Biological And Pathological Mechanism Of Ascosphaera Apis Infecting Honeybee Larvae

Bee Chalkbrood Disease, caused by a pathological fungi (Ascosphaera apis), mainly infect the honeybee brood and lead to death. The present research on chalkbrood are mainly on Ascosphaera apis classification, biological character, the optimal cultivating conditions, prevalence situation and resistant bee breeding etc. But the pathogenic mechanism is still not clear so far and no effective control measures developed. Therefore, the disease was serious and prevalent around the world now. In order to provide theoretical basis for the disease control, the pathogenic mechanism of Ascosphaera apis were studied by feeding larvae with spores of Ascosphaera apis in laboratory under different situation such as suffering chilling in different times. The results showed that the bee larvae would be successfully infected as long as enough spors injested. Larvae with different ages suffering from the chilling have no effect on the infection. Mycelia eventually breaking out the posterior end of the larva at 7~8 days. Ascosphaera apis infection of honeybee larvae from the resistant colony and the susceptible colony is same. The infection mainly relates to the development stage of larvae, especially the transition stage from larvae to pupal period.In ordder to futher explore the infection process of Ascosphaera apis, the bee larvae pathological section was prepared. The results showed that the spors of Ascosphaera apis germinated in lumen of the midgut, and then grew slowly. During pre-pupal period, the midgut is connected with hindgut. Mycelia together with the peritrophic membrane (PM) moved to hindgut. At that time, honeybee larvae is undergoing a tissue reconstruction, the larvae tissue is disintergrating and the pupae tissue is constructing. There is only stroma layer remained in gut with no peritrophic membrane secreted. Hyphae in the lumen of hindgut penetrated the stroma layer and then proliferated quickly in bee larvae. In a day, hyphae were full in bee larvae, and eventually breaking out the posterior end of the larva.Moreover, roles of the extracellular enzymes of Ascosphaera apis in infecting bee larvae was delved. The hemolymph from the healthy larvae and the Ascosphaera apis infected larvae were collected and subjected to SDS-PAGE and zymogram analysis of protease, esterase, diastase. The result showed that protein content in healthy bee larvae hemolymph is high and composed of four main proteins with high molecular weight, while the hemolymph protein content of Ascosphaera apis infected larvae is low and composed of low molecular weight protein with molecular weight less than 30kDa. And the same time, the activity of protease and esterase of Ascosphaera apis were detected, but no diastase activity detected. Such results shows that Ascosphaera apis could secreted protease and esterase to degrade the bee larvae tissue for its growth. The process affected the normal development of bees and eventually led the larvae to die.In conclusion, the fungi Ascosphaera apis fully take advantage of the immune deficiency and high nutrition in metamorphosis from larvae to pupae to sucessfully infect honeybee larvae, which is also the reason of Ascosphaera apis specifically infecting honeybee. It is also the results of long time competition between the Ascosphaera apis and its host honeybee.