RESUMEN

Hodgkin lymphoma is histologically characterised by the presence of Hodgkin (H) and Reed-Sternberg (RS) cells originating from germinal centre B-cells rearranged in the IgV gene. The formation of multinucleated RS cells is a product of telomere organisation in a process initiated by telomere aggregate accumulation in mononuclear H cells and may be mediated by latent membrane protein 1 (LMP-1) expression. LMP-1 is the main oncoprotein of EBV and supports several tumourigenic processes. LMP-1 may rescue proapoptotic B-cells through downregulation of B-cell receptor (BCR) components, mimicking and inducing multiple distinct B-cell signalling pathways to promote proliferation and survival, such as Janus kinase-signal transducer and activator of transcription (JAK-STAT), nuclear factor-kappa b (NF-кB), and cellular MYC (c-MYC), and inducing telomere instability mainly through Telomere repeat binding factor 2 (TRF2) downregulation to promote the formation of multinucleated RS cells. This review presents recent discoveries regarding the influence of LMP-1 on the surviving cellular signalling, genomic instability and mecanical formation of HRS cells.

Asunto(s)

Herpesvirus Humano 4 , Enfermedad de Hodgkin , Proteínas de la Matriz Viral , Enfermedad de Hodgkin/virología , Enfermedad de Hodgkin/patología , Enfermedad de Hodgkin/metabolismo , Humanos , Proteínas de la Matriz Viral/metabolismo , Proteínas de la Matriz Viral/genética , Herpesvirus Humano 4/genética , Transducción de Señal , Infecciones por Virus de Epstein-Barr/virología , Infecciones por Virus de Epstein-Barr/metabolismo , Infecciones por Virus de Epstein-Barr/patología , Inestabilidad Genómica , Células de Reed-Sternberg/metabolismo , Células de Reed-Sternberg/patología , Células de Reed-Sternberg/virología

RESUMEN

Epstein-Barr virus (EBV) is a ubiquitous gammaherpesvirus etiologically associated with benign and malignant diseases. Since the pathogenic mechanisms of EBV are not fully understood, understanding EBV genetic diversity is an ongoing goal. Therefore, the present work describes the genetic diversity of the lytic gene BZLF1 in a sampling of 70 EBV-positive cases from southeastern Brazil. Additionally, together with the genetic regions previously characterized, the aim of the present study was to determine the impact of viral genetic factors that may influence EBV genetic diversity. Accordingly, the phylogenetic analysis of the BZLF1 indicated two main clades with high support, BZ-A and BZ-B (PP > 0.85). Thus, the BZ-A clade was the most diverse clade associated with the main polymorphisms investigated, including the haplotype Type 1 + V3 (p < 0.001). Furthermore, the multigene phylogenetic analysis (MLA) between BZLF1 and the oncogene LMP1 showed specific clusters, revealing haplotypic segregation that previous single-gene phylogenies from both genes failed to demonstrate. Surprisingly, the LMP1 Raji-related variant clusters were shown to be more diverse, associated with BZ-A/B and the Type 2/1 + V3 haplotypes. Finally, due to the high haplotypic diversity of the Raji-related variants, the number of DNA recombination-inducing motifs (DRIMs) was evaluated within the different clusters defined by the MLA. Similarly, the haplotype BZ-A + Raji was shown to harbor a greater number of DRIMs (p < 0.001). These results call attention to the high haplotype diversity of EBV in southeast Brazil and strengthen the hypothesis of the recombinant potential of South American Raji-related variants via the LMP1 oncogene.

Asunto(s)

Infecciones por Virus de Epstein-Barr , Variación Genética , Herpesvirus Humano 4 , Filogenia , Recombinación Genética , Herpesvirus Humano 4/genética , Humanos , Brasil , Infecciones por Virus de Epstein-Barr/virología , Infecciones por Virus de Epstein-Barr/genética , Transactivadores/genética , Masculino , Femenino , Haplotipos/genética , Adulto , Proteínas de la Matriz Viral/genética , Niño , Persona de Mediana Edad , Adolescente , Latencia del Virus/genética , Preescolar , Adulto Joven

RESUMEN

In 2017, the World Health Organization (WHO) confirmed a new entity, Epstein Barr virus (EBV) + Diffuse large B cell lymphoma (DLBCL), not otherwise specified (NOS). Traces of EBV transcripts were described in lymphomas, including DLBCL, that were diagnosed as EBV negative by conventional methods. The aim of this study was to detect viral genome by qPCR, as well as LMP1 and EBNA2 transcripts, with a more sensitive method in DLBCL cases from Argentina. Fourteen cases originally considered as EBV negative expressed LMP1 and/or EBNA2 transcripts. In addition, LMP1 and/or EBNA2 transcripts were also observed in bystander cells. However, EBERs+ cells cases by conventional ISH showed higher numbers of cells with LMP1 transcripts and LMP1 protein. In the cases that were EBERS- in tumor cells but with expression of LMP1 and/or EBNA2 transcripts, the viral load was below the limit of detection. This study provides further evidence that EBV could be detected in tumor cells by more sensitive methods. However, higher expression of the most important oncogenic protein, LMP1, as well as increased viral load, are only observed in cases with EBERs+ cells by conventional ISH, suggesting that traces of EBV might not display a key role in DLBCL pathogenesis.

Asunto(s)

Infecciones por Virus de Epstein-Barr , Linfoma de Células B Grandes Difuso , Humanos , Adulto , Niño , Herpesvirus Humano 4/genética , Linfoma de Células B Grandes Difuso/patología , Argentina , Antígenos Nucleares del Virus de Epstein-Barr/genética , Proteínas de la Matriz Viral/genética

RESUMEN

Epstein−Barr virus (EBV) is a saliva-borne É£-herpesvirus associated with benign and malignant lymphoproliferation. EBV-mediated tumorigenic mechanisms are not fully understood and may be related to viral genetic variations. In this work, we characterize the genetic diversity of EBV from Brazil, assessing 82 samples derived from saliva from asymptomatic carriers (n = 45), biopsies of benign reactive hyperplasia (n = 4), and lymphomas (n = 33). Phylogenetic and phylogeographic analysis of the entire coding region of the LMP-1 was performed. Additionally, type 1/type 2 distinction by the EBNA3C gene and Zp variants were evaluated. Our results revealed a high diversity of EBV in Brazil, with the co-circulation of four main clades, described here as: Mediterranean (40.2%, n = 33), Raji/Argentine (39%, n = 32), B95-8 (6.1%, n = 5), and Asian II (1.2%, n = 1). The Raji/Argentine and Mediterranean clades were the most prevalent in South America (45% and 28%, respectively). The Raji/Argentine clade was associated with polymorphisms I124V/I152L, del30 bp, and ins15 bp (p < 0.0001, to all clades) and with a high haplotype diversity related to EBV type and Zp variants. We found that a Raji/Argentine subclade spread primarily from Brazil and later to other South American countries. Although no LMP1 variant has been directly associated with disease, the Raji/Argentine clade was predominantly clustered with lymphomas (61%) and the Mediterranean clade with non-malignant cases (59%) (p = 0.1). These data highlight the high genetic diversity of EBV circulating in Brazil, calling attention to a Raji-related variant with great recombination potential in Brazilian lymphomas.

Asunto(s)

Infecciones por Virus de Epstein-Barr , Linfoma , Negro o Afroamericano , Brasil/epidemiología , Herpesvirus Humano 4/genética , Humanos , Filogenia , Proteínas de la Matriz Viral/genética , Proteínas Virales/genética

RESUMEN

Psittaciform orthobornaviruses are currently considered to be a major threat to the psittacine bird population worldwide. Parrot bornavirus (PaBV) was identified recently in Brazil and, since then, few studies have been conducted to understand the epidemiology of PaBV in captive psittacine birds. In the present study, natural infections by PaBV in South American parrots were investigated in two breeding facilities: commercial (A) and conservationist (B). Thirty-eight psittacine of 21 different species were presented for postmortem examination. Tissue samples were collected and investigated for the presence of PaBV-RNA using RT-PCR. In addition, clinical information about these birds was used when available. PaBV infection was detected in 73.7% of all birds investigated, indicating a wide dissemination of this virus in both facilities. From birds investigated in aviary A, 66.7% showed clinical signs, 100% had typical lesions of proventricular dilatation disease (PDD), 100% had mild to severe proventricular dilatation and 88.9% were PaBV-positive. In birds from aviary B, 27.6% showed clinical signs, 65.5% had typical lesions of PDD, 62% had mild to severe proventricular dilatation and 69% were PaBV-positive. Neurological disease was observed more frequently than gastrointestinal disease. Sequencing analysis of the matrix gene fragment revealed the occurrence of genotype 4 (PaBV-4) in both places. About 15.8% of birds in this study are threatened species. We discussed the difficulties and challenges for controlling viral spread in these aviaries and implications for South American psittacine conservation. These results emphasize the urgent need to develop a national regulatory and health standard for breeding psittacine birds in the country.

Asunto(s)

Enfermedades de las Aves/patología , Bornaviridae/genética , Infecciones por Mononegavirales/patología , Animales , Enfermedades de las Aves/epidemiología , Enfermedades de las Aves/virología , Bornaviridae/clasificación , Bornaviridae/aislamiento & purificación , Brasil/epidemiología , Genotipo , Infecciones por Mononegavirales/complicaciones , Infecciones por Mononegavirales/epidemiología , Infecciones por Mononegavirales/virología , Enfermedades del Sistema Nervioso/complicaciones , Enfermedades del Sistema Nervioso/patología , Enfermedades del Sistema Nervioso/virología , Loros/virología , Filogenia , ARN Viral/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteínas de la Matriz Viral/clasificación , Proteínas de la Matriz Viral/genética

RESUMEN

Highly pathogenic H7N3 influenza A viruses have persisted in poultry in Mexico since 2012, diversifying into multiple lineages that have spread to three Mexican states, as of 2016. The H7N3 viruses segregate into three distinct clades that are geographically structured. All 2016 viruses are resistant to adamantane antiviral drugs and have an extended 24-nucleotide insertion at the HA cleavage site that was acquired from host 28S ribosomal RNA.

Asunto(s)

Evolución Biológica , Pollos , Subtipo H7N3 del Virus de la Influenza A/patogenicidad , Gripe Aviar/virología , Secuencia de Aminoácidos , Animales , Brotes de Enfermedades , Genoma Viral , Gripe Aviar/epidemiología , México/epidemiología , Proteínas de la Matriz Viral/genética , Proteínas de la Matriz Viral/metabolismo , Virulencia

RESUMEN

A new outbreak of equine Influenza A virus (IAV) was reported in Chile in January 2018, 6 years after its last report in 2012. Equine IAV was detected by rtRT-PCR, followed by virus isolation and full genome sequencing. Genetic characterization of equine IAV classified the virus within clade 1 of the Florida sublineage. Although this is the same sublineage that caused an outbreak in Chile in 2012, the virus has a high similarity to other cocirculating viruses that were recently identified in Europe and Asia. The Chilean 2018 equine influenza (EI) outbreak was caused by an H3N8 strain circulating globally that spread through horse movements.

Asunto(s)

Enfermedades Transmisibles Emergentes/veterinaria , Brotes de Enfermedades/veterinaria , Enfermedades de los Caballos/epidemiología , Subtipo H3N8 del Virus de la Influenza A/aislamiento & purificación , Infecciones por Orthomyxoviridae/veterinaria , Animales , Chile/epidemiología , Enfermedades Transmisibles Emergentes/epidemiología , Enfermedades Transmisibles Emergentes/virología , Enfermedades de los Caballos/virología , Caballos , Subtipo H3N8 del Virus de la Influenza A/genética , Infecciones por Orthomyxoviridae/epidemiología , Infecciones por Orthomyxoviridae/virología , Filogenia , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinaria , Proteínas de la Matriz Viral/genética

RESUMEN

BACKGROUND: There is insufficient knowledge about the relation of avian influenza virus (AIV) to migratory birds in South America. Accordingly, we studied samples obtained over a 4-year period (2009-2012) from wild birds at a major wintering site in southern Brazil. METHODS: We obtained 1212 oropharyngeal/cloacal samples from wild birds at Lagoa do Peixe National Park and screened them for influenza A virus by RT-PCR amplification of the matrix gene. Virus isolates were subjected to genomic sequencing and antigenic characterization. RESULTS: Forty-eight samples of 1212 (3.96%) contained detectable influenza virus RNA. Partial viral sequences were obtained from 12 of these samples, showing the presence of H2N2 (1), H6Nx (1), H6N1 (8), H9N2 (1), and H12N5 (1) viruses. As H6 viruses predominated, we generated complete genomes from all 9 H6 viruses. Phylogenetic analyses showed that they were most similar to viruses of South American lineage. The H6N1 viruses caused no disease signs in infected ferrets and, despite genetic differences, were antigenically similar to North American isolates. CONCLUSIONS: Lagoa do Peixe National Park is a source of multiple AIV subtypes, with the levels of influenza virus in birds being highest at the end of their wintering period in this region. H6N1 viruses were the predominant subtype identified. These viruses were more similar to viruses of South American lineage than to those of North American lineage.

Asunto(s)

Aves/virología , Variación Genética , Virus de la Influenza A/clasificación , Virus de la Influenza A/aislamiento & purificación , Gripe Aviar/virología , Animales , Antígenos Virales/análisis , Brasil , Cloaca/virología , Virus de la Influenza A/genética , Orofaringe/virología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Análisis de Secuencia de ADN , Proteínas de la Matriz Viral/genética

RESUMEN

MAIN CONCLUSION: The plant cell is able to produce the VP40 antigen from the Zaire ebolavirus , retaining the antigenicity and the ability to induce immune responses in BALB/c mice. The recent Ebola outbreak evidenced the need for having vaccines approved for human use. Herein we report the expression of the VP40 antigen from the Ebola virus as an initial effort in the development of a plant-made vaccine that could offer the advantages of being cheap and scalable, which is proposed to overcome the rapid need for having vaccines to deal with future outbreaks. Tobacco plants were transformed by stable DNA integration into the nuclear genome using the CaMV35S promoter and a signal peptide to access the endoplasmic reticulum, reaching accumulation levels up to 2.6 µg g-1 FW leaf tissues. The antigenicity of the plant-made VP40 antigen was evidenced by Western blot and an initial immunogenicity assessment in test animals that revealed the induction of immune responses in BALB/c mice following three weekly oral or subcutaneous immunizations at very low doses (125 and 25 ng, respectively) without accessory adjuvants. Therefore, this plant-based vaccination prototype is proposed as an attractive platform for the production of vaccines in the fight against Ebola virus disease outbreaks.

Asunto(s)

Ebolavirus/inmunología , Ebolavirus/metabolismo , Expresión Génica , Nicotiana/metabolismo , Plantas Modificadas Genéticamente/metabolismo , Proteínas de la Matriz Viral/metabolismo , Animales , Anticuerpos Antivirales/inmunología , Ebolavirus/genética , Fiebre Hemorrágica Ebola/inmunología , Fiebre Hemorrágica Ebola/prevención & control , Inmunización , Ratones , Ratones Endogámicos BALB C , Plantas Modificadas Genéticamente/genética , Nicotiana/genética , Proteínas de la Matriz Viral/genética

RESUMEN

AIM: To study LMP1 variants distribution among children with EBV+ malignant and benign conditions as well as in healthy carriers. METHODS: Oral secretions and blood cells from 31 children with IM, and biopsies from 14 EBV+ reactive lymphoid hyperplasia and 33 EBV+ lymphomas were included. LMP1 was amplified by nested PCR and sequenced. Phylogenetic reconstructions were made under Maximun Likelihood, Bayesian and coalescent algorithms. RESULTS: Six clades were defined (China1, China2, Med-, Alaskan, B95.8 and Argentine). Argentine variants, the most prevalent (46%), harbored 3 distinctive mutations and were a recombination between Raji and China1. Despite no pathology or compartment associations were observed for LMP1, the Argentine clade showed a phylogeographic association with our region. LMP1 estimated evolution rate was 8.591x10-5s/s/y and the estimated tMRCA for Raji and Argentine was 136ybp. CONCLUSIONS: An LMP1 Argentine clade was defined. LMP1 evolutionary rate was higher than expected for herpesviruses. The tMRCA for Raji and the Argentine agrees with African immigration and could explain the recombinant nature of the Argentine variant.

Asunto(s)

Herpesvirus Humano 4/genética , Polimorfismo Genético/genética , Proteínas de la Matriz Viral/genética , Proteínas Virales/genética , Adolescente , Argentina , Teorema de Bayes , Niño , Preescolar , Infecciones por Virus de Epstein-Barr/virología , Femenino , Humanos , Lactante , Linfoma/virología , Masculino , Mutación/genética , Filogenia , Filogeografía/métodos

RESUMEN

We investigated possible vaccinia virus (VACV) in urban house cats in Brazil. Serum samples from 6 cats were positive for VACV by PCR, indicating likely VACV circulation among house cats in urban areas of Brazil. This finding highlights the importance of epidemiologic surveillance to avoid outbreaks among urban human populations.

Asunto(s)

Enfermedades de los Gatos/epidemiología , Enfermedades de los Gatos/virología , Salud Urbana , Virus Vaccinia , Vaccinia/veterinaria , Animales , Animales Domésticos , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Brasil/epidemiología , Enfermedades de los Gatos/inmunología , Gatos , Filogenia , Vigilancia en Salud Pública , Virus Vaccinia/clasificación , Virus Vaccinia/genética , Virus Vaccinia/inmunología , Proteínas de la Matriz Viral/genética , Proteínas de la Matriz Viral/inmunología

RESUMEN

INTRODUCTION: EBV-associated Gastric Cancer (EBVaGC) comprises about 9% of all cases of GC and constitutes a distinct clinicopathological and molecular entity. The pattern of viral expression in EBVaGC cannot be set to any of the previously EBV-associated malignancies. Several lines of evidence support that viral expression in EBVaGC is characterized by high transcription of the BamH1- A rightward transcript (BART), low-levels of EBNA-1 and lack of LMP1. The high transcription activity of the BamH1-A region is importantly directed to express BART miRNAs, supporting a critical role for these miRNAs during epithelial cell infection and carcinogenesis. Several studies have shown that promoter hypermethylation is also a prominent feature of EBVaGC. Based on the recent TCGA report, the specific fingerprint of genomic alterations in EBVaGC is marked by mutations in PIK3CA, ARID1A and BCOR genes, and amplification of 9p24.1 that harbors the genes for the JAK2, PD-L1 and PD-L2 proteins. The specific programs of viral gene expression, promoter methylation and genomic mutations found in EBVaGC target cell signaling pathways leading to increased proliferation, increased cell survival, immune evasion, augmented EMT and acquisition of stemness features. Less understood is the participation of EBV in chronic gastric inflammation, but some studies argue that EBV, similar to and together with Helicobacter pylori, is an early participant in the GC oncogenic process through promoting chronic inflammation and increased tissue damage. CONCLUSION: Here, we discuss the principal and distinctive carcinogenic routes promoted by EBV in the gastric epithelium.

Asunto(s)

Infecciones por Virus de Epstein-Barr/complicaciones , Herpesvirus Humano 4/patogenicidad , Neoplasias Gástricas/virología , Proteínas Portadoras/genética , Islas de CpG , Metilación de ADN , Antígenos Nucleares del Virus de Epstein-Barr/genética , Femenino , Regulación Viral de la Expresión Génica , Humanos , Masculino , MicroARNs , ARN Viral/genética , Neoplasias Gástricas/etiología , Factores de Transcripción , Proteínas de la Matriz Viral/genética , Proteínas de la Matriz Viral/metabolismo

RESUMEN

Epstein Barr virus (EBV) sequence variation is thought to contribute to Burkitt lymphoma (BL), but lack of data from primary BL tumors hampers efforts to test this hypothesis. We directly sequenced EBV from 12 BL biopsies from Ghana, Brazil, and Argentina, aligned the obtained reads to the wild-type (WT) EBV reference sequence, and compared them with 100 published EBV genomes from normal and diseased people from around the world. The 12 BL EBVs were Type 1. Eleven clustered close to each other and to EBV from Raji BL cell line, but away from 12 EBVs reported from other BL-derived cell lines and away from EBV from NPC and healthy people from Asia. We discovered 23 shared novel nucleotide-base changes in the latent membrane protein (LMP)-1 promoter and gene (associated with 9 novel amino acid changes in the LMP-1 protein) of the 11 BL EBVs. Alignment of this region for the 112 EBV genomes revealed four distinct patterns, tentatively termed patterns A to D. The distribution of BL EBVs was 48%, 8%, 24% and 20% for patterns A to D, respectively; the NPC EBV's were Pattern B, and EBV-WT was pattern D. Further work is needed to investigate the association between EBV LMP-1 patterns with BL.

Asunto(s)

Linfoma de Burkitt/virología , Infecciones por Virus de Epstein-Barr/virología , Regulación Viral de la Expresión Génica , Genoma Viral , Herpesvirus Humano 4/genética , Proteínas de la Matriz Viral/genética , África , Secuencia de Aminoácidos , Biopsia , Linfoma de Burkitt/etiología , Linfoma de Burkitt/patología , Niño , Preescolar , Infecciones por Virus de Epstein-Barr/complicaciones , Infecciones por Virus de Epstein-Barr/patología , Femenino , Variación Genética , Genotipo , Herpesvirus Humano 4/patogenicidad , Humanos , Masculino , Datos de Secuencia Molecular , Filogenia , Regiones Promotoras Genéticas , Alineación de Secuencia , Análisis de Secuencia de ADN , América del Sur

RESUMEN

BACKGROUND: Epstein-Barr virus (EBV) is considered to be closely associated with nasopharyngeal carcinoma (NPC), in which EBV-encoded latent membrane protein 1 (LMP1) was found to have an oncogenic role. However, the results published on the LMP1 polymorphism are inconsistent. In the present study, we performed a meta-analysis to determine the frequency of the associations and a more precise association between NPC and EBV LMP1 gene variants (30-bp deletion (del)/XhoI-loss). METHODS: Eligible articles met the inclusion/exclusion criteria and were identified in the following electronic databases: PubMed, ScienceDirect, and SciELO. Consequently, the data of interest were extracted and plotted in a table to calculate the frequency and odds ratio (OR) of the outcomes of interest (30-bp del-LMP1/XhoI-loss) in patients with NPC. Study quality (Newcastle-Ottawa Scale (NOS)), publication bias, and heterogeneity were assessed. RESULTS: Thirty-one observational studies were included with a total of 2,846 individuals (NPC, n = 1,855; control, n = 991). The risk of bias in relation to study quality evaluated by NOS was considered low. The pooled estimate of the frequency of 30-bp del-LMP1 and XhoI-loss in patients with NPC was 77% (95% confidence interval (CI): 72 to 82) and 82% (95% CI: 71 to 92), respectively. There was an association between 30-bp del-LMP1 and NPC susceptibility (OR = 2.86, 95% CI: 1.35 to 6.07, P = 0.00). Similarly, there was an association between XhoI-loss and NPC (OR = 8.5, 95% CI: 1.7 to 41, P = 0.00). However, when we analyze the co-existence of the 30-bp del-LMP1 and XhoI-loss in patients with NPC, there was no association (OR = 1.09, 95% CI: 0.06 to 18.79, P = 0.002). CONCLUSIONS: Our results suggest an association between the 30-bp del-LMP1 and XhoI-loss with NPC susceptibility. However, our data should be interpreted with caution because the sample size was small, and there was heterogeneity between the studies. Thus, future studies are needed with adjusted estimates to simultaneously evaluate multiple factors involved in the development of NPC. SYSTEMATIC REVIEW REGISTRATION: PROSPERO CRD42014013496 .

Asunto(s)

ADN Viral , Herpesvirus Humano 4/genética , Proteínas de la Membrana/genética , Neoplasias Nasofaríngeas/virología , Polimorfismo Genético , Proteínas de la Matriz Viral/genética , Adulto , Carcinoma , Femenino , Humanos , Masculino , Persona de Mediana Edad , Carcinoma Nasofaríngeo

RESUMEN

Influenza virus exhibits two morphologies - spherical and filamentous. Strains that have been grown extensively in laboratory substrates are comprised predominantly of spherical virions while clinical or low passage isolates produce a mixture of spheres and filamentous virions of varying lengths. The filamentous morphology can be lost upon continued passage in embryonated chicken eggs, a common laboratory substrate for influenza viruses. The fact that the filamentous morphology is maintained in nature but lost in favor of a spherical morphology in ovo suggests that filaments confer a selective advantage within the infected host that is not necessary for growth in laboratory substrates. Indeed, we have recently shown that filament-producing variant viruses are selected upon passage of the spherical laboratory strain A/Puerto Rico/8/1934 (H1N1) [PR8] in guinea pigs. Toward determining the nature of the selective advantage conferred by filaments, we sought to identify functional differences between spherical and filamentous particles. We compared the wild-type PR8 virus to two previously characterized recombinant PR8 viruses in which single point mutations within M1 confer a filamentous morphology. Our results indicate that these filamentous PR8 mutants have higher neuraminidase activities than the spherical PR8 virus. Conversely, no differences were observed in HAU:PFU or HAU:RNA ratios, binding avidity, sensitivity to immune serum in hemagglutination inhibition assays, or virion stability at elevated temperatures. Based on these results, we propose that the pleomorphic nature of influenza virus particles is important for the optimization of neuraminidase functions in vivo.

Asunto(s)

Subtipo H1N1 del Virus de la Influenza A/fisiología , Gripe Aviar/virología , Neuraminidasa/metabolismo , Proteínas de la Matriz Viral/genética , Proteínas Virales/metabolismo , Animales , Embrión de Pollo , Pollos , Subtipo H1N1 del Virus de la Influenza A/enzimología , Subtipo H1N1 del Virus de la Influenza A/genética , Mutación Puntual , Virión/metabolismo

RESUMEN

Characterization of canine coronavirus (CCoV) strains currently in circulation is essential for understanding viral evolution. The aim of this study was to determine the presence of pantropic CCoV type IIa in tissue samples from five puppies that died in Southern Brazil as a result of severe gastroenteritis. Reverse-transcriptase PCR was used to generate amplicons for sequence analysis. Phylogenetic analysis of the CCoV-IIa strains indicated that they were similar to those found in other countries, suggesting a common ancestor of these Brazilian isolates. This is the first report of pantropic CCoV-II in puppies from Latin America and our findings highlight that CCoV should be included as a differential diagnosis when dogs present with clinical signs and lesions typically seen with canine parvovirus infection.

Asunto(s)

Infecciones por Coronavirus/veterinaria , Coronavirus Canino/genética , Enfermedades de los Perros/virología , Proteínas de la Matriz Viral/genética , Animales , Brasil , Infecciones por Coronavirus/virología , Coronavirus Canino/clasificación , Coronavirus Canino/aislamiento & purificación , Coronavirus Canino/metabolismo , Perros , Heces/virología , Genotipo , Datos de Secuencia Molecular , Filogenia , ARN Viral/genética , Análisis de Secuencia de ARN/veterinaria

RESUMEN

The Gag polyprotein of feline immunodeficiency virus (FIV) assembles at the plasma membrane of the infected cells. We aimed to identify the FIV Gag domains that interact and promote Gag multimerization. To do this we generated a series of Gag subdomains and tested their ability to associate with full-length Gag and be recruited into extracellular virus-like particles (VLPs). Removal of 37 residues from the C-terminus of FIV Gag and deletion of the N-terminal and central regions of the nucleocapsid (NC) domain attenuated but did not abrogate association with wild-type Gag, whereas a Gag mutant protein encompassing the matrix (MA) and capsid (CA) domains interacted poorly with full-length Gag. Association with wild-type Gag was abolished by deleting most of the NC together with the N-terminal 40 residues of the MA, which most likely reflects the inability of this Gag mutant to bind RNA. Notably, the CA-NC Gag subdomain both associated with wild-type Gag and was recruited into particles in a proportion close to 50 % of the total Gag-related protein mass of VLPs. Moreover, both a Gag protein lacking the C-terminal p2 peptide and a nonmyristoylated version of the polyprotein exhibited a transdominant-negative effect on the assembly of wild-type Gag. Analysis of Gag mutants carrying internal deletions within the CA revealed that the N-terminal and the C-terminal domains of the CA are necessary for Gag assembly. Our results demonstrate that the FIV CA-NC region constitutes the principal self-interaction domain of Gag and that the RNA-binding capacity of Gag is necessary for its multimerization.

Asunto(s)

Productos del Gen gag/genética , Virus de la Inmunodeficiencia Felina/genética , Multimerización de Proteína/genética , Secuencia de Aminoácidos , Animales , Sitios de Unión/genética , Células COS , Cápside/metabolismo , Proteínas de la Cápside/genética , Línea Celular , Membrana Celular/virología , Chlorocebus aethiops , Productos del Gen gag/biosíntesis , Productos del Gen gag/metabolismo , Virus de la Inmunodeficiencia Felina/patogenicidad , Datos de Secuencia Molecular , Nucleocápside/genética , Unión Proteica/genética , Estructura Terciaria de Proteína/genética , Proteínas de Unión al ARN/genética , Ratas , Alineación de Secuencia , Virus Vaccinia/genética , Virus Vaccinia/patogenicidad , Proteínas de la Matriz Viral/genética , Ensamble de Virus/genética

RESUMEN

UNLABELLED: The 2009 H1N1 lineage represented the first detection of a novel, highly transmissible influenza A virus genotype: six gene segments originated from the North American triple-reassortant swine lineage, and two segments, NA and M, derived from the Eurasian avian-like swine lineage. As neither parental lineage transmits efficiently between humans, the adaptations and mechanisms underlying the pandemic spread of the swine-origin 2009 strain are not clear. To help identify determinants of transmission, we used reverse genetics to introduce gene segments of an early pandemic isolate, A/Netherlands/602/2009 [H1N1] (NL602), into the background of A/Puerto Rico/8/1934 [H1N1] (PR8) and evaluated the resultant viruses in a guinea pig transmission model. Whereas the NL602 virus spread efficiently, the PR8 virus did not transmit. Swapping of the HA, NA, and M segments of NL602 into the PR8 background yielded a virus with indistinguishable contact transmissibility to the wild-type pandemic strain. Consistent with earlier reports, the pandemic M segment alone accounted for much of the improvement in transmission. To aid in understanding how the M segment might affect transmission, we evaluated neuraminidase activity and virion morphology of reassortant viruses. Transmission was found to correlate with higher neuraminidase activity and a more filamentous morphology. Importantly, we found that introduction of the pandemic M segment alone resulted in an increase in the neuraminidase activity of two pairs of otherwise isogenic PR8-based viruses. Thus, our data demonstrate the surprising result that functions encoded by the influenza A virus M segment impact neuraminidase activity and, perhaps through this mechanism, have a potent effect on transmissibility. IMPORTANCE: Our work uncovers a previously unappreciated mechanism through which the influenza A virus M segment can alter the receptor-destroying activity of an influenza virus. Concomitant with changes to neuraminidase activity, the M segment impacts the morphology of the influenza A virion and transmissibility of the virus in the guinea pig model. We suggest that changes in NA activity underlie the ability of the influenza M segment to influence virus transmissibility. Furthermore, we show that coadapted M, NA, and HA segments are required to provide optimal transmissibility to an influenza virus. The M-NA functional interaction we describe appears to underlie the prominent role of the 2009 pandemic M segment in supporting efficient transmission and may be a highly important means by which influenza A viruses restore HA/NA balance following reassortment or transfer to new host environments.

Asunto(s)

Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H1N1 del Virus de la Influenza A/fisiología , Neuraminidasa/metabolismo , Infecciones por Orthomyxoviridae/transmisión , Virus Reordenados/genética , Virus Reordenados/fisiología , Proteínas de la Matriz Viral/metabolismo , Proteínas Virales/metabolismo , Animales , Modelos Animales de Enfermedad , Femenino , Cobayas , Glicoproteínas Hemaglutininas del Virus de la Influenza/genética , Glicoproteínas Hemaglutininas del Virus de la Influenza/metabolismo , Subtipo H1N1 del Virus de la Influenza A/aislamiento & purificación , Subtipo H1N1 del Virus de la Influenza A/ultraestructura , Países Bajos , Neuraminidasa/genética , Infecciones por Orthomyxoviridae/virología , Puerto Rico , Genética Inversa , Proteínas de la Matriz Viral/genética , Proteínas Virales/genética , Virión/ultraestructura

RESUMEN

To characterize canine coronavirus (CCoV) circulating in diarrheic puppies in Brazil, 250 fecal samples collected between 2006 and 2012 were tested. By using RT-PCR to partially amplify the M gene, CCoV RNA was detected in 30 samples. Sequence analysis of the M protein grouped eight strains with CCoV-I and another 19 with CCoV-II prototypes. To genotype/subtype the CCoV strains and assess the occurrence of single or multiple CCoV infections, RT-PCR of the S gene was performed, and 25/30 CCoV-positive strains amplified with one or two primer pairs. For 17/25 samples, single infections were detected as follows: six CCoV-I, nine CCoV-IIa and two CCoV-IIb. Eight samples were positive for more than one genotype/subtype as follows: seven CCoV-I/IIa and one CCoV-I/IIb. Sequence analysis revealed that the CCoV-I and IIa strains shared high genetic similarity to each other and to the prototypes. The Brazilian strains of CCoV-IIb displayed an aminoacid insertion that was also described in CCoV-IIb-UCD-1 and TGEV strains. Among the 25 CCoV-positive puppies, five had a fatal outcome, all but one of which were cases of mixed infection. The current study is the first reported molecular characterization of CCoV-I, IIa and IIb strains in Brazil.

Asunto(s)

Infecciones por Coronavirus/veterinaria , Coronavirus Canino/genética , Enfermedades de los Perros/virología , Animales , Brasil , Coinfección , Infecciones por Coronavirus/epidemiología , Infecciones por Coronavirus/mortalidad , Infecciones por Coronavirus/virología , Proteínas M de Coronavirus , Enfermedades de los Perros/epidemiología , Enfermedades de los Perros/mortalidad , Perros , Heces/virología , Genotipo , Datos de Secuencia Molecular , Filogenia , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Análisis de Secuencia , Glicoproteína de la Espiga del Coronavirus/genética , Proteínas de la Matriz Viral/genética

RESUMEN

Patients infected with the human immunodeficiency virus (HIV) are at higher risk of developing Epstein-Barr Virus (EBV)-associated lymphomas. The usefulness of monitoring EBV in peripheral blood mononuclear cells (PBMCs) of patients infected with HIV has not been established. The aim of this study was to evaluate the EBV viral load in PBMCs, the frequency of viral genotypes, and the presence of the 30-bp deletion in the BNLF-1 gene. DNA samples from 156 patients attending the HIV/AIDS Day Clinic at Botucatu School of Medicine, Sao Paulo State University were evaluated. The EBV viral load was detectable by real time PCR in 123/156 (78.8%) cases and was higher in patients not receiving antiretroviral treatment or under therapeutic failure than in patients under successful highly active antiretroviral therapy (HAART) (P = 0.0076). Overall, the profile of patients with high EBV viral load included elevated HIV viremia (P = 0.0005), longer time of HIV diagnosis (P = 0.0026), and increased levels of T CD8 (+) lymphocytes (P = 0.0159). The successful amplification of the EBNA-2 gene by nested-PCR was achieved in 95 of 123 (77.2%) cases, of which 75.8% were EBV-1, 9.5% EBV-2, and 14.7% were co-infected with both EBV-1 and -2. The analysis of the BNLF-1 gene was possible in 99 of 123 (80.5%) cases, of which 50.5% had the 30-bp deletion. EBV-1 was more common than EBV-2, which may reflect the fact that the cohort was predominantly Caucasian and heterosexual.

Asunto(s)

Coinfección , Infecciones por Virus de Epstein-Barr/virología , Genotipo , Infecciones por VIH , Herpesvirus Humano 4/genética , Eliminación de Secuencia , Proteínas de la Matriz Viral/genética , Terapia Antirretroviral Altamente Activa , Recuento de Linfocito CD4 , Femenino , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/inmunología , Infecciones por VIH/virología , Humanos , Leucocitos Mononucleares/virología , Masculino , Carga Viral