Anoxia tolerance is strongly correlated with tolerance to heat, desiccation, hyperosmotic shock, freezing, and other general stressors, suggesting that anoxia tolerance is broadly related to stress tolerance. Age affects the capacity of many animals to survive anoxia, but the basis to this ontogenic variation is poorly understood. We exposed adult Drosophila, 1, 3, 5, 7, 9, and 12 days past eclosion, to six hours of anoxia and assessed survival 24-hours post-treatment. Survival of anoxia declined strongly with age (from 80% survival for one-day-old flies to 10% survival for 12 day-old-flies), a surprising result since adult fly senescence in Drosophila is usually observed much later. In anoxia, adenosine triphosphate (ATP) levels declined rapidly (< 30 min) to near-zero levels in both 1 and 12-day old adults; thus the higher anoxia-tolerance of young adults is not due to a better capacity to keep ATP elevated. Relatively few physiological parameters are reported to change over this age range in D. melanogaster, but gut bacterial content increases strongly. As a partial test for a causal link between bacterial load and anoxia tolerance, we replaced food daily, every third day, or every sixth day, and assayed survival of six hours of anoxia and bacterial load at 12 days of age. Anoxia tolerance for 12-day old flies was improved by more food changes and was strongly and negatively affected by bacterial load. These data suggest that increasing bacterial load may play an important role in the age-related decline of anoxia tolerance in Drosophila.

The interaction between a virus and its host is a constant competition for supremacy. Both the virus and the host immune system constantly evolve mechanisms to circumvent one another. Vaccinia virus (VACV) infections are a prime example of this. VACV contains a highly conserved innate immune evasion gene, E3L, which encodes the E3 protein composed of a Z-NA-binding domain (Z-NA BD) in the N terminus and a highly characterized dsRNA binding domain in the C-terminus. Both domains of E3 have been found to be essential for the inhibition of antiviral states initiated by host type 1 IFNs. However, the mechanism by which the Z-NA-BD of E3’s N-terminus confers IFN resistance has yet to be established. This is partially due to conflicting evidence showing that the Z-NA-BD is dispensable in most cell culture systems, yet essential for pathogenicity in mice. Recently it has been demonstrated that programmed necrosis is an alternative form of cell death that can be initiated by viral infections as part of the host’s innate immune response to control infection. The work presented here reveals that VACV has developed a mechanism to inhibit programmed necrosis. This inhibition occurs through utilizing E3’s N-terminus to prevent the initiation of programmed necrosis involving the host-encoded cellular proteins RIP3 and Z-NA-binding protein DAI. The inhibition of programmed necrosis has been shown to involve regions of both the viral and host proteins responsible for Z-NA binding through in vivo studies demonstrating that deletions of the Z-NA-BD in E3 correspond to an attenuation of pathogenicity in wild type mice that is restored in RIP3- and DAI-deficient models. Together these findings provide novel insight into the elusive function of the Z-NA-binding domain of the N-terminus and its role in preventing host recognition of viral infections. Furthermore, it is demonstrated that a unique mechanism for resisting virally induced programmed necrosis exists. This mechanism, specific to Z-NA binding, involves the inhibition of a DAI dependent form of programmed necrosis possibly by preventing host recognition of viral infections, and hints at the possible biological role of Z-NA in regulating viral infections.