Nosema ceranae contamination in bee keeping material: the use of ozone as disinfection method

The spore-forming microsporidian Nosema ceranae is considered an emergent and relevant parasite of western honey bees, Apis mellifera. It has been present in Europe since at least 1993 and has been spreading worldwide through commercial movements of bees, hive products, and bee keeping material. Wax and combs contaminated with spores of N. ceranae are an important source of infection. The goal of this work was to evaluate the resistance of N. ceranae spores to various ozone exposure times, both for experimentally inoculated and naturally contaminated wax combs. Spore viability was evaluated, by fluorescent microscopy, using Sytox-Green. The change in spore viability in artificially contaminated wax combs reached 78.18% after 10 h of ozone exposure. A significant reduction in spore viability was observed after the first 60 min of treatment (viability reduction of 20.25%). In naturally contaminated combs, 17.92% of spores were nonviable after a 6-h treatment period. Ozone has proven to be effective in reducing N. ceranae contamination on wax combs and could be proposed as method to minimize the risk of N. ceranae introduction into parasite-free areas. Future studies should focus on optimizing ozonation efficacy by maximizing the surface of combs directly exposed to ozone and by increasing the exposure time until a plateau in spore viability can be observed.