Generation and Biological Properties of a Recombinant Dodecahedron Containing the Short Fiber Protein of the Human Adenovirus 41

Objective: In order to gain further insight into the function of the enteric adenovirus short fiber (SF), we have constructed a recombinant dodecahedron containing the SF protein of HAdV-41 and the HAdV-3 penton base. Methods: Recombinant baculoviruses expressing the HAdV-41 SF protein and HAdV-3 penton base were cloned and amplified in Sf9 insect cells. Recombinant dodecahedra were expressed by coinfection of High Five (TM) cells with both baculoviruses, 72 h post-infection. Cell lysate was centrifuged on sucrose density gradient and the purified recombinant dodecahedra were recovered. Results: Analysis by negative staining electron microscopy demonstrated that chimeric dodecahedra made of the HAdV-3 penton base and decorated with the HAdV-41 SF were successfully generated. Next, recombinant dodecahedra were digested with pepsin and analyzed by Western blot. A 'site-specific' proteolysis of the HAdV-41 SF was observed, while the HAdV-3 penton base core was completely digested. Conclusion: These results show that, in vitro, the HAdV-41 SF likely undergoes proteolysis in the gastrointestinal tract, its natural environment, which may facilitate the recognition of receptors in intestinal cells. The results obtained in the present study may be the basis for the development of gene therapy vectors towards the intestinal epithelium, as well as orally administered vaccine vectors, but also for the HAdV-41 SF partner identification. Copyright (C) 2011 S. Karger AG, BaselFundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP) [2002/13859-5]Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)Conselho nacional de desenvolvimento cientifico e tecnologico [509838/2010-3]Conselho Nacional de Desenvolvimento Cientifico e Tecnologico'Fond d'intervention' of the Joseph Fourier University (Grenoble)Fond dintervention of the Joseph Fourier University (Grenoble

Gravidade das coinfecções virais em lactentes hospitalizados com infecção por vírus sincicial respiratório

OBJETIVO: Comparar a gravidade de infecções causadas por um único vírus (VSR) com a gravidade de coinfecções. MÉTODOS: Este estudo avaliou uma coorte histórica de lactentes com infecção aguda por VSR. Secreção de nasofaringe foi coletada de todos os pacientes rotineiramente para pesquisa viral usando técnicas de biologia molecular. Os seguintes desfechos foram analisados: tempo total de internação, duração da oxigenioterapia, admissão em unidade de terapia intensiva e uso de ventilação mecânica. Os resultados foram ajustados para os fatores confundidores (prematuridade, idade e aleitamento materno). RESULTADOS: Foram incluídos no estudo 176 lactentes com idade média de 4,5 meses e diagnósticos de bronquiolite e/ou pneumonia. Cento e vinte e um tinham infecção única por VSR, e 55 tinham coinfecções (24 VSR + adenovírus, 16 VSR + metapneumovírus humano e 15 outras associações menos frequentes). Os quatro desfechos de gravidade avaliados foram semelhantes entre o grupo com infecção única por VSR e os grupos com coinfecções, independente do tipo de vírus associado com o VSR. CONCLUSÃO: As coinfecções virais não parecem alterar o prognóstico de lactentes hospitalizados com infecção aguda por VSR

Duplex-PCR assay for the detection of adenovirus and respiratory syncytial virus in nasopharyngeal samples

Human adenovirus (HAdV) and human respiratory syncytial virus (HRSV) are important etiologic agents of acute respiratory infections. In this study, a duplex polymerase chain reaction (PCR) assay was developed for the simultaneous detection of HAdV and HRSV in clinical samples. Sixty previously screened nasopharyngeal aspirates were used: 20 HAdV-positive, 20 HRSV-positive and 20 double-negative controls. Eight samples were positive for both viruses. The duplex PCR assay proved to be as sensitive and specific as single-target assays and also detected the mixed infections with certainty. The identification of both viruses in a single reaction offers a reduction in both cost and laboratory diagnostic time

Infection kinetics of human adenovirus serotype 41 in HEK 293 cells

The purpose of this work was to acquire an overview of the infectious cycle of HAdV-41 in permissive HEK 293 cells and compare it to that observed with the prototype of the genus, Human adenovirus C HAdV-2. HEK 293 cells were infected with each virus separately and were harvested every 12 h for seven days. Infection kinetics were analysed using confocal and electronic microscopy. The results show that, when properly cultivated, HAdV-41 was not fastidious. It had a longer multiplication cycle, which resulted in the release of complete viral particles and viral stocks reached high titres. After 60 h of infection, the export of viral proteins from the infected cell to the extracellular milieu was observed, with a pattern similar to that previously described for HAdV-2 penton-base trafficking after 30 h of infection. HAdV-41 had a non-lytic cycle and the infection spread from the first infected cell to its neighbours. The release process of the viral particles is unknown. The results observed for HAdV-41 infection in HEK 293 cells show how different this virus is from the prototype HAdV-2 and provides information for the development of this vector for use in gene therapy