Date Published: April 18, 2016
Publisher: Public Library of Science
Author(s): Stephanie M. Rainey, Julien Martinez, Melanie McFarlane, Punita Juneja, Peter Sarkies, Aleksei Lulla, Esther Schnettler, Margus Varjak, Andres Merits, Eric A. Miska, Francis M. Jiggins, Alain Kohl, Elizabeth Ann McGraw.
The intracellular endosymbiotic bacterium Wolbachia can protect insects against viral infection, and is being introduced into mosquito populations in the wild to block the transmission of arboviruses that infect humans and are a major public health concern. To investigate the mechanisms underlying this antiviral protection, we have developed a new model system combining Wolbachia-infected Drosophila melanogaster cell culture with the model mosquito-borne Semliki Forest virus (SFV; Togaviridae, Alphavirus). Wolbachia provides strong antiviral protection rapidly after infection, suggesting that an early stage post-infection is being blocked. Wolbachia does appear to have major effects on events distinct from entry, assembly or exit as it inhibits the replication of an SFV replicon transfected into the cells. Furthermore, it causes a far greater reduction in the expression of proteins from the 3´ open reading frame than the 5´ non-structural protein open reading frame, indicating that it is blocking the replication of viral RNA. Further to this separation of the replicase proteins and viral RNA in transreplication assays shows that uncoupling of viral RNA and replicase proteins does not overcome Wolbachia’s antiviral activity. This further suggests that replicative processes are disrupted, such as translation or replication, by Wolbachia infection. This may occur by Wolbachia mounting an active antiviral response, but the virus did not cause any transcriptional response by the bacterium, suggesting that this is not the case. Host microRNAs (miRNAs) have been implicated in protection, but again we found that host cell miRNA expression was unaffected by the bacterium and neither do our findings suggest any involvement of the antiviral siRNA pathway. We conclude that Wolbachia may directly interfere with early events in virus replication such as translation of incoming viral RNA or RNA transcription, and this likely involves an intrinsic (as opposed to an induced) mechanism.
Arthropod-borne viruses (arboviruses) pose a considerable threat to human and animal health, yet effective control measures have proven difficult to implement [1, 2]. In recent years novel means of reducing their replication in arthropod vectors have been suggested as an alternative way to reduce the prevalence of these viruses. One of the most exciting approaches is the use of the endosymbiotic intracellular bacterium Wolbachia to control arbovirus transmission from mosquito to vertebrate from within the arthropod vector [3, 4]. Wolbachia was first found to confer resistance to viruses in Drosophila melanogaster [5, 6]. When it was transferred to the mosquito Aedes aegypti it made the mosquitoes resistant to two important human pathogenic arboviruses, dengue virus (DENV) and chikungunya virus (CHIKV) [7, 8]. Importantly, Wolbachia can also invade and be stably maintained in natural populations thanks to a trait called cytoplasmic incompatibility, which causes embryos to die when uninfected females mate with infected males . This allows Wolbachia to spread through mosquito populations by providing a reproductive advantage to the Wolbachia-infected females that transmit the bacterium . Field trials have shown that releasing Wolbachia-infected mosquitoes allows the bacterium to invade Ae. aegypti populations [11, 12] and reduces the susceptibility of the mosquitoes to DENV .
The bacterial symbiont Wolbachia offers an exciting opportunity in the fight against arbovirus transmission by mosquitoes. Several studies have found that it has antiviral activity in key arbovirus mosquito vectors [8, 28, 44, 45]. However, the exact mechanisms behind this activity are poorly understood. In order for Wolbachia to be used as a long term and sustainable system to control arbovirus spread, it is critical that we understand these mechanisms. By combining two powerful model systems–the model arbovirus SFV, and a Wolbachia-infected D. melanogaster cell line–we were able to show that Wolbachia may protect against virus at a very early stage of infection and appeared to block replication and/or translation of viral RNA. This did not involve an active transcriptional response from either the host, the small RNA pathways or Wolbachia itself.