Some insights on honey bee immune response to Deformed Wing Virus, Black Queen Cell Virus and Sacbrood Virus infection

Colony Collapse Disorder (CCD) of honey bee has been widely studied in the past decade and it is nowadays well recognized as a syndrome with multifactorial origin that leads to sudden depopulation of the hives. Among factors leading to CCD, pathogens and parasites may play a major role. According to the literature, the chronic exposure of bee colonies to biotic stressors, such as viruses and parasites, acts in an indirect way by deregulating the immune pathways linked to the antimicrobial response (Toll and Imd pathways). Nevertheless, a better comprehension of the impact of every actor involved is necessary to elucidate the mechanisms that underlie these interactions. In the frame of the Interreg-Alcotra project "Innov'api", which was monitoring the health status of several honey bee colonies managed with two alternative methods for varroa mite control, we selected a subset of samples collected from Piedmontese apiaries in July and September 2019 (before and after the summer acaricide treatment). The selected colonies were characterized by a wide range of viral loads, previously assessed by qPCR analyses carried out on pooled adult bee samples. We then investigated the variability of infection level among individual bees belonging to the same colonies, by focusing on the three most widespread ssRNA+ viruses (Deformed Wing virus, Black Queen Cell virus, Sacbrood virus). The microsporidian Nosema ceranae, another major stressor involved in CCD, was also included in the analysis. On the same samples, we quantified the expression levels of five genes belonging to the Toll and Imd pathways (three antimicrobial peptides, a peptidoglycan receptor and a NF-kB factor) and of vitellogenin, a protein involved in the immunocompetence, in order to investigate their correlations with biotic stress level. Viral loads as well as gene expression values showed very different levels among individuals within the same colony, allowing us to study how these pathogens may affect the individual immune responses. Our multivariate analysis suggests that DWV and BQCV loads positively correlated with the production of antimicrobial peptides such as apidaecin and hymenoptaecin and with the expression level of the NF-kB factor dorsal-1A. Moreover, the type of varroa control treatment had an impact on viral loads and gene expression levels, whereas vitellogenin transcript levels were mostly affected by the sampling time. As the microsporidian N. ceranae was seldom detected, we were not able to evaluate the impact of this pathogen on the regulation of the investigated pathways.