Constitutive and therapeutic benefits of plant resins and a propolis envelope to honey bee, Apis mellifera L., immunity and health

Abstract

Honey bees (Apis mellifera L.), like many social insects, have collective behavioral defenses called “social immunity” to help defend and protect the colony against pathogens and parasites. One example of social immunity is the collection of plant resins by honey bees and the placement of the resins on the interior walls of the nest cavity, where it is called a propolis envelope. Propolis is known to have many antimicrobial proprieties against bacteria, fungi, and viruses and has been harvested from bee hives for use in human medicine since antiquity. However the benefit of propolis to honey bees has not been studied until recently. This dissertation research focused on how bees collect and use the antimicrobial properties of plant resins within the hive as a form of social immunity and defense against infectious bacterial and fungal pathogens. In the first experiment, the benefit of a naturally constructed propolis envelope to individual bee health was assessed by quantifying the gene expression of immune-related genes via real-time PCR, and to colony health by measuring colony strength, pathogen and parasite load of large field colonies. The presence of a propolis envelope within hives of apparently healthy colonies directly affected individual bee health by decreasing the baseline and variability in expression of immune-related genes (such as hymenoptaecin and abaecin) throughout the active foraging season. As the immune system is one of the most costly physiological systems to maintain in animals, a decrease in energetic costs associated with the maintenance of an up-regulated immune system helps bees allocate their energy toward vital tasks (e.g. foraging, rearing brood) and maintain higher storage protein levels in their bodies required for overwintering success. The propolis envelope also benefited colony strength in the spring and increased colony survivorship in one year of the study. In a second experiment, after colonies were challenged with a highly infectious brood bacterial pathogen, Paenibacillus larvae, nurse bee immune system activity and the antimicrobial activity of larval food (fed to young larvae by nurse bees), were both higher when challenged colonies had a propolis envelope compared to when they did not have the envelope. The immune system activity of nurse bees was measured via real-time PCR, using primers for 3 honey bee antimicrobial peptides (hymenoptaecin, apidaecin and defensin-1). A bacterial growth assay was performed to assess the inhibitory activity of larval food from 1-2 day old larvae against the growth of P. larvae. Colonies with a propolis envelope had reduced level of American foulbrood clinical signs (caused by P. larvae) two months following challenge, which was likely due to a combination of the effects of propolis on both the collective and individual behavioral responses (larval food bioactivity and individual bee immune response). The results of this experiment reveal a novel therapeutic effect of the propolis envelope and a protective physiological response of nurse bees towards the brood. The third experiment explored the role of resin collection by honey bees as a general vs. specialized immune defense against the two highly infectious brood pathogens, Ascosphaera apis (a fungal pathogen that causes chalkbrood) and P. larvae (a bacterial pathogen that causes American foulbrood). The hypothesis was tested that bees self-medicate with resin in response to infection with either pathogen. Results from three years of data suggested that bees significantly increased resin collection, that is, self-medicated the colony with resin, in response to A. apis challenge, but not in response to challenge with P. larvae. We also tested the hypothesis that bees may shift their selection of resin sources at the colony-level after challenge with the fungal or bacterial pathogen, and, if so, how the antimicrobial activity might differ between the pre- and post-challenge resin plant sources. Resin loads from each bee were analyzed by reverse-phase liquid chromatography mass spectrometry (LC-MS) to identify the plant sources of resin. The inhibitory activity of each resin source against A. apis and P. larvae was quantified using dilution assays for each pathogen. After challenge with either pathogen, colonies increased the number of foragers collecting resin from the plants they were already visiting, and not necessarily from the most inhibitory resin. This study sheds light on the complex way in which colony-level behavioral defenses contribute to diminish pathogen infection, and on the role of resins as pharmacological agents in the ecology and evolution of plant-animal interactions. Further research will be necessary to investigate whether bees self-medicate with resin based on resin quality or available quantity. In all, these studies demonstrate the significance of the propolis envelope as a crucial component of the nest architecture in honey bee colonies. The collection and deposition of resins into the nest architecture impacts individual immunity, colony health, and induces honey bees antimicrobial defenses. These results emphasize the importance of resin to bees and show that plants are not only a source of food but can also be "pharmacies."