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ARS Home Pacific West Area Tucson, Arizona Carl Hayden Bee Research Center Research





- Meikle, William
- Carroll, Mark
- Corby-Harris, Vanessa
- DeGrandi-Hoffman, Gloria
- 2023
- 2022
- 2021
- Publications
- Crop Production (305)

Research Project: Quantifying and Reducing Colony Losses from Nutritional, Pathogen/Parasite, and Pesticide Stress by Improving Colony Management Practices **Location:
                            Carl Hayden Bee Research Center** 2023 Annual Report **Objectives**
                                

Obj. 1: Reduce bee colony losses from nutritional stress by determining the nutrients required to support optimum colony growth and queen health in the spring and summer, and in the fall to prepare bees for overwintering. 
Sub-obj. 1A:Determine nutrient use and storage in bees through the year.
Sub-obj. 1B:Determine the seasonal balances of fatty acid nutrients required to support queen health and physiology, and worker-queen interactions linked to queen productivity and retention in the spring and summer, and in the fall prior to overwintering.

Obj. 2: Determine plant growth conditions and practices that affect the nutrient composition of nectar and pollen for bees.
Sub-obj. 2A:Determine whether cultivars of pollinator-dependent plants with known lipid profiles produce pollens with similar lipid profiles.
Sub-obj. 2B:Determine the effects of plant growth conditions on nectar and pollen secondary metabolites.
Sub-obj. 2C:Determine the effects of growth condition-dependent secondary metabolites on bees.

Obj. 3: Develop methodological and statistical tools for extracting information on bee colony health and activity from continuous sensor data, apply those methods to manipulative field experiments, and relate sensor output to colony performance.
Sub-obj. 3A:Correlate daily patterns of hive weight change, thermoregulation and CO2 levels with colony pest and disease status, environmental factors, gene expression and protein metrics.

Obj. 4: Develop Best Management Practices for placing hives in cold storage that consider nutritional needs, parasite and pathogen spread, and optimal timing to reduce colony losses.
Sub-obj. 4A:Evaluate the impact of placing bee colonies in cold storage for short periods in Sept. to Oct. with respect to colony health, survival, behavior, stress, and pest and pathogen levels.
Sub-obj. 4B:Identify the optimum timing for placing colonies in cold storage to minimize winter losses and maximize populations in spring.
Sub-obj. 4C:Compare survival and growth in the spring for colonies from different latitudes overwintered in cold storage.

Obj. 5: Determine if improved nutrition and overwintering in cold storage reduces the impact of Varroa migration on mite population and virus levels in bee colonies.
Sub-obj. 5A:Determine the relationship between mite populations and Varroa-transmitted virus levels through the year in colonies with and without supplemental pollen feeding.
Sub-obj. 5B:Determine whether a Sept. miticide treatment followed by placing colonies in cold storage in Oct. affects Varroa populations, deformed-wing virus levels, and overwintering survival.

Obj. 6: Quantify the impacts of exposure to agrochemicals on worker bees and queens, and the interactions of those agrochemicals with in-hive treatments against pests and diseases.
Sub-obj. 6A:Measure the impact of the interaction of agricultural pesticides with in-hive pest treatments on colony growth, foraging, thermoregulation and CO2 regulation.
Sub-obj. 6B:Quantify the impacts of agrochemicals and their interactions with in-hive pest treatments on queen physiology, development and replacement, and on worker-queen interactions linked to queen productivity. **Approach**
                                

Objective 1: The components of an artificial diet healthy for bees in the long term remains elusive. This project will address the roles of lipids and secondary metabolites in queen and worker bee health. Lipids are a diverse nutrient class with roles in energy production and physiological homeostasis. Essential fatty acids such as linoleic and linolenic acid, are obtained solely from pollen and needed for gland development, production of worker and royal jelly, and brood and queen rearing. How these essential lipids move through the colony and how that flux is influenced by diet will be explored.
Objective 2: Pollen and nectar contain a variety of secondary metabolites such as thymol and eugenol that are produced in response to biotic and abiotic stressors. Thymol and eugenol are broad-spectrum antimicrobials that inhibit growth of bee pathogens, and both occur in flowers, including nectar. Thymol is also used as a miticide against bee pests. This project will explore how the concentrations of these secondary compounds can be manipulated via environmental conditions, and the effects of those compounds on bee diseases and colony microbiota.
Objective 3: Monitoring colonies using sensors can reveal information on colony genetics, phenology, pesticide exposure and nutrition. Interest in monitoring colonies using sensors is increasing and with it the need to identify which kinds of data, such as weight, temperature and CO2 concentration, are most informative, and the most effective models and methods for extracting information from the data.
Objective 4: The use of cold storage is a common method to preserve and protect colonies. This project will determine whether cold storage has the potential to induce the production of diutinus bees needed for winter survival. Cold storage may also be used to control Varroa mites by inducing colonies to reduce brood production and thus allow more effective treatment of the Varroa. The project will focus on whether the value of the improved treatment efficacy exceeds the cost of the stress on the colonies by monitoring bees on the colony and individual level.
Objective 5: Varroa mites remain a major cause of bee stress worldwide; modern commercial beekeeping often involves treating colonies frequently with miticides, and placing colonies in high densities during pollination events, which puts them in contact with pests and diseases of other colonies. This project will develop recommendations to help reduce Varroa and diseases they transmit by improving colony nutrition and by exploring the application of miticides prior to cold storage to isolate colonies.
Objective 6: Agrochemical exposure is considered an important stressor for honey bees, and have been shown to affect colony growth and activity, as well as queen health. In addition, beekeepers typically apply miticides against bee pests. How these different agrochemicals interact within the hive has seldom been explored. This project will focus on monitoring queen health and worker-queen interactions, as well as sensor-based measures of colony health and behavior, in controlled field studies with field-realistic concentrations of pesticides and miticides. **Progress Report**
                                

This report documents fiscal year (FY) 2023 progress for project 2022-21000-022-000D; titled, “Quantifying and Reducing Colony Losses from Nutritional, Pathogen/Parasite, and Pesticide Stress by Improving Colony Management Practices.”

In support of Sub-objective 1A, year 2 of the planned field experiments were conducted in FY22 (spring 2022) by ARS researchers in Tucson, Arizona, and completed in FY23 (spring 2023). Fifty colonies were established in two different apiaries and were measured monthly for brood and adult population size. Samples were obtained to measure nutrients (total protein and lipid) in brood nest bee (BNB) and forager (F) fat bodies, corbicular pollen (CP) and stored pollen (SP), larvae, drones, and supersedure queens. Colony population data from years 1 and 2 are analyzed. Nutrient levels were measured in the BNB and F fat bodies, SP, CP, and larvae for all of the year 2 samples and the data are analyzed. For year 3, another 25 colonies were established to join the 25 colonies that overwintered from year 3. They are being monitored as above. Monthly sampling is on schedule. With respect to Experiment 2, planned for the end of FY22, researchers did not have the personnel or funds for this experiment. With funding from stakeholders, they were, however, able to conduct field trials in North Dakota on commercial pollen supplements, measuring colony health before and after feeding in the fall and again in the spring after cold storage. Their work in Tucson, Arizona, and in  North Dakota, suggests that later summer and fall (August-Nov) and spring (January-March) are crucial times for colony contraction and expansion, respectively. Lipids are a crucial nutrient for colonies moving into the fall, while protein is more important during colony expansion.

In support of Sub-objective 1B, field colonies were treated (+/-) with supplemental pollen patty during the seasonally critical early fall overwintering preparation period and assessed for queen performance through the early spring.  Supplemental food was not a factor due to plentiful fall forage after a very wet summer monsoon and due to unexpected brood rearing problems. All colonies experienced an unusually sharp reduction in brood production during a prolonged cold period from mid-October to early November during which colonies reared very few replacement workers, despite abundant food stores in all colonies. Half of the colonies in both treatments failed by mid-November with most of the remainder left with small worker populations for overwintering. Only 21% of the colonies survived through spring. Surviving colonies did not differ by treatment in population size, brood rearing, worker quality, queen care (queen retinue size) or queen supersedure attempts, suggesting that supplemental feeding was not a major factor. Colonies did not differ by treatment in how queens were attended by retinue workers, how queens were retained, and how many colonies survived. Queen pheromone and physiological metrics are currently being analyzed. Experiment 1 will be repeated this year under normal-to-dry monsoon conditions and the manuscript for Experiment 1 delayed until Year 4. Experiment 2 will be delayed and run concurrent with Experiment 3 in Years 4 and 5.

In support of Sub-objective 2A, pollen and seed from the year 2 planting (30 Brassica sp. (canola) cultivars) were collected and are being analyzed by ARS researchers for fatty acid content. They will repeat the year 3 planting in FY24, as the project was shifted forward by one year due to a freezer failure in FY22. ARS researchers have obtained pilot data on visitation and handling time for the cultivars; that experiment will be repeated in FY24.

In support of Sub-objective 3A, a second experiment monitoring colony-level behavior of commercial queenlines and the USDA Pol-Line and Russian lines, was completed in collaboration with ARS researchers in Baton Rouge, Louisiana. Hive weight, temperature, and carbor dioxide (CO2) concentrations were monitored for nine months, with colonies being regularly assessed for adult bees and brood, and worker bee samples taken for physiological parameters. A subset of each line was placed in cold storage and monitored over winter. In addition, three cage studies were conducted in the laboratory, comparing the Russian, the Pol-Line and one of the commercial queenlines, and cages monitored for adult bee longevity, food consumption and cluster temperature. A hive monitoring device, developed by researchers at Utah State University, Logan, Utah, was tested on two queenlines to assess flight activity as a queenline-specific character. Another novel device, a photonic fence, was donated to our project for collaborative work with an ARS researcher from Ft. Pierce, Florida. A photonic fence tracks and records all flying insects in a given volume of space. A field experiment was conducted using the photonic fence to compare the flight activity to two different queenlines in the field, and those data are being analyzed. 

In support of Sub-objective 4A, studies to determine how a treatment of 3 weeks cold storage combined with miticide might affect colony health and mite levels, were detailed in a formal paper and submitted for publication. The studies, conducted in collaboration with ARS researchers in Baton Rouge, Louisiana, showed that the combination of 3 weeks of cold storage in October followed by a miticide treatment did not result in lower mite levels, but did show that environmental differences between the two years of the study were more important than the experimental treatments in affecting colony health. That study is being repeated but with the cold storage period moved from October to August, to harmonize the experiment with the methods used by beekeepers in California. Finally, experiments are being conducted with bee colonies in cold storage to explore circadian rhythms in hive temperature and CO2 concentration, and brood production, when a light cycle is introduced. Data on hive temperature and CO2 concentration in cold storage, collected during the original two experiments on Varroa control, have been analyzed and will be included in that work.

In support of Sub-objective 4B, having determined the timing for placing hives in cold storage, ARS researchers examined the role of queen line in cold storage overwintering. The researchers placed Russian and Italian bee colonies located in North Dakota in cold storage in October, and in February used them for almond pollination. Colony sizes were measured before and after cold storage and after almond bloom. THey found that Russian bees were similar to Italian bees in colony size after cold storage and almond bloom. As well, the researchers also found that Russian bees begin rearing brood at comparable levels to Italian bees while in cold storage.

In support of Sub-objective 4C, ARS researchers evaluated the effects of Nosema infections on colonies overwintered in cold storage and used for almond pollination. They tested if diet alone or in combination with antibiotic (Fumagillin) treatment for Nosema prior to cold storage overwintering improved colony survival. Colonies were fed either pollen or protein supplement with 4% added pollen. Fumagillin reduced Nosema spore numbers below the treatment threshold of 1 million spores per bee in all treated colonies. More than 40% of untreated colonies averaged > 1 million spores, and fewer of these were large enough to rent for almond pollination. Fewer of the colonies survived after almond bloom. The researcher's economic analysis showed that treating with Fumagillin can reduce Nosema spore numbers and increase numbers of colonies that are available for almond pollination especially those fed protein supplement with 4% pollen. However, if colonies were fed pollen prior to overwintering, there was no economic advantage to treating them with Fumagillin, indicating that diet can prevent the growth of Nosema to levels that impact bee health and perhaps be used instead of antibiotic treatments. ARS researchers also examined the role of queen line in cold storage overwintering by comparing Russian and Italian bee colony sizes before and after cold storage. They found that Russian bees were similar to Italian bees in colony size after cold storage and almond bloom. The researchers also found that Russian bees begin rearing brood at comparable levels to Italian bees while in cold storage.

In support of Sub-objective 6B, ARS researchers further examined the sublethal effects of amitraz and chlorpyrifos on queens and their attendant worker's behaviors in small scale observation experiments to address recent literature findings. The researchers developed new observation methods to separate sublethal effects on queen productivity (oviposition) from worker brood care, thereby establishing where the most deleterious effects occur. Retinue worker activity and worker brood rearing activities, but not queen oviposition rates, are significantly reduced at high exposures to both amitraz and chlorpyrifos. Workers may partially shield queens from dietary agrochemical exposure since queens receive all their food materials from worker glandular secretions (“jellies”). ARS reserchers are currently preparing a large scale longitudinal study to assess sublethal effects of these agrochemicals on queens and retinue workers in the colony environment later this summer and fall. This modified experiment will include new behavioral observation comparisons intended to capture sublethal effects. After an initial delay, Experiment 1 will be conducted in Years 3 and 4 and the manuscript for Experiment 1 will be delayed until Year 4. **Accomplishments**
                                

1. Cold storage evaluated as part of a Varroa management strategy. Varroa mites feed on developing bees and transmit disease, making them one of the most serious problems in beekeeping. Placing honey bee colonies in cold storage has been proposed as a way to induce a pause in brood production and produce bee colonies with low Varroa mite levels in the spring. In a two-year study colonies were exposed to an October cold storage period and/or a subsequent miticide application. ARS researchers in Tucson, Arizona, found that the full treatment did not reduce spring Varroa levels, but cold storage did stop brood production.  Long-term impacts of cold storage on adult bee populations, colony thermoregulation and stress biomarkers in bees were low compared to, for example, nutritional effects due to yearly differences in bee forage availability.  Although further work is needed to optimize the treatment timing in the late summer and fall, cold storage may be an effective way to reduce brood as part of a Varroa management plan. 2. Increased knowledge of how honey bee colony nutrients change over time. Natural and supplemental diets that are available to hives throughout the year must meet the colony’s nutritional needs, as poor nutrition compromises colony health. ARS researchers in Maricopa, Arizona, characterized the nutrients that were stored and utilized by the bee during key seasonal changes in the colony and conducted field trials with colonies in the Upper Midwest to test the adequacy of fall commercial pollen supplements. For colonies in both the Upper Midwest and Tucson, macronutrients like total lipid and protein correlate with seasonal patterns of hive growth and contraction. The direction and magnitude of these changes are similar, but their timing differs. Our results suggest that lipid storage is key for colony health and survival through the winter, so colonies should be fed more lipids in the fall than are typically available in commercial diets. Colonies that are expanding in the spring need more protein, so they should be fed diets high in protein, which is typical of most commercial diet supplements. 3. Antibiotic treatment for Nosema prior to cold storage overwintering can increase the number of colonies available for almond pollination. Nosema is a gut parasite of honey bees that can reduce overwintering colony size and survival and limit the number of colonies available for almond pollination. ARS scientists in Tucson, Arizona, conducted a study to determine the effects of diet and antibiotic treatment on Nosema infections in colonies overwintered in cold storage and used for almond pollination. Colonies were fed diets of pollen or commercial protein supplement prior to overwintering. Colonies fed each diet were divided into groups that were either treated with antibiotic or were left untreated. Nosema levels in colonies fed either diet were significantly higher in untreated compared with treated colonies, and 40-55% of the untreated colonies had infection levels that reduced colony size and survival after overwintering. The results indicate that diet alone cannot mitigate the effects of Nosema infections, and beekeepers should treat for Nosema prior to cold storage overwintering or risk having fewer colonies to rent for almond pollination. 4. Removal of Varroa mites from whole honey bee colonies by powdered sugar shake dusting. Varroa mites are susceptible to control by dusting treatments that cause mites to lose their grip on their adult worker hosts. ARS researchers in Tucson, Arizona, developed novel powdered sugar dusting techniques that provided a fast, effective, and low impact treatment for mite removal from whole bee colonies. Phoretic mites were removed by shaking a mass of worker bees into a screen box, dusting them with powdered sugar, then shaking the honey bees to dislodge the mites. This powdered sugar shake method removed 92% of phoretic mites from worker honey bees in three weekly applications, a control rate comparable to commercial miticide treatments without chemical residues from the miticides. Powdered sugar applications had no deleterious effects on brood rearing, adult populations, or queens, and as a non-toxic agent, can be used during honey flows. This method removes the mites more rapidly from the honey bees compared to chemical treatments, but is labor intensive. **Review Publications**
                            *Meikle, W.G., Corby-Harris, V.L., Ricigliano, V.A., Snyder, L.A., Weiss, M. 2023. Cold storage as part of a Varroa management strategy: Effects on honey bee colony performance, mite levels and stress biomarkers. Scientific Reports. 13. Article 11842. https://doi.org/10.1038/s41598-023-39095-5.*
                                *Chen, J., Rodriguez Rincon, J., Hoffman, G.D., Fewell, Harrison, J., Kang, Y. 2023. Impacts of seasonality and parasitism on honey bee population dynamics. Journal of Mathematical Biology. 87. Article 19. https://doi.org/10.1007/s00285-023-01952-2.*
                                *Fisher, A., Glass, J.R., Ozturk, C., DesJardins, N., Raka, Y., Hoffman, G.D., Smith, B.H., Fewell, J.H., Harrison, J.F. 2022. Seasonal variability in physiology and behavior affect the impact of fungicide exposure on honey bee (Apis mellifera) health. Environmental Pollution. 311. Article 120010. https://doi.org/10.1016/j.envpol.2022.120010.*


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