Avaliação da infecção experimental de neurônios e células da glia do gânglio cervical superior por uma cepa brasileira atípica de Toxoplasma gondii

Abstract

Approximately two billion people are infected chronically with Toxoplasma gondii worldwide, showing unknown consequences. The infection is extremely successful. The parasite is able to cross the biological brain barrier inducing a chronic stage after the establishment of an immune response. The reactivation of the disease in immunocompromised patients causes toxoplasmic encephalitis that can lead to death. The toxoplasmosis treatment is limited and can be toxic for the host. In Brazil, T. gondii shows an uncommon genetic population structure, which have shown to correlate to more severe manifestations of the disease. We used a newly isolated and genotyped virulent strain obtained from the peripheral blood of human newborns diagnosed with congenital toxoplasmosis from 12 regions of Minas Gerais/ Brazil. In order to analyze the in vitro infection of neurons and glial cells by the virulent TgCTBr9 strain we used primary neuron and glial cell cultures, obtained by the removal of the superior cervical ganglia from 1 to 2 days C57BL/6 mice. The cultures were infected with a MOI of 10 and followed for 24, 48, 72, 96 and 192 hours post-infection and then fixed for analysis of neuronal damage, infection rate and immunofluorescence staining for structural and functional markers. We found the parasite in neurons and glial cells in all times analyzed, and the rate of infection in these cells increased with time. At 72 hours of infection we started to detect the expression of components of the cyst wall and this deposition was completed 192 hours post-infection. At this time we also found bradyzoites. From 24 to 96 hours, the neuronal survival rate and the neurite density of infected cultures were not significantly different from their intraexperimental controls. However, evidence for neuronal damage was qualitatively observed at 96 hours post-infection when infection rate was significantly increased. Regarding neuronal function and morphology, we have found that tubulin is expressed around tachyzoites in infected neurons, which express progressive decrease in tyrosine hydroxylase staining intensity over time. Our data shows that the virulent TgCTBr9 strain is not only able to infect neurons and glial cells in vitro, but also undergoes spontaneous encystmen 192hs post-infection. This was an unprecedented result since there are no studies correlating this unusual strain and the pathogenesis of Toxoplasmosis in Brazil.