided by an active T-cell response. Subsequently, the serum and cardiac levels of IFN-γ and tumor necrosis factor alpha and infiltration of inflammatory infiltrate in the heart were decreased in vaccinated mice during the course of infection and chronic disease development. Taken together, these results demonstrate the identification of novel vaccine candidates that provided protection fromT. cruzi-induced immunopathology in experimental mice.
Development of a Fluorescent-Bead-Based Multiplex Immunoassay To Determine Immunoglobulin G Subclass Responses to
Neisseria meningitidis
Serogroup A and C Polysaccharides
Authors: de Voer Richarda M. , van der Klis Fiona R. M. , Engels Carla W. A. M. , Rijkers Ger T. , Sanders Elisabeth A. , Berbers Guy A. M. 
Published: June 2008

ABSTRACT

A fluorescent-particle-based multiplex flow cytometric immunoassay (MIA) for the detection of serum immunoglobulin G (IgG) and two IgG subclasses, IgG1 and IgG2, specific for
Neisseria meningitidis
serogroup A (MenA) and C (MenC) polysaccharides (PS) was developed. The assay comprised three separate duplex assays, one for the detection of the IgG response to MenA and MenC PS, another for the detection of the IgG1 response to MenA and MenC PS, and a third for the detection of the IgG2 response to MenA and MenC PS. Next, the three separate duplex assays were combined and analyzed as a hexaplex assay. No interference between monoplex, duplex, and hexaplex assays was observed, and the assay was found to have low intra- and interassay variation (<9.0% and <27%, respectively). Comparison of the meningococcal subclass MIA to the in-house enzyme-linked inmmunosorbent assays showed a good correlation (
R
≥ 0.85) for each of the subclasses. We conclude that the hexaplex meningococcal subclass MIA is an easy and specific assay for the determination of anti-MenA and anti-MenC PS subclass IgG, requiring minimal amounts of serum to study IgG subclass responses to vaccines.
Mitogen-Activated Protein Kinase Hog1 Mediates Adaptation to G1Checkpoint Arrest during Arsenite and Hyperosmotic Stress
Authors: Migdal Iwona , Ilina Yulia , Tamás Markus J. , Wysocki Robert 
Published: June 2008

ABSTRACTCells slow down cell cycle progression in order to adapt to unfavorable stress conditions. Yeast (Saccharomyces cerevisiae) responds to osmotic stress by triggering G1and G2checkpoint delays that are dependent on the mitogen-activated protein kinase (MAPK) Hog1. The high-osmolarity glycerol (HOG) pathway is also activated by arsenite, and thehog1Δ mutant is highly sensitive to arsenite, partly due to increased arsenite influx intohog1Δ cells. Yeast cell cycle regulation in response to arsenite and the role of Hog1 in this process have not yet been analyzed. Here, we found that long-term exposure to arsenite led to transient G1and G2delays in wild-type cells, whereas cells that lack theHOG1gene or are defective in Hog1 kinase activity displayed persistent G1cell cycle arrest. Elevated levels of intracellular arsenite and “cross talk” between the HOG and pheromone response pathways, observed in arsenite-treatedhog1Δ cells, prolonged the G1delay but did not cause a persistent G1arrest. In contrast, deletion of theSIC1gene encoding a cyclin-dependent kinase inhibitor fully suppressed the observed block of G1exit inhog1Δ cells. Moreover, the Sic1 protein was stabilized in arsenite-treatedhog1Δ cells. Interestingly, Sic1-dependent persistent G1arrest was also observed inhog1Δ cells during hyperosmotic stress. Taken together, our data point to an important role of the Hog1 kinase in adaptation to stress-induced G1cell cycle arrest.
Expression, Purification, and Structural Characterization of CfrA, a Putative Iron Transporter from
Campylobacter jejuni
Authors: Carswell Casey L. , Rigden Marc D. , Baenziger John E. 
Published: June 2008

ABSTRACT

The gene for the
Campylobacter
ferric receptor (CfrA), a putative iron-siderophore transporter in the enteric food-borne pathogen
Campylobacter jejuni
, was cloned, and the membrane protein was expressed in
Escherichia coli
, affinity purified, and then reconstituted into model lipid membranes. Fourier transform infrared spectra recorded from the membrane-reconstituted CfrA are similar to spectra that have been recorded from other iron-siderophore transporters and are highly characteristic of a β-sheet protein (∼44% β-sheet and ∼10% α-helix). CfrA undergoes relatively extensive peptide hydrogen-deuterium exchange upon exposure to
2
H
2
O and yet is resistant to thermal denaturation at temperatures up to 95°C. The secondary structure, relatively high aqueous solvent exposure, and high thermal stability are all consistent with a transmembrane β-barrel structure containing a plug domain. Sequence alignments indicate that CfrA contains many of the structural motifs conserved in other iron-siderophore transporters, including the Ton box, PGV, IRG, RP, and LIDG motifs of the plug domain. Surprisingly, a homology model reveals that regions of CfrA that are expected to play a role in enterobactin binding exhibit sequences that differ substantially from the sequences of the corresponding regions that play an essential role in binding/transport by the
E. coli
enterobactin transporter, FepA. The sequence variations suggest that there are differences in the mechanisms used by CfrA and FepA to interact with bacterial siderophores. It may be possible to exploit these structural differences to develop CfrA-specific therapeutics.
Genetic Analysis of Spirochete Flagellin Proteins and Their Involvement in Motility, Filament Assembly, and Flagellar Morphology
Authors: Li Chunhao , Wolgemuth Charles W. , Marko Michael , Morgan David G. , Charon Nyles W. 
Published: June 2008

ABSTRACT

The filaments of spirochete periplasmic flagella (PFs) have a unique structure and protein composition. In most spirochetes, the PFs consist of a core of at least three related proteins (FlaB1, FlaB2, and FlaB3) and a sheath of FlaA protein. The functions of these filament proteins remain unknown. In this study, we used a multidisciplinary approach to examine the role of these proteins in determining the composition, shape, and stiffness of the PFs and how these proteins impact motility by using the spirochete
Brachyspira
(formerly
Treponema
,
Serpulina
)
hyodysenteriae
as a genetic model. A series of double mutants lacking combinations of these PF proteins was constructed and analyzed. The results show the following. First, the diameters of PFs are primarily determined by the sheath protein FlaA, and that FlaA can form a sheath in the absence of an intact PF core. Although the sheath is important to the PF structure and motility, it is not essential. Second, the three core proteins play unequal roles in determining PF structure and swimming speed. The functions of the core proteins FlaB1 and FlaB2 overlap such that either one of these proteins is essential for the spirochete to maintain the intact PF structure and for cell motility. Finally, linear elasticity theory indicates that flagellar stiffness directly affects the spirochete's swimming speed.
Duplication of Teichoic Acid Biosynthetic Genes in
Staphylococcus aureus
Leads to Functionally Redundant Poly(Ribitol Phosphate) Polymerases
Authors: Pereira Mark P. , D'Elia Michael A. , Troczynska Justyna , Brown Eric D. 
Published: June 2008

ABSTRACT

Wall teichoic acids are anionic phosphate-rich polymers that are part of the complex meshwork of carbohydrates that make up the gram-positive cell wall. These polymers are essential to the proper rod-shaped morphology of
Bacillus subtilis
and have been shown to be an important virulence determinant in the nosocomial opportunistic pathogen
Staphylococcus aureus
. Together, sequence-based studies, in vitro experiments with biosynthetic proteins, and analyses of the chemical structure of wall teichoic acid have begun to shed considerable light on our understanding of the biogenesis of this polymer. Nevertheless, some paradoxes remain unresolved. One of these involves a putative duplication of genes linked to CDP-ribitol synthesis (
tarI
′
J
′ and
tarIJ
) as well as poly(ribitol phosphate) polymerization (
tarK
and
tarL
) in
S. aureus
. In the work reported here, we performed careful studies of the dispensability of each gene and discovered a functional redundancy in the duplicated gene clusters. We were able to create mutants in either of the putative ribitol phosphate polymerases (encoded by
tarK
and
tarL
) without affecting teichoic acid levels in the
S. aureus
cell wall. Although genes linked to CDP-ribitol synthesis are also duplicated, a null mutant in only one of these (
tarI
′
J
′) could be obtained, while
tarIJ
remained essential. Suppression analysis of the
tarIJ
null mutant indicated that the mechanism of dysfunction in
tarI
′
J
′ is due to poor translation of the TarJ′ enzyme, which catalyzes the rate-limiting step in CDP-ribitol formation. This work provides new insights into understanding the complex synthetic steps of the ribitol phosphate polymer in
S. aureus
and has implications on specifically targeting enzymes involved in polymer biosynthesis for antimicrobial design.
Evaluation of Linear Array Human Papillomavirus Genotyping Using Automatic Optical Imaging Software
Authors: Jeronimo J. , Wentzensen N. , Long R. , Schiffman M. , Dunn S. T. , Allen R. A. , Walker J. L. , Gold M. A. , Zuna R. E. , Sherman M. E. , Wacholder S. , Wang S. S. 
Published: June 2008

ABSTRACT

Variations in biological behavior suggest that each carcinogenic human papillomavirus (HPV) type should be considered individually in etiologic studies. HPV genotyping assays might have clinical applications if they are approved for use by the FDA. A widely used genotyping assay is the Roche Linear Array HPV genotyping test (LA). We used LA to genotype the HPV isolates from cervical specimens from women with the full spectrum of cervical disease: cervical cancer, cervical intraepithelial neoplasia (CIN), and HPV infections. To explore the feasibility and value of the automated reading of the LA results, we custom-designed novel, optical imaging software that provides optical density measurements of LA bands. We compared unmagnified visual examination with the automated measurements. The two measurements were highly associated. By either method, the threshold between a negative and a positive result was fairly sharp, with a clear bimodal distribution. Visually, most positive results were judged to be strong or medium, with fewer equivocal results categorized as weak (9.5% of positive samples), very weak (6.5% of positive samples), or extremely weak (7.7% of positive samples). The automated measurements of the intensities were significantly associated with the strength of the visual categories (
P
< 0.001). At the extremes of the automated signal intensities (≤20 units or ≥120 units), the bands were almost always categorized visually as negative and positive, respectively. In the equivocal zone (20 to 119 units), specimens were more increasingly likely to be judged to be visually positive as the number of other, definite infections on the same strip increased (
P
for trend < 0.001). Multiple, concurrent infections comprise ≥25% of HPV infections; thus, any systematic visual tendency that influences their evaluation when the result is equivocal should be minimized. Therefore, automated reading is probably worth development if easy-to-calibrate hardware and software can be optimized.
Prevalence of Toxic Shock Syndrome Toxin 1 (TSST-1)-Producing Strains of
Staphylococcus aureus
and Antibody to TSST-1 among Healthy Japanese Women
Authors: Parsonnet Jeffrey , Goering Richard V. , Hansmann Melanie A. , Jones Michaelle B. , Ohtagaki Kumiko , Davis Catherine C. , Totsuka Kyoichi 
Published: June 2008

ABSTRACT

Many cases of neonatal toxic shock syndrome (TSS)-like exanthematous disease but few cases of menstrual TSS (mTSS) have been reported in Japan. We determined the prevalence of mucosal colonization with
Staphylococcus aureus
and of positive antibodies to TSS toxin 1 (TSST-1) among 209 healthy Japanese women in Tokyo.
S. aureus
isolates from mucosal sites were characterized with respect to TSST-1 production and resistance genotype. Antibody titers were determined for test subjects and for 133 Japanese and 137 Caucasian control women living in the United States.
S. aureus
was isolated from at least one site in 108 of 209 women (52%) in Tokyo. Of the 159
S. aureus
isolates recovered, 14 (9%) were TSST-1 positive (12 unique strains). Twelve of 209 women (6%) were colonized with a TSST-1-producing strain; two (<1%) had vaginal colonization. Only 2 of 12 unique toxigenic strains (14%) were methicillin resistant. Of the 12 TSST-1-positive strains isolated, 6 (50%) were pulsed-field gel electrophoresis type USA200, multilocus sequence type clonal complex 30. Fewer Japanese women in Tokyo (47%) than Caucasian and Japanese women in the United States (89% and 75%, respectively) had TSST-1 antibodies. The prevalences of colonization with TSST-1-producing
S. aureus
were comparable in Japan and the United States, despite low seropositivity to TSST-1 in Japan. Environmental factors appear to be important in promoting the development of anti-TSST-1 antibodies, as there was a significant difference in titers between Japanese women living in Tokyo and those living in the United States. Most colonizing TSST-1-producing
S. aureus
strains in Japan were genotypically similar to mTSS strains found in the United States.