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ARS Home Pacific West Area Albany, California Western Regional Research Center Invasive Species and Pollinator Health Research





- Fine, Julia
- Pratt, Paul
- 2023
- 2022
- 2021
- Publications
- Crop Production (305)

Research Project: Conduct Longitudinal Studies on Colony Performance and Explore Near-term Effects of Nutritional and Agrochemical Stressors on Honey Bee Health **Location:
                            Invasive Species and Pollinator Health** 2023 Annual Report **Objectives**
                                

As pollinators, honey bees (Apis mellifera) are the third most important agricultural livestock after cattle and pork, indispensable for crop production and global food security. Despite their significant economic importance, honey bees continue to face debilitating challenges from several interactive factors including poor nutrition, exposure to agrochemicals, reduced genetic diversity, devastating pests and pathogens and ongoing changes to climatic conditions. A common underlying thread is that these factors vary across seasons and across years, and their build-up leads to compounding impacts that result in patterns not easily discerned with short-term datasets. Depending on agricultural practices, some of the factors may go through cyclical trends that affect long-term performance of apiaries. To get a comprehensive understanding of factors determining honey bee health, it is necessary to obtain continuous recording of data on colony performance, survivorship, environmental factors surrounding the apiaries, agrochemical residues, and availability of nutritional forage, over long periods of time. This project will establish a Long-Term Honey Bee Research (LTHBR) system in California at the Davis ARS location, associated with the Invasive Species and Pollinator Health Research (ISPHR) unit. This LTHBR system will elucidate how key components involved in honey bee health and pollinator sustainability interact with the demand for pollination and agricultural practices. The data collected will generate insights into whether sustainable intensification of beekeeping can occur through better mitigation of stress-induced hive declines. This LTHBR system will also support the development of research projects that monitor a range of environmental conditions and correlated episodic events such as changes to nutrient flow, pest-pathogen cycles, and changing weather patterns affecting hive performance. The data collected over several years will enable the development and validation of theoretical and empirical models to forecast colony performance across various scenarios, allowing for strategies to improve pollinator health and sustainability.
The following are the objectives and sub-objectives of the project plan: 

Objective 1: Establish longitudinal monitoring of apiaries to collect long-term data on colony performance and evaluate relative benefits of new management discoveries to improve honey bee health. 

Sub-objective 1A: Establish a Long-Term Honey Bee Research (LTHBR) system in California. 

Sub-objective 1B: Quantify the expression of hygienic behavior across the cooperator apiaries in the LTHBR system. 

Objective 2: Understand the effects of nutritional and agrochemical stressors on honey bee health and develop hive management strategies. 

Sub-objective 2A: Determine performance of honey bees under nutrient conditions relevant to California agriculture. 

Sub-objective 2B: Characterize how exposure to agrochemical stressors like IGRs affect honey bee reproduction, development and long-term colony stability to help beekeepers predict and mitigate the long-term consequences of agrochemical exposure. **Approach**
                                

The primary objective of this project plan is to conduct longitudinal monitoring of apiaries and collect long-term data on colony performance. As a part of Sub-objective 1A, the LTHBR system will be established in California in collaboration with commercial beekeepers, such that the different monitoring locations will be spread over the different beekeeping regions within the state. Research colonies will be established in cooperator apiaries and monitored by the Davis, California, ARS Bee lab scientists. Using a combination of laboratory and field methods, the following parameters will be recorded over the entire duration of the project plan: (1) colony performance parameters including weight, brood and food storage areas, adult bee population, queen laying patterns, honey and pollen storage areas, and prevalence of pests and pathogens including viruses, (2) reproductive performance parameters measured on queens and drones including viability of the sperm in the spermathecae of queens and in the semen of drones and (3) apiary parameters including beekeeper operating costs, their profit margins and available floral resources and their bloom time in the vicinity of the apiaries. To determine the efficacy of automated hive monitoring technologies, inhive sensors will be installed in the experimental hives. Hive performance measures recorded by the automated devices will be compared with the parameters recorded by researchers during the same time periods in the same apiaries. As a part of Sub-objective 1B, the expression of hygienic behavior, an important a form of behavioral resistance to American Foul Brood (AFB) and a behavioral defense against chalkbrood, will be quantified in the colonies of the participating stakeholder apiaries and the impact of nutrition on behavioral expression will be determined using the established Freeze Killed Brood assay. The second objective of the project plan is to determine the effects of nutritional and agrochemical stressors on honey bee health. Towards this goal, field and laboratory studies in Sub-objective 2A will determine the impact of monocrop and multi-floral pollen diets on colony-level performance measures and individual bee-level behavioral parameters. To determine the effects of agrochemical stressors on honey bee reproduction and development, laboratory studies in Sub-objective 2B will explore how Insect Growth Regulators used in almond orchards affect honey bee queen fecundity and the survival and performance of offspring. **Progress Report**
                                

For Sub-objective 1A aimed at establishing long term honey bee research (LTHBR) sites, two additional beekeeper cooperators were added to the monitoring network. Additional apiary locations for research colonies were identified in Davis, California. All colonies continue to be monitored for their health and performance by recording frames of bees, egg laying, colony growth and other health parameters in each of the experimental colonies. These efforts will continue through the year adding to the long-term data being collected. Queens continue to be analyzed for efficacy through factors including total number of sperms in spermatheca, percent sperm viability and percent spermatheca filled, which yield information on queen quality. Preliminary analyses of data collected in years one and two are also underway.

For Sub-objective 1B that aims to determine whether nutrition impacts the expression of hygienic behavior and colony performance, we have identified the apiary where experiments will be conducted. In addition to supplementing colonies with the treatment diets, experiments to monitor survival of adult bees on different treatment diets, in a laboratory setting are in progress. For evaluating hygienic behavior as affected by nutrition, we are continuing to standardize the protocols used to conduct the test.

Experiment 1 of Sub-objective 2A aims to determine forager preference of pollen from different crops grown in California. ARS researchers in Davis, California, compared behavioral preferences of honey bee foragers provided with a choice of mixed cover crop pollen and pollen from monocrop fields. Scientists observed that foragers prefer pollen from mixed cover crop habitat that includes a variety of different plant species. Scientists have compiled behavioral data and nutritional composition of the different pollens used in the choice assay. A manuscript describing the findings and its implications for honey bee nutrition has been submitted to journal for peer review. 

Experiment 2 of Sub-objective 2A aims to determine the impact of monocrop diet and a mixed source pollen diet on behavior in honey bees. ARS researchers have adapted protocol from Experiment 1 above to raise adult bees in hives provided with controlled diet - mixed cover crop pollen and pollen from monocrop fields. Laboratory setup and standardization of protocols for learning and retention assays are underway. Experiment 2 of Sub-objective 2B aims to assess the effects of insect growth disrupting chemicals on queens and their offspring. Based on previous results, insect growth disrupting chemicals known to affect the hatching rates of eggs laid by exposed queens were selected for further study. Cages containing workers and one queen each were exposed to lower doses of the chemicals. Scientists did not observe changes in the behavior of exposed workers towards queens, but treating queens with pyriproxyfen increased the hatching rates of her eggs, and her offspring displayed higher rates of queen attendance behavior. Proteomics analysis of the exposed queens’ ovaries demonstrated significant changes in the expression of protein groups in the queens treated with pyriproxyfen relative to control. An experiment is currently underway to further study the unexpected positive effects of this treatment and identify any changes in the physiology of the resultant offspring.

Progress with sample analyses continues in the subordinate project 2030-21000-055-005S (Modeling honey bee exposure to pesticides in pollination dependent crops of California). DNA metabarcoding to identify plant sources of pollen has been performed and pesticide residue analysis is ongoing. The findings of this subordinate project relate to main project objectives that target long-term factors affecting honey bee health in California. Prevalence of pesticides and diversity of pollen sources are two critical long-term factors impacting honey bee colony performance. Satisfactory progress on the subordinate project 2030-21000-055-003S (Analyzing Factors Contributing to Long-term Honey Bee Health and Hive Performance) was achieved. This includes standardizing protocol for collecting samples for histopathological analyses and recording performance metrics for colonies placed in areas prone to wildfires in California. ARS scientists in Davis, California, completed the comparison of different protocols for collecting and storing samples for histopathological analysis. Routine formalin fixation was found to be adequate, and immobilizing bees with indirect dry ice exposure aids in sample collection without negatively impacting the quality of histologic sections. This protocol was further utilized to determine the overlap between histopathological findings and molecular studies demonstrating gene expression and the analyses suggest that poor air quality from wildfires has a positive association with higher maximum daily temperatures and lower expression level of genes tied to honey bee immune system strength. Furthermore, poor air quality is associated with higher expression level of genes tied to oxidative stress. The results of these studies will add to the baseline data for scientists to compare the effects of wildfires and climate change-related extreme temperature events on honey bee colony performance. Satisfactory progress has been made on the subordinate project 2030-21000-055-001S (Analyzing Nutritional Content of Pollen in Relation to Long-term Honey Bee Colony Health). The findings from the metabolomic analyses indicate that the pollen from mixed species inter row plantings (MSP) preferred by foragers was chemically diverse and richest in cholesterol, vitamins, and phytochemicals, but was not consistently high for essential amino acids and polyunsaturated fatty acids. Although diversity in chemical profiles may not directly relate to plant species diversity, these results suggest that preference for MSP pollen adds credence to the importance of diverse floral resources in the habitat. These results relate to the changes in the landscape nutritional quality as beehives are placed in monocrop fields to render pollination services, thus restricting dietary diversity of bee hives. **Accomplishments**
                                

1. Juvenile hormone positively affects aspects of honey bee reproduction. Exposing queen honey bees to field-relevant doses of insect growth disruptors through worker diet does not negatively affect the queen’s reproductive output or the health and behavior of her worker offspring. However, a positive effect on egg hatching rates and queen acceptance behaviors was observed by ARS researchers in Albany, California, in the offspring of queens exposed to a juvenile hormone mimic, suggesting juvenile hormone may have a beneficial effect on honey bee queen fecundity. This finding may lead to the development of a novel queen treatment that will enhance the longevity of queens and the stability of honey bee colonies which could be used by beekeepers to promote healthy hives. 2. Poor air quality is linked to lowered immune system functions in honey bees. Higher daily maximum temperatures in central California are related to poorer air quality. Honey bees, essential for crop pollination, are likely to be impacted. ARS researchers in Davis, California, in collaboration with university scientists, report that in honey bees, poor air quality was associated low expression of genes tied to immune system strength and high expression level of genes tied to oxidative stress. There was a negative correlation between Varroa mite loads and heat shock protein gene expression, suggesting that mite-infested colonies may have a limited ability to buffer against extreme temperatures. This finding provides insights into aspects of honey bee health that may be directly compromised with the ongoing climate change related stressors. 3. Honey bee foragers prefer mixed species pollen that is rich in vitamins, essential amino acids and phytochemicals. Honeybees providing pollination services are regularly exposed to a monoculture diet. ARS researchers in Davis, California, have shown that, when given a choice, bees prefer pollen from mixed species intercrops. Analyses of pollen from almond orchards, sunflower fields and from mixed species intercrops showed that mixed species pollen is rich in important dietary compounds known to promote healthy honey bees. This result lends support to the need for pollinator habitats and intercrop plantings in almond orchards and other agricultural fields. **Review Publications**
                            *Bernklau, E., Seshadri, A.H. 2023. Seasonal patterns of beneficial phytochemical availability in honey and stored pollen from honey bee colonies in large apiaries. Journal of Economic Entomology. 116(4):1069-1077. https://doi.org/10.1093/jee/toad096.*
                                *Seshadri, A.H., Smith, T. 2023. Drought and temperature stresses impact pollen production and autonomous selfing in a California wildflower, Collinsia heterophylla. Ecology and Evolution. 13(7). Article e10324. https://doi.org/10.1002/ece3.10324.*
                                *Lastro-Nino, E., Yokota, S., Stacy, W.H., Seshadri, A.H. 2022. Dietary phytochemicals alter hypopharyngeal gland size in honey bee (Apis mellifera L.) workers. Heliyon. 8(9). Article e10452. https://doi.org/10.1016/j.heliyon.2022.e10452.*
                                *Heller, S., Fine, J.D., Phan, N., Rajotte, E., Biddinger, D., Joshi, N. 2022. Toxicity of formulated systemic insecticides used in apple orchard pest management programs to the honey bee (Apis mellifera (L.)). Environments. 9(7). Article 90. https://doi.org/10.3390/environments9070090.*


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