Production of juvenile hormone acid o-methyltransferase (JHAMT) knockout mutants using CRISPR-Cas9 in apis mellifera anatoliaca honey bee and investigation of its effect on varroa destructor

Honeybee colony losses, which have been observed in different countries recently, are seen as a problem that may affect the ecological balance of the world in the future. Although many methods of struggle are applied against Varroa destructor, which causes high colony loss, this mite is still the most dangerous threat to the honey bee. For these reasons, scientists and beekeepers continue to search for alternative and definitive solutions in the fight against Varroa. Varroa is an ectoparasite with high host specificity. This mite can distinguish drone larvae from worker larvae, and forager bees from nurse bees. It is known that pheromones and other low-density volatile compounds play a role in larval preference. However, the molecular relationship underlying Varroa's host preference is not yet resolved. Juvenile hormone acid O-methyltransferase (JHAMT) is an enzyme that converts JH acids or inactive precursors of JHs to active JHs in the final step of the Juvenile hormone (JH) biosynthesis pathway in insects. The fact that it is also responsible to produce methyl farnesoate, which can act as a pheromone, has led to the idea that there may be a relationship between the expression of the JHAMT gene and the host preference of Varroa. Based on this idea, it is aimed to develop a new sustainable method in the fight against Varroa in this thesis project. In order to achieve this aim, pedigree family sets were created in Muğla ecotype of Anatolian honey bee (Apis mellifera anatoliaca) within the Muğla Beekeepers' Association, and two generations of knockout mutant queens and worker bees were raised by CRISPR-Cas9 method. In terms of host preference of Varroa, mutant larvae with JHAMT gene silenced and wild-type control larvae were compared in vitro and in vivo. The fact that Varroa did not prefer highly mutant larvae in both in vitro (p=0.002) and in vivo (p=0.037) trials revealed the effect of JHAMT gene on host preference. The results obtained with gene manipulations in the study enabled the development of a new, effective and sustainable solution against the Varroa mite using CRISPR-Cas9 technology.