Replication of Rocio virus in primary cultures of mouse neural cells

ABSTRACT Objective This study verified the replication efficiency of the Rocio virus in a primary culture of mouse neural cells. Methods Mixed primary cultures (neurons/glia) obtained from the brains of newborn isogenic BALB/c mice were inoculated with Rocio virus on the 7 th day of culture, and the development of cytopathogenic effects was monitored. The infection was confirmed via immunocytochemistry (anti-ROCV), while viral replication was quantified in infected primary cultures. The titration method used depended on the infection period. Results Rocio virus efficiently infected primary cultured neural cells, with the highest viral titer causing cytopathic changes was observed at 2 days post infection. The virus-infected primary culture survived for up to 7 days post infection, and viral load quantitation showed viral replication kinetics compatible with the cell death kinetics of cultures. Conclusion The findings of this study suggest that mouse neural cell primary cultures support Rocio virus replication and could be used as an alternative system for studying Flavivirus infection in the central nervous system.

Persistence of experimental Rocio virus infection in the golden hamster (Mesocricetus auratus)

Rocio virus (ROCV) is an encephalitic flavivirus endemic to Brazil. Experimental flavivirus infections have previously demonstrated a persistent infection and, in this study, we investigated the persistence of ROCV infection in golden hamsters (Mesocricetus auratus). The hamsters were infected intraperitoneally with 9.8 LD50/0.02 mL of ROCV and later anaesthetised and sacrificed at various time points over a 120-day period to collect of blood, urine and organ samples. The viral titres were quantified by real-time-polymerase chain reaction (qRT-PCR). The specimens were used to infect Vero cells and ROCV antigens in the cells were detected by immunefluorescence assay. The levels of antibodies were determined by the haemagglutination inhibition technique. A histopathological examination was performed on the tissues by staining with haematoxylin-eosin and detecting viral antigens by immunohistochemistry (IHC). ROCV induced a strong immune response and was pathogenic in hamsters through neuroinvasion. ROCV was recovered from Vero cells exposed to samples from the viscera, brain, blood, serum and urine and was detected by qRT-PCR in the brain, liver and blood for three months after infection. ROCV induced histopathological changes and the expression of viral antigens, which were detected by IHC in the liver, kidney, lung and brain up to four months after infection. These findings show that ROCV is pathogenic to golden hamsters and has the capacity to cause persistent infection in animals after intraperitoneal infection.

Eco-epidemiologia dos arbovírus na área de influência da rodovia Cuiabá-Santarém (BR 163), Estado do Pará, Brasil / Arbovirus eco-epidemiology in the area affected by the Cuiabá-Santarém Highway (BR-163), Pará State, Brazil

O presente estudo descreve os aspectos eco-epidemiológicos sobre arbovírus nos Municípios de Novo Progresso e Trairão, Estado do Pará, Brasil, na área de influência da BR 163. Anticorpos IH foram detectados para diferentes arbovírus, com reações monotípicas para os VMAY e VORO, dois importantes arbovírus associados a epidemias na Amazônia. Anticorpos IgM para o VORO e VMAY foram detectados em soros humanos, sugerindo infecção recente por esses arbovírus. Duas cepas do VDEN-3 foram isoladas de pacientes febris residentes em Novo Progresso e identificadas como genótipo III. Em termos gerais, os dados obtidos sugerem uma área propícia para a circulação e manutenção de arbovírus e uma população pouco imunizada. Portanto, é importante um monitoramento dinâmico das populações locais e de imigrantes e de animais silvestres quanto à presença de anticorpos e isolamentos de arbovírus, o que permitirá um efetivo controle das infecções por esses agentes virais em residentes da área da rodovia dentro do território paraense.

The current study describes the eco-epidemiological aspects of arbovirus diseases in the municipalities (counties) of Novo Progresso and Trairão, Para State, Brazil, in the area affected by highway BR-163. Hemagglutination inhibition (HI) antibodies to different arboviruses were detected, with monotypic reactions to MAYV and OROV, two important arboviruses associated with epidemics in the Amazon. IgM antibodies to OROV and MAYV were found in human sera, suggesting recent infections by these viruses. Two DENV-3 strains were isolated from febrile patients in Novo Progresso and identified as genotype III strains. In general, the data suggest that the area displays ideal conditions for maintenance and circulation of arboviruses, plus a population with low immunization levels. Dynamic surveillance of local immigrants and wild animals is thus important, focusing on antibody prevalence and isolation of arboviruses, thereby allowing effective control of infections by these viral agents in the resident population along highway BR-163 in Pará State.