Wolbachia-Mediated Antiviral Protection in Drosophila Larvae and Adults following Oral Infection

Abstract

Understanding viral dynamics in arthropods is of great importance when designing models to describe how viral spread can influence arthropod populations. The endosymbiotic bacterium Wolbachia spp., which is present in up to 40% of all insect species, has the ability to alter viral dynamics in both Drosophila spp. and mosquitoes, a feature that in mosquitoes may be utilized to limit spread of important arboviruses. To understand the potential effect of Wolbachia on viral dynamics in nature, it is important to consider the impact of natural routes of virus infection on Wolbachia antiviral effects. Using adult Drosophila strains, we show here that Drosophila-Wolbachia associations that have previously been shown to confer antiviral protection following systemic viral infection also confer protection against virus-induced mortality following oral exposure to Drosophila C virus in adults. Interestingly, a different pattern was observed when the same fly lines were challenged with the virus when still larvae. Analysis of the four Drosophila-Wolbachia associations that were protective in adults indicated that only the w1118-wMelPop association conferred protection in larvae following oral delivery of the virus. Analysis of Wolbachia density using quantitative PCR (qPCR) showed that a high Wolbachia density was congruent with antiviral protection in both adults and larvae. This study indicates that Wolbachia-mediated protection may vary between larval and adult stages of a given Wolbachia-host combination and that the variations in susceptibility by life stage correspond with Wolbachia density. The differences in the outcome of virus infection are likely to influence viral dynamics in Wolbachia-infected insect populations in nature and could also have important implications for the transmission of arboviruses in mosquito populations.A rthropods harbor a wide range of viruses that can be transmitted between individuals or populations of the same species or can bridge the interspecies gap to infect plants or other animals. The outcome of viral infections can be modulated by tripartite interactions between arthropods, viruses, and bacteria (1). One such interaction is the tripartite interaction between insects, viruses, and the endosymbyotic bacterium Wolbachia pipientis.Wolbachia spp. have gained much attention due to the antiviral effects they confer to their host. The impact of Wolbachia spp. on virus infection was first described in the Drosophila melanogaster host, where it was shown to protect against mortality induced by diverse viruses, including Drosophila C virus (DCV), cricket paralysis virus, and Flock House virus (2, 3). Since that discovery, Wolbachia-mediated antiviral effects have been demonstrated in a number of insect hosts and are being investigated as a way of limiting spread of arboviruses (reviewed in references 1, 4, 5, and 6). Notably, Wolbachia-mediated antiviral effects have been demonstrated in adult mosquitoes artificially infected with Wolbachia; in mosquitoes, Wolbachia can interfere with accu...

Impact of ERK activation on fly survival and Wolbachia-mediated protection during virus infection

“…While Wolbachia-mediated antiviral protection has been previously demonstrated for Wolbachia-infected WT flies employed in this study (Drosophila melanogaster w 1118-wMel) (Stevanovic & Johnson, 2015), whether the Wolbachia strain used in this study (wMel) (Fig. 2a), which is consistent with previous findings of elevated ROS in Drosophila infected by protective Wolbachia strains (Wong et al, 2015).…”

“…In contrast, survival of Wolbachia-infected WT flies was significantly higher than Wolbachia-free WT flies [log-rank (Mantel-Cox) test; P<0.05] (Fig. 3b), supporting the results shown in Stevanovic & Johnson (2015). Interestingly, following DCV challenge by injection, mortality of Wolbachia-infected ERK (À) and Wolbachia-infected WT flies was significantly delayed compared with Wolbachia-free ERK(À) and Wolbachia-free WT flies, respectively [log-rank (MantelCox) test; ****P<0.0001] (Fig.…”

“…Paired fly lines cured of Wolbachia infection were generated by treating the flies with 0.03 % tetracycline (Hoffmann et al, 1986). Following tetracycline treatment, the gut flora was normalised across all fly lines using previously described methods (Chrostek et al, 2014;Stevanovic & Johnson, 2015). All experiments were conducted a minimum of two generations post-tetracycline treatment.…”

“…Virus used in systemic infection was purified as previously described Johnson & Christian, 1998), suspended in Tris pH 7.4 and maintained in single use aliquots at À20 C. Virus for oral infection was prepared as described in Stevanovic & Johnson (2015). Briefly, flies were either injected with DCV or PBS, collected at 4 days p.i.…”

“…Virus was fed to 4-7-day-old male adult flies as previously described (Stevanovic & Johnson, 2015). Briefly, oral infections were performed in plastic vials containing a piece of filter paper (Whatman) soaked with 150 µl of 75 % virus extract (the DCV titre ranged between 4.4Â10 8 and 2Â10 9 IU ml…”