bluuebunny/crossref_metadata_2025_split huggingface dataset:
PB2 Mutations D701N and S714R Promote Adaptation of an Influenza H5N1 Virus to a Mammalian Host
Authors: Czudai-Matwich Volker , Otte Anna , Matrosovich Mikhail , Gabriel Gülsah , Klenk Hans-Dieter 
Published: June 2014

ABSTRACT
Mutation D701N in the PB2 protein is known to play a prominent role in the adaptation of avian influenza A viruses to mammalian hosts. In contrast, little is known about the nearby mutations S714I and S714R, which have been observed in some avian influenza viruses highly pathogenic for mammals. We have generated recombinant H5N1 viruses with PB2 displaying the avian signature 701D or the mammalian signature 701N and serine, isoleucine, and arginine at position 714 and compared them for polymerase activity and virus growth in avian and mammalian cells, as well as for pathogenicity in mice. Mutation D701N led to an increase in polymerase activity and replication efficiency in mammalian cells and in mouse pathogenicity, and this increase was significantly enhanced when mutation D701N was combined with mutation S714R. Stimulation by mutation S714I was less distinct. These observations indicate that PB2 mutation S714R, in combination with the mammalian signature at position 701, has the potential to promote the adaptation of an H5N1 virus to a mammalian host.

IMPORTANCE
Influenza A/H5N1 viruses are avian pathogens that have pandemic potential, since they are spread over large parts of Asia, Africa, and Europe and are occasionally transmitted to humans. It is therefore of high scientific interest to understand the mechanisms that determine the host specificity and pathogenicity of these viruses. It is well known that the PB2 subunit of the viral polymerase is an important host range determinant and that PB2 mutation D701N plays an important role in virus adaptation to mammalian cells. In the present study, we show that mutation S714R is also involved in adaptation and that it cooperates with D701N in exposing a nuclear localization signal that mediates importin-α binding and entry of PB2 into the nucleus, where virus replication and transcription take place.
HIV-1 Vpr Redirects Host Ubiquitination Pathway
Authors: Arora Sakshi , Verma Sachin , Banerjea Akhil C. 
Published: June 2014

ABSTRACTHIV-1 modulates key host cellular pathways for successful replication and pathogenesis through viral proteins. By evaluating the hijacking of the host ubiquitination pathway by HIV-1 at the whole-cell level, we now show major perturbations in the ubiquitinated pool of the host proteins post-HIV-1 infection. Our overexpression- and infection-based studies of T cells with wild-type and mutant HIV-1 proviral constructs showed that Vpr is necessary and sufficient for reducing whole-cell ubiquitination. Mutagenic analysis revealed that the three leucine-rich helical regions of Vpr are critical for this novel function of Vpr, which was independent of its other known cellular functions. We also validated that this effect of Vpr was conserved among different subtypes (subtypes B and C) and circulating recombinants from Northern India. Finally, we establish that this phenomenon is involved in HIV-1-mediated diversion of host ubiquitination machinery specifically toward the degradation of various restriction factors during viral pathogenesis.IMPORTANCEHIV-1 is known to rely heavily on modulation of the host ubiquitin pathway, particularly for counteraction of antiretroviral restriction factors, i.e., APOBEC3G, UNG2, and BST-2, etc.; viral assembly; and release. Reports to date have focused on the molecular hijacking of the ubiquitin machinery by HIV-1 at the level of E3 ligases. Interaction of a viral protein with an E3 ligase alters its specificity to bring about selective protein ubiquitination. However, in the case of infection, multiple viral proteins can interact with this multienzyme pathway at various levels, making it much more complicated. Here, we have addressed the manipulation of ubiquitination at the whole-cell level post-HIV-1 infection. Our results show that HIV-1 Vpr is necessary and sufficient to bring about the redirection of the host ubiquitin pathway toward HIV-1-specific outcomes. We also show that the three leucine-rich helical regions of Vpr are critical for this effect and that this ability of Vpr is conserved across circulating recombinants. Our work, the first of its kind, provides novel insight into the regulation of the ubiquitin system at the whole-cell level by HIV-1.
Isolation and Characterization of the DNA and Protein Binding Activities of Adenovirus Core Protein V
Authors: Pérez-Vargas Jimena , Vaughan Robert C. , Houser Carolyn , Hastie Kathryn M. , Kao C. Cheng , Nemerow Glen R. 
Published: June 2014

ABSTRACT

The structure of adenovirus outer capsid was revealed recently at 3- to 4-Å resolution (V. Reddy, S. Natchiar, P. Stewart, and G. Nemerow, Science 329:1071–1075, 2010,
http://dx.doi.org/10.1126/science.1187292
); however, precise details on the function and biochemical and structural features for the inner core still are lacking. Protein V is one the most important components of the adenovirus core, as it links the outer capsid via association with protein VI with the inner DNA core. Protein V is a highly basic protein that strongly binds to DNA in a nonspecific manner. We report the expression of a soluble protein V that exists in monomer-dimer equilibrium. Using reversible cross-linking affinity purification in combination with mass spectrometry, we found that protein V contains multiple DNA binding sites. The binding sites from protein V mediate heat-stable nucleic acid associations, with some of the binding sites possibly masked in the virus by other core proteins. We also demonstrate direct interaction between soluble proteins V and VI, thereby revealing the bridging of the inner DNA core with the outer capsid proteins. These findings are consistent with a model of nucleosome-like structures proposed for the adenovirus core and encapsidated DNA. They also suggest an additional role for protein V in linking the inner nucleic acid core with protein VI on the inner capsid shell.

IMPORTANCE
Scant knowledge exists of how the inner core of adenovirus containing its double-stranded DNA (dsDNA) genome and associated proteins is organized. Here, we report a purification scheme for a recombinant form of protein V that allowed analysis of its interactions with the nucleic acid core region. We demonstrate that protein V exhibits stable associations with dsDNA due to the presence of multiple nucleic acid binding sites identified both in the isolated recombinant protein and in virus particles. As protein V also binds to the membrane lytic protein VI molecules, this core protein may serve as a bridge from the inner dsDNA core to the inner capsid shell.
Functional Analyses of the Three Simian Hemorrhagic Fever Virus Nonstructural Protein 1 Papain-Like Proteases
Authors: Vatter Heather A. , Di Han , Donaldson Eric F. , Radu Gertrud U. , Maines Taronna R. , Brinton Margo A. 
Published: June 2014

ABSTRACT

The N-terminal region of simian hemorrhagic fever virus (SHFV) nonstructural polyprotein 1a is predicted to encode three papain-like proteases (PLP1α, PLP1β, and PLP1γ). Catalytic residues and cleavage sites for each of the SHFV PLP1s were predicted by alignment of the SHFV PLP1 region sequences with each other as well as with those of other arteriviruses, and the predicted catalytic residues were shown to be proximal by homology modeling of the SHFV nsp1s on porcine respiratory and reproductive syndrome virus (PRRSV) nsp1 crystal structures. The functionality of the predicted catalytic Cys residues and cleavage sites was tested by analysis of the autoproteolytic products generated in
in vitro
transcription/translation reactions done with wild-type or mutant SHFV nsp1 constructs. Cleavage sites were also analyzed by mass spectroscopy analysis of selected immunoprecipitated cleavage products. The data showed that each of the three SHFV PLP1s is an active protease. Cys63 was identified as the catalytic Cys of SHFV PLP1α and is adjacent to an Ala instead of the canonical Tyr observed in other arterivirus PLP1s. SHFV PLP1γ is able to cleave at both downstream and upstream nsp1 junction sites. Although intermediate precursor polyproteins as well as alternative products generated by each of the SHFV PLP1s cleaving at sites within the N-terminal region of nsp1β were produced in the
in vitro
reactions, Western blotting of SHFV-infected, MA104 cell lysates with SHFV nsp1 protein-specific antibodies detected only the three mature nsp1 proteins.

IMPORTANCE
SHFV is unique among arteriviruses in having three N-terminal papain-like protease 1 (PLP1) domains. Other arteriviruses encode one or two active PLP1s. This is the first functional study of the SHFV PLP1s. Analysis of the products of
in vitro
autoprocessing of an N-terminal SHFV nonstructural 1a polypeptide fragment showed that each of the three SHFV PLP1s is active, and the predicted catalytic Cys residues and cleavage sites for each PLP1 were confirmed by testing mutant constructs. Several unique features of the SHFV PLP1s were discovered. The SHFV PLP1α catalytic Cys63 is unique among arterivirus PLP1s in being adjacent to an Ala instead of a Trp. Other arterivirus PLP1s cleave only in
cis
at a single downstream site, but SHFV PLP1γ can cleave at both the downstream nsp1γ-nsp2 and upstream nsp1β-nsp1γ junctions. The three mature nsp1 proteins were produced both in the
in vitro
reactions and in infected cells.
Avian Influenza H7N9/13 and H7N7/13: a Comparative Virulence Study in Chickens, Pigeons, and Ferrets
Authors: Kalthoff Donata , Bogs Jessica , Grund Christian , Tauscher Kerstin , Teifke Jens P. , Starick Elke , Harder Timm , Beer Martin 
Published: June 2014

ABSTRACT
Human influenza cases caused by a novel avian H7N9 virus in China emphasize the zoonotic potential of that subtype. We compared the infectivity and pathogenicity of the novel H7N9 virus with those of a recent European avian H7N7 strain in chickens, pigeons, and ferrets. Neither virus induced signs of disease despite substantial replication in inoculated chickens and rapid transmission to contact chickens. Evidence of the replication of both viruses in pigeons, albeit at lower levels of RNA excretion, was also detected. No clear-cut differences between the two H7 isolates emerged regarding replication and antibody development in avian hosts. In ferrets, in contrast, greater replication of the avian H7N9 virus than of the H7N7 strain was observed with significant differences in viral presence, e.g., in nasal wash, lung, and cerebellum samples. Importantly, both viruses showed the potential to spread to the mammal brain. We conclude that efficient asymptomatic viral replication and shedding, as shown in chickens, facilitate the spread of H7 viruses that may harbor zoonotic potential. Biosafety measures are required for the handling of poultry infected with avian influenza viruses of the H7 subtype, independently of their pathogenicity for gallinaceous poultry.

IMPORTANCE
This study is important to the field since it provides data about the behavior of the novel H7N9 avian influenza virus in chickens, pigeons, and ferrets in comparison with that of a recent low-pathogenicity H7N7 strain isolated from poultry. We clearly show that chickens, but not pigeons, are highly permissive hosts of both H7 viruses, allowing high-titer replication and virus shedding without any relevant clinical signs. In the ferret model, the potential of both viruses to infect mammals could be demonstrated, including infection of the brain. However, the replication efficiency of the H7N9 virus in ferrets was higher than that of the H7N7 strain. In conclusion, valuable data for the risk analysis of low-pathogenicity avian influenza viruses of the H7 subtype are provided that could also be used for the risk assessment of zoonotic potentials and necessary biosafety measures.
Hepatitis C Virus Entry Is Impaired by Claudin-1 Downregulation in Diacylglycerol Acyltransferase-1-Deficient Cells
Authors: Sung Pil Soo , Murayama Asako , Kang Wonseok , Kim Myung-Sun , Yoon Seung Kew , Fukasawa Masayoshi , Kondoh Masuo , Kim Jin-Soo , Kim Hyongbum , Kato Takanobu , Shin Eui-Cheol 
Published: June 2014

ABSTRACT

Diacylglycerol acyltransferase-1 (DGAT1) is involved in the assembly of hepatitis C virus (HCV) by facilitating the trafficking of the HCV core protein to the lipid droplet. Here, we abrogated DGAT1 expression in Huh-7.5 cells by using either the transcription activator-like effector nuclease (TALEN) or lentivirus vector short hairpin RNA (shRNA) and achieved complete long-term silencing of DGAT1. HCV entry was severely impaired in DGAT1-silenced Huh-7.5 cell lines, which showed markedly diminished claudin-1 (CLDN1) expression. In DGAT1-silenced cell lines, the forced expression of CLDN1 restored HCV entry, implying that the downregulation of CLDN1 is a critical factor underlying defective HCV entry. The expression of the gene coding for hepatocyte nuclear factor 4α (HNF4α) and other hepatocyte-specific genes was also reduced in DGAT1-silenced cell lines. After
DGAT1
gene rescue, CLDN1 expression was preserved, and HCV entry was restored. Strikingly, after DGAT1 silencing, CLDN1 expression and HCV entry were also restored by low-dose palmitic acid treatment, indicating that the downregulation of CLDN1 was associated with altered fatty acid homeostasis in the absence of DGAT1. Our findings provide novel insight into the role of DGAT1 in the life cycle of HCV.

IMPORTANCE
In this study, we report the novel effect of complete silencing of DGAT1 on the entry of HCV. DGAT1 was recently reported as a host factor of HCV, involved in the assembly of HCV by facilitating the trafficking of the HCV core protein to lipid droplets. We achieved complete and long-term silencing of DGAT1 by either TALEN or repeated transduction of lentivirus shRNA. We found that HCV entry was severely impaired in DGAT1-silenced cell lines. The impairment of HCV entry was caused by CLDN1 downregulation, and the expression of HNF4α and other hepatocyte-specific genes was also downregulated in DGAT1-silenced cell lines. Our results suggest new roles of DGAT1 in human liver-derived cells: maintaining intracellular lipid homeostasis and affecting HCV entry by modulating CLDN1 expression.
Changes in addressing inequalities in access to hospital care in Andhra Pradesh and Maharashtra states of India: a difference-in-differences study using repeated cross-sectional surveys
Authors: Rao Mala , Katyal Anuradha , Singh Prabal V , Samarth Amit , Bergkvist Sofi , Kancharla Manjusha , Wagstaff Adam , Netuveli Gopalakrishnan , Renton Adrian 
Published: June 2014

ObjectivesTo compare the effects of the Rajiv Aarogyasri Health Insurance Scheme of Andhra Pradesh (AP) with health financing innovations including the Rashtriya Swasthya Bima Yojana (RSBY) in Maharashtra (MH) over time on access to and out-of-pocket expenditure (OOPE) on hospital inpatient care.Study designA difference-in-differences (DID) study using repeated cross-sectional surveys with parallel control.SettingNational Sample Survey Organisation of India (NSSO) urban and rural ‘first stratum units’, 863 in AP and 1008 in MH.MethodsWe used two cross-sectional surveys: as a baseline, the data from the NSSO 2004 survey collected before the Aarogyasri and RSBY schemes were launched; and as postintervention, a survey using the same methodology conducted in 2012.Participants8623 households in AP and 10 073 in MH.Main outcome measuresAverage OOPE, large OOPE and large borrowing per household per year for inpatient care, hospitalisation rate per 1000 population per year.ResultsAverage expenditure, large expenditures and large borrowings on inpatient care had increased in MH and AP, but the increase was smaller in AP across these three measures. DIDs for average expenditure and large borrowings were significant and in favour of AP for the rural and the poorest households. Hospitalisation rates also increased in both states but more so in AP, although the DID was not significant and the subgroup analysis presented a mixed picture.ConclusionsHealth innovations in AP had a greater beneficial effect on inpatient care-related expenditures than innovations in MH. The Aarogyasri scheme is likely to have contributed to these impacts in AP, at least in part. However, OOPE increased in both states over time. Schemes such as the Aarogyasri and RSBY may result in some positive outcomes, but additional interventions may be required to improve access to care for the most vulnerable sections of the population.
Conhecimentos sobre saúde ocular entre profissionais de um hospital universitário
Authors: Temporini Edméa Rita , Kara-José Newton , Gondim Éverton Lima , Dantas Fernando Justino 
Published: June 2014

Observacional exploratório. OBJETIVO: Verificar conhecimentos sobre saúde ocular entre profissionais de diferentes categorias e áreas de especialidade em um hospital universitário. MÉTODOS: Aplicou-se questionário não-estruturado sobre afecções oculares (catarata, glaucoma e retinopatia diabética) e conduta em queimadura química ocular. RESULTADOS: A amostra foi formada por 122 sujeitos: 30 enfermeiros (24,6%), 52 auxiliares de enfermagem (42,6%), 24 auxiliares administrativos (19,7%) e 16 funcionários do serviço de oftalmologia (13,1%) que se dispuseram a participar da pesquisa. Afirmaram saber o que é catarata 83,3% dos enfermeiros e 75,0% do pessoal de oftalmologia. Entre os grupos não atuantes em oftalmologia observou-se definição incorreta dessa afecção. Resumo . Acrescentar: . O glaucoma é desconhecido por 79,1% dos funcionários administrativos do hospital; 37,6% do pessoal do serviço de oftalmologia declaram não saber a respeito de cura ou controle dessa afecção. O desconhecimento acerca das conseqüências do diabetes para o sistema visual foi observado em todos os grupos. CONCLUSÃO: O desconhecimento a respeito das afecções oculares ocorreu em todas as categorias. Sugere-se o desenvolvimento de programas de treinamento de funcionários hospitalares, visando à multiplicação de orientações à clientela e no meio sóciocultural.
Resultados visuais no tratamento cirúrgico da subluxação de cristalino em crianças
Authors: Oliveira Denise Fornazari , Marchi Pollyanna Hueb , Arieta Carlos Eduardo Leite 
Published: June 2014

OBJETIVO: descrever os resultados das cirurgias de facectomia via pars plana realizadas em 40 pacientes (71 olhos) com sudescrevebluxação de cristalino. MATERIAL E MÉTODOS: quarenta pacientes com subluxação de cristalino, submetidos a facectomia-vitrectomia via pars plana, no período de Janeiro de 1990 a Dezembro de 2000, foram avaliados quanto ao diagnóstico sistêmico, doenças oculares associadas, idade do paciente na cirurgia, acuidade visual pré e pós-operatória, e complicações da cirurgia. As indicações para a cirurgia foram acuidade visual menor ou igual a 0,3 ou complicações relacionadas a subluxação. RESULTADOS: o seguimento pós-operatório variou de 6 a 120 meses (média 42,3 meses). A acuidade visual melhorou em cinqüenta e sete olhos (80,3%), e em quarenta e sete olhos (66,2%) foi melhor do que 0,5. Em quatro olhos, a acuidade manteve-se a mesma do pré-operatório, e um olho teve piora da acuidade. Complicações ocorreram em dois olhos: um paciente teve descolamento de retina no pós-operatório e outro apresentou sangramento intravítreo no trans-operatório. CONCLUSÕES: Os resultados da série e o tempo de seguimento dos pacientes demonstram que a facectomia-vitrectomia,em um sistema fechado, pode ser um procedimento seguro e efetivo no tratamento da subluxação de cristalino, em crianças, quando o erro refrativo não pode ser corrigido por outros meios, com melhora da acuidade visual e baixo número de complicações.
Necessidades no cuidado hospitalar do lesado medular
Authors: Sartori Nely R. , Melo Márcia RAC 
Published: June 2014

Modelo de estudo: Estudo retrospectivo e documental. Objetivo: Identificar a incidência das complicações e quais as ações assistenciais a serem desenvolvidas pela equipe de enfermagem ao paciente com lesão medular. Método: Foram analisados os prontuários dos pacientes com lesão medular, segundo a classificação do CID 10, no período compreendido entre 1997 a 2000; os dados foram obtidos através do Serviço de Arquivo Médico (SAME) da instituição, a partir da solicitação de um relatório de morbidade; elaborou-se um instrumento que continha a identificação do paciente, dados básicos da história clínica e as alterações fisiológicas apresentadas, bem como os dias após a internação em que elas ocorreram e quais os cuidados prestados pela equipe de enfermagem; os dados foram agrupados e analisados através de tabela e quadro. Observou-se que os pacientes com lesão medular ficam realmente susceptíveis às complicações, como infecções, bexiga neurogênica, íleo paralítico, priapismo, trombose venosa profunda, entre outras; também foi constatada, através das anotações da equipe de enfermagem, a inexistência de cuidados específicos a tais complicações, ocorrendo apenas os cuidados básicos e as ações de natureza propedêutica e terapêutica complementares ao ato médico. Neste estudo, também se observou que as anotações das orientações da alta hospitalar, não apareceram em seis prontuários, e que, em nove, as anotações não eram especificas aos cuidados que o paciente e seus cuidadores necessitariam em seu domicilio a fim de prevenir novas intercorrências. Considerações: A realização deste estudo possibilitou-nos uma visão mais ampla do contexto que envolve o cuidado ao paciente com lesão medular. Acredita-se ser importante um programa de educação continuada em enfermagem, de modo a garantir a atualização do conhecimento para toda a equipe, das necessidades de cada paciente e etapa pela qual está passando.
Niveles de apoproteina-B y LDL- colesterol en una población joven de la ciudad de maracaibo
Authors: Bonezzi S Eleonora , Casanova R Ilya , Gomez G María , Casanova Angel , Villalobos Ana Ortega 
Published: June 2014

El presente estudio se propuso determinar si las concentraciones de apoproteína-B (g/L) se correlacionan con el LDL-colesterol (mg/dL) y al mismo tiempo, investigar la posibilidad de aplicarlas a la población de Maracaibo, en substitución de la fórmula de Friedewald de LDL-Colesterol. Las concentraciones séricas de la apoproteína-B fueron estudiadas en una muestra de 41 sujetos (27 hombres y 14 mujeres) con edades entre 20 y 39 años, usuarios del Laboratorio Clínico de la Escuela de Bioánalisis de La Universidad del Zulia, de la ciudad de Maracaibo. Se realizaron mediciones de glicemia, colesterol total, triacilglicéridos y el cálculo de LDL-C. La apoproteína-B fue determinada por un método inmunoturbidimétrico. Los valores de apoproteína-B ± DS fueron de 0.93 ± 0.3 g/L en el sexo femenino y de 1.06 ± 0.31 g/L en el sexo masculino. La correlación entre la apoproteína-B y el LDL-C fue de 0.8640. Al aplicar el análisis de regresión linear se obtuvieron fórmulas para el cálculo del LCL-C a partir de la concentración de apoproteína-B, representando un cálculo estadístico que deberá ser corroborado de manera experimental. El valor de riesgo de apoproteína -B
³ 1.2 g/L se ubicó en los percentiles 95 th para para el sexo femenino y 75 th para el masculino.