bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential Rescue of Light- and Food-Entrainable Circadian Rhythms
Authors: Fuller Patrick M. , Lu Jun , Saper Clifford B.
Published: May 2008

When food is plentiful, circadian rhythms of animals are powerfully entrained by the light-dark cycle. However, if animals have access to food only during their normal sleep cycle, they will shift most of their circadian rhythms to match the food availability. We studied the basis for entrainment of circadian rhythms by food and light in mice with targeted disruption of the clock gene
Bmal1
, which lack circadian rhythmicity. Injection of a viral vector containing the
Bmal1
gene into the suprachiasmatic nuclei of the hypothalamus restored light-entrainable, but not food-entrainable, circadian rhythms. In contrast, restoration of the
Bmal1
gene only in the dorsomedial hypothalamic nucleus restored the ability of animals to entrain to food but not to light. These results demonstrate that the dorsomedial hypothalamus contains a
Bmal1
-based oscillator that can drive food entrainment of circadian rhythms.
Microbial Biogeography: From Taxonomy to Traits
Authors: Green Jessica L. , Bohannan Brendan J. M. , Whitaker Rachel J.
Published: May 2008

The biogeographic variation of life has predominantly been studied using taxonomy, but this focus is changing. There is a resurging interest in understanding patterns in the distribution not only of taxa but also of the traits those taxa possess. Patterns of trait variation shed light on fundamental questions in biology, including why organisms live where they do and how they will respond to environmental change. Technological advances such as environmental genomics place microbial ecology in a unique position to move trait-based biogeography forward. We anticipate that as trait-based biogeography continues to evolve, micro- and macroorganisms will be studied in concert, establishing a science that is informed by and relevant to all domains of life.
Microbial Ecology of Ocean Biogeochemistry: A Community Perspective
Authors: Strom Suzanne L.
Published: May 2008

The oceans harbor a tremendous diversity of marine microbes. Different functional groups of bacteria, archaea, and protists arise from this diversity to dominate various habitats and drive globally important biogeochemical cycles. Explanations for the distribution of microbial taxa and their associated activity often focus on resource availability and abiotic conditions. However, the continual reshaping of communities by mortality, allelopathy, symbiosis, and other processes shows that community interactions exert strong selective pressure on marine microbes. Deeper exploration of microbial interactions is now possible via molecular prospecting and taxon-specific experimental approaches. A holistic outlook that encompasses the full array of selective pressures on individuals will help elucidate the maintenance of microbial diversity and the regulation of biogeochemical reactions by planktonic communities.
Detection of Silica-Rich Deposits on Mars
Authors: Squyres S. W. , Arvidson R. E. , Ruff S. , Gellert R. , Morris R. V. , Ming D. W. , Crumpler L. , Farmer J. D. , Marais D. J. Des , Yen A. , McLennan S. M. , Calvin W. , Bell J. F. , Clark B. C. , Wang A. , McCoy T. J. , Schmidt M. E. , de Souza P. A.
Published: May 2008

Mineral deposits on the martian surface can elucidate ancient environmental conditions on the planet. Opaline silica deposits (as much as 91 weight percent SiO
2
) have been found in association with volcanic materials by the Mars rover Spirit. The deposits are present both as light-toned soils and as bedrock. We interpret these materials to have formed under hydrothermal conditions and therefore to be strong indicators of a former aqueous environment. This discovery is important for understanding the past habitability of Mars because hydrothermal environments on Earth support thriving microbial ecosystems.
Structural Diversity of Sodium
Authors: Gregoryanz Eugene , Lundegaard Lars F. , McMahon Malcolm I. , Guillaume Christophe , Nelmes Richard J. , Mezouar Mohamed
Published: May 2008

Sodium exhibits a pronounced minimum of the melting temperature at ∼118 gigapascals and 300 kelvin. Using single-crystal high-pressure diffraction techniques, we found that the minimum of the sodium melting curve is associated with a concentration of seven different crystalline phases. Slight changes in pressure and/or temperature induce transitions between numerous structural modifications, several of which are highly complex. The complexity of the phase behavior above 100 gigapascals suggests extraordinary liquid and solid states of sodium at extreme conditions and has implications for other seemingly simple metals.
Anticorrelated Seismic Velocity Anomalies from Post-Perovskite in the Lowermost Mantle
Authors: Hutko Alexander R. , Lay Thorne , Revenaugh Justin , Garnero Edward J.
Published: May 2008

Earth's lowermost mantle has thermal, chemical, and mineralogical complexities that require precise seismological characterization. Stacking, migration, and modeling of over 10,000
P
and
S
waves that traverse the deep mantle under the Cocos plate resolve structures above the core-mantle boundary. A small –0.07 ± 0.15% decrease of
P
wave velocity (
V
p
) is accompanied by a 1.5 ± 0.5% increase in
S
wave velocity (
V
s
) near a depth of 2570 km. Bulk-sound velocity

\batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \([V_{\mathrm{b}}=(V_{\mathrm{p}}^{2}-\frac{4}{3}V_{\mathrm{s}}^{2})^{\frac{1}{2}}]\) \end{document}

decreases by –1.0 ± 0.5% at this depth. Transition of the primary lower-mantle mineral, (Mg
1-x-y
Fe
x
Al
y
)(Si,Al)O
3
perovskite, to denser post-perovskite is expected to have a negligible effect on the bulk modulus while increasing the shear modulus by ∼6%, resulting in local anticorrelation of
V
b
and
V
s
anomalies; this behavior explains the data well.
Formation of Box Canyon, Idaho, by Megaflood: Implications for Seepage Erosion on Earth and Mars
Authors: Lamb Michael P. , Dietrich William E. , Aciego Sarah M. , DePaolo Donald J. , Manga Michael
Published: May 2008

Amphitheater-headed canyons have been used as diagnostic indicators of erosion by groundwater seepage, which has important implications for landscape evolution on Earth and astrobiology on Mars. Of perhaps any canyon studied, Box Canyon, Idaho, most strongly meets the proposed morphologic criteria for groundwater sapping because it is incised into a basaltic plain with no drainage network upstream, and approximately 10 cubic meters per second of seepage emanates from its vertical headwall. However, sediment transport constraints,
4
He and
14
C dates, plunge pools, and scoured rock indicate that a megaflood (greater than 220 cubic meters per second) carved the canyon about 45,000 years ago. These results add to a growing recognition of Quaternary catastrophic flooding in the American northwest, and may imply that similar features on Mars also formed by floods rather than seepage erosion.
Virus Population Dynamics and Acquired Virus Resistance in Natural Microbial Communities
Authors: Andersson Anders F. , Banfield Jillian F.
Published: May 2008

Viruses shape microbial community structure and function by altering the fitness of their hosts and by promoting genetic exchange. The complexity of most natural ecosystems has precluded detailed studies of virus-host interactions. We reconstructed virus and host bacterial and archaeal genome sequences from community genomic data from two natural acidophilic biofilms. Viruses were matched to their hosts by analyzing spacer sequences that occur among clustered regularly interspaced short palindromic repeats (CRISPRs) that are a hallmark of virus resistance. Virus population genomic analyses provided evidence that extensive recombination shuffles sequence motifs sufficiently to evade CRISPR spacers. Only the most recently acquired spacers match coexisting viruses, which suggests that community stability is achieved by rapid but compensatory shifts in host resistance levels and virus population structure.
What Systems Biology Is (Not, Yet)
Authors: Cain Corey J. , Conte Debra A. , García-Ojeda Marcos E. , Daglio Liza Gómez , Johnson Larry , Lau Eric H. , Manilay Jennifer O. , Phillips Julie Baker , Rogers Nabora Soledad , Stolberg Sarah E. , Swift Holly F. , Dawson Michael N.
Published: May 2008

Systems Biology
. Philosophical Foundations.
Fred C. Boogerd, Frank J. Bruggeman, Jan-Hendrik S. Hofmeyr, and Hans V. Westerhoff, Eds.
. Elsevier, Amsterdam, 2007. 360 pp. $120, £68.99, €99.95. ISBN 9780444520852.

An Introduction to Systems Biology
. Design Principles of Biological Circuits.
By Uri Alon
. Chapman and Hall/CRC, Boca Raton, FL, 2006. 317 pp. $54.95, £30.99. ISBN 9781584886426. Mathematical and Computational Biology, vol. 10.

Students and faculty from an interdisciplinary graduate class on systems biology review two books that offer a variety of perspectives on the field. Alon provides an analytical approach, whereas Boogerd et al. focus on philosophical aspects.
Resource Partitioning and Sympatric Differentiation Among Closely Related Bacterioplankton
Authors: Hunt Dana E. , David Lawrence A. , Gevers Dirk , Preheim Sarah P. , Alm Eric J. , Polz Martin F.
Published: May 2008

Identifying ecologically differentiated populations within complex microbial communities remains challenging, yet is critical for interpreting the evolution and ecology of microbes in the wild. Here we describe spatial and temporal resource partitioning among
Vibrionaceae
strains coexisting in coastal bacterioplankton. A quantitative model (AdaptML) establishes the evolutionary history of ecological differentiation, thus revealing populations specific for seasons and life-styles (combinations of free-living, particle, or zooplankton associations). These ecological population boundaries frequently occur at deep phylogenetic levels (consistent with named species); however, recent and perhaps ongoing adaptive radiation is evident in
Vibrio splendidus
, which comprises numerous ecologically distinct populations at different levels of phylogenetic differentiation. Thus, environmental specialization may be an important correlate or even trigger of speciation among sympatric microbes.
Hard Facts About Soft Animals
Authors: Glaubrecht Matthias
Published: May 2008

Phylogeny and Evolution of the Mollusca.Winston F. Ponder and David R. Lindberg, Eds.. University of California Press, Berkeley, 2008. 481 pp. $49.95, £29.95. ISBN 9780520250925.

The contributors combine data from anatomy, ecology, paleontology, and genomics to provide a summary of what we do and do not know about the evolutionary relationships of molluscs.
Regulated Protein Denitrosylation by Cytosolic and Mitochondrial Thioredoxins
Authors: Benhar Moran , Forrester Michael T. , Hess Douglas T. , Stamler Jonathan S.
Published: May 2008

Nitric oxide acts substantially in cellular signal transduction through stimulus-coupled
S
-nitrosylation of cysteine residues. The mechanisms that might subserve protein denitrosylation in cellular signaling remain uncharacterized. Our search for denitrosylase activities focused on caspase-3, an exemplar of stimulus-dependent denitrosylation, and identified thioredoxin and thioredoxin reductase in a biochemical screen. In resting human lymphocytes, thioredoxin-1 actively denitrosylated cytosolic caspase-3 and thereby maintained a low steady-state amount of
S
-nitrosylation. Upon stimulation of Fas, thioredoxin-2 mediated denitrosylation of mitochondria-associated caspase-3, a process required for caspase-3 activation, and promoted apoptosis. Inhibition of thioredoxin-thioredoxin reductases enabled identification of additional substrates subject to endogenous
S
-nitrosylation. Thus, specific enzymatic mechanisms may regulate basal and stimulus-induced denitrosylation in mammalian cells.
Appraisal of Interpretation Criteria for the Comparative Intradermal Tuberculin Test for Diagnosis of Tuberculosis in Cattle in Central Ethiopia
Authors: Ameni Gobena , Hewinson Glyn , Aseffa Abraham , Young Douglas , Vordermeier Martin
Published: May 2008

Accurate detection and removal of infected cattle, using immunodiagnostic tests such as the comparative intradermal tuberculin (CIDT) test, are the basis of control strategies for bovine tuberculosis (TB). According to the Office des Internationale Epizooties recommendation, the cutoff point for positivity of the CIDT test, calculated as the difference between skin thicknesses after bovine tuberculin (B) and avian tuberculin (A) injections (B−A), is >4 mm. This cutoff point is used worldwide, although it is likely that local conditions influence test performance. Thus, this study was formulated to determine CIDT test cutoff points applicable to cattle in central Ethiopia. Receiver operating characteristic analysis was performed for the CIDT test, using data from 186Bos indicus(zebu) andBos taurus(Holstein) cattle. Detailed postmortem examination for the presence of TB lesions was used to define disease status. At a cutoff of >2 mm, CIDT test sensitivity was 69% (95% confidence interval [95% CI], 58.5 to 79%), while it was 59% (95% CI, 49 to 69%) at a cutoff of >4 mm. In contrast, specificities of the CIDT test at these two cutoff values were identical, at 97% (95% CI, 89 to 100%). Thus, the maximum sensitivity of the CIDT test can be realized using a >2-mm cutoff without affecting specificity. The apparent prevalence was significantly (χ2= 13.56;P< 0.001) higher at a cutoff of >2 mm (16.0%;n= 5,424) than at a >4-mm cutoff (13.5%;n= 5,424). Nonetheless, no significant difference (χ2= 2.15;P= 0.14) in true prevalence was observed at a cutoff of >2 mm (19.6%) and at a cutoff of >4 mm (18.5%). Thus, our study demonstrates the importance of defining local, relevant cutoff values to maximize test sensitivity, and we suggest the application of the >2-mm cutoff for testing of cattle in central Ethiopia.
Antibodies to Proteinase 3 Prime Human Oral, Lung, and Kidney Epithelial Cells To Secrete Proinflammatory Cytokines upon Stimulation with Agonists to Various Toll-Like Receptors, NOD1, and NOD2
Authors: Uehara Akiko , Hirabayashi Yasuhiko , Takada Haruhiko
Published: May 2008

Antineutrophil cytoplasmic antibodies (ANCA) are autoantibodies, the detection of which in serum can be used in the diagnosis of Wegener's granulomatosis (WG). Proteinase 3 (PR3) is a major target antigen of ANCA in WG patients, and the interaction of PR3 ANCA with leukocytes causes a debilitating autoimmune disease. The first signs and symptoms in WG patients are observed in the oral cavity, lungs, and kidneys. Human epithelial cells generally do not secrete proinflammatory cytokines upon stimulation with pathogen-associated molecular patterns (PAMPs). In this study, anti-PR3 antibodies (Abs) and PR3 ANCA-containing sera from WG patients endowed human oral, lung, and kidney epithelial cells with responsiveness to PAMPs in terms of the production of proinflammatory cytokines, such as interleukin-6 (IL-6), IL-8, monocyte chemoattractant protein-1, and tumor necrosis factor alpha. Protease-activated receptor-2 (PAR-2) agonist peptides mimicked the priming effects of PR3 ANCA against PAMPs. Furthermore, the anti-PR3 Ab-mediated cell activation was significantly abolished by RNA interference targeting PAR-2 and NF-κB. This is the first report of priming effects of anti-PR3 Abs (PR3 ANCA) on epithelial cells. The results suggest that anti-PR3 Abs (PR3 ANCA) prime human epithelial cells to produce cytokines upon stimulation with various PAMPs, and these mechanisms may be involved in severe chronic inflammation in WG.
Double-Antigen Enzyme-Linked Immunosorbent Assay for Detection of Hepatitis E Virus-Specific Antibodies in Human or Swine Sera
Authors: Hu Wei Ping , Lu Yang , Precioso Nestor Amadeo , Chen Hsiao Ying , Howard Teresa , Anderson David , Guan Ming
Published: May 2008

A new double-antigen sandwich-based enzyme-linked immunosorbent assay (ELISA) for the detection of total antibodies (immunoglobulin G [IgG] and IgM) specific for hepatitis E virus (HEV) was developed by utilizing well-characterized recombinant protein ET2.1 and its peroxidase-labeled counterpart. Our study showed that the ELISA detected all the positive patient samples (
n
= 265) regardless of whether they contained IgM or IgG antibodies, or both, while it maintained an excellent specificity of 98.8% with samples from various patient or healthy control groups (total number of samples, 424). The test had a detection limit for anti-HEV IgG antibodies that was equivalent to 62 mIU/ml of the international reference. Compared with the serological status of the specimens determined on the basis of tests performed at the individual collection sites, the testing outcome generated by the new ELISA had a good agreement of 99.3%, with a kappa value of 0.985. The positive predictive value and the negative predictive value for the new test reached 98.1% and 100%, respectively. This ELISA had a positive delta value of 4.836 and a negative delta value of 3.314 (where delta is a measure of the number of standard deviations by which the cutoff is separated from the mean of the sample groups) (N. Crofts, W. Maskill, and I. D. Gust, J. Virol. Methods 22:51-59, 1988), indicating that it had an excellent ability to differentiate the infected and noninfected cohorts. Furthermore, the new design enables the detection of antibodies not only in human samples but also in pig samples. Our preliminary data showed that the ELISA could detect seroconversion in samples from pigs at as early as 14 days postinoculation. The potential utility of detecting specific antibodies in pigs will be an added advantage for managing the disease, with suggested zoonotic implications.
A Second Pilus Type in
Streptococcus pneumoniae
Is Prevalent in Emerging Serotypes and Mediates Adhesion to Host Cells
Authors: Bagnoli Fabio , Moschioni Monica , Donati Claudio , Dimitrovska Valentina , Ferlenghi Ilaria , Facciotti Claudia , Muzzi Alessandro , Giusti Fabiola , Emolo Carla , Sinisi Antonella , Hilleringmann Markus , Pansegrau Werner , Censini Stefano , Rappuoli Rino , Covacci Antonello , Masignani Vega , Barocchi Michele A.
Published: May 2008

Analysis of publicly available genomes of
Streptococcus pneumoniae
has led to the identification of a new genomic element containing genes typical of gram-positive pilus islets (PIs). Here, we demonstrate that this genomic region, herein referred to as PI-2 (consisting of
pitA
,
sipA
,
pitB
,
srtG1
, and
srtG2
) codes for a second functional pilus in pneumococcus. Polymerization of the PI-2 pilus requires the backbone protein PitB as well as the sortase SrtG1 and the signal peptidase-like protein SipA. Presence of PI-2 correlates with the genotype as defined by multilocus sequence typing and clonal complex (CC). The PI-2-positive CCs are associated with serotypes 1, 2, 7F, 19A, and 19F, considered to be emerging serotypes in both industrialized and developing countries. Interestingly, strains belonging to CC271 (where sequence type 271 is the predicted founder of the CC) contain both PI-1 and PI-2, as revealed by genome analyses. In these strains both pili are surface exposed and independently assembled. Furthermore, in vitro experiments provide evidence that the pilus encoded by PI-2 of
S. pneumoniae
is involved in adherence. Thus, pneumococci encode at least two types of pili that play a role in the initial host cell contact to the respiratory tract and are potential antigens for inclusion in a new generation of pneumococcal vaccines.
Direct Comparison of the BD Phoenix System with the MicroScan WalkAway System for Identification and Antimicrobial Susceptibility Testing of
Enterobacteriaceae
and Nonfermentative Gram-Negative Organisms
Authors: Snyder J. W. , Munier G. K. , Johnson C. L.
Published: May 2008

The Phoenix automated microbiology system (BD Diagnostics, Sparks, MD) is designed for the rapid identification (ID) and antimicrobial susceptibility testing (AST) of clinically significant human bacterial pathogens. We evaluated the performance of the Phoenix instrument in comparison with that of the MicroScan WalkAway system (Dade Behring, West Sacramento, CA) in the ID and AST of gram-negative clinical strains and challenge isolates of
Enterobacteriaceae
(
n
= 150) and nonfermentative gram-negative bacilli (NFGNB; 45 clinical isolates and 8 challenge isolates). ID discrepancies were resolved with the API 20E and API 20NE conventional biochemical ID systems (bioMerieux, Durham, NC). The standard disk diffusion method was used to resolve discordant AST results. The overall percentages of agreement between the Phoenix ID results and the MicroScan results at the genus and species levels for clinical isolates of
Enterobacteriaceae
were 98.7 and 97.7%, respectively; following resolution with conventional biochemical testing, the accuracy of the Phoenix system was determined to be 100%. For NFGNB, the levels of agreement were 100 and 97.7%, respectively. Both systems incorrectly identified the majority of the uncommon nonfermentative nonpseudomonal challenge isolates recovered from cystic fibrosis patients; these isolates are not included in the databases of the respective systems. For AST of
Enterobacteriaceae
, the rate of complete agreement between the Phoenix results and the MicroScan results was 97%; the rates of very major, major, and minor errors were 0.3, 0.2, and 2.7%, respectively. For NFGNB, the rate of complete agreement between the Phoenix results and the MicroScan results was 89.1%; the rates of very major, major, and minor errors were 0, 0.5, and 7.7%, respectively. Following the confirmatory testing of nine clinical isolates initially screened by the MicroScan system as possible extended-spectrum-β-lactamase (ESBL)-producing organisms (seven
Klebsiella pneumoniae
isolates and two
Escherichia coli
isolates), complete agreement was achieved for eight isolates (one ESBL positive and seven negative); one false positive was obtained with the Phoenix instrument. The MicroScan system correctly detected the 10 ESBL challenge isolates, versus the 6 detected by the Phoenix system. Overall, there was good correlation between the Phoenix instrument and the MicroScan system for the ID and AST of
Enterobacteriaceae
and common NFGNB. The Phoenix system is a reliable method for the ID and AST of the majority of clinical strains encountered in the clinical microbiology laboratory. Until additional performance data are available, results for all
Klebsiella pneumoniae
or
Klebsiella oxytoca
and
E. coli
isolates screened and confirmed as ESBL producers by any automated system should be confirmed by alternate methods prior to the release of final results.
DNA Typing of Cytological Samples for Retrospective Identification of an Early Case of Panton-Valentine Leucocidin-Positive, Community-Associated Methicillin-Resistant
Staphylococcus aureus
Pneumonia
Authors: Ho P. L. , Wong M. P. , Lai Eileen L. , Chan K. H. , Chiu Susan S.
Published: May 2008

This paper describes a fatal case of culture-confirmed, community-associated methicillin-resistant
Staphylococcus aureus
(MRSA) pneumonia in an 8-month-old child in Hong Kong in 2001. Stored cytological materials prepared from the pleural fluid were retrieved for molecular analysis. The result indicates the presence of a Panton-Valentine leucocidin-positive,
spa
type 019 MRSA.
First Report of Isolation and Characterization of
Aurantimonas altamirensis
from Clinical Samples
Authors: Luong Me-Linh , Békal Sadjia , Vinh Donald C. , Lauzon Dominique , Leung Vicki , Al-Rawahi Ghada N. , Ng Betty , Burdz Tamara , Bernard Kathryn
Published: May 2008

The genus
Aurantimonas
, proposed in 2003, encompasses four species from environmental sources, including
Aurantimonas altamirensis
, isolated from a cave wall in Spain. Here, we report what we believe are the first cases of the recovery of
A. altamirensis
from human clinical materials.
Prevalence and Sequence Diversity of a Factor Required for Actin-Based Motility in Natural Populations of
Burkholderia
Species
Authors: Sitthidet Chayada , Stevens Joanne M. , Chantratita Narisara , Currie Bart J. , Peacock Sharon J. , Korbsrisate Sunee , Stevens Mark P.
Published: May 2008

Actin-based motility of the melioidosis pathogen
Burkholderia pseudomallei
requires BimA. We report a high degree of conservation of
bimA
in 99
B. pseudomallei
isolates from the area of endemicity. A geographically restricted subset of
B. pseudomallei
isolates harbored a
B. mallei
-like
bimA
allele (12.1%), confounding a differential diagnostic test based on amplification of species-specific
bimA
regions.
Accuracy of Phenotypic Methods for Identification of
Streptococcus pneumoniae
Isolates Included in Surveillance Programs
Authors: Richter Sandra S. , Heilmann Kristopher P. , Dohrn Cassie L. , Riahi Fathollah , Beekmann Susan E. , Doern Gary V.
Published: May 2008

Similarities between
Streptococcus pneumoniae
and viridans group streptococci may result in misidentification of these organisms. In surveillance programs which assess antimicrobial resistance rates among respiratory tract pathogens, such identification errors could lead to overestimates of pneumococcal resistance rates. DNA probe analysis (Gen-Probe, San Diego, CA), the bile solubility test, optochin susceptibility, colony morphology, and the capsular swelling reaction with Omni serum (Staten Serum Institut, Copenhagen, Denmark) were used to characterize 1,733 organisms provisionally identified as
S. pneumoniae
in a 2004 to 2005 antimicrobial resistance surveillance program. These organisms were obtained in 41 U.S. medical centers. Among these, 1,647 (95%) were determined to be
S. pneumoniae
by DNA probe. Elimination of those isolates found not to be
S. pneumoniae
resulted in 1 to 2% decreases in resistance rate estimates with penicillin, erythromycin, tetracycline, and trimethoprim-sulfamethoxazole. With AccuProbe as a reference standard, the sensitivities and specificities of each phenotypic method for the identification of
S. pneumoniae
were, respectively, 98.8% and 82.6% for bile solubility, 99.3% and 74.4% for the capsular swelling reaction with Omni serum, and 87.9% and 59.3% for optochin susceptibility. Colony morphology was of limited value, as 391 (23.7%) isolates lacked the typical button or mucoid colony appearance of
S. pneumoniae
.
Human Coronavirus NL63 and 229E Seroconversion in Children
Authors: Dijkman Ronald , Jebbink Maarten F. , El Idrissi Nawal Bahia , Pyrc Krzysztof , Müller Marcel A. , Kuijpers Taco W. , Zaaijer Hans L. , van der Hoek Lia
Published: May 2008

In 2004, the novel respiratory human coronavirus NL63 (HCoV-NL63) was identified, and subsequent research revealed that the virus has spread worldwide. HCoV-229E is a close relative of HCoV-NL63, and infection with either virus can lead to the hospitalization of young children, immunocompromised persons, and the elderly. Children infected with HCoV-NL63 often develop croup, with obstruction of the airway. In this study we investigated at which age children are confronted for the first time with an HCoV-NL63 infection and, thus, at which age they seroconvert to HCoV-NL63 positivity. We designed a recombinant HCoV-229E and a recombinant HCoV-NL63 nucleocapsid protein enzyme-linked immunosorbent assay and performed a seroepidemiology survey on longitudinal and cross-sectional serum samples. The longitudinal serum samples were collected from 13 newborns, and data for those newborns were available from multiple time points spanning a period of at least 18 months. For the cross-sectional survey we tested serum samples of 139 children, including newborns to children 16 years of age. In examinations of the longitudinal serum samples we observed that all of the children had maternal anti-NL63 and anti-229E antibodies at birth that disappeared within 3 months. Seven of the 13 children became HCoV-NL63 seropositive during follow-up, whereas only 2 became HCoV-229E seropositive. The serology data of the cross-sectional serum samples revealed that 75% and 65% of the children in the age group 2.5 to 3.5 years were HCoV-NL63 and HCoV-229E seropositive, respectively. We conclude that on average, HCoV-NL63 and HCoV-229E seroconversion occurs before children reach the age of 3.5 years.
Isolation and Characterization of a Novel
Francisella
sp. from Human Cerebrospinal Fluid and Blood
Authors: Kugeler Kiersten J. , Mead Paul S. , McGowan Karin L. , Burnham Jon M. , Hogarty Michael D. , Ruchelli Eduardo , Pollard Kerry , Husband Brigitte , Conley Caryn , Rivera Tanya , Kelesidis Theodoros , Lee Walter M. , Mabey Walburga , Winchell Jonas M. , Stang Heather L. , Staples J. Erin , Chalcraft Linda J. , Petersen Jeannine M.
Published: May 2008

We describe the isolation of a
Francisella
sp. from normally sterile sites in acutely ill patients in two different states within 2 years. Microbiologic and molecular analyses indicate that this organism represents a novel
Francisella
sp. Clinicians and microbiologists should be aware of this new potential pathogen, as infection may be more common than recognized.
Relationship between
Escherichia coli
Strains Causing Acute Cystitis in Women and the Fecal
E. coli
Population of the Host
Authors: Moreno Eva , Andreu Antonia , Pigrau Carles , Kuskowski Michael A. , Johnson James R. , Prats Guillem
Published: May 2008

Previous epidemiological assessments of the prevalence versus special-pathogenicity hypothesis for urinary tract infection (UTI) pathogenesis in women may have been confounded by underlying host population differences between women with UTI and healthy controls and have not considered the clonal complexity of the fecal
Escherichia coli
population of the host. In the present study, 42 women with acute uncomplicated cystitis served as their own controls for an analysis of the causative
E. coli
strain and the concurrent intestinal
E. coli
population. Clonality among the urine isolate and 30 fecal colonies per subject was assessed by repetitive-element PCR and macrorestriction analysis. Each unique clone underwent PCR-based phylotyping and virulence genotyping. Molecular analysis resolved 109 unique clones (4 urine-only, 38 urine-fecal, and 67 fecal-only clones). Urine clones exhibited a significantly higher prevalence of group B2 than fecal-only clones (69% versus 10%;
P
< 0.001) and higher aggregate virulence scores (mean, 6.2 versus 2.9;
P
< 0.001). In multilevel regression models for predicting urine clone status, significant positive predictors included group B2, 10 individual virulence traits, the aggregate virulence score, fecal dominance, relative fecal abundance, and (unique to the present study) a pauciclonal fecal sample. In summary, within the fecal
E. coli
populations of women with acute cystitis, pauciclonality, clonal dominance, virulence, and group B2 status are closely intertwined. Phylogenetic group B2 status and/or associated virulence factors may promote fecal abundance and pauciclonality, thereby contributing to upstream steps in UTI pathogenesis. This relationship suggests a possible reconciliation of the prevalence and special-pathogenicity hypotheses.
Epidemic of Genotype GII.2 Noroviruses during Spring 2004 in Osaka City, Japan
Authors: Iritani Nobuhiro , Kaida Atsushi , Kubo Hideyuki , Abe Niichiro , Murakami Tsukasa , Vennema Harry , Koopmans Marion , Takeda Naokazu , Ogura Hisashi , Seto Yoshiyuki
Published: May 2008

Between March and May 2004, a GII.2 genotype norovirus strain caused an epidemic of acute gastroenteritis in Osaka, Japan. Phylogenetic analysis showed that this strain was distinct from all other GII.2 strains detected in Osaka City between April 1996 and March 2005.
In Situ Detection of
Anaplasma
spp. by DNA Target-Primed Rolling-Circle Amplification of a Padlock Probe and Intracellular Colocalization with Immunofluorescently Labeled Host Cell von Willebrand Factor
Authors: Wamsley Heather L. , Barbet Anthony F.
Published: May 2008

Endothelial cell culture and preliminary immunofluorescent staining of
Anaplasma
-infected tissues suggest that endothelial cells may be an in vivo nidus of mammalian infection. To investigate endothelial cells and other potentially cryptic sites of
Anaplasma
sp. infection in mammalian tissues, a sensitive and specific isothermal in situ technique to detect localized
Anaplasma
gene sequences by using rolling-circle amplification of circularizable, linear, oligonucleotide probes (padlock probes) was developed. Cytospin preparations of uninfected or
Anaplasma
-infected cell cultures were examined using this technique. Via fluorescence microscopy, the technique described here, and a combination of differential interference contrast microscopy and von Willebrand factor immunofluorescence,
Anaplasma phagocytophilum
and
Anaplasma marginale
were successfully localized in situ within intact cultured mammalian cells. This work represents the first application of this in situ method for the detection of a microorganism and forms the foundation for future applications of this technique to detect, localize, and analyze
Anaplasma
nucleotide sequences in the tissues of infected mammalian and arthropod hosts and in cell cultures.
Development of Multiplex PCR-Ligase Detection Reaction Assay for Detection of West Nile Virus
Authors: Rondini S. , Pingle M. R. , Das S. , Tesh R. , Rundell M. S. , Hom J. , Stramer S. , Turner K. , Rossmann S. N. , Lanciotti R. , Spier E. G. , Muñoz-Jordán J. , Larone D. , Spitzer E. , Barany F. , Golightly L. M.
Published: May 2008

We have developed a novel multiplex reverse transcription-PCR ligase detection reaction (RT-PCR/LDR) assay for the detection of West Nile virus (WNV) in both clinical and mosquito pool samples. The method relies on the amplification of three different genomic regions, one in the coding sequence of nonstructural protein NS2a and two in nonstructural protein NS5, to minimize the risk of detection failure due to genetic variation. The sensitivity of the PCR is complemented by the high specificity of the LDR step, and the detection of the LDR products can be achieved with capillary electrophoresis (CE) or a universal DNA microarray. We evaluated the limit of detection by both one-step and two-step multiplex RT-PCR/LDR/CE approaches, which reached, respectively, 0.005 and 0.017 PFU. The assay demonstrated 99% sensitivity when mosquito pool samples were tested and 100% sensitivity with clinical samples when the one-step approach was used. The broad strain coverage was confirmed by testing 34 WNV isolates belonging to lineages 1 and 2, and the high specificity of the assay was determined by testing other flaviviruses, as well as negative mosquito pool and clinical samples. In summary, the multiplex RT-PCR/LDR assay could represent a valuable complement to WNV serological diagnosis, especially in early symptomatic patients. In addition, the multiplexing capacity of the technique, which can be coupled to universal DNA microarray detection, makes it an amenable tool to develop a more comprehensive assay for viral pathogens.
Early Increases in Superantigen-Specific Foxp3+Regulatory T Cells during Mouse Mammary Tumor Virus Infection
Authors: Cabrera Gabriel , Burzyn Dalia , Mundiñano Juliana , Courreges M. Cecilia , Camicia Gabriela , Lorenzo Daniela , Costa Héctor , Ross Susan R. , Nepomnaschy Irene , Piazzon Isabel
Published: May 2008

Mouse mammary tumor virus (MMTV) is a milk-borne betaretrovirus that has developed strategies to exploit and subvert the host immune system. Here, we show in a natural model of MMTV infection that the virus causes early and progressive increases in superantigen (SAg)-specific Foxp3+regulatory T cells (Treg) in Peyer's patches (PP). These increases were shown to be dependent on the presence of dendritic cells. CD4+CD25+T cells from the PP of infected mice preferentially suppress the proliferative response of T cells to SAg-expressing antigen-presenting cells ex vivo. We investigated the influence of the depletion of CD25+cells at different stages of the infection. When CD25+cells were depleted before MMTV infection, an increase in the number of PP SAg-cognate Foxp3−T cells was found at day 6 of infection. Since the SAg response is associated with viral amplification, the possibility exists that Tregcells attenuate the increase in viral load at the beginning of the infection. In contrast, depletion of CD25+cells once the initial SAg response has developed caused a lower viral load, suggesting that at later stages Tregcells may favor viral persistence. Thus, our results indicated that Tregcells play an important and complex role during MMTV infection.
Compensatory Role of Human Immunodeficiency Virus Central Polypurine Tract Sequence in Kinetically Disrupted Reverse Transcription
Authors: Skasko Mark , Kim Baek
Published: May 2008

We tested whether the additional positive-strand DNA synthesis initiation of human immunodeficiency virus type 1 (HIV-1) from the central polypurine tract (cPPT) facilitates efficient completion of kinetically disturbed proviral DNA synthesis induced by dysfunctional reverse transcriptase (RT) mutants or limited cellular deoxynucleoside triphosphate (dNTP) pools. Indeed, the cPPT enabled the HIV-1 vectors harboring RT mutants with reduced dNTP binding affinity to transduce human lung fibroblasts (HLFs), without which these mutant vectors normally fail to transduce. The cPPT showed little effect on wild-type HIV-1 vector transduction in HLF, whereas it significantly enhanced vector transduction in HLFs engineered to contain reduced dNTP pools, suggesting a novel compensatory role for cPPT in viruses harboring kinetically impaired RT.
Role for Conserved Residues of Sindbis Virus Nonstructural Protein 2 Methyltransferase-Like Domain in Regulation of Minus-Strand Synthesis and Development of Cytopathic Infection
Authors: Mayuri  , Geders Todd W. , Smith Janet L. , Kuhn Richard J.
Published: May 2008

The plus-strand RNA genome of Sindbis virus (SINV) encodes four nonstructural proteins (nsP1 to nsP4) that are involved in the replication of the viral RNA. The ∼800-amino-acid nsP2 consists of an N-terminal domain with nucleoside triphosphatase and helicase activities and a C-terminal protease domain. Recently, the structure determined for Venezuelan equine encephalitis virus nsP2 indicated the presence of a previously unrecognized methyltransferase (MTase)-like domain within the C-terminal ∼200 residues and raised a question about its functional importance. To assess the role of this MTase-like region in viral replication, highly conserved arginine and lysine residues were mutated to alanine. The plaque phenotypes of these mutants ranged from large/wild-type to small plaques with selected mutations demonstrating temperature sensitive lethality. The proteolytic polyprotein processing activity of nsP2 was unaffected in most of the mutants. Some of the temperature-sensitive mutants showed reduction in the minus-strand RNA synthesis, a function that has not yet been ascribed to nsP2. Mutation of SINV residue R615 rendered the virus noncytopathic and incapable of inhibiting the host cell translation but with no effects on the transcriptional inhibition. This property differentiated the mutation at R615 from previously described noncytopathic mutations. These results implicate nsP2 in regulation of minus-strand synthesis and suggest that different regions of the nsP2 MTase-like domain differentially modulate host defense mechanisms, independent of its role as the viral protease.
UL36p Is Required for Efficient Transport of Membrane-Associated Herpes Simplex Virus Type 1 along Microtubules
Authors: Shanda Sara K. , Wilson Duncan W.
Published: May 2008

Microtubule-mediated anterograde transport is essential for the transport of herpes simplex virus type 1 (HSV-1) along axons, yet little is known regarding the mechanism and the machinery required for this process. Previously, we were able to reconstitute anterograde transport of HSV-1 on microtubules in an in vitro microchamber assay. Here we report that the large tegument protein UL36p is essential for this trafficking. Using a fluorescently labeled UL36 null HSV-1 strain, KΔUL36GFP, we found that it is possible to isolate a membrane-associated population of this virus. Although these viral particles contained normal amounts of tegument proteins VP16, vhs, and VP22, they displayed a 3-log decrease in infectivity and showed a different morphology compared to UL36p-containing virions. Membrane-associated KΔUL36GFP also displayed a slightly decreased binding to microtubules in our microchamber assay and a two-thirds decrease in the frequency of motility. This decrease in binding and motility was restored when UL36p was supplied in
trans
by a complementing cell line. These findings suggest that UL36p is necessary for HSV-1 anterograde transport.
A Single Amino Acid Substitution in the Murine Norovirus Capsid Protein Is Sufficient for Attenuation In Vivo
Authors: Bailey D. , Thackray L. B. , Goodfellow I. G.
Published: May 2008

Murine norovirus (MNV), a prevalent pathogen of laboratory mice, shares many characteristics with human noroviruses. Previous results indicated that passage of MNV1 in the macrophage cell line RAW 264.7 results in attenuation in STAT1-deficient mice (C. E. Wobus, S. M. Karst, L. B. Thackray, K. O. Chang, S. V. Sosnovtsev, G. Belliot, A. Krug, J. M. Mackenzie, K. Y. Green, and H. W. Virgin, PLoS. Biol. 2:e432, 2004). Sequence analysis revealed two amino acid differences between the virulent and attenuated viruses. Using an infectious cDNA clone of the attenuated virus, we demonstrated that a glutamate-to-lysine substitution at position 296 in the capsid protein (VP1) is sufficient to restore virulence in vivo, identifying, for the first time, a virus-encoded molecular determinant of norovirus virulence.
Varicella-Zoster Virus Open Reading Frame 66 Protein Kinase Is Required for Efficient Viral Growth in Primary Human Corneal Stromal Fibroblast Cells
Authors: Erazo Angela , Yee Michael B. , Osterrieder Nikolaus , Kinchington Paul R.
Published: May 2008

Varicella-zoster virus (VZV) open reading frame 66 (ORF66) encodes a serine/threonine protein kinase that is not required for VZV growth in most cell types but is needed for efficient growth in T cells. The ORF66 kinase affects nuclear import and virion packaging of IE62, the major regulatory protein, and is known to regulate apoptosis in T cells. Here, we further examined the importance of ORF66 using VZV recombinants expressing green fluorescent protein (GFP)-tagged functional and kinase-negative ORF66 proteins. VZV virions with truncated or kinase-inactivated ORF66 protein were marginally reduced for growth and progeny yields in MRC-5 fibroblasts but were severely growth and replication impaired in low-passage primary human corneal stromal fibroblasts (PCF). To determine if the growth impairment was due to ORF66 kinase regulation of IE62 nuclear import, recombinant VZVs that expressed IE62 with alanine residues at S686, the suspected target by which ORF66 kinase blocks IE62 nuclear import, were made. IE62 S686A expressed by the VZV recombinant remained nuclear throughout infection and was not packaged into virions. However, the mutant virus still replicated efficiently in PCF cells. We also show that inactivation of the ORF66 kinase resulted in only marginally increased levels of apoptosis in PCF cells, which could not fully account for the cell-specific growth requirement of ORF66 kinase. Thus, the unique short region VZV kinase has important cell-type-specific functions that are separate from those affecting IE62 and apoptosis.
Formation of Guanosine(5′)Tetraphospho(5′)Adenosine Cap Structure by an Unconventional mRNA Capping Enzyme of Vesicular Stomatitis Virus
Authors: Ogino Tomoaki , Banerjee Amiya K.
Published: May 2008

The RNA-dependent RNA polymerase L protein of vesicular stomatitis virus (VSV) elicits GTPase and RNA:GDP polyribonucleotidyltransferase (PRNTase) activities to produce a 5′-cap core structure, guanosine(5′)triphospho(5′)adenosine (GpppA), on viral mRNAs. Here, we report that the L protein produces an unusual cap structure, guanosine(5′)tetraphospho(5′)adenosine (GppppA), that is formed by the transfer of the 5′-monophosphorylated viral mRNA start sequence to GTP by the PRNTase activity before the removal of the γ-phosphate from GTP by GTPase. Interestingly, GppppA-capped and polyadenylated full-length mRNAs were also found to be synthesized by an in vitro transcription system with the native VSV RNP.
Herpes Simplex Virus Type 1 ICP27 Regulates Expression of a Variant, Secreted Form of Glycoprotein C by an Intron Retention Mechanism
Authors: Sedlackova Lenka , Perkins Keith D. , Lengyel Joy , Strain Anna K. , van Santen Vicky L. , Rice Stephen A.
Published: May 2008

We previously showed that herpes simplex virus type 1 (HSV-1) immediate-early (IE) protein ICP27 can posttranscriptionally stimulate mRNA accumulation from a transfected viral late gene encoding glycoprotein C (gC) (K. D. Perkins, J. Gregonis, S. Borge, and S. A. Rice, J. Virol. 77:9872-9884, 2003). We began this study by asking whether ICP27 homologs from other herpesviruses can also mediate this activity. Although the homologs from varicella-zoster virus (VZV) and human cytomegalovirus (HCMV) were inactive, the homolog from bovine herpesvirus 4 (BHV-4), termed HORF1/2, was a very efficient transactivator. Surprisingly, most of the mRNA produced via HORF1/2 transactivation was 225 nucleotides shorter than expected due to the removal of a previously undescribed intron from the gC transcript. We found that the gC mRNA produced in the absence of transactivation was also mostly spliced. In contrast, gC mRNA produced by ICP27 transactivation was predominantly unspliced. Based on these results, we conclude that ICP27 has two distinct effects on the transfected gC gene: it (i) stimulates mRNA accumulation and (ii) promotes the retention of an intron. Interestingly, the spliced transcript encodes a variant of gC that lacks its transmembrane domain and is secreted from transfected cells. As the gC splicing signals are conserved among several HSV-1 strains, we investigated whether the variant gC is expressed during viral infection. We report here that both the spliced transcript and its encoded protein are readily detected in Vero cells infected with three different laboratory strains of wild-type HSV-1. Moreover, the variant gC is efficiently secreted from infected cells. We have designated this alternate form of the protein as gCsec. As the extracellular domain of gC is known to bind heparan sulfate-containing proteoglycans and to inhibit the complement cascade via an interaction with complement component C3b, we speculate that gCsec could function as a secreted virulence factor.