Engineered symbiotic bacteria interfering Nosema redox system inhibit microsporidia parasitism in honeybees

Nosema ceranae is an emergent microsporidia parasite of the European honey bee (Apis mellifera), which causes serious nosemosis implicated in honeybee colony losses worldwide. N. ceranae is an obligate intracellular eukaryotic parasite that mainly invades the midgut of honeybees. Recent studies find that bee gut microbiota is potentially involved in protecting against parasitism. Here, using laboratory-generated bees mono-associated with gut members, we find that Snodgrassella alvi inhibited microsporidia proliferation, potentially via the stimulation of host oxidant-mediated immune response. Accordingly, N. ceranae employs the thioredoxin and glutathione systems to defend against oxidative stress and maintain a balanced redox equilibrium, which is essential for the infection process. We knocked down the gene expression using nanoparticle-mediated RNA interference, which targets the {gamma}-glutamyl-cysteine synthetase and thioredoxin reductase genes of microsporidia. It significantly reduces the spore load, confirming the importance of the antioxidant mechanism for the intracellular invasion of the N. ceranae parasite. Finally, we genetically modified the symbiotic S. alvi to deliver dsRNA corresponding to the genes involved in the redox system of the microsporidia. The engineered S. alvi induces RNA interference and represses parasite gene expression, thereby inhibits the parasitism by up to 99.8%. Specifically, N. ceranae was most suppressed by the recombinant strain corresponding to the glutathione synthetase or by a mixture of bacteria expressing variable dsRNA. Our findings extend our previous understanding of the protection of gut symbionts against N. ceranae and provide a symbiont-mediated RNAi system for inhibiting microsporidia infection in honeybees.