RESUMO
St. Louis encephalitis virus (SLEV), a flavivirus transmitted to humans by Culex mosquitoes, causes clinical symptoms ranging from acute febrile disorder to encephalitis. To reach the central nervous system (CNS) from circulating blood, the pathogen must cross the blood-brain barrier formed by endothelial cells and astrocytes. Because astrocytes play an essential role in CNS homeostasis, in this study these cells were infected with SLEV and investigated for astrogliosis, major histocompatibility complex (MHC)-I-dependent immune response, and apoptosis by caspase-3 activation. Cultures of Vero cells were used as a positive control for the viral infection. Cytopathic effects were observed in both types of cell cultures, and the cytotoxicity levels of the two were compared. Astrocytes infected with a dilution of 1E-01 (7.7E+08 PFU/mL) had a reduced mortality rate of more than 50% compared to the Vero cells. In addition, the astrocytes responded to the flavivirus infection with increased MHC-I expression and astrogliosis, characterized by intense glial fibrillary acidic protein expression and an increase in the number and length of cytoplasmic processes. When the astrocytes were exposed to higher viral concentrations, a proportional increase in caspase-3 expression was observed, as well as nuclear membrane destruction. SLEV immunostaining revealed a perinuclear location of the virus during the replication process. Together, these results suggest that mechanisms other than SLEV infection in astrocytes must be associated with the development of the neuroinvasive form of the disease.
Assuntos
Astrócitos/virologia , Vírus da Encefalite de St. Louis/fisiologia , Animais , Morte Celular , Células Cultivadas , Chlorocebus aethiops , Camundongos , Células VeroRESUMO
BACKGROUND: Influenza remains a major health problem due to the seasonal epidemics that occur every year caused by the emergence of new influenza virus strains. Hemagglutinin (HA) and neuraminidase (NA) glycoproteins are under selective pressure and subjected to frequent changes by antigenic drift. Therefore, our main objective was to investigate the influenza cases in Uberlândia city, Midwestern Brazil, in order to monitor the appearance of new viral strains, despite the availability of a prophylactic vaccine. METHODS: Nasopharyngeal samples were collected from 605 children less than five years of age presenting with acute respiratory disease and tested by immunofluorescence assay (IFA) for detection of adenovirus, respiratory syncytial virus, parainfluenza virus types 1, 2, and 3 and influenza virus types A and B. A reverse transcription-PCR (RT-PCR) for influenza viruses A and B was carried out to amplify partial segments of the HA and NA genes. The nucleotide sequences were analyzed and compared with sequences of the virus strains of the vaccine available in the same year of sample collection. RESULTS: Forty samples (6.6%) were tested positive for influenza virus by IFA and RT-PCR, with 39 samples containing virus of type A and one of type B. By RT-PCR, the type A viruses were further characterized in subtypes H3N2, H1N2 and H1N1 (41.0%, 17.9%, and 2.6%, respectively). Deduced amino acid sequence analysis of the partial hemagglutinin sequence compared to sequences from vaccine strains, revealed that all strains found in Uberlândia had variations in the antigenic sites. The sequences of the receptor binding sites were preserved, although substitutions with similar amino acids were observed in few cases. The neuraminidase sequences did not show significant changes. All the H3 isolates detected in the 2001-2003 period had drifted from vaccine strain, unlike the isolates of the 2004-2007 period. CONCLUSIONS: These results suggest that the seasonal influenza vaccine effectiveness could be reduced because of A H3N2 variants that circulated in 2001-2003 years. Thus, an early monitoring of variants circulating in the country or in a region may provide important information about the probable efficacy of the vaccine that will be administered in an influenza season.