Efeitos de primagem sobre a resposta antiviral da mosca Drosophila melanogaster

Abstract

Insects have the capacity to act as vectors for arbovirus transmission to vertebrate hosts. In recent years, mosquitoes transmitting arboviruses, such as the dengue and Zika viruses have become the focus of studies. During an outbreak insect can be expose to multiples infections by the same arbovirus or other distinct species. Arbovirus may interact with the immune system of the vector for efficient transmission to host. Insects lack adaptive immunity and rely exclusively on innate immune mechanisms. Nevertheless, previous exposure to bacteria and parasites can increase protection to a subsequent challenge in insects and other invertebrates. This phenomenon of memory associated with the innate immune system has been termed immune priming. Here we used Drosophila melanogaster as an insect model to evaluate the priming effect generated by viral infections and identify molecular mechanisms involving on this effect. In our model, we chose Vesicular stomatitis virus (VSV) and Sindibis virus (SINV) because they have distinct strategies of replication. We reported that primary infection by VSV can negatively interfere with secondary infection by the same virus, but not to a different one. Inactived virus did not induces consistently priming effect. In contrast, first infection by live virus strongly interferes with virus from second infection. Since thet viral replication is essential to promotes interference effect, we thought that dsRNA could be the main trigger of this phenomenon. A prior treatment with dsRNA corresponding to essential genes for VSV and SINV strongly inhibits their replication. We also used dsRNA corresponding to the GFP sequence, a reporter gene that is present in the transgenic viruses, VSV-GFP and SINV-GFP, but is dispensable for virus replication. Prior treatment with dsGFP did not interfere with both virus replication. This indicates that the RNA genome is not directly silenced by siRNAs onto AGO2, and others RNA species may be the targets, like transcripts. siRNAi deficient flies were still capable of controlling second virus replication after primary infection by VSV. This result suggested that interference effect is not a kind of innate immune priming and is likely a viral competition. Although we did not detect immune priming in flies, our model allows us to characterize the silencing step of siRNAi pathway. It is possible that the inhibitory mechanism by AGO2 to control RNA virus infection in insects is mediated by silence of viral transcripts rather than viral genome. The study of molecular biology of the main antiviral response in insects can be eventually used to design new strategies for arbovirus controlling in the insect vector.