Flupyradifurone (FPF) is a novel systemic nAChR agonist that interferes with signal transduction in the central nervous system of sucking pests. Despite claims that FPF is potentially “bee-safe” by risk assessments, laboratory data have suggested that FPF has multiple sub-lethal effects on individual honey bees. Our study aimed to expand the studies to the effects of field-realistic concentration of FPF. We found a statistically significant decrease in the survival rate of honey bees exposed to FPF, whereas there were no significantly negative effects on larvae development durations nor foraging activity. In addition, we found that the exposed foragers showed significantly higher expression of ApidNT, CYP9Q2, CYP9Q3, and AmInR-2 compared to the CK group (control group), but no alteration in the gene expression was observed in larvae. The exposed newly emerged bees showed significantly higher expression of Defensin and ApidNT. These results indicate that the chronic exposure to the field-realistic concentration of FPF has negligible effects, but more important synergistic and behavioral effects that can affect colony fitness should be explored in the future, considering the wide use of FPF on crops pollinated and visited by honey bees.

Ascosphaera apis and some Aspergillus species are the main pathogenic fungi of honey bee, and A. apis is the pathogen of chalkbrood disease. However, the infection mechanism of them is incompletely known and it is still unclear whether other factors impact their pathogenesis. In this study, Aspergillus tubingensis were obtained from the chalkbrood bee samples for the first time. Our results showed that A. tubingensis could promote the accumulation of the spores of A. apis. Pathogenicity test found that inoculation of the spores of the two fungi alone or their combination could induce disease characterization of chalkbrood and stonebrood but the extent was less than those in field. To further identify other pathogens impacted the pathogenesis, we found several honey bee viruses presented in the pathogenic fungi A. apis and A. tubingensis, which were different from previous reported. Our results indicated that acute bee paralysis virus (ABPV) and chronic bee paralysis virus (CBPV) could replicate in these two fungi and increased in titer with the going of cultivation time. In addition, CBPV could not only transmit vertically to the next generation by spores, but also spread horizontally to different fungi through hyphal anastomosis. These results suggested that the honey bee chalkbrood contained the other pathogenic fungi besides A. apis, the interactions between different pathogens of chalkbrood microbial communities may influence the prevalence of chalkbrood. Moreover, the discovery of honey bee viruses and their transmission mode in these two fungi enhanced the potential of exploring fungi virus as valuable factors that cause fungal disease outbreak.