Managed honey bee (Apis mellifera) populations play a crucial role in supporting pollination of food crops but are facing unsustainable colony losses, largely due to rampant disease spread within agricultural environments. While mounting evidence suggests that select lactobacilli strains (some being natural symbionts of honey bees) can protect against multiple infections, there has been limited validation at the field-level and few methods exist for applying viable microorganisms to the hive. Here, we compare how two different delivery systems—standard pollen patty infusion and a novel spray-based formulation—affect supplementation of a three-strain lactobacilli consortium (LX3). Hives in a pathogen-dense region of California are supplemented for 4 weeks and then monitored over a 20-week period for health outcomes. Results show both delivery methods facilitate viable uptake of LX3 in adult bees, although the strains do not colonize long-term. Despite this, LX3 treatments induce transcriptional immune responses leading to sustained decreases in many opportunistic bacterial and fungal pathogens, as well as selective enrichment of core symbionts including Bombilactobacillus, Bifidobacterium, Lactobacillus, and Bartonella spp. These changes are ultimately associated with greater brood production and colony growth relative to vehicle controls, and with no apparent trade-offs in ectoparasitic Varroa mite burdens. Furthermore, spray-LX3 exerts potent activities against Ascosphaera apis (a deadly brood pathogen) likely stemming from in-hive dispersal differences, whereas patty-LX3 promotes synergistic brood development via unique nutritional benefits. These findings provide a foundational basis for spray-based probiotic application in apiculture and collectively highlight the importance of considering delivery method in disease management strategies.

Honey bees are major pollinators of agricultural and non-agricultural landscapes. In recent years, honey bee colonies have exhibited high annual losses and commercial beekeepers frequently report poor queen quality and queen failure as the primary causes. Honey bee colonies are highly vulnerable to compromised queen fertility, as each hive is headed by one reproductive queen. Queens mate with multiple drones (male bees) during a single mating period early in life in which they obtain enough spermatozoa to fertilize their eggs for the rest of their reproductive life span. The process of mating initiates numerous behavioral, physiological, and molecular changes that shape the fertility of the queen and her influence on the colony. For example, receipt of drone semen can modulate queen ovary activation, pheromone production, and subsequent worker retinue behavior. In addition, seminal fluid is a major component of semen that is primarily derived from drone accessory glands. It also contains a complex mixture of proteins such as proteases, antioxidants, and antimicrobial proteins. Seminal fluid proteins are essential for inducing post-mating changes in other insects such as Drosophila and thus they may also impact honey bee queen fertility and health. However, the specific molecules in semen and seminal fluid that initiate post-mating changes in queens are still unidentified. Herein, we summarize the mating biology of honey bees, the changes queens undergo during and after copulation, and the role of drone semen and seminal fluid in post-mating changes in queens. We then review the effects of seminal fluid proteins in insect reproduction and potential roles for honey bee drone seminal fluid proteins in queen reproduction and health. We finish by proposing future avenues of research. Further elucidating the role of drone fertility in queen reproductive health may contribute towards reducing colony losses and advancing honey bee stock development.

Climate change-related extreme weather events have manifested in the western United States as warmer and drier conditions with an increased risk of wildfires. Honeybees, essential for crop pollination in California, are at the center of these extreme weather events. We associated the maximum daily temperature and air quality index values with the performance of colonies placed in wildfire-prone areas and determined the impact of these abiotic stressors on gene expression and histopathology. Our results indicate that poor air quality was associated with higher maximum daily temperatures and a lower gene expression level of Prophenoloxidase (ProPO), which is tied to immune system strength; however, a higher gene expression level of Vitellogenin (Vg) is tied to oxidative stress. There was a positive relationship between Varroa mites and N. ceranae pathogen loads, and a negative correlation between Varroa mites and Heat Shock Protein 70 (HSP70) gene expression, suggesting the limited ability of mite-infested colonies to buffer against extreme temperatures. Histological analyses did not reveal overt signs of interaction between pathology and abiotic stressors, but N. ceranae infections were evident. Our study provides insights into interactions between abiotic stressors, their relation to common biotic stressors, and the expression of genes related to immunity and oxidative stress in bees.

Honey bee queens undergo dramatic behavioral (e.g., reduced sexual receptivity), physiological (e.g., ovary activation, ovulation, and modulation of pheromone production) and transcriptional changes after they complete mating. To elucidate how queen post-mating changes are influenced by seminal fluid, the non-spermatozoa-containing component of semen, we injected queens with semen or seminal fluid alone. We assessed queen sexual receptivity (as measured by likelihood to take mating flights), ovary activation, worker retinue response (which is influenced by queen pheromone production), and transcriptional changes in queen abdominal fat body and brain tissues. Injection with either seminal fluid or semen resulted in decreased sexual receptivity, increased attractiveness of queens to workers, and altered expression of several genes that are also regulated by natural mating in queens. The post-mating and transcriptional changes of queens receiving seminal fluid were not significantly different from queens injected with semen, suggesting that components in seminal fluid, such as seminal fluid proteins, are largely responsible for stimulating post-mating changes in queens.

Honey bees are the most efficient pollinators of several important fruits, nuts and vegetables and are indispensable for the profitable production of these crops. Health and performance of honey bee colonies have been declining for decades due to a combination of factors including poor nutrition, agrochemicals, pests and diseases. Bees depend on a diversity of plants for nutrition as pollen is the predominant protein and lipid source, and nectar, the source of carbohydrates for larval development. Additionally, pollen and nectar also contain small amounts of plant secondary metabolites or phytochemicals that are primarily plant defense compounds. Bees have coevolved to benefit from these compounds as seen by the improved longevity, pathogen tolerance and gut microbiome abundance in worker bees whose diets were supplemented with select phytochemicals. Here we investigate the impact of four phytochemicals, known to benefit bees, – caffeine, kaempferol, gallic acid and p-coumaric acid, on hypopharyngeal gland (HPG) size of nurse bees. Newly emerged bees were provided with 25 ppm of each of the four phytochemicals in 20% (w/v) sucrose solution and the size of HPGs were measured after a 10 d period. Bees that received p-coumaric acid or kaempferol showed a significant increase in HPG size. A significant decrease in HPG size was seen in bees receiving caffeine or gallic acid. The implication of our findings on worker bee ontogeny, transitioning from nurses to foragers and relevance to foraging related competencies are discussed. It is critical that bees have access to phytochemicals to ensure colony health and performance. Such access could be through natural habitats that provide a diversity of pollen and nectar sources or through dietary supplements for bee colonies.