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1.
BMC Infect Dis ; 19(1): 595, 2019 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-31288749

RESUMO

BACKGROUND: Noroviruses (NVs) are an important cause of acute gastroenteritis (AGE) worldwide. There are limited data on the prevalence and molecular characterization of NVs in children in Hohhot, China. METHODS: Between January 2012 and December 2017, 1863 stool samples were collected at Maternal and Child Health Hospital in Hohhot. All samples were screened for NVs by real-time reverse transcription polymerase chain reaction (real-time RT-PCR). RESULTS: NVs were detected in 24.15% of these inpatient cases, ranging from 12.78 to 32.92% in different years. NV was detected throughout the year, with a peak in winter. Based on sequence analysis of the partial VP1 gene, the 306 identified NV strains were divided into six genotypes: GII.3 (71.24%), GII.4 (23.53%), and GII.2, GII.5, GII.6, and GII.13 (total 5.23%). Based on further sequence analysis of the RNA-dependent RNA polymerase (RdRp), GII.P12/GII.3, GII.Pe/GII.4, and GII.P4/GII.4 were identified as predominant genotypes, accounting for 92.6% of genotyped strains. The median age of the children with NV infection was 8.0 (range 0-59) months. However, children infected with GII.3 were younger (median 7.0 months) than GII.4-positive patients (median 10.0 months). CONCLUSION: NV contributed greatly to AGE among hospitalized children in Hohhot in China. Continuous surveillance is important for understanding the local prevalence and characterization of NV.


Assuntos
Infecções por Caliciviridae/diagnóstico , Gastroenterite/diagnóstico , Norovirus/genética , Doença Aguda , Infecções por Caliciviridae/epidemiologia , Criança Hospitalizada , Pré-Escolar , China/epidemiologia , Fezes/virologia , Feminino , Gastroenterite/epidemiologia , Genótipo , Humanos , Lactente , Recém-Nascido , Masculino , Norovirus/classificação , Norovirus/isolamento & purificação , Filogenia , Prevalência , RNA Viral/isolamento & purificação , RNA Viral/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Estações do Ano
2.
BMC Infect Dis ; 19(1): 613, 2019 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-31299924

RESUMO

BACKGROUND: The Respiratory Syncytial Virus (RSV) A genotype ON1, which was first detected in Ontario (Canada) in 2010/11, appeared in Germany in 2011/12. Preliminary observations suggested a higher clinical severity in children infected with this new genotype. We investigated spread and disease severity of RSV-A ON1 in pediatric in- and outpatient settings. METHODS: During 2010/11 to 2016/17, clinical characteristics and respiratory samples from children with acute respiratory tract infections (RTI) were obtained from ongoing surveillance studies in 33 pediatric practices (PP), one pediatric hospital ward (PW) and 23 pediatric intensive care units (PICU) in Germany. RSV was detected in the respiratory samples by PCR; genotypes were identified by sequencing. Within each setting, clinical severity markers were compared between RSV-A ON1 and RSV-A non-ON1 genotypes. RESULTS: A total of 603 children with RSV-RTI were included (132 children in PP, 288 in PW, and 183 in PICU). Of these children, 341 (56.6%) were infected with RSV-A, 235 (39.0%) with RSV-B, and one child (0.2%) with both RSV-A and RSV-B; in 26 (4.3%) children, the subtype could not be identified. In the 341 RSV-A positive samples, genotype ON1 was detected in 247 (72.4%), NA1 in 92 (26.9%), and GA5 in 2 children (0.6%). RSV-A ON1, rarely observed in 2011/12, was the predominant RSV-A genotype in all settings by 2012/13 and remained predominant until 2016/17. Children in PP or PW infected with RSV-A ON1 did not show a more severe clinical course of disease compared with RSV-A non-ON1 infections. In the PICU group, hospital stay was one day longer (median 8 days, inter-quartile range (IQR) 7-12 vs. 7 days, IQR 5-9; p = 0.02) and duration of oxygen treatment two days longer (median 6 days, IQR 4-9 vs. 4 days, IQR 2-6; p = 0.03) for children infected with RSV-A ON1. CONCLUSIONS: In children, RSV-A ON1 largely replaced RSV-A non-ON1 genotypes within two seasons and remained the predominant RSV-A genotype in Germany during subsequent seasons. A higher clinical severity of RSV-A ON1 was observed within the group of children receiving PICU treatment, whereas in other settings clinical severity of RSV-A ON1 and non-ON1 genotypes was largely similar.


Assuntos
Infecções por Vírus Respiratório Sincicial/patologia , Vírus Sincicial Respiratório Humano/genética , Infecções Respiratórias/patologia , Pré-Escolar , Feminino , Genótipo , Alemanha/epidemiologia , Hospitais Pediátricos , Humanos , Lactente , Unidades de Terapia Intensiva Pediátrica , Tempo de Internação , Masculino , Filogenia , RNA Viral/metabolismo , Infecções por Vírus Respiratório Sincicial/epidemiologia , Infecções por Vírus Respiratório Sincicial/virologia , Vírus Sincicial Respiratório Humano/isolamento & purificação , Infecções Respiratórias/epidemiologia , Infecções Respiratórias/virologia , Estações do Ano , Índice de Gravidade de Doença
3.
BMC Infect Dis ; 19(1): 562, 2019 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-31248372

RESUMO

BACKGROUND: The proportion of older HIV-1 infected people in China has increased rapidly in recent years. Elucidation of the transmission characteristics of this high-risk population subgroup is helpful for the development of tailored interventions. METHODS: A phylogenetic analysis was performed that uses available HIV-1 pol sequences amplified with nested RT-PCR from plasma samples of all newly diagnosed participants spanning from October 2017 to September 2018 in Fuyang, Anhui Province. Transmission clusters were identified as two or more sequences that shared a corresponding node with an aLRT-SH value ≥90 in the maximum-likelihood phylogenetic tree and had an overall mean genetic distance of ≤1.5%. A local transmission cluster was defined as a cluster that had more than 80% of its sequences from Fuyang. The role of older people in local HIV-1 transmission was determined using an integration of molecular and demographic data. RESULTS: Of 362 available sequences, 14 subtypes, and 28 local transmission clusters were identified. It was found that the proportion of older people in the local transmission cluster (69/77, 89.61%) was much higher than that of younger people (46/114, 40.35%) (χ2 test, P < 0.001). In the pretreatment drug resistance analysis, the proportion of sequences with PDRMs in the local transmission cluster was not significantly different between the older people group (57.14%, 4/7) and non-old-aged group (11.11%, 1/9) (Fisher's exact test, P > 0.05). CONCLUSION: By combining phylogenetic analyses with demographic data, more detailed information was provided about the local transmission structure in Fuyang. These findings suggested that older people play an important role in local transmission, and more tailored interventions for this population subgroup are urgently needed.


Assuntos
Infecções por HIV/diagnóstico , HIV-1/classificação , Adulto , Idoso , Antirretrovirais/uso terapêutico , China/epidemiologia , Farmacorresistência Viral/genética , Feminino , Genótipo , Infecções por HIV/tratamento farmacológico , Infecções por HIV/epidemiologia , Infecções por HIV/transmissão , HIV-1/genética , HIV-1/isolamento & purificação , Humanos , Masculino , Pessoa de Meia-Idade , Filogenia , Reação em Cadeia da Polimerase , RNA Viral/isolamento & purificação , RNA Viral/metabolismo
4.
BMC Infect Dis ; 19(1): 566, 2019 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-31253127

RESUMO

BACKGROUND: To infer transmission direction of a HIV transmission chain is helpful not only in legal jurisdiction but also in precise intervention to prevent HIV spread. Recently, the direction of transmission is inferred by whether paraphyletic-monophyletic (PM) or a combination of paraphyletic and polyphyletic (PP) topologies is observed or not between the sequences of source and recipient in the phylogenetic tree. However, paraphyly between them often declines over time and may disappear between spouses due to bidirectional transmission after primary infection. In this study, our aim is to test the reliability of inferring HIV transmission direction between epidemiologically linked HIV-1 positive couples using whether or not paraphyly is observed in phylogenetic tree. METHODS: HIV quasi-species were sequenced using PCR product clones, and then Bayesian analysis of molecular sequences with MCMC was employed to construct phylogenetic relationship of env, gag, pol gene fragments of HIV-1 positive couples using BEAST software. RESULTS: Our results showed that all sequences of seven couples except pol sequences of couple 12 and 13 form their own monophyletic cluster in phylogenetic tree including the closest control sequences from GenBank or other studies on local samples, which are supported by significant Bayesian posterior probabilities more than 0.9932. Of seven couples, paraphyly is only observed in phylogenetic tree constructed with env and pol gene sequences of three couples and gag gene sequences of four couples. Paraphyly is not observed in half of HIV positive couples. Pol sequences of couple 13 is separated by Blast selected controls; pol sequences of couple 12 in phylogenetic tree is supported by a lower Bayesian posterior value. CONCLUSION: Paraphyly relationship between sequences of donator and recipient is only observed among partial HIV-1 positive couples with epidemiological link. Phylogenetic relationship is not always the same when various gene regions of HIV are used to conduct phylogenetic analysis. The combination of phylogenetic analysis based on various gene regions of HIV and enough epidemiology investigation is essential when inferring transmission direction of HIV in a transmission chain or in one couple. However, while observed paraphyly can be used to infer transmission direction in HIV-1 positive couple, no observed paraphyly cannot deny it.


Assuntos
Infecções por HIV/transmissão , HIV-1/genética , Quase-Espécies , Teorema de Bayes , Feminino , Infecções por HIV/epidemiologia , Infecções por HIV/virologia , HIV-1/classificação , Humanos , Masculino , Filogenia , Reação em Cadeia da Polimerase , RNA Viral/genética , RNA Viral/isolamento & purificação , RNA Viral/metabolismo , Comportamento Sexual , Produtos do Gene gag do Vírus da Imunodeficiência Humana/classificação , Produtos do Gene pol do Vírus da Imunodeficiência Humana/classificação
5.
Nat Commun ; 10(1): 2453, 2019 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-31165735

RESUMO

RNA chaperones are proteins that aid in the folding of nucleic acids, but remarkably, many of these proteins are intrinsically disordered. How can these proteins function without a well-defined three-dimensional structure? Here, we address this question by studying the hepatitis C virus core protein, a chaperone that promotes viral genome dimerization. Using single-molecule fluorescence spectroscopy, we find that this positively charged disordered protein facilitates the formation of compact nucleic acid conformations by acting as a flexible macromolecular counterion that locally screens repulsive electrostatic interactions with an efficiency equivalent to molar salt concentrations. The resulting compaction can bias unfolded nucleic acids towards folding, resulting in faster folding kinetics. This potentially widespread mechanism is supported by molecular simulations that rationalize the experimental findings by describing the chaperone as an unstructured polyelectrolyte.


Assuntos
Hepacivirus/metabolismo , Proteínas do Nucleocapsídeo/metabolismo , Dobramento de RNA , RNA Viral/metabolismo , Proteínas do Core Viral/metabolismo , Dimerização , Genoma Viral , Chaperonas Moleculares/metabolismo , Ácidos Nucleicos/metabolismo , Imagem Individual de Molécula , Espectrometria de Fluorescência , Eletricidade Estática
6.
Nat Commun ; 10(1): 2300, 2019 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-31127091

RESUMO

Single-stranded circular RNAs (circRNAs), generated through 'backsplicing', occur more extensively than initially anticipated. The possible functions of the vast majority of circRNAs remain unknown. Virus-derived circRNAs have recently been described in gamma-herpesviruses. We report that oncogenic human papillomaviruses (HPVs) generate circRNAs, some of which encompass the E7 oncogene (circE7). HPV16 circE7 is detectable by both inverse RT-PCR and northern blotting of HPV16-transformed cells. CircE7 is N6-methyladenosine (m6A) modified, preferentially localized to the cytoplasm, associated with polysomes, and translated to produce E7 oncoprotein. Specific disruption of circE7 in CaSki cervical carcinoma cells reduces E7 protein levels and inhibits cancer cell growth both in vitro and in tumor xenografts. CircE7 is present in TCGA RNA-Seq data from HPV-positive cancers and in cell lines with only episomal HPVs. These results provide evidence that virus-derived, protein-encoding circular RNAs are biologically functional and linked to the transforming properties of some HPV.


Assuntos
Transformação Celular Neoplásica/patologia , Interações Hospedeiro-Patógeno/genética , RNA Viral/metabolismo , RNA/metabolismo , Neoplasias do Colo do Útero/virologia , Animais , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Conjuntos de Dados como Assunto , Feminino , Técnicas de Silenciamento de Genes , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/patogenicidade , Humanos , Camundongos , Camundongos Endogâmicos NOD , Proteínas E7 de Papillomavirus/genética , Polirribossomos/genética , Polirribossomos/metabolismo , RNA/genética , RNA/isolamento & purificação , RNA Interferente Pequeno/metabolismo , RNA Viral/genética , RNA Viral/isolamento & purificação , Análise de Sequência de RNA , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
7.
Nat Commun ; 10(1): 2216, 2019 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-31101900

RESUMO

Transcribing and replicating a double-stranded genome require protein modules to unwind, transcribe/replicate nucleic acid substrates, and release products. Here we present in situ cryo-electron microscopy structures of rotavirus dsRNA-dependent RNA polymerase (RdRp) in two states pertaining to transcription. In addition to the previously discovered universal "hand-shaped" polymerase core domain shared by DNA polymerases and telomerases, our results show the function of N- and C-terminal domains of RdRp: the former opens the genome duplex to isolate the template strand; the latter splits the emerging template-transcript hybrid, guides genome reannealing to form a transcription bubble, and opens a capsid shell protein (CSP) to release the transcript. These two "helicase" domains also extensively interact with CSP, which has a switchable N-terminal helix that, like cellular transcriptional factors, either inhibits or promotes RdRp activity. The in situ structures of RdRp, CSP, and RNA in action inform mechanisms of not only transcription, but also replication.


Assuntos
Replicação do DNA/fisiologia , RNA Replicase/ultraestrutura , RNA Mensageiro/ultraestrutura , Rotavirus/fisiologia , Transcrição Genética/fisiologia , Animais , Proteínas do Capsídeo/metabolismo , Proteínas do Capsídeo/ultraestrutura , Linhagem Celular , Cercopithecus aethiops , Microscopia Crioeletrônica , Modelos Moleculares , Domínios Proteicos/genética , RNA Replicase/genética , RNA Replicase/metabolismo , RNA de Cadeia Dupla/metabolismo , RNA Mensageiro/metabolismo , RNA Viral/metabolismo , Rotavirus/ultraestrutura , Replicação Viral/fisiologia
8.
Int J Mol Sci ; 20(9)2019 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-31083507

RESUMO

Some single-stranded positive-sense RNA [ssRNA(+)] viruses, including Flavivirus, generate specific organelle-like structures in the host endoplasmic reticulum (ER). These structures are called virus replication organelles and consist of two distinct subdomains, the vesicle packets (VPs) and the convoluted membranes (CMs). The VPs are clusters of small vesicle compartments and are considered to be the site of viral genome replication. The CMs are electron-dense amorphous structures observed in proximity to the VPs, but the exact roles of CMs are mostly unknown. Several recent studies have revealed that flaviviruses recruit several host factors that are usually used for the biogenesis of other conventional organelles and usurp their function to generate virus replication organelles. In the current review, we summarize recent studies focusing on the role of host factors in the formation of virus replication organelles and discuss how these intricate membrane structures are organized.


Assuntos
Retículo Endoplasmático/metabolismo , Flavivirus/fisiologia , Biogênese de Organelas , RNA Viral/metabolismo , Replicação Viral/fisiologia , Autofagossomos/metabolismo , Autofagossomos/ultraestrutura , Retículo Endoplasmático/ultraestrutura
9.
Acta Crystallogr F Struct Biol Commun ; 75(Pt 5): 340-347, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-31045563

RESUMO

Ebola virus is an emerging virus that is capable of causing a deadly disease in humans. Replication, transcription and packaging of the viral genome are carried out by the viral nucleocapsid. The nucleocapsid is a complex of the viral nucleoprotein, RNA and several other viral proteins. The nucleoprotein forms large, RNA-bound, helical filaments and acts as a scaffold for additional viral proteins. The 3.1 Šresolution single-particle cryo-electron microscopy structure of the nucleoprotein-RNA helical filament presented here resembles previous structures determined at lower resolution, while providing improved molecular details of protein-protein and protein-RNA interactions. The higher resolution of the structure presented here will facilitate the design and characterization of novel and specific Ebola virus therapeutics targeting the nucleocapsid.


Assuntos
Ebolavirus/química , Nucleocapsídeo/química , Nucleoproteínas/química , RNA Viral/química , Proteínas Virais/química , Sequência de Aminoácidos , Sítios de Ligação , Clonagem Molecular , Microscopia Crioeletrônica , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Células HEK293 , Humanos , Modelos Moleculares , Conformação de Ácido Nucleico , Nucleocapsídeo/ultraestrutura , Nucleoproteínas/genética , Nucleoproteínas/metabolismo , Ligação Proteica , Conformação Proteica em alfa-Hélice , Domínios e Motivos de Interação entre Proteínas , RNA Viral/genética , RNA Viral/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Proteínas Virais/genética , Proteínas Virais/metabolismo
10.
Nat Commun ; 10(1): 2184, 2019 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-31097716

RESUMO

Chronic hepatitis B virus (HBV) infection can cause cirrhosis and hepatocellular carcinoma and is therefore a serious public health problem. Infected patients are currently treated with nucleoside/nucleotide analogs and interferon α, but this approach is not curative. Here, we screen 978 FDA-approved compounds for their ability to inhibit HBV replication in HBV-expressing HepG2.2.15 cells. We find that ciclopirox, a synthetic antifungal agent, strongly inhibits HBV replication in cells and in mice by blocking HBV capsid assembly. The crystal structure of the HBV core protein and ciclopirox complex reveals a unique binding mode at dimer-dimer interfaces. Ciclopirox synergizes with nucleoside/nucleotide analogs to prevent HBV replication in cells and in a humanized liver mouse model. Therefore, orally-administered ciclopirox may provide a novel opportunity to combat chronic HBV infection by blocking HBV capsid assembly.


Assuntos
Antivirais/farmacologia , Ciclopirox/farmacologia , Vírus da Hepatite B/fisiologia , Hepatite B Crônica/tratamento farmacológico , Montagem de Vírus/efeitos dos fármacos , Animais , Antivirais/uso terapêutico , Capsídeo/efeitos dos fármacos , Capsídeo/metabolismo , Ciclopirox/química , Ciclopirox/uso terapêutico , Cristalografia por Raios X , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Sinergismo Farmacológico , Células Hep G2 , Vírus da Hepatite B/efeitos dos fármacos , Hepatite B Crônica/patologia , Hepatite B Crônica/virologia , Hepatócitos/transplante , Hepatócitos/virologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos SCID , RNA Viral/metabolismo , Quimeras de Transplante , Resultado do Tratamento , Proteínas do Core Viral/química , Proteínas do Core Viral/metabolismo , Replicação Viral/efeitos dos fármacos
11.
BMC Infect Dis ; 19(1): 479, 2019 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-31142265

RESUMO

BACKGROUND: The increasing availability of high-throughput sequencing data provides researchers with unprecedented opportunities to investigate viral genetic elements in host genomes that contribute to virus-linked cancers. Almost all of the available computational tools for secondary analysis of sequencing data detect viral infection or genome integration events. However, viral oncogenes expression is likely of importance in carcinoma. We therefore developed a new software, DisV-HPV16, for the evaluation of HPV16 oncogenes expression. RESULTS: HPV16 virus and viral oncogenes expression was detected more rapidly using DisV-HPV16 compared to other software. DisV-HPV16 was proved highly convenient for detecting candidate virus after modification of the reference file. The accuracy of DisV-HPV16 was empirically confirmed in laboratory experiments. DisV-HPV16 exhibited greater reliability than other software. CONCLUSIONS: DisV-HPV16 is a new, dependable software to detect virus and viral oncogenes expression through analysis of RNA sequencing data. Use of DisV-HPV16 can yield deeper, more comprehensive insights into virus infection status and viral and host cell gene expression.


Assuntos
Papillomavirus Humano 16/genética , Proteínas Oncogênicas Virais/genética , Software , Sequência de Bases , Linhagem Celular Tumoral , Genoma Viral , Sequenciamento de Nucleotídeos em Larga Escala , Papillomavirus Humano 16/isolamento & purificação , Humanos , Infecções por Papillomavirus/diagnóstico , Infecções por Papillomavirus/virologia , RNA Viral/química , RNA Viral/isolamento & purificação , RNA Viral/metabolismo , Reprodutibilidade dos Testes , Análise de Sequência de RNA
12.
BMC Infect Dis ; 19(1): 418, 2019 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-31088375

RESUMO

BACKGROUND: The global expansion of dengue (DENV), chikungunya (CHIKV), and Zika viruses (ZIKV) is having a serious impact on public health. Because these arboviruses are transmitted by the same mosquito species and co-circulate in the same area, a sensitive diagnostic assay that detects them together, with discrimination, is needed. METHODS: We present here a diagnostics panel based on reverse transcription-PCR amplification of viral RNA and an xMap Luminex architecture involving direct hybridization of PCRamplicons and virus-specific probes. Two DNA innovations ("artificially expanded genetic information systems", AEGIS, and "self-avoiding molecular recognition systems", SAMRS) increase the hybridization sensitivity on Luminex microspheres and PCR specificity of the multiplex assay compared to the standard approach (standard nucleotides). RESULTS: The diagnostics panel detects, if they are present, these viruses with a resolution of 20 genome equivalents (DENV1), or 10 (DENV3-4, CHIKV) and 80 (DENV2, ZIKV) genome equivalents per assay. It identifies ZIKV, CHIKV and DENV RNAs in a single infected mosquito, in mosquito pools comprised of 5 to 50 individuals, and mosquito saliva (ZIKV, CHIKV, and DENV2). Infected mosquitoes and saliva were also collected on a cationic surface (Q-paper), which binds mosquito and viral nucleic acids electrostatically. All samples from infected mosquitoes displayed only target-specific signals; signals from non-infected samples were at background levels. CONCLUSIONS: Our results provide an efficient and multiplex tool that may be used for surveillance of emerging mosquito-borne pathogens which aids targeted mosquito control in areas at high risk for transmission.


Assuntos
Vírus Chikungunya/genética , Culicidae/virologia , Vírus da Dengue/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Zika virus/genética , Animais , Febre de Chikungunya/diagnóstico , Febre de Chikungunya/virologia , Vírus Chikungunya/isolamento & purificação , Dengue/diagnóstico , Dengue/virologia , Vírus da Dengue/isolamento & purificação , Humanos , Hibridização de Ácido Nucleico , RNA Viral/genética , RNA Viral/metabolismo , Kit de Reagentes para Diagnóstico , Saliva/virologia , Zika virus/isolamento & purificação , Infecção por Zika virus/diagnóstico , Infecção por Zika virus/virologia
13.
BMC Infect Dis ; 19(1): 370, 2019 May 02.
Artigo em Inglês | MEDLINE | ID: mdl-31046684

RESUMO

BACKGROUND: Several inactivated enterovirus-A71 (EV-A71) vaccines are currently licensed in China; however, the development of additional EV-A71 vaccines is ongoing, necessitating extensive analysis of the molecular epidemiology of the virus worldwide. Until 2012, laboratory confirmation of EV-A71 for hand, foot, and mouth disease (HFMD) and other associated diseases had not occurred in the Philippines. Because EV-A71 has been linked with cases of acute flaccid paralysis (AFP), AFP surveillance is one strategy for documenting its possible circulation in the country. To expand current knowledge on EV-A71, molecular epidemiologic analysis and genetic characterization of EV-A71 isolates were performed in this study. METHODS: A retrospective study was performed to identify and characterize nonpolio enteroviruses (NPEVs) associated with AFP in the Philippines, and nine samples were found to be EV-A71-positive. Following characterization of these EV-A71 isolates, the complete viral protein 1 (VP1) gene was targeted for phylogenetic analysis. RESULTS: Nine EV-A71 isolates detected in 2000 (n = 2), 2002 (n = 4), 2005 (n = 2), and 2010 (n = 1) were characterized using molecular methods. Genomic regions spanning the complete VP1 region were amplified and sequenced using specific primers. Phylogenetic analysis of the full-length VP1 region identified all nine EV-A71 Philippine isolates as belonging to the genogroup C lineage, specifically the C2 cluster. The result indicated a genetic linkage with several strains isolated in Japan and Taiwan, suggesting that strains in the C2 cluster identified in the Asia-Pacific region were circulating in the Philippines. CONCLUSION: The study presents the genetic analysis of EV-A71 in the Philippines. Despite some limitations, the study provides additional genetic data on the circulating EV-A71 strains in the Asia-Pacific region, in which information on EV-A71 molecular epidemiology is incomplete. Considering that EV-A71 has a significant public health impact in the region, knowledge of its circulation in each country is important, especially for formulating vaccines covering a wide variety of strains.


Assuntos
Infecções por Enterovirus/diagnóstico , Paralisia/diagnóstico , Doença Aguda , Animais , Proteínas do Capsídeo/genética , Criança , Pré-Escolar , Enterovirus Humano A/classificação , Enterovirus Humano A/genética , Enterovirus Humano A/isolamento & purificação , Infecções por Enterovirus/virologia , Fezes/virologia , Febre Aftosa/diagnóstico , Febre Aftosa/virologia , Genótipo , Humanos , Lactente , Paralisia/virologia , Filipinas , Filogenia , RNA Viral/isolamento & purificação , RNA Viral/metabolismo , Estudos Retrospectivos
14.
BMC Infect Dis ; 19(1): 373, 2019 May 02.
Artigo em Inglês | MEDLINE | ID: mdl-31046702

RESUMO

BACKGROUND: Body mass index (BMI) may contribute somewhat to drug metabolism, and thus affecting the efficacy of highly active antiretroviral therapy (HAART). This study aimed to determine the frequencies of underweight, normal and overweight/obesity at pre-HAART in a large cohort of HIV-infected Chinese patients, and investigate the prospective effects of BMI on immune reconstitution after HAART initiation. METHODS: A longitudinal cohort study was performed to analyze the effects of BMI on immune reconstitution in HIV-infected patients treated with HAART. Multiple linear regression was used to evaluate the relationship between baseline BMI and increased CD4+ T lymphocyte levels at 12 and 30 months after initiating HAART. In addition, Cox proportional hazard model was used to assess the relationship between BMI and time to achieve immunologic reconstitution (CD4+ T lymphocytes>500cells/µL) during the follow-up period. RESULTS: Among the 1612 enrolled patients, 283 (17.6%) were overweight/obese (BMI ≥ 25 kg/m2), 173 (10.7%) were underweight (BMI < 18.5 kg/m2) and the remaining were normal weight. Prior to HAART initiating, overweight HIV-infected patients were mostly males, older ages, exhibited higher CD4+ T lymphocytes and lower viral loads (p < 0.01 for all). Patients with higher baseline BMI had an independently positive effect on 30-month CD4+ T lymphocyte recovery (p = 0.028), but not 12-month CD4+ T lymphocyte gain (p = 0.104). In addition, a Cox proportional hazard model with baseline BMI as an independent variable indicated that BMI was correlated with an increased likelihood of achieving immunologic reconstitution over time (hazard ratios [HR] 1.03; 95% confidence intervals [CI] 1.01-1.06; p = 0.011), after adjusting for baseline age, gender, CD4+ T lymphocytes, CD4/CD8 ratio, viral load and WHO stage. CONCLUSIONS: Higher baseline BMI could predict better immune reconstitution in HIV-infected patients after HAART initiating.


Assuntos
Terapia Antirretroviral de Alta Atividade , Infecções por HIV/tratamento farmacológico , Adulto , Fatores Etários , Índice de Massa Corporal , Contagem de Linfócito CD4 , Linfócitos T CD4-Positivos/citologia , China , Feminino , HIV/genética , HIV/isolamento & purificação , Infecções por HIV/patologia , Infecções por HIV/virologia , Humanos , Modelos Lineares , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Modelos de Riscos Proporcionais , RNA Viral/genética , RNA Viral/metabolismo , Carga Viral
15.
Arch Virol ; 164(7): 1851-1855, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31055651

RESUMO

The RNA genome of human parainfluenza virus type 2 (hPIV2) is encapsidated by nucleoprotein (NP) to act as a template for RNA synthesis. We examined the importance of individual amino acids in the RNA-binding domain of hPIV2 NP for polymerase activity using a mini-replicon assay. We showed that substitution of tyrosine at amino acid position 260, located in the RNA-binding pocket of NP, severely reduced polymerase activity. The aromatic side-chain of Y260 may be required for the formation of stable contacts between nucleotides and basic amino acids, thereby affecting promoter recognition by the viral polymerase.


Assuntos
Nucleoproteínas/genética , Vírus da Parainfluenza 2 Humana/genética , RNA Replicase/metabolismo , RNA Viral/metabolismo , Motivos de Ligação ao RNA/genética , Sequência de Aminoácidos , Cristalografia por Raios X , Genoma Viral/genética , Humanos , Tirosina/genética , Replicação Viral/genética
16.
Virus Res ; 266: 15-24, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30951791

RESUMO

Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiological agent of Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), and multicentric Castleman Disease (MCD). Recent mechanistic advances have discerned the importance of microRNAs in the virus-host relationship. KSHV has two modes of replication: lytic and latent phase. KSHV entry into permissive cells, establishment of infection, and maintenance of latency are contingent upon successful modulation of the host miRNA transcriptome. Apart from host cell miRNAs, KSHV also encodes viral miRNAs. Among various cellular and molecular targets, miRNAs are appearing to be key players in regulating viral pathogenesis. Therefore, the use of miRNAs as novel therapeutics has gained considerable attention as of late. This innovative approach relies on either mimicking miRNA species by identical oligonucleotides, or selective silencing of miRNA with specific oligonucleotide inhibitors. Here, we provide an overview of KSHV pathogenesis at the molecular level with special emphasis on the various roles miRNAs play during virus infection.


Assuntos
Herpesvirus Humano 8/genética , Herpesvirus Humano 8/patogenicidade , MicroRNAs/metabolismo , RNA Viral/metabolismo , Hiperplasia do Linfonodo Gigante/virologia , Regulação Viral da Expressão Gênica , Herpesvirus Humano 8/fisiologia , Humanos , Linfoma de Efusão Primária/virologia , MicroRNAs/genética , RNA Viral/genética , Sarcoma de Kaposi/virologia , Internalização do Vírus , Latência Viral , Replicação Viral
17.
Nat Commun ; 10(1): 1629, 2019 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-30967547

RESUMO

Influenza A virus has an eight-partite RNA genome that during viral assembly forms a complex containing one copy of each RNA. Genome assembly is a selective process driven by RNA-RNA interactions and is hypothesized to lead to discrete punctate structures scattered through the cytosol. Here, we show that contrary to the accepted view, formation of these structures precedes RNA-RNA interactions among distinct viral ribonucleoproteins (vRNPs), as they assemble in cells expressing only one vRNP type. We demonstrate that these viral inclusions display characteristics of liquid organelles, segregating from the cytosol without a delimitating membrane, dynamically exchanging material and adapting fast to environmental changes. We provide evidence that viral inclusions develop close to endoplasmic reticulum (ER) exit sites, depend on continuous ER-Golgi vesicular cycling and do not promote escape to interferon response. We propose that viral inclusions segregate vRNPs from the cytosol and facilitate selected RNA-RNA interactions in a liquid environment.


Assuntos
Retículo Endoplasmático/virologia , Vírus da Influenza A/fisiologia , Influenza Humana/patologia , Ribonucleoproteínas/metabolismo , Proteínas Virais/metabolismo , Montagem de Vírus , Células A549 , Animais , Citosol/metabolismo , Citosol/virologia , Cães , Retículo Endoplasmático/metabolismo , Complexo de Golgi/metabolismo , Complexo de Golgi/virologia , Células HEK293 , Células HeLa , Interações Hospedeiro-Patógeno , Humanos , Vírus da Influenza A/patogenicidade , Influenza Humana/virologia , Células Madin Darby de Rim Canino , Ligação Proteica , RNA Viral/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo
18.
BMC Infect Dis ; 19(1): 308, 2019 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-30947693

RESUMO

BACKGROUND: Despite World Health Organization recommendations, in many countries young children are not targeted for influenza vaccination. To help inform influenza vaccination policy, we examined the occurrence and burden of influenza in healthy children aged 6 to 35 months using data from a recent phase III placebo-controlled influenza vaccine trial conducted in countries in the Northern and Southern Hemispheres. METHODS: This was an analysis of data from participants included in the placebo arm of a phase III clinical trial in healthy children aged 6 to 35 months (EudraCT no. 2013-001231-51). Included children had never been vaccinated for influenza and were observed for one influenza season. Outcome measures included the occurrence of influenza-like illness (ILI), laboratory-confirmed influenza, virus types/subtypes, severe symptoms and complications of confirmed influenza, and healthcare use associated with confirmed influenza. RESULTS: Data from 2210 participants were analysed. ILI was reported for 811 participants (36.7%). Of these, 255 participants (31.4%) had 263 virologically confirmed episodes of influenza. The overall influenza attack rate was 11.5%. The most common influenza virus detected was A(H3N2) (40.7%), followed by B/Yamagata (23.6%), A(H1N1) (18.6%), and B/Victoria (8.0%). Grade 3 fever was reported in 24.3% of confirmed episodes, acute lower respiratory infection in 8.7%, acute otitis media in 6.1%, and pneumonia in 1.9%. In most influenza episodes (93.2%), antipyretics, analgesics, or non-steroidal anti-inflammatory drugs were taken. Antibiotics were prescribed for 41.4% of influenza episodes. More than half of the influenza episodes (57.0%) resulted in outpatient visits. Influenza resulted in overnight hospitalisation in 1.1% of episodes. CONCLUSIONS: Influenza is associated with a significant burden of disease in healthy children. This analysis also revealed that antibiotics continue to be frequently used for young children with influenza. TRIAL REGISTRATION: EudraCT no. 2013-001231-51 .


Assuntos
Influenza Humana/epidemiologia , Avaliação de Resultados (Cuidados de Saúde) , Antibacterianos/uso terapêutico , Antipiréticos/uso terapêutico , Pré-Escolar , Efeitos Psicossociais da Doença , Feminino , Humanos , Lactente , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Vírus da Influenza A Subtipo H3N2/genética , Vírus da Influenza A Subtipo H3N2/isolamento & purificação , Vacinas contra Influenza/imunologia , Influenza Humana/economia , Influenza Humana/patologia , Influenza Humana/virologia , Masculino , Efeito Placebo , RNA Viral/genética , RNA Viral/metabolismo , Infecções Respiratórias/diagnóstico , Infecções Respiratórias/tratamento farmacológico , Infecções Respiratórias/epidemiologia , Índice de Gravidade de Doença
19.
BMC Bioinformatics ; 20(1): 213, 2019 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-31029080

RESUMO

BACKGROUND: Next Generation Sequencing (NGS) is a commonly used technology for studying the genetic basis of biological processes and it underpins the aspirations of precision medicine. However, there are significant challenges when dealing with NGS data. Firstly, a huge number of bioinformatics tools for a wide range of uses exist, therefore it is challenging to design an analysis pipeline. Secondly, NGS analysis is computationally intensive, requiring expensive infrastructure, and many medical and research centres do not have adequate high performance computing facilities and cloud computing is not always an option due to privacy and ownership issues. Finally, the interpretation of the results is not trivial and most available pipelines lack the utilities to favour this crucial step. RESULTS: We have therefore developed a fast and efficient bioinformatics pipeline that allows for the analysis of DNA sequencing data, while requiring little computational effort and memory usage. DNAscan can analyse a whole exome sequencing sample in 1 h and a 40x whole genome sequencing sample in 13 h, on a midrange computer. The pipeline can look for single nucleotide variants, small indels, structural variants, repeat expansions and viral genetic material (or any other organism). Its results are annotated using a customisable variety of databases and are available for an on-the-fly visualisation with a local deployment of the gene.iobio platform. DNAscan is implemented in Python. Its code and documentation are available on GitHub: https://github.com/KHP-Informatics/DNAscan . Instructions for an easy and fast deployment with Docker and Singularity are also provided on GitHub. CONCLUSIONS: DNAscan is an extremely fast and computationally efficient pipeline for analysis, visualization and interpretation of NGS data. It is designed to provide a powerful and easy-to-use tool for applications in biomedical research and diagnostic medicine, at minimal computational cost. Its comprehensive approach will maximise the potential audience of users, bringing such analyses within the reach of non-specialist laboratories, and those from centres with limited funding available.


Assuntos
Biologia Computacional/métodos , Sequenciamento de Nucleotídeos em Larga Escala , Interface Usuário-Computador , Esclerose Amiotrófica Lateral/genética , Esclerose Amiotrófica Lateral/patologia , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Bacteriano/metabolismo , Bases de Dados Factuais , HIV-1/genética , Humanos , Mutação INDEL , Polimorfismo de Nucleotídeo Único , RNA Viral/química , RNA Viral/genética , RNA Viral/metabolismo , Sequenciamento Completo do Genoma
20.
Mol Med Rep ; 19(5): 4441-4448, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30896873

RESUMO

Extra­hepatic manifestations are frequently observed in hepatitis C virus (HCV)­infected patients; however the underlying mechanisms remain largely unknown. In the present study, the human glioblastoma SF268 cell line (the precise origin of the cell type is not clear) was infected with HCV using HCV­positive serum, and viral replication was assessed by immunofluorescence, reverse transcription­polymerase chain reaction (PCR), quantitative PCR and western blotting following infection. HCV core protein and HCV RNA were detected in HCV­positive serum­infected SF268 cells at day 4 post­infection, while no infection was observed in cells exposed to HCV­negative serum. The mean HCV RNA levels at day 4 post­infection were up to 5.00 IU/ml log10; however, HCV RNA and immunostaining for core protein were negative when cultured to day 6 or longer. The data suggest that human glioblastoma SF268 cells were transiently infected with HCV. AKT serine/threonine kinase phosphorylation was also detected in HCV­infected SF268 cells at day 4 post­infection. To the best of our knowledge, this is the first demonstration that a human glioblastoma cell line can be infected with serum­derived HCV. The results provide evidence that HCV infection can occur in cells of the central nervous system. Neurological disorder­associated phosphoinositide 3­kinase­AKT signaling pathway was activated in parallel with HCV infection, suggesting that SF268 may serve as an in vitro model for investigating HCV­nervous system cell interactions.


Assuntos
Hepacivirus/fisiologia , Hepatite C/patologia , Adulto , Linhagem Celular Tumoral , Feminino , Glioblastoma/metabolismo , Glioblastoma/patologia , Glicogênio Sintase Quinase 3 beta/metabolismo , Hepacivirus/genética , Hepacivirus/isolamento & purificação , Hepatite C/sangue , Hepatite C/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Viral/metabolismo , Transdução de Sinais
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