bluuebunny/crossref_metadata_2025 huggingface dataset:
MCM3 could be a better marker than Ki‐67 for evaluation of dysplastic oral lesions: an immunohistochemical study
Authors: Lameira Aladim Gomes , Pontes Flávia Sirotheau Corrêa , Guimarães Douglas Magno , Alves Ana Carolina Garcia , de Jesus Adriana Souza , Pontes Hélder Antônio Rebelo , Pinto Décio dos Santos
Published: January 2014

Background

The aim of this study was to identify the expression of MCM3, Ki‐67 and p27 in normal mucosa, leucoplakia and oral squamous cell carcinoma (OSCC) and determine whether altered expression could serve as a prognostic marker of a malignant progression of dysplastic lesions.

Methods

The samples were collected from 37 patients with oral leucoplakia (13 with mild dysplasia – MLD, 12 with moderate dysplasia – MD and 12 with severe dysplasia – SD). Eleven samples of mouth floor mucocele (M) and 50 floor mouth and tongue samples OSCC of untreated patients were included in this study. Immunohistochemical expression of MCM3, Ki‐67 and p27 of all the groups was analysed. Kruskal–Wallis and Dunn's test were used to determine differences among groups, and a Pearson's correlation test was used to evaluate the correlation between the proteins.

Results

Ki‐67 expression was higher in OSCC than M (P < 0.001) and MLD (P < 0.01) groups, and there was a lower expression in M compared with MD and SD (P < 0.05). Regarding p27, its expression was lower in OSCC compared with M, MD and SD. MCM3 expression was lower in M compared with SD and OSCC (P < 0.001), and MLD showed a lower expression when compared SD (P < 0.01) and OSCC (P < 0.001). Moreover, a better correlation was observed between the proteins MCM3 and p27 than between Ki‐67 and p27 proteins when all lesions were examined together.

Conclusions

This study showed that MCM3 could be a better marker than Ki‐67 for evaluation of dysplastic oral lesions.
Correlation of ALDH1, CD44, OCT4 and SOX2 in tongue squamous cell carcinoma and their association with disease progression and prognosis
Authors: Huang Cong‐Fa , Xu Xiao‐Rong , Wu Tian‐Fu , Sun Zhi‐Jun , Zhang Wen‐Feng
Published: January 2014

Background

Recently, studies indicated that cancer stem cell plays a key role in cancer development and progression. However, the role of cancer stem cell has not been well elucidated in tongue squamous cell carcinoma (TSCC). The objective of this study was to investigate the relationships between the expressions of stem cell markers and the prognosis of TSCC.

Materials and methods

Immunohistochemistry was employed to analyse the protein expression levels of ALDH1, CD44, OCT4 and SOX2 in 66 TSCC tissue samples. The results were then evaluated semiquantitatively and compared with other clinicopathological variables.

Results

Immunohistochemistry revealed that the ALDH1, CD44, OCT4 and SOX2 proteins were overexpressed in the 66 TSCC specimens used in this study. Spearman's correlation analysis showed that the expressions of ALDH1 and CD44 were significantly correlated with SOX2 except other proteins (P < 0.05) and that OCT4 and SOX2 were significantly related (P < 0.01). Kaplan–Meier analysis revealed that T category, node metastasis, TNM stage, differentiation and distant metastasis were associated with poor patient survival (P < 0.05). Multivariate Cox regression analysis demonstrated that SOX2, recurrence and distant metastasis were independent prognostic factors of overall survival in patients with TSCC.

Conclusion

Taken together, these data suggest that the stem cell markers ALDH1, CD44, OCT4 and SOX2 are closely related in TSCC, and the expression of SOX2 can be used as a prognostic indicator of TSCC.
Prosecutorial Retention: Signaling by Trial
Authors: BANDYOPADHYAY SIDDHARTHA , McCANNON BRYAN C.
Published: January 2014

We examine how retention motives affect prosecutor behavior under different evaluation criteria. In particular, we analyze how prosecutors of differing capabilities respond in choosing which cases to take to trial and which to plea bargain. We show how different criteria distort the mix of cases chosen for trial and that the direction of the distortion depends crucially on the evaluation tool used. Optimal evaluation metrics are derived that combine multiple signals of performance and are shown to achieve the first‐best outcome.
The importance of temporal heterothermy in bats
Authors: Stawski C. , Willis C. K. R. , Geiser F.
Published: January 2014

Animals must balance their energy budgets even when confronted with periodic food shortages and/or adverse environmental conditions. Especially, small endothermic animals require large amounts of energy to maintain high and stable body temperatures (Tb) via endogenous heat production. To deal with energetic challenges, many small endotherms are heterothermic, abandon regulation of highTband enter a state of torpor resulting in large energy savings. Torpor is used by many bat species because they are small, have high rates of heat loss and rely on fluctuating food resources (e.g. insects, fruit, nectar). Many bats use torpor all year, but the expression of temporal heterothermy can be strongly seasonal especially for temperate and subtropical species, which may hibernate for long periods. Recent advances in our understanding of torpor expression in bats have been made using temperature telemetry for remote data collection ofTbin free‐ranging wild individuals from all climate zones. This new knowledge on free‐ranging bats has revealed the importance of torpor expression not only for energy conservation but also for other benefits, such as reduction of extrinsic mortality (e.g. predation). On the contrary, dense clustering during hibernation, important for minimizing energy and water loss, may also expose bats to infectious disease. An emerging, cold‐tolerant fungal pathogen of bats causes a new disease called white‐nose syndrome (WNS), which is devastating populations of multiple species in easternNorthAmerica. Given the importance of temporal heterothermy to their biology, and links between torpor expression and mortality fromWNS, it is becoming increasingly important to understand the ecology and physiology of torpor in this largely understudied and cryptic mammalian group. Here, we review past and current literature to summarize the importance and evolution of heterothermy in bats.
Individual Preferences and Job Characteristics: An Analysis of Cooperative Credit Banks
Authors: Nese Annamaria , Troisi Roberta
Published: January 2014

The aim of this paper is to analyse individual preferences in relation to different job characteristics. More specifically, this work focuses on the case of employees of cooperative credit banks (CCBs) in Campania and accounts for certain fundamental institutional features: CCBs are designed to pursue specific member interests rather than profit maximization, and most employees are both owners and consumers. The research is conducted by applying a conjoint analysis approach with stated preference data. Novel features of the analysis include the application of this approach to empirical research on worker incentives and the use of a mixed logit model.
Right Node Raising
Authors: Sabbagh Joseph
Published: January 2014

Right Node Raising is the term used by linguists to refer to a construction in which a shared argument surfaces at the right periphery of a coordinate structure (e.g.Phil loves, but Mary hates, Greek tragedies). This paper begins by introducing some of the basic properties of this construction and then moves on to present and compare the main analytical approaches to this construction.
Assembly and stoichiometry of FliF and FlhA in Salmonella flagellar basal body
Authors: Morimoto Yusuke V. , Ito Mariko , Hiraoka Koichi D. , Che Yong‐Suk , Bai Fan , Kami‐ike Nobunori , Namba Keiichi , Minamino Tohru
Published: January 2014

The bacterial flagellar export apparatus is required for the construction of the bacterial flagella beyond the cytoplasmic membrane. The membrane‐embedded part of the export apparatus, which consists of FlhA, FlhB, FliO, FliP, FliQ and FliR, is located in the central pore of the MS ring formed by 26 copies of FliF. The C‐terminal cytoplasmic domain of FlhA is located in the centre of the cavity within the C ring made of FliG, FliM and FliN. FlhA interacts with FliF, but its assembly mechanism remains unclear. Here, we fused yellow fluorescent protein (YFP) and cyan fluorescent protein (CFP) to the C‐termini of FliF and FlhA and investigated their subcellular localization by fluorescence microscopy. The punctate pattern of FliF–YFP localization required FliG but neither FliM, FliN, FlhA, FlhB, FliO, FliP, FliQ nor FliR. In contrast, FlhA–CFP localization required FliF, FliG, FliO, FliP, FliQ and FliR. The number of FlhA–YFP molecules associated with the MS ring was estimated to be about nine. We suggest that FlhA assembles into the export gate along with other membrane components during the MS ring complex formation in a co‐ordinated manner.
Combined Approach Improves the Outcomes of Catheter Ablation of Idiopathic Ventricular Arrhythmias Originating from the Vicinity of Tricuspid Annulus
Authors: TENG LI , XIAN‐ZHANG ZHAN , YU‐MEI XUE , XIAN‐HONG FANG , HONG‐TAO LIAO , HAI DENG , WEI WEI , SHU‐LIN WU
Published: January 2014

Background

Ablation of idiopathic ventricular arrhythmias (VAs) originating from the vicinity of tricuspid annulus (TA) is often unsuccessful via inferior approach. We report the initial experience with catheter ablation of VAs arising from the vicinity of the TA via superior approach.

Methods

We retrospectively studied 36 patients with VAs arising from the vicinity of TA who underwent ablation via transfemoral vein approach first. If patients had a failed prior ablation or VAs recurred during follow‐up, they were referred for repeat ablation via transsubclavian vein approach.

Results

Among 36 patients, 11 (30.6%) patients (five failed during the index procedure and six recurred during the follow‐up) were assigned to perform repeat ablation via the transsubclavian vein approach. After the final procedure two patients recurred again, and success rate increased from 69.5% (25/36) to 94.4% (34/36). Amplitudes of the atrial electrograms of all successful ablation sites via the transsubclavian vein approach was <0.036 mV.

Conclusions

The transsubclavian vein approach plus transfemoral vein approach improve the outcomes of catheter ablation of idiopathic VAs originating from the vicinity of TA. The transsubclavian vein approach is a feasible alternative for VAs, which has been refractory to ablation via the inferior approach.
The Nicotiana attenuata GLA1 lipase controls the accumulation of Phytophthora parasitica‐induced oxylipins and defensive secondary metabolites
Authors: SCHUCK STEFAN , KALLENBACH MARIO , BALDWIN IAN T. , BONAVENTURE GUSTAVO
Published: January 2014

Nicotiana attenuata plants silenced in the expression of GLYCEROLIPASE A1 (ir‐gla1 plants) are compromised in the herbivore‐ and wound‐induced accumulation of jasmonic acid (JA). However, these plants accumulate wild‐type (WT) levels of JA and divinyl‐ethers during Phytophthora parasitica infection. By profiling oxylipin‐enriched fractions with targeted and untargeted liquid chromatography‐tandem time‐of‐flight mass spectrometry approaches, we demonstrate that the accumulation of 9‐hydroxy‐10E,12Z‐octadecadienoic acid (9‐OH‐18:2) and additional C18 and C19 oxylipins is reduced by ca. 20‐fold in P. parasitica‐infected ir‐gla1 leaves compared with WT. This reduced accumulation of oxylipins was accompanied by a reduced accumulation of unsaturated free fatty acids and specific lysolipid species. Untargeted metabolic profiling of total leaf extracts showed that 87 metabolites accumulated differentially in leaves of P. parasitica‐infected ir‐gla1 plants with glycerolipids, hydroxylated‐diterpene glycosides and phenylpropanoid derivatives accounting together for ca. 20% of these 87 metabolites. Thus, P. parasitica‐induced oxylipins may participate in the regulation of metabolic changes during infection. Together, the results demonstrate that GLA1 plays a distinct role in the production of oxylipins during biotic stress responses, supplying substrates for 9‐OH‐18:2 and additional C18 and C19 oxylipin formation during P. parasitica infection, whereas supplying substrates for the biogenesis of JA during herbivory and mechanical wounding.
Colloid Centrifugation Selects Normal Spermatozoa from Polymorphic Bull Ejaculates: A Case Study
Authors: Morrell JM , Rodriguez‐Martinez H , Andersson M
Published: January 2014

Contents

Semen from a Western Finncattle bull exhibiting a highly polymorphic spermiogram was processed by colloid centrifugation using Androcoll‐B, a species‐specific silane‐coated silica colloid. In the first experiment, Single Layer Centrifugation (SLC) was used to identify which density colloids were needed to separate different cell populations. Colloids of the two chosen densities were then used in a density gradient resulting in two sperm subpopulations, one containing nearly all normally sized spermatozoa and the other enriched for the macrocephalic spermatozoa. Microcephalic spermatozoa did not appear in either of the selected subpopulations. Using a combination of SLC and DGC with this species‐specific colloid, it was possible to separate the spermatozoa into different subpopulations, that is, a subpopulation containing nearly all normally sized spermatozoa, and another one enriched for the macrocephalic spermatozoa. Thus, colloid centrifugation could be used to select sufficient normal spermatozoa from a highly polymorphic ejaculate for AI, if desired.
Influence of ion-to-atom ratio on the microstructure of evaporated molybdenum thin films grown using low energy argon ions
Authors: Yadav Praveen Kumar , Sant Tushar , Mukherjee Chandrachur , Nayak Maheswar , Rai Sanjay Kumar , Lodha Gyanendra Singh , Sharma Surinder Mohan
Published: January 2014

The authors report the effect of argon ion to molybdenum atom ratio (r) on the microstructure of low energy (70 eV) argon ion assisted electron beam evaporated Mo thin films. Surface roughness, morphology, and crystallinity of Mo films are found to strongly depend on “r.” Increase of “r” from 0 to 100 induces gradual loss in crystallinity, reduction in surface roughness and systematic increase in density of the film. For “r” ∼ 100, average atomic density of the film approaches the bulk value (97%) with lowest surface roughness. Further, increasing “r” up to 170 reduces the atomic density, increases roughness, and increase in crystallinity induced by low energy Ar ion beam. The observed surface roughness and grain size determined by x-ray reflectivity and glancing incidence x-ray diffraction correlate well with atomic force microscopy measurements. This study demonstrates that for r = 100 one gets lowest roughness Mo film with highest density and nearly amorphous microstructure. The growth model is discussed by structural zone model.
Sensitive surface-enhanced Raman scattering active substrate based on gap surface plasmon polaritons
Authors: Jiang Youwei , Wang Binhao , Cheng Xing
Published: January 2014

The authors fabricate three types of surface-enhanced Raman scattering (SERS) active substrates by nanoimprint lithography and experimentally investigated the intensity enhancement of Raman scattering induced by gap surface plasmon polaritons (GSPPs). An area-averaged Raman signal enhancement factor of 1.1 × 107 can be achieved by the GSPPs structures for monolayer thiophenol molecules absorbed on silver surfaces, compared with 6.2 × 105 of conventional SERS-active substrates. The exceptional field enhancement originates not only from the localized surface plasmon polaritons but also from the coupling between the GSPPs at two metal/dielectric interfaces. These structures are expected to find promising applications as SERS devices in trace-level molecular detection and identification.
Development of atomic layer deposition-activated microchannel plates for single particle detection at cryogenic temperatures
Authors: Gorelikov Dmitry , Sullivan Neal , de Rouffignac Philippe , Li Huazhi , Narayanamoorthy Jayasri , Tremsin Anton S.
Published: January 2014

Atomic layer deposition (ALD) technology is used to nanoengineer functional films inside the pores of microchannel plate (MCP) electron multipliers, enabling a novel MCP manufacturing technology that substantially improves performance and opens novel applications. The authors have developed custom tools and recipes for the growth of conformal films, with optimized conductance and secondary electron emission inside very long channels (∼6–20 μm diameter and >600 μm length, with tens of millions of channels per single MCP) by ALD. The unique ability to tune the characteristics of these ALD films enables their optimization to applications where time-resolved single particle imaging can be performed in extreme conditions, such as high counting rates at cryogenic temperatures. Adhesion of the conductive and emissive nanofilms to the 20 μm pore MCP glass substrates and their mechanical stability over a very wide range of temperatures (10–700 K) were confirmed experimentally. Resistance of ALD MCPs was reproducible during multiple cool-down cycles with no film degradation observed. Optimizing resistance of novel MCPs for operation at cryogenic temperature should enable high count rate event detection at temperatures below 20 K.
Introducing many-body physics using atomic spectroscopy
Authors: Krebs Dietrich , Pabst Stefan , Santra Robin
Published: January 2014

Atoms constitute relatively simple many-body systems, making them suitable objects for developing an understanding of basic aspects of many-body physics. Photoabsorption spectroscopy is a prominent method to study the electronic structure of atoms and the inherent many-body interactions. In this article, the impact of many-body effects on well-known spectroscopic features, such as Rydberg series, Fano resonances, Cooper minima, and giant resonances, is studied and related many-body phenomena in other fields are outlined. To calculate photoabsorption cross sections, the time-dependent configuration interaction singles (TDCIS) model is employed. The conceptual clearness of TDCIS in combination with the compactness of atomic systems allows for a pedagogical introduction to many-body phenomena.
Onsager symmetry relations and ideal gas effusion: A detailed example
Authors: Patitsas S. N.
Published: January 2014

Onsager coefficients are calculated for the approach of a gas to equilibrium by effusion between two chambers. Using kinetic gas theory, the Onsager symmetry relation is explicitly verified. The approach to equilibrium is determined by two time scales that are explicitly calculated; this is followed by example calculations for dynamics of the system approaching equilibrium in several ways. Also, calculations for the cross-correlation functions for this system are presented, which are used to calculate various noise spectral functions. This study provides students of statistical mechanics and thermodynamics with a good example to aid in understanding some of the general concepts encountered in studies of non-equilibrium systems.
Sixteen years of collaborative learning through active sense-making in physics (CLASP) at UC Davis
Authors: Potter Wendell , Webb David , Paul Cassandra , West Emily , Bowen Mark , Weiss Brenda , Coleman Lawrence , De Leone Charles
Published: January 2014

This paper describes our large reformed introductory physics course at UC Davis, which bioscience students have been taking since 1996. The central feature of this course is a focus on sense-making by the students during the 5 h per week discussion/labs in which the students take part in activities emphasizing peer-peer discussions, argumentation, and presentations of ideas. The course differs in many fundamental ways from traditionally taught introductory physics courses. After discussing the unique features of CLASP and its implementation at UC Davis, various student outcome measures are presented that show increased performance by students who took the CLASP course compared to students who took a traditionally taught introductory physics course. Measures we use include upper-division GPAs, MCAT scores, FCI gains, and MPEX-II scores.
Unraveling the mystery of the “Maoshan Bugle”
Authors: Chen Xu , Qin Ming , Wang Sihui , Zhou Huijun
Published: January 2014

The mystery of the “Maoshan Bugle,” first classified in 1997, involves the surprising production of clearly audible bugle-like sounds in response to a launched firecracker blast in the square in front of the Southern Jiangsu Victory Monument in Jiangsu, China. In this paper, we analyze the origin of these sounds, attributing them to interference due to six groups of stairs leading to a monument in the square. Our analysis indicates that the stairs act as a reflection grating for the sounds produced by a firecracker blast. Based on Kirchhoff's diffraction formula, we calculate the spectra and the fundamental frequencies of the bugle-like echoes using the parameters of the actual structures. The calculated spectra fits well with the experimental results derived from the audio samples recorded on site.
Single β-Actin mRNA Detection in Neurons Reveals a Mechanism for Regulating Its Translatability
Authors: Buxbaum Adina R. , Wu Bin , Singer Robert H.
Published: January 2014

The physical manifestation of learning and memory formation in the brain can be expressed by strengthening or weakening of synaptic connections through morphological changes. Local actin remodeling underlies some forms of plasticity and may be facilitated by local β-actin synthesis, but dynamic information is lacking. In this work, we use single-molecule in situ hybridization to demonstrate that dendritic β-actin messenger RNA (mRNA) and ribosomes are in a masked, neuron-specific form. Chemically induced long-term potentiation prompts transient mRNA unmasking, which depends on factors active during synaptic activity. Ribosomes and single β-actin mRNA motility increase after stimulation, indicative of release from complexes. Hence, the single-molecule assays we developed allow for the quantification of activity-induced unmasking and availability for active translation. Further, our work demonstrates that β-actin mRNA and ribosomes are in a masked state that is alleviated by stimulation.
Transmissible Dog Cancer Genome Reveals the Origin and History of an Ancient Cell Lineage
Authors: Murchison Elizabeth P. , Wedge David C. , Alexandrov Ludmil B. , Fu Beiyuan , Martincorena Inigo , Ning Zemin , Tubio Jose M. C. , Werner Emma I. , Allen Jan , De Nardi Andrigo Barboza , Donelan Edward M. , Marino Gabriele , Fassati Ariberto , Campbell Peter J. , Yang Fengtang , Burt Austin , Weiss Robin A. , Stratton Michael R.
Published: January 2014

Canine transmissible venereal tumor (CTVT) is the oldest known somatic cell lineage. It is a transmissible cancer that propagates naturally in dogs. We sequenced the genomes of two CTVT tumors and found that CTVT has acquired 1.9 million somatic substitution mutations and bears evidence of exposure to ultraviolet light. CTVT is remarkably stable and lacks subclonal heterogeneity despite thousands of rearrangements, copy-number changes, and retrotransposon insertions. More than 10,000 genes carry nonsynonymous variants, and 646 genes have been lost. CTVT first arose in a dog with low genomic heterozygosity that may have lived about 11,000 years ago. The cancer spawned by this individual dispersed across continents about 500 years ago. Our results provide a genetic identikit of an ancient dog and demonstrate the robustness of mammalian somatic cells to survive for millennia despite a massive mutation burden.
Ancient Aqueous Environments at Endeavour Crater, Mars
Authors: Arvidson R. E. , Squyres S. W. , Bell J. F. , Catalano J. G. , Clark B. C. , Crumpler L. S. , de Souza P. A. , Fairén A. G. , Farrand W. H. , Fox V. K. , Gellert R. , Ghosh A. , Golombek M. P. , Grotzinger J. P. , Guinness E. A. , Herkenhoff K. E. , Jolliff B. L. , Knoll A. H. , Li R. , McLennan S. M. , Ming D. W. , Mittlefehldt D. W. , Moore J. M. , Morris R. V. , Murchie S. L. , Parker T. J. , Paulsen G. , Rice J. W. , Ruff S. W. , Smith M. D. , Wolff M. J.
Published: January 2014

Opportunity has investigated in detail rocks on the rim of the Noachian age Endeavour crater, where orbital spectral reflectance signatures indicate the presence of Fe
+3
-rich smectites. The signatures are associated with fine-grained, layered rocks containing spherules of diagenetic or impact origin. The layered rocks are overlain by breccias, and both units are cut by calcium sulfate veins precipitated from fluids that circulated after the Endeavour impact. Compositional data for fractures in the layered rocks suggest formation of Al-rich smectites by aqueous leaching. Evidence is thus preserved for water-rock interactions before and after the impact, with aqueous environments of slightly acidic to circum-neutral pH that would have been more favorable for prebiotic chemistry and microorganisms than those recorded by younger sulfate-rich rocks at Meridiani Planum.
An Economist's Journey
The Climate Casino
Risk, Uncertainty, and Economics for a Warming World

by William Nordhaus

Yale University Press, New Haven, CT, 2013. 392 pp. $30, £20. ISBN 9780300189773.
Authors: Jaccard Mark
Published: January 2014

Nordhaus offers an economic perspective of the challenges and possible responses to anthropogenic climate change.
A Lifeline for Syria's Science Exiles
Authors: Stone Richard
Published: January 2014

As violence in Syria escalates and the regime increasingly targets academics, an international effort to support Syria's beleaguered scholars with visas, fellowships, and guest appointments is gaining momentum.
An Attenuated Duck Plague Virus (DPV) Vaccine Induces both Systemic and Mucosal Immune Responses To Protect Ducks against Virulent DPV Infection
Authors: Huang Juan , Jia Renyong , Wang Mingshu , Shu Bing , Yu Xia , Zhu Dekang , Chen Shun , Yin Zhongqiong , Chen Xiaoyue , Cheng Anchun
Published: January 2014

Duck plague (DP) is a severe disease caused by DP virus (DPV). Control of the disease is recognized as one of the biggest challenges in avian medicine. Vaccination is an efficient way to control DPV, and an attenuated vaccine is the main routine vaccine. The attenuated DPV vaccine strain CHa is a modified live vaccine, but the systemic and mucosal immune responses induced by this vaccine have been poorly understood. In this study, the immunogenicity and efficacy of the vaccine were evaluated after subcutaneous immunization of ducks. CD4+and CD8+T cells were counted by flow cytometry, and humoral and mucosal Ig antibodies were analyzed by enzyme-linked immunosorbent assay (ELISA). The results showed that high levels of T cells and Ig antibodies were present postimmunization and that there were more CD4+T cells than CD8+T cells. Titers of humoral IgG were higher than those of humoral IgA. Local IgA was found in each sample, whereas local IgG was found only in the spleen, thymus, bursa of Fabricius, harderian gland, liver, bile, and lung. In a protection assay, the attenuated DPV vaccine completely protected ducks against 1,000 50% lethal doses (LD50) of the lethal DPV strain CHv via oral infection. These data suggest that this subcutaneous vaccine elicits sufficient systemic and mucosal immune responses against lethal DPV challenge to be protective in ducks. This study provides broad insights into understanding the immune responses to the attenuated DPV vaccine strain CHa through subcutaneous immunization in ducks.
MyD88- and TRIF-Independent Induction of Type I Interferon Drives Naive B Cell Accumulation but Not Loss of Lymph Node Architecture in Lyme Disease
Authors: Hastey Christine J. , Ochoa Jennine , Olsen Kimberley J. , Barthold Stephen W. , Baumgarth Nicole
Published: January 2014

Rapidly after infection, live
Borrelia burgdorferi
, the causative agent of Lyme disease, is found within lymph nodes, causing rapid and strong tissue enlargement, a loss of demarcation between B cell follicles and T cell zones, and an unusually large accumulation of B cells. We sought to explore the mechanisms underlying these changes, as lymph tissue disruption could be detrimental for the development of robust
Borrelia
-specific immunity. A time course study demonstrated that the loss of the normal lymph node structure was a distinct process that preceded the strong increases in B cells at the site. The selective increases in B cell frequencies were due not to proliferation but rather to cytokine-mediated repositioning of B cells to the lymph nodes, as shown with various gene-targeted and bone marrow irradiation chimeras. These studies demonstrated that
B. burgdorferi
infection induced type I interferon receptor (IFNR) signaling in lymph nodes in a MyD88- and TRIF-independent manner and that type I IFNR indirect signaling was required for the excessive increases of naive B cells at those sites. It did not, however, drive the observed histopathological changes, which occurred independently also from major shifts in the lymphocyte-homing chemokines, CXCL12, CXCL13, and CCL19/21, as shown by quantitative reverse transcription-PCR (qRT-PCR), flow cytometry, and transwell migration experiments. Thus,
B. burgdorferi
infection drives the production of type I IFN in lymph nodes and in so doing strongly alters the cellular composition of the lymph nodes, with potential detrimental effects for the development of robust
Borrelia
-specific immunity.
Targeting Pili in Enterococcal Pathogenesis
Authors: Pinkston Kenneth L. , Singh Kavindra V. , Gao Peng , Wilganowski Nathaniel , Robinson Holly , Ghosh Sukhen , Azhdarinia Ali , Sevick-Muraca Eva M. , Murray Barbara E. , Harvey Barrett R.
Published: January 2014

Passive protection, the administration of antibodies to prevent infection, has garnered significant interest in recent years as a potential prophylactic countermeasure to decrease the prevalence of hospital-acquired infections. Pili, polymerized protein structures covalently anchored to the peptidoglycan wall of many Gram-positive pathogens, are ideal targets for antibody intervention, given their importance in establishing infection and their accessibility to antibody interactions. In this work, we demonstrated that a monoclonal antibody to the major component of
Enterococcus faecalis
pili, EbpC, labels polymerized pilus structures, diminishes biofilm formation, and significantly prevents the establishment of a rat endocarditis infection. The effectiveness of this anti-EbpC monoclonal provides strong evidence in support of its potential as a preventative. In addition, after radiolabeling, this monoclonal identified the site of enterococcal infection, providing a rare example of molecularly specific imaging of an established bacterial infection and demonstrating the versatility of this agent for use in future diagnostic and therapeutic applications.
Francisella tularensis Schu S4 Lipopolysaccharide Core Sugar and O-Antigen Mutants Are Attenuated in a Mouse Model of Tularemia
Authors: Rasmussen Jed A. , Post Deborah M. B. , Gibson Bradford W. , Lindemann Stephen R. , Apicella Michael A. , Meyerholz David K. , Jones Bradley D.
Published: January 2014

The virulence factors mediating
Francisella
pathogenesis are being investigated, with an emphasis on understanding how the organism evades innate immunity mechanisms.
Francisella tularensis
produces a lipopolysaccharide (LPS) that is essentially inert and a polysaccharide capsule that helps the organism to evade detection by components of innate immunity. Using an
F. tularensis
Schu S4 mutant library, we identified strains that are disrupted for capsule and O-antigen production. These serum-sensitive strains lack both capsule production and O-antigen laddering. Analysis of the predicted protein sequences for the disrupted genes (
FTT1236
and
FTT1238c
) revealed similarity to those for
waa
(
rfa
) biosynthetic genes in other bacteria. Mass spectrometry further revealed that these proteins are involved in LPS core sugar biosynthesis and the ligation of O antigen to the LPS core sugars. The 50% lethal dose (LD
50
) values of these strains are increased 100- to 1,000-fold for mice. Histopathology revealed that the immune response to the
F. tularensis
mutant strains was significantly different from that observed with wild-type-infected mice. The lung tissue from mutant-infected mice had widespread necrotic debris, but the spleens lacked necrosis and displayed neutrophilia. In contrast, the lungs of wild-type-infected mice had nominal necrosis, but the spleens had widespread necrosis. These data indicate that murine death caused by wild-type strains occurs by a mechanism different from that by which the mutant strains kill mice. Mice immunized with these mutant strains displayed >10-fold protective effects against virulent type A
F. tularensis
challenge.
Small Colony Variant of Methicillin-Resistant Staphylococcus pseudintermedius ST71 Presenting as a Sticky Phenotype
Authors: Savini Vincenzo , Carretto Edoardo , Polilli Ennio , Marrollo Roberta , Santarone Stella , Fazii Paolo , D'Antonio Domenico , Rossano Alexandra , Perreten Vincent
Published: January 2014

We first observed the phenomenon of small colony variants (SCVs) in a
Staphylococcus pseudintermedius
sequence type 71 (ST71) strain, isolated from a non-pet owner. Although we found that small-sized colonies share main features with
Staphylococcus aureus
SCVs, they nevertheless show a novel, particular, and sticky phenotype, whose expression was extremely stable, even after subcultivation.
Mycobacterium paraffinicum Causing Symptomatic Pulmonary Infection
Authors: Chan Austin W. , Kabbani Sarah , Staton Gerald , Kraft Colleen S.
Published: January 2014

Mycobacterium paraffinicum
has been newly recognized as a species. A case of symptomatic pulmonary infection caused by
M. paraffinicum
is described, and as far as we know, this is the first case of the organism as a human pathogen.
Twenty-Five-Year Outbreak of Pseudomonas aeruginosa Infecting Individuals with Cystic Fibrosis: Identification of the Prairie Epidemic Strain
Authors: Parkins Michael D. , Glezerson Bryan A. , Sibley Christopher D. , Sibley Kristen A. , Duong Jessica , Purighalla Swathi , Mody Christopher H. , Workentine Matthew L. , Storey Douglas G. , Surette Michael G. , Rabin Harvey R.
Published: January 2014

Transmissible strains of
Pseudomonas aeruginosa
have been described for cystic fibrosis (CF) and may be associated with a worse prognosis. Using a comprehensive strain biobank spanning 3 decades, we sought to determine the prevalence and stability of chronic
P. aeruginosa
infection in an adult population.
P. aeruginosa
isolates from sputum samples collected at initial enrollment in our adult clinic and at the most recent clinic visit were examined by a combination of pulsed-field gel electrophoresis and multilocus sequence typing and compared against a collection of established transmissible and local non-CF bronchiectasis (nCFB) isolates. A total of 372 isolates from 107 patients, spanning 674 patient-years, including 66 patients with matched isolates from initial and final encounters, were screened. A novel clone with increased antibacterial resistance, termed the prairie epidemic strain (PES), was found in 29% (31/107 patients) of chronically infected patients referred from multiple prairie-based CF centers. This isolate was not found in those diagnosed with CF as adults or in a control population with nCFB. While 90% (60/66 patients) of patients had stable infection over a mean of 10.8 years, five patients experienced strain displacement of unique isolates, with PES occurring within 2 years of transitioning to adult care. PES has been present in our cohort since at least 1987, is unique to CF, generally establishes chronic infection during childhood, and has been found in patients at the time of transition of patients from multiple prairie-based CF clinics, suggesting broad endemicity. Studies are under way to evaluate the clinical implications of PES infection.
PCR-BasedIn VitroSynthesis of Hepatitis C Virus NS3 Protease for Rapid Phenotypic Resistance Testing of Protease Inhibitors
Authors: Qiao Jinjuan , Yu Junping , Yang Hang , Wei Hongping
Published: January 2014

Protease inhibitors (PIs) targeting the hepatitis C virus (HCV) NS3 protease, such as telaprevir, have significantly improved the sustained virologic response (SVR) rates of HCV genotype 1 antiviral therapy. Given the expanding antiviral therapy regimen, fast HCV PI resistance assays are urgently needed. In this view, we have developed a novel phenotypic resistance test for HCV PIs based onin vitrosynthesis of patient-derived HCV NS3 protease and subsequent enzymatic testing in a fluorescent readout. The enzymatically active HCV NS3 proteases were synthesized from PCR-derived templates by anEscherichia coliS30 extract system. Tests of the protease genes with known mutations for telaprevir resistance showed that the phenotypic resistance test was fast, with a total turnaround time of <10 h, and was fully in agreement with the previous resistance results. The initial tests with 38 treatment-naive serum samples showed that the method was significantly less laborious and faster than currently available phenotypic resistance assays of HCV NS3 PIs.
Comparative Assessment of Automated Nucleic Acid Sample Extraction Equipment for Biothreat Agents
Authors: Kalina Warren Vincent , Douglas Christina Elizabeth , Coyne Susan Rajnik , Minogue Timothy Devin
Published: January 2014

Magnetic beads offer superior impurity removal and nucleic acid selection over older extraction methods. The performances of nucleic acid extraction of biothreat agents in blood or buffer by easyMAG, MagNA Pure, EZ1 Advanced XL, and Nordiag Arrow were evaluated. All instruments showed excellent performance in blood; however, the easyMAG had the best precision and versatility.
Inefficient Type I Interferon-Mediated Antiviral Protection of Primary Mouse Neurons Is Associated with the Lack of Apolipoprotein L9 Expression
Authors: Kreit Marguerite , Paul Sophie , Knoops Laurent , De Cock Aurélie , Sorgeloos Frédéric , Michiels Thomas
Published: January 2014

We examined the antiviral response promoted by type I interferons (IFN) in primary mouse neurons. IFN treatment of neuron cultures strongly upregulated the transcription of IFN-stimulated genes but conferred a surprisingly low resistance to infection by neurotropic viruses such as Theiler's murine encephalomyelitis virus (TMEV) or vesicular stomatitis virus (VSV). Response of primary mouse neurons to IFN treatment was heterogeneous, as many neurons failed to express the typical IFN response marker Mx1 after IFN treatment. This heterogeneous response of primary neurons correlated with a low level of basal expression of IFN-stimulated genes, such asStat1, that are involved in signal transduction of the IFN response. In addition, transcriptomic analysis identified 15 IFN-responsive genes whose expression was low in IFN-treated primary neurons compared to that of primary fibroblasts derived from the same mice (Dhx58,Gvin1,Sp100,Ifi203isoforms 1 and 2,Irgm2,Lgals3bp,Ifi205,Apol9b,Ifi204,Ifi202b,Tor3a,Slfn2,Ifi35,Lgals9). Among these genes, the gene coding for apolipoprotein L9b (Apol9b) displayed antiviral activity against Theiler's virus when overexpressed in L929 cells or in primary neurons. Accordingly, knocking downApol9bexpression in L929 cells increased viral replication. Therefore, we identified a new antiviral protein induced by interferon, ApoL9b, whose lack of expression in primary neurons likely contributes to the high sensitivity of these cells to viral infection.

IMPORTANCEThe type I interferon (IFN) response is an innate immune defense mechanism that is critical to contain viral infection in the host until an adaptive immune response can be mounted. Neurons are a paradigm for postmitotic, highly differentiated cells. Our data show that primary mouse neurons that are exposed to type I interferon remain surprisingly susceptible to viral infection. On one hand, the low level of basal expression of some factors in neurons might prevent a rapid response of these cells. On the other hand, some genes that are typically activated by type I interferon in other cell types are expressed at much lower levels in neurons. Among these genes is the gene encoding apolipoprotein L9, a protein that proved to have antiviral activity against the neurotropic Theiler's murine encephalomyelitis virus. Our data suggest important functional differences in the IFN response mounted by specific cell populations.
Human Cytomegalovirus Infection of Human Embryonic Stem Cell-Derived Primitive Neural Stem Cells Is Restricted at Several Steps but Leads to the Persistence of Viral DNA
Authors: Belzile Jean-Philippe , Stark Thomas J. , Yeo Gene W. , Spector Deborah H.
Published: January 2014

Congenital human cytomegalovirus (HCMV) infection is a major cause of central nervous system structural anomalies and sensory impairments. It is likely that the stage of fetal development, as well as the state of differentiation of susceptible cells at the time of infection, affects the severity of the disease. We used human embryonic stem (ES) cell-derived primitive prerosette neural stem cells (pNSCs) and neural progenitor cells (NPCs) maintained in chemically defined conditions to study HCMV replication in cells at the early stages of neural development. In contrast to what was observed previously using fetus-derived NPCs, infection of ES cell-derived pNSCs with HCMV was nonprogressive. At a low multiplicity of infection, we observed only a small percentage of cells expressing immediate-early genes (IE) and early genes. IE expression was found to be restricted to cells negative for the anterior marker FORSE-1, and treatment of pNSCs with retinoic acid restored IE expression. Differentiation of pNSCs into NPCs restored IE expression but not the transactivation of early genes. Virions produced in NPCs and pNSCs were exclusively cell associated and were mostly non-neural tropic. Finally, we found that viral genomes could persist in pNSC cultures for up to a month after infection despite the absence of detectable IE expression by immunofluorescence, and infectious virus could be produced upon differentiation of pNSCs to neurons. In conclusion, our results highlight the complex array of hurdles that HCMV must overcome in order to infect primitive neural stem cells and suggest that these cells might act as a reservoir for the virus.

IMPORTANCE
Human cytomegalovirus (HCMV) is a betaherpesvirus that is highly prevalent in the population. HCMV infection is usually asymptomatic but can lead to severe consequences in immunosuppressed individuals. HCMV is also the most important infectious cause of congenital developmental birth defects. Manifestations of fetal HCMV disease range from deafness and learning disabilities to more severe symptoms such as microcephaly. In this study, we have used embryonic stem cells to generate primitive neural stem cells and have used these to model HCMV infection of the fetal central nervous system (CNS)
in vitro
. Our results reveal that these cells, which are similar to those present in the developing neural tube, do not support viral replication but instead likely constitute a viral reservoir. Future work will define the effect of viral persistence on cellular functions as well as the exogenous signals leading to the reactivation of viral replication in the CNS.
Ribavirin-Resistant Variants of Foot-and-Mouth Disease Virus: the Effect of Restricted Quasispecies Diversity on Viral Virulence
Authors: Zeng Jianxiong , Wang Haiwei , Xie Xiaochun , Li Chen , Zhou Guohui , Yang Decheng , Yu Li
Published: January 2014

Mutagenic nucleoside analogues can be used to isolate RNA virus high-fidelity RNA-dependent RNA polymerase (RdRp) variants, the majority of which are attenuated
in vivo
. However, attenuated foot-and-mouth disease virus (FMDV) high-fidelity RdRp variants have not been isolated, and the correlations between RdRp fidelity and virulence remain unclear. Here, the mutagen ribavirin was used to select a ribavirin-resistant population of FMDV, and 4 amino acid substitutions (D5N, A38V, M194I, and M296V) were identified in the RdRp-coding region of the population. Through single or combined mutagenesis using a reverse genetics system, we generated direct experimental evidence that the rescued D5N, A38V, and DAMM mutants but not the M194I and M296V mutants are high-fidelity RdRp variants. Mutagen resistance assays revealed that the higher replication fidelity was associated with higher-level resistance to ribavirin. In addition, significantly attenuated fitness and virulence phenotypes were observed for the D5N, A38V, and DAMM mutants. Based on a systematic quantitative analysis of fidelity and virulence, we concluded that higher replication fidelity is associated with a more attenuated virus. These data suggest that the resulting restricted quasispecies diversity compromises the adaptability and virulence of an RNA virus population. The modulation of replication fidelity to attenuate virulence may represent a general strategy for the rational design of new types of live, attenuated vaccine strains.

IMPORTANCE
The ribavirin-isolated poliovirus (PV) RdRp G64S variant, the polymerases of which were of high replication fidelity, was attenuated
in vivo
. It has been proposed (M. Vignuzzi, E. Wendt, and R. Andino, Nat. Med. 14:154–161,
http://dx.doi.org/10.1038/nm1726
) that modulation of replication fidelity is a promising approach for engineering attenuated virus vaccines. The subsequently mutagen-isolated RdRp variants also expressed the high-fidelity polymerase, but not all of them were attenuated. Few studies have shown the exact correlation between fidelity and virulence. The present study investigates the effect of restricted quasispecies diversity on viral virulence via several attenuated FMDV high-fidelity RdRp variants. Our findings may aid in the rational design of a new type of vaccine strain.