Inferring plant-bee-microbe associations: Foragers, hive workers, and honey tell complementary stories.

Pollinators, both wild and managed, form diverse associations with plants and microbes which affect the wellbeing of the plants and the pollinators. The method by which these associations are sampled impacts our understanding of the system. The common ways to understand pollinator-plant or pollinator-plant-microbe associations are to observe flower visits of insects, or to collect foraging individuals and identify the pollen and microbes they carry. Honey bees offer a test case for methods of sampling these associations. Hives of managed honey bees host thousands of pollinator individuals together with jointly-collected nectar which is turned into honey. Previous studies have used DNA preserved in honey to infer honey bee associations with plants and microbes. Here, we sampled honey, individual bees while they were foraging, and groups of bees from inside the hive. We identified plants and microbes on the surface of the bees or in the honey using DNA metabarcoding - expecting that bees sampled singly or in groups would reveal a subset of the associations recorded in the communal honey deposits. However, we found that each sample type revealed different aspects of the richness and community composition of plants and microbes encountered by bees. Both honey samples and hive bees had more plant and microbial taxa per sample than samples of individual bees. Though individual bees are subsets of the larger colony, pollen and microbe associations recovered from individual bees did not represent a subsample of associations recovered from groups of hive bees or from honey. Thus, while each sampling technique provides information about honey bee ecology, they are not equivalent. DNA in honey represents time-integrated associations between bees and the surrounding ecosystem; the hive bees provide a snapshot of current colony-level associations, and individual foraging bees capture intraspecific variation in foraging preferences and microbe exposure.