bluuebunny/crossref_metadata_2025 huggingface dataset:
Ropivacaine in neonates and infants: a population pharmacokinetic evaluation following single caudal block
Authors: Rapp Hans‐Jürgen , Molnár Valeria , Austin Stephen , Krohn Stefan , Gädeke Veronika , Motsch Johann , Boos Karin , Williams D.G. , Gustafsson Urban , Huledal Gunilla , Larsson Lars E.
Published: August 2004

Background : The aims of this study were to evaluate pharmacokinetics, efficacy and safety of ropivacaine in infants aged 0–12 months following a single caudal injection.

Methods : Term ASA I–III patients, scheduled for surgery, with a body weight of ≥2500 g received a caudal block with ropivacaine 2 mg·ml−1, 1.0 ml·kg−1. Plasma samples were collected at different time intervals up to 30 h, for analysis of total and unbound ropivacaine and alpha‐1‐acid glycoprotein (AAG). Pharmacokinetic data were characterized by population analysis. Unbound and total concentrations from 35 patients, median (min–max) postnatal age of 66 (4–351) days, were included in the nonlinear mixed effects modeling to provide estimates of pharmacokinetic parameters and the exploration of covariate relationships. Simulations were made to test the predictive performance of the final model and to describe the effect of significant covariates on systemic exposure.

Results : The mean (min–max) peak plasma concentration of total ropivacaine was 0.83 (0.05–1.57) mg·l−1 at 0.5–5.7 h (median: 1.0 h) and the plasma concentration of unbound ropivacaine was 0.042 (0.012–0.081) mg·l−1 within 0.5–1 h. The observed unbound fraction in plasma was 6% (1%–14%). A one‐compartment open model with first‐order absorption and elimination, incorporating a linear‐binding model of ropivacaine to AAG best described the data. The only significant covariate relationship was that of age on Clu/F according to the following relationship Clu/F = 3.01 × e0.00474 × Age. This predicts a Clu/F of 3.5 l·h−1·kg−1 at 30 days and 10.8 l·h−1·kg−1 at 270 days with corresponding terminal half‐lives of 6.7 and 2.2 h. The interindividual variability (coefficient of variation, CV) in Clu/F was 39%. The population estimate (CV) of ka was 1.65 h−1 (30%), Vu/F was 33.6 (l·kg−1) (45%) and Ka was 1.78 l·mg−1 (14%). Thirty‐five infants received supplementary analgesics (mostly paracetamol). The median time to first supplementary analgesic (based on all 37 patients) was 3.9 h. No safety concerns or signs of systemic toxicity were observed.

Conclusions : Following a caudal block with ropivacaine 2 mg·kg−1 plasma concentrations of unbound ropivacaine were well below threshold levels for toxicity in adults. Apparent volume of distribution is unchanged, apparent unbound clearance increases and the terminal half‐life decreases with age in 0–12‐month‐old neonates and infants. The postoperative pain management provided adequate analgesia and was well tolerated.
Mental Illness and Length of Inpatient Stay for Medicaid Recipients with AIDS
Authors: Hoover Donald R. , Sambamoorthi Usha , Walkup James T. , Crystal Stephen
Published: August 2004

Objective. To examine the associations between comorbid mental illness and length of hospital stays (LOS) among Medicaid beneficiaries with AIDS.

Data Source and Collection/Study Setting. Merged 1992–1998 Medicaid claims and AIDS surveillance data obtained from the State of New Jersey for adults with ≥1 inpatient stay after an AIDS diagnosis from 1992 to 1996.

Study Design. Observational study of 6,247 AIDS patients with 24,975 inpatient visits. Severe mental illness (SMI) and other less severe mental illness (OMI) diagnoses at visits were ascertained from ICD–9 Codes. About 4 percent of visits had an SMI diagnosis; 5 percent had an OMI diagnosis; 43 percent did not have a mental illness diagnosis, but were patients who had been identified as having an SMI or OMI history; and 48 percent were from patients with no identified history of mental illness.

Principal Findings. The overall mean hospital LOS was 12.7 days. After adjusting for measures of HIV disease severity and health care access in multivariate models, patients presenting with primary and secondary severe mental illness (SMI) diagnoses had ∼32 percent and ∼11 percent longer LOS, respectively, than did similar patients without a mental illness history (p<0.001 for each). But in these adjusted models of length of stay: (1) diagnosis of OMI was not related to LOS, and (2) in the absence of a mental illness diagnosed at the visit, an identified history of either SMI or OMI was also not related to LOS. In adjusted models of time to readmission for a new visit, current diagnosis of SMI or OMI and in the absences of a current diagnosis, history of SMI or OMI all tended to be associated with quicker readmission.

Conclusions. This study finds greater (adjusted) LOS for AIDS patients diagnosed with severe mental illness (but not for those diagnosed with less severe mental comorbidity) at a visit. The effect of acute severe mental illness on hospitalization time may be comparable to that of an acute AIDS opportunistic illness. While previous research raises concerns that mental illness increases LOS by interfering with treatment of HIV conditions, the associations here may simply indicate that extra time is needed to treat severe mental illnesses or arrange for discharge of afflicted patients.
Cost Shifting to Jails after a Change to Managed Mental Health Care
Authors: Domino Marisa Elena , Norton Edward C. , Morrissey Joseph P. , Thakur Neil
Published: August 2004

Objective. To determine whether managed mental health care for Medicaid enrollees in King County, Washington, has led to indirect cost‐shifting to substitute treatments, such as jails and state mental hospitals that are free goods to providers.

Data Sources. Complete service records for 47,300 adults who used at least one of the following systems from 1993 to 1998: King County jail system, Medicaid, or the King County mental health system. Data were also obtained from the Washington State Hospital System.

Study Design. A quasi‐experimental analysis that compares the difference in outcomes between the pre‐ and post‐managed care periods for Medicaid enrollees compared to non‐Medicaid enrollees. The outcomes—jail costs, state hospital costs, and county outpatient mental health costs—were estimated with two‐part difference‐in‐differences models. The regressions control for person‐level fixed effects on up to 66 months of data per person.

Data Collection Methods. Administrative data were collected from the jail, Medicaid, and mental health systems, then merged and cleaned. Additional data on costs were obtained in interviews.

Principal Findings. There is a striking increase in the probability of jail use for persons on Medicaid following the introduction of managed care. There was a significant decrease in expenditures in the county mental health system for outpatient care.

Conclusions. Managed care led to indirect cost‐shifting, probably through poor access to services, which may have led to an increased probability of jail detention.
Communities and Hospitals: Social Capital, Community Accountability, and Service Provision in U.S. Community Hospitals
Authors: Lee Shoou‐Yih D. , Chen Wendy L. , Weiner Bryan J.
Published: August 2004

Objectives.The study related community social capital to the level of community accountability and provision of community‐oriented services in U.S. community hospitals.

Study Setting.The sample included 1,383 community hospitals that participated in the 1997 American Hospital Association's (AHA) Hospital Annual and Governance Surveys.

Data Sources.(1) The 1997 AHA Annual Hospital Survey, (2) the 1997 AHA Hospital Governance Survey, (3) the DDB Needham Market Facts Survey, (4) the 1996 County Election Data File, and (5) the 1998 Area Resource File.

Research Design.The study used a mix of longitudinal and cross‐sectional data.

Key Findings.We identified two distinct indicators of social capital—community participation and voting participation. Community accountability in hospitals was unrelated to either indicator. Hospitals' provision of community‐oriented health services was negatively associated with community participation but unrelated with voting participation. The interaction between voting participation and community representation on hospital governance was positively associated with community accountability and provision of community‐oriented health services.

Conclusion.Neither community participation nor voting participation was sufficient to influence hospital behavior. The positive finding associated with the interaction between voting participation and community representation on hospital governance underscored the importance of an active political culture in influencing hospital behavior, without which the installation of community representatives on hospital governance might be more symbolic than actually serving the health concerns of community residents.
Availability of Safety Net Providers and Access to Care of Uninsured Persons
Authors: Hadley Jack , Cunningham Peter
Published: August 2004

Objective. To understand how proximity to safety net clinics and hospitals affects a variety of measures of access to care and service use by uninsured persons.

Data Sources. The 1998–1999 Community Tracking Study household survey, administered primarily by telephone survey to households in 60 randomly selected communities, linked to data on community health centers, other free clinics, and safety net hospitals.

Study Design. Instrumental variable estimation of multivariate regression models of several measures of access to care (having a usual source of care, unmet or delayed medical care needs, ambulatory service use, and overnight hospital stays) against endogenous measures of distances to the nearest community health center and safety net hospital, controlling for characteristics of uninsured persons and other area characteristics that are related to access to care. The models are estimated with data from a nationally representative sample of uninsured people.

Principal Findings. Shorter distances to the nearest safety net providers increase access to care for uninsured persons. Failure to account for the endogeneity of distance to safety net providers on access to care generally leads to finding little or no safety net effects on access.

Conclusions. Closer proximity to the safety net increases access to care for uninsured persons. However, the improvements in access to care are relatively small compared with similar measures of access to care for insured persons. Modest expansion of the safety net is unlikely to provide a full substitute for insurance coverage expansions.
The Alignment and Blending of Payment Incentives within Physician Organizations
Authors: Robinson James C. , Shortell Stephen M. , Li Rui , Casalino Lawrence P. , Rundall Thomas
Published: August 2004

Objective.To analyze the blend of retrospective (fee‐for‐service, productivity‐based salary) and prospective (capitation, nonproductivity‐based salary) methods for compensating individual physicians within medical groups and independent practice associations (IPAs) and the influence of managed care on the compensation blend used by these physician organizations.

Data Sources.Of the 1,587 medical groups and IPAs with 20 or more physicians in the United States, 1,104 responded to a one‐hour telephone survey, with 627 providing detailed information on physician payment methods.

Study Design.We calculated the distribution of compensation methods for primary care and specialty physicians, separately, in both medical groups and IPAs. Multivariate regression methods were used to analyze the influence of market and organizational factors on the payment method developed by physician organizations for individual physicians.

Principal Findings.Within physician organizations, approximately one‐quarter of physicians are paid on a purely retrospective (fee‐for‐service) basis, approximately one‐quarter are paid on a purely prospective (capitation, nonproductivity‐based salary) basis, and approximately one‐half on blends of retrospective and prospective methods. Medical groups and IPAs in heavily penetrated managed care markets are significantly less likely to pay their individual physicians based on fee‐for‐service than are organizations in less heavily penetrated markets.

Conclusions.Physician organizations rely on a wide range of prospective, retrospective, and blended payment methods and seek to align the incentives faced by individual physicians with the market incentives faced by the physician organization.
A Call for a Collective
Authors: Oreskes Naomi
Published: August 2004

Politics of Nature
How to Bring the Sciences into Democracy.
By Bruno Latour
. Harvard University Press, Cambridge, MA, 2004. 319 pp. $55, £35.95, €50.70. ISBN 0-674-01289-5. Paper, $24.95, £16.95, €23.10. ISBN 0-674-01347-6. Translated by Catherine Porter ().

Rejecting a dichotomy between nature and society, Latour argues for a "political ecology," one that would provide the basis for tackling the problems of environmental damage.
The Neural Basis of Altruistic Punishment
Authors: de Quervain Dominique J.-F. , Fischbacher Urs , Treyer Valerie , Schellhammer Melanie , Schnyder Ulrich , Buck Alfred , Fehr Ernst
Published: August 2004

Many people voluntarily incur costs to punish violations of social norms. Evolutionary models and empirical evidence indicate that such altruistic punishment has been a decisive force in the evolution of human cooperation. We used H
2
15
O positron emission tomography to examine the neural basis for altruistic punishment of defectors in an economic exchange. Subjects could punish defection either symbolically or effectively. Symbolic punishment did not reduce the defector's economic payoff, whereas effective punishment did reduce the payoff. We scanned the subjects' brains while they learned about the defector's abuse of trust and determined the punishment. Effective punishment, as compared with symbolic punishment, activated the dorsal striatum, which has been implicated in the processing of rewards that accrue as a result of goal-directed actions. Moreover, subjects with stronger activations in the dorsal striatum were willing to incur greater costs in order to punish. Our findings support the hypothesis that people derive satisfaction from punishing norm violations and that the activation in the dorsal striatum reflects the anticipated satisfaction from punishing defectors.
A Decktop View of Overfishing
Authors: Ash Caroline
Published: August 2004

Trawler
A Journey Through the North Atlantic.
By Redmond O'Hanlon
. Hamish Hamilton, London, 2003. 352 pp. £20. ISBN 0-241-14014-5. Paper, Penguin, London, 2004. £7.99. ISBN 0-140-27668-8. Forthcoming from Knopf, New York. ISBN 1-4000-4275-5.

This account of trawlermen at work also provides a perspective on developments that now threaten the future of the world's fisheries.
Discrimination among Rhinovirus Serotypes for a Variant ICAM-1 Receptor Molecule
Authors: Xiao Chuan , Tuthill Tobias J. , Kelly Carol M. Bator , Challinor Lisa J. , Chipman Paul R. , Killington Richard A. , Rowlands David J. , Craig Alister , Rossmann Michael G.
Published: August 2004

Intercellular adhesion molecule 1 (ICAM-1) is the cellular receptor for the major group of human rhinovirus serotypes, including human rhinovirus 14 (HRV14) and HRV16. A naturally occurring variant of ICAM-1, ICAM-1
Kilifi
, has altered binding characteristics with respect to different HRV serotypes. HRV14 binds to ICAM-1 only transiently at physiological temperatures but forms a stable complex with ICAM-1
Kilifi
. Conversely, HRV16 forms a stable complex with ICAM-1 but does not bind to ICAM-1
Kilifi
. The three-dimensional structures of HRV14 and HRV16, complexed with ICAM-1, and the structure of HRV14, complexed with ICAM-1
Kilifi
, have been determined by cryoelectron microscopy (cryoEM) image reconstruction to a resolution of approximately 10 Å. Structures determined by X-ray crystallography of both viruses and of ICAM-1 were fitted into the cryoEM density maps. The interfaces between the viruses and receptors contain extensive ionic networks. However, the interactions between the viruses and ICAM-1
Kilifi
contain one less salt bridge than between the viruses and ICAM-1. As HRV16 has fewer overall interactions with ICAM-1 than HRV14, the absence of this charge interaction has a greater impact on the binding of ICAM-1
Kilifi
to HRV16 than to HRV14.
A Recombinant Newcastle Disease Virus (NDV) Expressing VP2 Protein of Infectious Bursal Disease Virus (IBDV) Protects against NDV and IBDV
Authors: Huang Zhuhui , Elankumaran Subbiah , Yunus Abdul S. , Samal Siba K.
Published: August 2004

Infectious bursal disease virus (IBDV) causes a highly immunosuppressive disease in chickens. Currently available, live IBDV vaccines can lead to generation of variant viruses. We have developed an alternative vaccine that will not create variant IBDV. By using the reverse genetics approach, we devised a recombinant Newcastle disease virus (NDV) vector from a commonly used vaccine strain LaSota to express the host-protective immunogen VP2 of a variant IBDV strain GLS-5. The gene encoding the VP2 protein of the IBDV was inserted into the most 3′-proximal locus of a full-length NDV cDNA for high-level expression. We successfully recovered the recombinant virus, rLaSota/VP2. The rLaSota/VP2 was genetically stable, at least up to 12 serial passages in chicken embryos, and was shown to express the VP2 protein. The VP2 protein was not incorporated into the virions of recombinant virus. Recombinant rLaSota/VP2 replicated to a titer similar to that of parental NDV strain LaSota in chicken embryos and cell cultures. To assess protective efficacy of the rLaSota/VP2, 2-day-old specific-pathogen-free chickens were vaccinated with the recombinant virus and challenged with a highly virulent NDV strain Texas GB or IBDV variant strain GLS-5 at 3 weeks postvaccination. Vaccination with rLaSota/VP2 generated antibody responses against both NDV and IBDV and provided 90% protection against NDV and IBDV. Booster immunization induced higher levels of antibody responses against both NDV and IBDV and conferred complete protection against both viruses. These results indicate that the recombinant NDV can be used as a vaccine vector for other avian pathogens.
Cytoplasmic Dynein Mediates Adenovirus Binding to Microtubules
Authors: Kelkar Samir A. , Pfister K. Kevin , Crystal Ronald G. , Leopold Philip L.
Published: August 2004

During infection, adenovirus (Ad) capsids undergo microtubule-dependent retrograde transport as part of a program of vectorial transport of the viral genome to the nucleus. The microtubule-associated molecular motor, cytoplasmic dynein, has been implicated in the retrograde movement of Ad. We hypothesized that cytoplasmic dynein constituted the primary mode of association of Ad with microtubules. To evaluate this hypothesis, an Ad-microtubule binding assay was established in which microtubules were polymerized with taxol, combined with Ad in the presence or absence of microtubule-associated proteins (MAPs), and centrifuged through a glycerol cushion. The addition of purified bovine brain MAPs increased the fraction of Ad in the microtubule pellet from 17.3% ± 3.5% to 80.7% ± 3.8% (P< 0.01). In the absence of tubulin polymerization or in the presence of high salt, no Ad was found in the pellet. Ad binding to microtubules was not enhanced by bovine brain MAPs enriched for tau protein or by the addition of bovine serum albumin. Enhanced Ad-microtubule binding was also observed by using a fraction of MAPs purified from lung A549 epithelial cell lysate which contained cytoplasmic dynein. Ad-microtubule interaction was sensitive to the addition of ATP, a hallmark of cytoplasmic dynein-dependent microtubule interactions. Immunodepletion of cytoplasmic dynein from the A549 cell lysate abolished the MAP-enhanced Ad-microtubule binding. The interaction of Ad with both dynein and dynactin complexes was demonstrated by coimmunoprecipitation. Partially uncoated capsids isolated from cells 40 min after infection also exhibited microtubule binding. In summary, the primary mode of Ad attachment to microtubules occurs though cytoplasmic dynein-mediated binding.
Independent Evolution of Human Immunodeficiency Virus (HIV) Drug Resistance Mutations in Diverse Areas of the Brain in HIV-Infected Patients, with and without Dementia, on Antiretroviral Treatment
Authors: Smit Theresa K. , Brew Bruce J. , Tourtellotte Wallace , Morgello Susan , Gelman Benjamin B. , Saksena Nitin K.
Published: August 2004

AIDS dementia complex (ADC) in human immunodeficiency virus (HIV)-infected patients continues to be a problem in the era of highly active antiretroviral therapy (HAART). A better understanding of the drug resistance mutation patterns that emerge in the central nervous system (CNS) during HAART is of paramount importance as these differences in drug resistance mutations may explain underlying reasons for poor penetration of antiretroviral drugs into the CNS and suboptimal concentrations of the drugs that may reside in the brains of HIV-infected individuals during therapy. Thus, we provide a detailed analysis of HIV type 1 (HIV-1) protease and reverse transcriptase (RT) genes derived from different regions of the brains of 20 HIV-1-infected patients (5 without ADC, 2 with probable ADC, and 13 with various stages of ADC) on antiretroviral therapy. We show the compartmentalization and independent evolution of both primary and secondary drug resistance mutations to both RT and protease inhibitors in diverse regions of the CNS of HIV-infected patients, with and without dementia, on antiretroviral therapy. Our results suggest that the independent evolution of drug resistance mutations in diverse areas of the CNS may emerge as a consequence of incomplete suppression of HIV, probably related to suboptimal drug levels in the CNS and drug selection pressure. The emergence of resistant virus in the CNS may have considerable influence on the outcome of neurologic disease and also the reseeding of HIV in the systemic circulation upon failure of therapy.
Role of Protein Kinase C δ in Reactivation of Kaposi's Sarcoma-Associated Herpesvirus
Authors: Deutsch Einat , Cohen Adina , Kazimirsky Gila , Dovrat Sara , Rubinfeld Hadara , Brodie Chaya , Sarid Ronit
Published: August 2004

TPA (12-
O
-tetradecanoylphorbol-13-acetate), a well-known activator of protein kinase C (PKC), can experimentally induce reactivation of Kaposi's sarcoma-associated herpesvirus (KSHV) in certain latently infected cells. We selectively blocked the activity of PKC isoforms by using GF 109203X or rottlerin and demonstrated that this inhibition largely decreased lytic KSHV reactivation by TPA. Translocation of the PKCδ isoform was evident shortly after TPA stimulation. Overexpression of the dominant-negative PKCδ mutant supported an essential role for the PKCδ isoform in virus reactivation, yet overexpression of PKCδ alone was not sufficient to induce lytic reactivation of KSHV, suggesting that additional signaling molecules participate in this pathway.
Full Resistance of Herpes Simplex Virus Type 1-Infected Primary Human Cells to Alpha Interferon Requires both the Us11 and γ
1
34.5 Gene Products
Authors: Mulvey Matthew , Camarena Vladimir , Mohr Ian
Published: August 2004

The γ
1
34.5 gene product is important for the resistance of herpes simplex virus type 1 (HSV-1) to interferon. However, since the inhibition of protein synthesis observed in cells infected with a γ
1
34.5 mutant virus results from the combined loss of the γ
1
34.5 gene product and the failure to translate the late Us11 mRNA, we sought to characterize the relative interferon sensitivity of mutants unable to produce either the Us11 or the γ
1
34.5 polypeptide. We now demonstrate that primary human cells infected with a Us11 mutant virus are hypersensitive to alpha interferon, arresting translation upon entry into the late phase of the viral life cycle. Furthermore, immediate-early expression of Us11 by a γ
1
34.5 deletion mutant is sufficient to render translation resistant to alpha interferon. Finally, we establish that the Us11 gene product is required for wild-type levels of replication in alpha interferon-treated cells and, along with the γ
1
34.5 gene, is an HSV-1-encoded interferon resistance determinant.
Retroviral Capsid Determinants of
Fv1
NB and NR Tropism
Authors: Stevens Anthony , Bock Michael , Ellis Scott , LeTissier Paul , Bishop Kate N. , Yap Melvyn W. , Taylor Willie , Stoye Jonathan P.
Published: August 2004

The specificity determinants for susceptibility to resistance by the
Fv1
n and b alleles map to amino acid 110 of the murine leukemia virus CA protein. To study the interaction between
Fv1
and CA, we examined changes in CA resulting in the loss of susceptibility to
Fv1
resistance in naturally occurring NB- and NR-tropic viruses. A variety of amino acid changes affecting Fv1 tropism were identified, at CA positions 82, 92 to 95, 105, 114, and 117, and they all were mapped to the apparent exterior of virion-associated CA. These amino acids may form a binding surface for Fv1.
Herpes Simplex Virus Type 1 Regulatory Protein ICP0 Does Not Protect Cyclins D1 and D3 from Degradation during Infection
Authors: Everett Roger D.
Published: August 2004

Previous reports have suggested that herpes simplex virus type 1 (HSV-1) immediate-early regulatory protein ICP0 stabilizes cyclins D1 and D3 during infection by inducing the degradation of cdc34, the E2-conjugating enzyme that is responsible for regulating the stability of these cyclins. Since ICP0 has complex effects on the progress of viral infection that vary greatly with cell type and viral dose, it can be difficult to distinguish between direct effects caused by ICP0 itself and indirect effects caused by the rate of the progression of infection in the absence of ICP0 at the chosen multiplicity of infection. This report describes the fates of cdc34 and cyclins D1 and D3 during HSV-1 infection under conditions that ensured that viral infection and gene expression were proceeding at equivalent rates in the presence and absence of ICP0. It was confirmed that both D-type cyclins were unstable during HSV-1 infection of a variety of cell types, but no effect on cdc34 was observed, even when high levels of ICP0 were expressed. Furthermore, there was no evidence that ICP0 protected either cyclin D1 or cyclin D3 from degradation. Reconstruction of the conditions of the experiments in the previous studies, using the stated cell type and multiplicities of infection, indicated that the original results could be explained by differences in the rate of progression of infection rather than by the presence or absence of ICP0. The data presented in this report are incompatible with the hypothesis that ICP0 induces the degradation of cdc34 and thereby stabilizes cyclins D1 and D3 during HSV-1 infection.
Involvement of a Bovine Viral Diarrhea Virus NS5B Locus in Virion Assembly
Authors: Ansari Israrul H. , Chen Li-Mei , Liang Delin , Gil Laura H. , Zhong Weidong , Donis Ruben O.
Published: August 2004

A novel mutant of bovine viral diarrhea virus (BVDV) was found with a virion assembly phenotype attributable to an insertion into the NS5B polymerase locus. This mutant, termed 5B-741, was engineered by reverse genetics to express NS5B with a C-terminal peptide tag of 22 amino acids. Electroporation of bovine cells with genomic RNA from this mutant showed levels RNA synthesis which were regarded as sufficient for infectivity, yet infectious virions were not produced. Pseudorevertants of mutant 5B-741 that released infectious virions and formed plaques revealed a single nucleotide change (T12369C). This change resulted in a leucine-to-proline substitution within the NS5B tag (L726P). Genetic analysis revealed that indeed a single nucleotide change encoding proline at NS5B position 726 in the pseudorevertant polyprotein mediated recovery of virion assembly function without improving genomic RNA accumulation levels. A subgenomic BVDV reporter replicon (rNS3-5B) was used to analyze the consequences of alterations of the genomic region encoding the NS5B C terminus on replication and assembly. Interestingly, rNS3-5B-L726P (revertant) replicated with the same efficiency as the rNS3-5B-741 mutant but produced 10 times more virions in atrans-packaging assay. These results indicated that impairment of assembly function in 5B-741 was independent of RNA accumulation levels and agreed with the observations from the full-length mutant and revertant genomes. Finally, we recapitulated the packaging defect of 5B-741 with a vaccinia virus expression system to eliminate possible unwanted interactions between the helper virus and the packaged replicon. Taken together, these studies revealed an unexpected role of NS5B in infectious virion assembly.
A Mutant of Sindbis Virus Which Is Able To Replicate in Cells with Reduced CTP Makes a Replicase/Transcriptase with a Decreased

K
m

for CTP
Authors: Li Mei-Ling , Lin Yen-Huei , Simmonds H. Anne , Stollar Victor
Published: August 2004

We reported earlier the isolation and characterization of a Sindbis virus mutant, SV
PZF
, that can grow in mosquito cells treated with pyrazofurin (PZF), a compound that interferes with pyrimidine biosynthesis (Y. H. Lin, P. Yadav, R. Ravatn, and V. Stollar, Virology 272:61-71, 2000; Y. H. Lin, H. A. Simmonds, and V. Stollar, Virology 292:78-86, 2002). Three amino acid changes in nsP4, the viral RNA polymerase, were required to produce this phenotype. We now describe a mutant of Sindbis virus, SV
CPC
, that is resistant to cyclopentenylcytosine (CPC), a compound that interferes only with the synthesis of CTP. Thus, in contrast to SV
PZF
, which was selected for its ability to grow in mosquito cells with low levels of UTP and CTP, SV
CPC
was selected for its ability to grow in cells in which only the level of CTP was reduced. Although SV
PZF
was cross-resistant to CPC, SV
CPC
was not resistant to PZF. Only one amino acid change in nsP4, Leu 585 to Phe, was required for the CPC resistance phenotype. The viral replicase/transcriptase generated in SV
CPC
-infected mosquito cells had a lower

K
m

for CTP (but not for UTP) than did the enzyme made in SV
STD
-infected mosquito cells. SV
PZF
and SV
CPC
represent the first examples of viral mutants selected for the ability to grow in cells with low levels of ribonucleoside triphosphates (rNTPs). Further study of these mutants and determination of the structure of nsP4 should demonstrate how alterations in an RNA-dependent RNA polymerase permit it to function in cells with abnormally low levels of rNTPs.
Genetic Relationships and Evolution of Genotypes of Yellow Fever Virus and Other Members of the Yellow Fever Virus Group within the
Flavivirus
Genus Based on the 3′ Noncoding Region
Authors: Mutebi John-Paul , Rijnbrand René C. A. , Wang Heiman , Ryman Kate D. , Wang Eryu , Fulop Lynda D. , Titball Rick , Barrett Alan D. T.
Published: August 2004

Genetic relationships among flaviviruses within the yellow fever (YF) virus genetic group were investigated by comparing nucleotide sequences of the 3′ noncoding region (3′NCR). Size heterogeneity was observed between members and even among strains of the same viral species. Size variation between YF strains was due to duplications and/or deletions of repeated nucleotide sequence elements (RYF). West African genotypes had three copies of the RYF (RYF1, RYF2, and RYF3); the Angola and the East and Central African genotypes had two copies (RYF1 and RYF3); and South American genotypes had only a single copy (RYF3). Nucleotide sequence analyses suggest a deletion within the 3′NCR of South American genotypes, including RYF1 and RYF2. Based on studies with the French neurotropic vaccine strain, passage of a YF virus strain in cell culture can result in deletion of RYF1 and RYF2. Taken together, these observations suggest that South American genotypes of YF virus evolved from West African genotypes and that the South American genotypes lost RYF1 and RYF2, possibly in a single event. Repeated sequence elements were found within the 3′NCR of other members of the YF virus genetic group, suggesting that it is probably characteristic for members of the YF virus genetic group. A core sequence of 15 nucleotides, containing two stem-loops, was found within the 3′NCR of all members of the YF genetic group and may represent the progenitor repeat sequence. Secondary structure predictions of the 3′NCR showed very similar structures for viruses that were closely related phylogenetically.
Interactions of Foot-and-Mouth Disease Virus with Soluble Bovine α
V
β
3
and α
V
β
6
Integrins
Authors: Duque Hernando , LaRocco Michael , Golde William T. , Baxt Barry
Published: August 2004

At least four members of the integrin family of receptors, α
V
β
1
, α
V
β
3
, α
V
β
6
, and α
V
β
8
, have been identified as receptors for foot-and-mouth disease virus (FMDV) in vitro. Our investigators have recently shown that the efficiency of receptor usage appears to be related to the viral serotype and may be influenced by structural differences on the viral surface (H. Duque and B. Baxt, J. Virol.
77:
2500-2511, 2003). To further examine these differences, we generated soluble α
V
β
3
and α
V
β
6
integrins. cDNA plasmids encoding the individual complete integrin α
V
, β
3
, and β
6
subunits were used to amplify sequences encoding the subunits' signal peptide and ectodomain, resulting in subunits lacking transmembrane and cytoplasmic domains. COS-1 cells were transfected with plasmids encoding the soluble α
V
subunit and either the soluble β
3
or β
6
subunit and labeled with [
35
S]methionine-cysteine. Complete subunit heterodimeric integrins were secreted into the medium, as determined by radioimmunoprecipitation with specific monoclonal and polyclonal antibodies. For the examination of the integrins' biological activities, stable cell lines producing the soluble integrins were generated in HEK 293A cells. In the presence of divalent cations, soluble α
V
β
6
bound to representatives of type A or O viruses, immobilized on plastic dishes, and significantly inhibited viral replication, as determined by plaque reduction assays. In contrast, soluble α
V
β
3
was unable to bind to immobilized virus of either serotype; however, virus bound to the immobilized integrin, suggesting that FMDV binding to α
V
β
3
is a low-affinity interaction. In addition, soluble α
V
β
3
did not neutralize virus infectivity. Incubation of soluble α
V
β
6
with labeled type A
12
or O
1
resulted in a significant inhibition of virus adsorption to BHK cells, while soluble α
V
β
3
caused a low (20 to 30%), but consistent, inhibition of virus adsorption. Virus incubated with soluble α
V
β
6
had a lower sedimentation rate than native virus on sucrose density gradients, but the particles retained all of their structural proteins and still contained bound integrin and, therefore, were not exhibiting characteristics of a picornavirus A particle.
Long-Term Persistence of Infection in Chimpanzees Inoculated with an Infectious Hepatitis C Virus Clone Is Associated with a Decrease in the Viral Amino Acid Substitution Rate and Low Levels of Heterogeneity
Authors: Fernandez Javier , Taylor Deborah , Morhardt Duncan R. , Mihalik Kathleen , Puig Montserrat , Rice Charles M. , Feinstone Stephen M. , Major Marian E.
Published: August 2004

Two chimpanzees, 1535 and 1536, became persistently infected following inoculation with RNA transcripts from cDNA clones of hepatitis C virus (HCV). Analysis of the HCV genomes from both animals showed an accumulation of amino acid substitutions over time. The appearance of substitutions in the envelope genes was associated with increased antienvelope antibody titers. However, extensive mutations were not incorporated into hypervariable region 1 (HVR1). A comparison of the nonsynonymous substitution rate/synonymous substitution rate was made at various time points to analyze selective pressure. The highest level of selective pressure occurred during the acute phase and decreased as the infection continued. The nonsynonymous substitution rate was initially higher than the synonymous substitution rate but decreased over time from 3.3 × 10−3(chimpanzee 1535) and 3.2 × 10−3(chimpanzee 1536) substitutions/site/year at week 26 to 1.4 × 10−3(chimpanzee 1535) and 1.7 × 10−3(chimpanzee 1536) at week 216, while the synonymous substitution rate remained steady at ∼1 × 10−3substitutions/site/year. Analysis of PCR products using single-stranded conformational polymorphism indicated a low level of heterogeneity in the viral genome. The results of these studies confirm that the persistence of infection is not solely due to changes in HVR1 or heterogeneity and that the majority of variants observed in natural infections could not arise simply through mutation during the time period most humans and chimpanzees are observed. These data also indicate that immune pressure and selection continue throughout the chronic phase.
Interspecific Recombination between Two Ruminant Alphaherpesviruses, Bovine Herpesviruses 1 and 5
Authors: Meurens François , Keil Günther M. , Muylkens Benoît , Gogev Sacha , Schynts Frédéric , Negro Sandra , Wiggers Laetitia , Thiry Etienne
Published: August 2004

Homologous recombination between different species of alphaherpesviruses has been described between herpes simplex viruses 1 and 2 but has not yet been observed between other alphaherpesviruses. In the present study we chose to assess to what extent in vitro recombination can occur between members of a well-defined group of closely related viruses such as ruminant alphaherpesviruses. At 24 h after infection of epithelial bovine kidney cells with a double-deleted mutant of bovine herpesvirus 1 (BoHV-1) (containing green fluorescent protein and red fluorescent protein genes) and different ruminant alphaherpesviruses, four types of progeny viruses were detected and distinguished according to their phenotype. Frequent recombination events between identical or different strains of BoHV-1 were observed (up to 30%), whereas only two BoHV-1/BoHV-5 recombinants were identified, and no recombinants between BoHV-1 and less closely related caprine and cervine herpesviruses were detected. Restriction analysis of the genomes of the two BoHV-1/BoHV-5 recombinants showed different genetic backgrounds. One possessed a restriction pattern close to BoHV-1, whereas the other one was close to BoHV-5. This exhaustive analysis of each combination of coinfection in a unique situation of five closely related alphaherpesviruses revealed the importance of a high degree of genetic relatedness and similar parental virus growth kinetics for successful interspecific recombination.
Molecular, Biological, and In Vivo Characterization of the Guinea Pig Cytomegalovirus (CMV) Homologs of the Human CMV Matrix Proteins pp71 (UL82) and pp65 (UL83)
Authors: McGregor Alistair , Liu Fenyong , Schleiss Mark R.
Published: August 2004

We recently identified the genes encoding the guinea pig cytomegalovirus (GPCMV) homologs of the upper and lower matrix proteins of human CMV, pp71 (UL82) and pp65 (UL83), which we designated GP82 and GP83, respectively. Transient-expression studies with a GP82 plasmid demonstrated that the encoded protein targets the nucleus and that the infectivity and plaquing efficiency of cotransfected GPCMV viral DNA was enhanced by GP82. The transactivation function of GP82 was not limited to GPCMV, but was also observed for a heterologous virus, herpes simplex virus type 1 (HSV-1). This was confirmed by its ability to complement the growth of an HSV-1 VP16 transactivation-defective mutant virus in an HSV viral DNA cotransfection assay. Study of a GP82 “knockout” virus (and its attendant rescuant), generated on a GPCMV bacterial artificial chromosome construct, confirmed the essential nature of the gene. Conventional homologous recombination was used to generate a GP83 mutant to examine the role of GP83 in the viral life cycle. Comparison of the one-step growth kinetics of the GP83 mutant (vAM409) and wild-type GPCMV indicated that GP83 protein is not required for viral replication in tissue culture. The role of GP83 in vivo was examined by comparing the pathogenesis of wild-type GPCMV, vAM409, and a control virus, vAM403, in guinea pigs. The vAM409 mutant was significantly attenuated for dissemination in immunocompromised strain 2 guinea pigs, suggesting that the GP83 protein is essential for full pathogenicity in vivo.
Current smoking and the risk of non-fatal myocardial infarction in the WHO MONICA Project populations
Authors: Mähönen M S , McElduff P , Dobson A J , Kuulasmaa K A , Evans A E
Published: August 2004

Background:Cohort studies have shown that smoking has a substantial influence on coronary heart disease mortality in young people. Population based data on non-fatal events have been sparse, however.

Objective:To study the impact of smoking on the risk of non-fatal acute myocardial infarction (MI) in young middle age people.

Methods:From 1985 to 1994 all non-fatal MI events in the age group 35–64 were registered in men and women in the WHO MONICA (multinational monitoring of trends and determinants in cardiovascular disease) project populations (18 762 events in men and 4047 in women from 32 populations from 21 countries). In the same populations and age groups 65 741 men and 66 717 women participated in the surveys of risk factors (overall response rate 72%). The relative risk of non-fatal MI for current smokers was compared with non-smokers, by sex and five year age group.

Results:The prevalence of smoking in people aged 35–39 years who experienced non-fatal MI events was 81% in men and 77% in women. It declined with increasing age to 45% in men aged 60–64 years and 36% in women, respectively. In the 35–39 years age group the relative risk of non-fatal MI for smokers was 4.9 (95% confidence interval (CI) 3.9 to 6.1) in men and 5.3 (95% CI 3.2 to 8.7) in women, and the population attributable fractions were 65% and 55%, respectively.

Conclusions:During the study period more than half of the non-fatal MIs occurring in young middle age people can be attributed to smoking.
Current smoking among young adolescents: assessing school based contextual norms: Table 1
Authors: Pokorny S B , Jason L A , Schoeny M E
Published: August 2004

Objective:To extend research on the relation of school based contextual norms to current smoking among adolescents by using three analytic techniques to test for contextual effects. It was hypothesised that significant contextual effects would be found in all three models, but that the strength of these effects would vary by the statistical rigor of the model.

Design:Three separate analytic approaches were conducted on baseline self report student survey data from a larger study to test the relation between school level perceived peer tobacco use and individual current smoking status.

Participants:A representative sample of 5399 sixth through eighth grade students in 14 midwestern middle schools completed the survey. All enrolled sixth through eighth grade students were eligible to participate in the survey. The student participation rate was 91.4% for the entire sample, and did not differ significantly between the schools (range 82–100%).

Main outcome measure:Thirty day cigarette smoking prevalence.

Results:A level 2 only model based on aggregated individual responses indicated that students in schools with higher average reported peer tobacco use were more likely to be current smokers than students in schools with lower average peer tobacco use. Using a level 1 only model based on individual responses indicated that the effect of school level perceived peer tobacco use on current smoking was significant when individual perceived peer tobacco use was excluded from the model but was non-significant when individual perceived peer tobacco use was added to the model. A multilevel model also indicated that the effect of school level perceived peer tobacco use on current smoking was not significant when individual perceived peer tobacco use was added to the model.

Conclusion:The analytic approach used to examine contextual effects using individuals’ reports of peer tobacco use norms that were aggregated to obtain a context measure of the school norms may produce statistical artefacts that distort the association of the school context in general, and peer tobacco use norms in particular, with increased risk for current smoking beyond the risk associated with individual factors.
Recent trends in home and work smoking bans
Authors: Levy D T , Romano E , Mumford E A
Published: August 2004

Objectives:Home and work smoking bans at the national and state level in the USA, and their relation to smoking prevalence and to tobacco control policies, are examined.

Data:The Current Population Survey’s 1992/93 and 1998/99 tobacco use supplement surveys are the primary data source, supplemented with information on state level tobacco control policies.

Methods:The national and state rate of bans are estimated, and changes over the course of the 1990s and their relation to smoking rates and tobacco control policies are examined.

Results:The prevalence of work and home bans has increased considerably between 1992/93 and 1998/99. By 1999, over 65% of the population age 15 and above work in places with smoking bans, and over 60% live in homes with bans. We found that states with lower than average ban rates in 1993 tended to have had larger increases in ban rates between 1993 and 1999. We also found that home and work bans became more prevalent in states with initially low smoking rates, and that the growth in home bans coincided with a declining prevalence of smoking. States with higher levels of bans by 1999 also tended to have higher cigarette taxes, stricter clean air laws, and media/comprehensive campaigns in place.

Conclusions:The results indicate that lower smoking rates are associated with higher rates of work and home bans, although substantial progress has also been made by those states with initially low rates of bans. While further work is needed to establish the direction of causality, it would appear that reductions in smoking rates, either through stronger tobacco control policies or otherwise, may reduce exposure to tobacco smoke not only by reducing the number of smokers, but also through increasing the number of firms and homes with smoking restrictions.
The impact of anti-tobacco industry prevention messages in tobacco producing regions: evidence from the US truth® campaign
Authors: Thrasher J F , Niederdeppe J , Farrelly M C , Davis K C , Ribisl K M , Haviland M L
Published: August 2004

Background:Adolescents who live in tobacco producing regions may not respond favourably to anti-industry ads.

Objective:To examine whether state level involvement in tobacco production appears to limit the effectiveness of anti-industry ads to prevent tobacco use among adolescents in the USA.

Design:Time trend analyses were done using repeated cross sectional data from six waves of the Legacy Media Tracking Survey, which were collected between 1999 and 2003.

Setting and participants:28 307 adolescents, ages 12–17 years, were classified as living in: tobacco producing states (TPS) (n  =  1929); non-tobacco producing states (non-TPS) with low tobacco control funding comparable to TPS (n  =  5323); non-TPS with relatively high funding (n  =  15 076); and non-TPS with established anti-industry ad campaigns (n  =  5979).

Main outcome measures:Reactions to anti-industry ads; strength of anti-industry attitudes/beliefs; changes in anti-industry attitudes/beliefs over time.

Results:Ad reactions did not differ by state type. Multivariate adjusted time trend analyses indicated significant, comparable increases in anti-industry attitudes/beliefs since the onset of the truth® campaign, in both TPS and non-TPS. Mediation analyses indicated that these increases were due, in part, to campaign exposure.

Conclusions:Adolescents who live in tobacco producing regions appear to be as responsive to anti-industry ads as their counterparts in non-tobacco producing regions. This study provides further evidence for the effectiveness of such ads.