Gag-Pol supplied in trans is efficiently packaged and supports viral function in human immunodeficiency virus type 1

The intracellular trafficking and subsequent incorporation of Gag-Pol into human immunodeficiency virus type 1 (HIV-1) remains poorly defined. Gag-Pol is encoded by the same mRNA as Gag and is generated by ribosomal frameshifting. The multimerization of Gag and Gag-Pol is an essential step in the formation of infectious viral particles. In this study, we examined whether the interaction between Gag and Gag-Pol is initiated during protein translation in order to facilitate the trafficking and subsequent packaging of Gag-Pol into the virion. A conditional cotransfection system was developed in which virion formation required the coexpression of two HIV-1-based plasmids, one that produces both Gag and Gag-Pol and one that only produces Gag-Pol. The Gag-Pol proteins were either immunotagged with a His epitope or functionally tagged with a mutation (K65R) in reverse transcriptase that is associated with drug resistance. Gag-Pol packaging was assessed to determine whether the Gag-Pol incorporated into the virion was preferentially packaged from the plasmid that expressed both Gag and Gag-Pol or whether it could be packaged from either plasmid. Our data show that translation of Gag and Gag-Pol from the same mRNA is not critical for virion packaging of the Gag-Pol polyprotein or for viral function.

Effects of concentrated viral communities on photosynthesis and community composition of co-occurring benthic microalgae and phytoplankton

Marine viruses have been shown to affect phytoplankton productivity; however, there are no reports on the effect of viruses on benthic microalgae (microphytobenthos). Hence, this study investigated the effects of elevated concentrations of virus-like particles on the photosynthetic physiology and community composition of benthic microalgae and phytoplankton. Virus populations were collected near the sediment surface and concentrated by tangential flow ultrafiltration, and the concentrate was added to benthic and water column samples that were obtained along a eutrophication gradient in the Brisbane River/Moreton Bay estuary, Australia. Photosynthetic and community responses of benthic microalgae, phytoplankton and bacteria were monitored over 7 d in aquaria and in situ. Benthic microalgal communities responded to viral enrichment in both eutrophic and oligotrophic sediments. In eutrophic sediments, Euglenophytes (Euglena sp.) and bacteria decreased in abundance by 20 to 60 and 26 to 66%, respectively, from seawater controls. In oligotrophic sediments, bacteria decreased in abundance by 30 to 42% from seawater controls but the dinoflagellate Gymnodinium sp. increased in abundance by 270 to 3600% from seawater controls, The increased abundance of Gymnodinium sp. may be related to increased availability of dissolved organic matter released from lysed bacteria. Increased (140 to 190% from seawater controls) initial chlorophyll a fluorescence measured with a pulse-amplitude modulated fluorometer was observed in eutrophic benthic microalgal incubations following virus enrichment, consistent with photosystem II damage. Virus enrichment in oligotrophic water significantly stimulated carbon fixation rates, perhaps due to increased nutrient availability by bacterial lysis. The interpretation of data from virus amendment experiments is difficult due to potential interaction with unidentified bioactive compounds within seawater concentrates. However, these results show that viruses are capable of influencing microbial dynamics in sediments.

Detection of dengue viral RNA in Aedes aegypti (Diptera : Culicidae) exposed to sticky lures using reverse-transcriptase polymerase chain reaction

Active surveillance for dengue (DEN) virus infected mosquitoes can be an effective way to predict the risk of dengue infection in a given area. However, doing so may pose logistical problems if mosquitoes must be kept alive or frozen fresh to detect DEN virus. In an attempt to simplify mosquito processing, we evaluated the usefulness of a sticky lure and a seminested reverse-transcriptase polymerase chain reaction assay (RT-PCR) for detecting DEN virus RNA under laboratory conditions using experimentally infected Aedes aegypti (L.) mosquitoes. In the first experiment, 40 male mosquitoes were inoculated with 0.13 mul of a 10(4) pfu/ml DEN-2 stock solution. After a 7-d incubation period, the mosquitoes were applied to the sticky lure and kept at room temperatures of 23-30 degreesC. Following 7,10,14, and 28 d application, 10 mosquitoes each were removed from the lure pooled and assayed for virus. DEN virus nucleic acid was clearly detectable in all pools up to 28 d after death. A second study evaluated sensitivity and specificity using one, two, and five DEN-infected mosquitoes removed after 7, 10, 14, 21 and 30 d application and tested by RT-PCR. All four DEN serotypes were individually inoculated in mosquitoes and evaluated using the same procedures as experiment 1. The four serotypes were detectable in as few as one mosquito 30 d after application to the lure with no evidence of cross-reactivity. The combination of sticky lures and RT-PCR show promise for mosquito and dengue virus surveillance and warrant further evaluation.

Promiscuous CTL recognition of viral epitopes on multiple human leukocyte antigens: Biological validation of the proposed HLA A24 supertype

Multiple HLA class I alleles can bind peptides with common sequence motifs due to structural similarities in the peptide binding cleft, and these groups of alleles have been classified into supertypes. Nine major HLA supertypes have been proposed, including an A24 supertype that includes A*2301, A*2402, and A*3001. Evidence for this A24 supertype is limited to HLA sequence homology and/or similarity in peptide binding motifs for the alleles. To investigate the immunological relevance of this proposed supertype, we have examined two viral epitopes (from EBV and CMV) initially defined as HLA-A*2301-binding peptides. The data clearly demonstrate that each peptide could be recognized by CTL clones in the context of A*2301 or A*2402; thus validating the inclusion of these three alleles within an A24 supertype. Furthermore, CTL responses to the EBV epitope were detectable in both A*2301(+) and A*2402(+) individuals who had been previously exposed to this virus. These data substantiate the biological relevance of the A24 supertype, and the identification of viral epitopes with the capacity to bind promiscuously across this supertype could aid efforts to develop CTL-based vaccines or immunotherapy. The degeneracy in HLA restriction displayed by some T cells in this study also suggests that the dogma of self-MHC restriction needs some refinement to accommodate foreign peptide recognition in the context of multiple supertype alleles.

Proteasomal targeting of a viral oncogene abrogates oncogenic phenotype and enhances immunogenicity

The ability of viral or mutated cellular oncogenes to initiate neoplastic events and their poor immunogenicity have considerably undermined their potential use as immunotherapeutic tools for the treatment of human cancers. Using an EpsteinBarr virus-encoded oncogene, latent membrane protein 1 (LMP1), as a model, we report a novel strategy that both deactivates cellular signaling pathways associated with the oncogenic phenotype and reverses poor immunogenicity. We show that cotranslational ubiquitination combined with Wend rule targeting of LMP1 enhanced the intracellular degradation of LMP1 and total blockade of LMP1-mediated nuclear factor-kappaB (NF-kappaB) and signal transducer and activator of transcription (STAT) activation in human cells. In addition, although murine cells expressing LMP1 were uniformly tumorigenic, this oncogenicity was completely abrogated by covalent linkage of LMP1 with ubiquitin, while an enhanced CD8(+) T cell response to a model epitope fused to the C-terminus of LMP1 was observed following immunization with ubiquitinated LMP1. These observations suggest that proteasomal targeting of tumor-associated oncogenes could be exploited therapeutically by either gene therapy or vaccination.

IL-10 Mediates Suppression of the CD8 T Cell IFN-γ Response to a Novel Viral Epitope in a Primed Host

Priming to Ag can inhibit subsequent induction of an immune response to a new epitope incorporated into that Ag, a phenomenon referred to as original antigenic sin. In this study, we show that prior immunity to a virus capsid can inhibit subsequent induction of the IFN-gamma effector T cell response to a novel CD8-restricted antigenic epitope associated with the virus capsid. Inhibition does not involve Ab to the virus capsid, as it is observed in animals lacking B cells. CD8-restricted virus-specific T cell responses are not required, as printing to virus without CTL induction is associated with inhibition. However, IL-10(-/-) mice, in contrast to IL-10(+/+) mice, generate CD8 T cell and Ab responses to novel epitopes incorporated into a virus capsid, even when priming to the capsid has resulted in high titer Ab to the capsid. Furthermore, capsid-primed mice, unable to mount a response to a novel epitope in the capsid protein, are nevertheless able to respond to the same novel epitope delivered independently of the capsid. Thus, inhibition of responsiveness to a novel epitope in a virus-primed animal is a consequence of secretion of IL-10 in response to presented Ag, which inhibits local generation of new CD8 IFN-gamma-secreting effector T cells. Induction of virus- or tumor Ag-specific CD8 effector T cells in the partially Ag-primed host may thus be facilitated by local neutralization of IL-10.

Animação Viral, Vanguarda Trash e o Capital Simbólico dos nossos Piores Defeitos: a Ideologia de Bruno Aleixo

"Bruno Aleixo" is a viral animation character, created by the Portuguese collective GANA, that surfaced online in 2008. Their animation works have meanwhile crossed onto the most diverse media, and have been branching out in multiple webs of narratives, constantly referring to each other, as well as constantly quoting disparate references such as film classics, chatrooms and TV ads for detergents. This paper attempts a triple analysis of this object of study: the ways in which technology has been fostering non-linear narratives while widening the available aesthetic spectrum, the ways in which processes of cultural consumerism are being reinvented in light of the web 2.0, and the use of "pseudo-nonsense" as a process of oblique cultural psychoanalysis. We will further attempt to demonstrate how new media and web networks have been contributing to a fragmentation of audiences, as well as a blurring between dominant cultures and sub-cultural phenomena; and we will end by positing that the structural principles behind the "Bruno Aleixo" series can be applied in social and cultural contexts situated at the opposite end of the spectrum of traditional expectations regarding Animation.

Aspectos epidemiológicos da meningite meningocócica no município de São Paulo (Brasil), no período de 1968 a 1974

Estudo epidemiológico da meningite meningocócica no município de São Paulo no período de janeiro de 1968 a junho de 1974. A partir de julho de 1971 uma epidemia causada pelo meningococo sorogrupo C ocorreu na área. Em abril de 1974 foi observado o início de uma segunda epidemia determinada pelo meningococo sorogrupo A. Os maiores coeficientes de morbidade durante o período estudado ocorreram: nos meses frios; nos distritos sanitários da periferia do município; no grupo etário de 0 a 4 anos, no sexo masculino. No primeiro mês das duas epidemias diferiu significantemente o grupo etário mais atingido. A letalidade decresceu de 15,0% em 1968-1970 para 7,9% em 1971-1974. Ela apresentou, durante o período epidêmico, uma variação sazonal no sentido inverso ao da morbidade.

Tendência da sub-notificação de casos no decorrer da epidemia de meningite meningocócica ocorrida no Estado de São Paulo, Brasil, no período de 1971/75

A sub-notificação de casos de meningite no decorrer de uma epidemia de meningite meningocócica foi estudada em uma região do Estado de S. Paulo, Brasil. Os casos conhecidos através dos atestados de óbito (não notificados em vida) foram usados como medida indireta da sub-notificação. Esta, que chegou a 37,6% em 1971, caiu para 2,1% em 1975. Foi possível quantificá-la e demonstrar que seguiu uma função exponencial negativa com o tempo. A aplicação da metodologia apresentada, em amostra casual e probabilística, proporcionará um melhor conhecimento da incidência da doença, mais adequado ao estudo da tendência da epidemia, o que permitirá estimar com maior segurança o impacto epidemiológico provocado pelas medidas adotadas para controle da mesma.

Etiologia viral e bacteriana de casos de gastroenterite infantil: uma caracterização clínica

Numa pesquisa sobre gastroenterites infantis, feita em crianças internadas no Instituto da Criança do Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Brasil, foram identificados rotavírus em 35,2% dos pacientes e bactérias patogênicas em 37,0%; em 38,9% não foi possível identificar rotavírus ou bactérias patogênicas; foram identificados rotavírus em associação com bactérias patogênicas em 11,1% dos casos. Esta pesquisa compreendeu, além do estudo etiológico, outros aspectos, como distribuição etária, estado nutricional, tipo e grau de desidratação e duração média da diarréia.

Avaliação de três cepas de vírus rábico, antigenicamente distintas, em camundongos: II - Estudo da disseminação viral por diferentes órgãos

Estudou-se, comparativamente, o grau de disseminação de três cepas de vírus rábico, duas de origem de cão, Jales e Nigéria, e uma de origem de morcego, DR 19, com perfis antigênicos do nucleocapside distintos. Estas cepas foram inoculadas por via intramuscular, na face interna da coxa, em dois grupos de camundongos, com 21 e 28 dias de idade. Os animais foram mantidos em observação por um período total de 30 dias, e dos animais vitimados pela infecção, foram coletados diferentes órgãos, músculo lingual, coração, pulmão, rim e fígado, além do cérebro e da medula espinal, para avaliar-se o grau de disseminação de cada cepa viral, através da prova de imunofluorescência direta (IFD). Os resultados obtidos evidenciaram que os decalques de cérebro e de medula espinal apresentaram total concordância na prova de IFD, constatando-se as maiores diferenças com as cepas Jales e Nigéria, situando-se a cepa DR 19, intermediariamente, a estas duas. O músculo lingual foi o órgão que apresentou maior freqüência de positividade para ambos os grupos etários e para as três cepas virais.

Aspectos epidemiológicos da meningite tuberculosa em menores de 15 anos de idade, na Grande São Paulo, Brasil, 1982 -1983

Foram estudadas algumas características epidemiológicas dos casos de meningite tuberculosa ocorridos em menores de 15 anos na Grande São Paulo (Brasil), nos anos de 1982 e 1983. O levantamento dos dados foi realizado em fontes oficiais de informação, complementado pela visitação domiciliária. Foram identificados 126 casos, analisados segundo distribuição etária, sexo, fonte de contágio, vacinação BCG, diagnóstico firmado, letalidade hospitalar, seqüelas e eventos ocorridos na seqüência do tratamento. Os resultados mostraram: demora no diagnóstico devido a prováveis falhas assistênciais, alta letalidade, identificação dos focos para a maioria dos casos. Houve dificuldade em avaliar a proteção conferida pela vacinação BCG e na ocorrência de seqüelas, o grupo de menores de 5 anos foi o mais comprometido (83,9%) enquanto que a maior letalidade ocorreu no grupo de 0 a 1 ano de idade (43,1%). Houve 38,9% de cura; 33,3% de óbito; 15,1% de abandono e em 12,7% dos casos alguns permaneceram sob controle; e o restante era desconhecido pelo sistema de controle de notificações.

Epidemiologia da meningite por Streptococcus pneumoniae em área metropolitana, Brasil, 1960-1977

Analisa-se o comportamento epidemiológico das meningites por S. pneumoniae no Município de São Paulo, Brasil, no período 1960-1977. Os dados foram coletados diretamente do prontuário dos pacientes e a confirmação da meningite por S. pneumoniae se fez pela bacterioscopia e/ou pela cultura do líquor. No período 1960-1977 foram confirmados 1.965 casos com um coeficiente médio de 1,9 por cem mil habitantes. As crianças menores de 5 anos contribuíram com 52% dos casos dos quais 38,5% eram menores de um ano. Os coeficientes médios por cem mil habitantes, para os menores de um ano, foram 37,1 e 30,1 para 1960-1969 e 1970-1977, respectivamente. A incidência por cem mil habitantes na zona periférica do município, na primeira década (2,3) foi o dobro daquela da zona central (1,1). Os coeficientes padronizados segundo idade foram 1,6, 1,5 e 2,0 para as zonas central, intermediária e periférica, respectiva-mente. No período seguinte estes valores foram 1,4, 1,5 e 2,0. A letalidade média no período foi de 44%, inversamente proporcional ao número de leucócitos no líquor de entrada. A letalidade entre os menores de um ano foi de 60%.