SUNFLOWERS AS TREATMENT AND PREVENTATIVE FOR BUMBLE AND HONEY BEE PATHOGENS

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: COMPLETE
• Funding Source: AFRI COMPETITIVE GRANT
• Reporting Frequency: Annual
• Accession No.: 1012092
• Grant No.: 2017-68004-26325
• Cumulative Award Amt.: $999,960.00
• Proposal No.: 2016-07962
• Project Start Date: Apr 15, 2017
• Project End Date: Apr 14, 2022
• Grant Year: 2017
• Program Code: [A5170]- New Frontiers in Pollinator Health: From Research to Application

Recipient Organization
UNIV OF MASSACHUSETTS
(N/A)
AMHERST,MA 01003

Performing Department
Biology

Non Technical Summary
Pollinators provide billions of dollars in crop pollination annually, and parasites have been implicated in declines of many bee species. Nutrition, including diet quality as well as quantity, is an important factor in bee health. We found that sunflower pollen dramatically reduced Crithidia infection in bumble bees and Nosema infection in honey bees, suggesting the potential for sunflower pollen supplements and plantings as easily-implemented approaches to improving bee health. Our research goals are to evaluate the benefits and costs of sunflower pollen supplements on bee colonies, assess the impacts of sunflower pollen supplements on pathogen resistance in the field, and determine the degree to which sunflower plantings affect parasite infection in bees. Each objective will include bumble and honey bees, and honey bee objectives will take advantage of Bee Informed Partnership data and directly include beekeepers in assessment. In addition to directly engaging beekeepers in our research,extension goals include hiring a Pollinator Extension Educator who will use a new state apiary, engaging an Advisory Board representing diverse stakeholders, and conducting a wide range of outreach activities to reach local, regional and national audiences. Our proposal is directly relevant to Priority 1, "Development and evaluation of strategies to reduce pollinator declines caused by factors such as habitat changes or loss, nutritional imbalances, pathogens..." This research will open up new mechanisms to manage bee health via diet manipulation and foraging options, using simple approaches that are accessible and applicable to a diversity of stakeholders.

Animal Health Component

75%

Research Effort Categories

Basic

25%

Applied

75%

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
211	0120	1070	100%

Knowledge Area
211 - Insects, Mites, and Other Arthropods Affecting Plants;

Subject Of Investigation
0120 - Land;

Field Of Science
1070 - Ecology;

Keywords

apis mellifera

bee informed partnership

bombus impatiens

crithidia bombi

nosema

bee parasites

Goals / Objectives
Research objectives and predictions. We will address the following linked objectives, ranging from lab to field experiments, to assess the effects of sunflower on bee health and disease:Objective 1. Evaluate the benefits and costs of supplementing bumble and honey bee colonies with sunflower pollen on bee disease and health. We will use laboratory experiments to test how sunflower vs. mixed wildflower pollen supplements affect bee disease and health. Experiments will be performed on commercial and wild bumble bees, and honey bees.Obj. 1A: Bombus (Year 1; Adler, Irwin). We will use commercial colonies of B. impatiens, and colonies of B. impatiens, B. griseocollis, and B. bimaculatus produced from wild-caught queens. Using commercial colonies of B. impatiens is of direct relevance to industry stakeholders, testing how pollen diet affects parasite infection and bee health in managed bees. By extending our experiments to multiple wild bumble bee species, we will assess the degree to which sunflower pollen affects disease and health in wild pollinators important for crop pollination.Obj 1B: Apis (May-Nov 2017; Evans). We will test the effect of four pollen treatments (no pollen, wildflower pollen, 2:1 wildflower:sunflower pollen, and 1:2 wildflower:sunflower pollen) on adult worker bees from each of three healthy colonies (i.e., capped and uncapped brood with normal brood pattern, bee bread and honey stores) of A. mellifera in which both Deformed wing virus and the trypanosomatid parasite Lotmaria passim have been identified.Objective 2. Assess the degree to which sunflower pollen supplements protect against parasite infection in field colonies. We will supplement field colonies of bumble and honey bees with sunflower vs. wildflower pollen or protein supplements, working with commercial beekeepers for honey bees. Colonies will be allowed to freely forage for nectar and additional pollen, naturally exposing them to parasites and viruses. We will measure the time-course of parasite and viral acquisition, the extent of parasite load, colony foraging and productivity. We predict that supplementing colonies with sunflower pollen will slow and reduce parasite acquisition, and, using our optimized ratios from Obj 1, that sunflower pollen will improve colony performance compared to wildflower pollen.Obj 2A: Bombus (Summer Year 2, Adler, Irwin, McFrederick). We will deploy 40 colonies each (n=20/treatment/location) of commercial bumble bees (Bombus impatiens) in the Northeastern US (South Deerfield Station, 250 acres, Amherst, MA) and southeastern US (Lake Wheeler Road Field Laboratory, 1500 acres, Raleigh, NC).Obj 2B: Apis (Years 1 and 2; Adler and Evans with BIP, Inc.). We will take advantage of the BIP network of commercial beekeepers and Tech Team standardized disease assessments (https://beeinformed.org/programs/tech-teams/) to test whether pollen supplements increase disease resistance and colony health in commercial honey bee operations. In Year 1, we will conduct a dose trial to ascertain the dose with the greatest medicinal benefit and colony performance. This trial will be conducted using 32 colonies within a single commercial apiary; there will be four treatment levels (0, 25%, 50% or 75% sunflower mixed with MegaBee (megabee.com), a standard commercial honey bee protein supplement. In Year 2, we will conduct a similar but expanded field trial comparing the selected optimal supplement dose (e.g., 50% sunflower), a negative control of no supplement, and a positive control of 100% MegaBee. This full trial will be conducted at three commercial apiaries, with three treatments and three pallets (12 colonies)/treatment/apiary, for a total of 36 colonies in each apiary.As an add-on study in Year 2, we will conduct a similar assessment with Massachusetts beekeepers, restricting inclusion to commercial beekeepers with at least 5 years of experience and at least 40 colonies, to test efficacy in multiple locations.Objective 3. Document whether growing sunflower on farms affects pathogen acquisition and colony productivity. We will use multiple approaches to address this objective.Obj 3A (Year 1, Baylis): We will take advantage of longer-term data from the Bee Informed Project (BIP) to assess how sunflower acreage around honey bee colonies correlates with pathogen loads. We predict that increasing acreage of sunflower will reduce pathogen acquisition and increase colony health, consistent with our pilot data (Obj 3), but the effect of sunflower acreage on colony health may be non-linear, with the largest health benefits at intermediate acreage and greater costs at large-scale sunflower mono-cultures.Obj 3B (Year 3, Adler, Irwin & McFrederick): We will experimentally plant sunflower on farms at varying acreage and measure pathogen acquisition and colony productivity for bumble and honey bee colonies in North Carolina. This approach will provide precise control over sunflower variety, acreage and cultivation. In Massachusetts, we will place bumble and honey bee colonies at a mixture of small-scale organic and conventional farms with varied sunflower acreage, and measure pathogen acquisition and colony production. This allows us to move from experimental to real-world farms and connect with end users.Extension Objectives and Integration. Because our proposal may provide simple, cost-effective solutions to reduce bee disease without the use of chemicals, we have an unparalleled opportunity to combine research and extension efforts to test effects while also communicating with end users. Directly including industry and beekeepers in our research is one of several extension objectives. Because we propose to mitigate bee diseases through the use of sunflower pollen supplements and plantings in pollinator habitat, there is the potential for engagement and outreach with a wide range of stakeholders, including beekeepers, the commercial bumble bee industry, vegetable and fruit crop growers and the general public interested in pollinator-friendly plantings to preserve pollination services. Although not required, we will create an Advisory Board representing these interests to solicit input on objectives and communicate results via regular meetings. Furthermore, in Massachusetts there is an unprecedented opportunity for extension impact due to the recent creation of a state apiary (MA Dept. of Agricultural Resources, or MDAR) on the UMass campus and the lack of a Pollinator Extension Educator at UMass. The state has a very high proportion of small-scale farms that utilize extension services, and currently the nearest pollinator extension educator is at Cornell University. Hiring a Pollinator Extension Educator who can take advantage of the new apiary will provide a state hub for pollinator extension and leadership on extension objectives. This new hire will conduct workshops, create and maintain a website, interface with the public and, with all members of the leadership team, participate in a wide range of outreach activities. Finally, our involvement with BIP provides a national platform for disseminating results, as well as through eXtension.org posts. Thus, our Extension Objectives are to (1) directly involve beekeepers in assessing the efficacy of sunflower pollen supplements and plantings for honey bee health, (2) create an Advisory Board that engages with diverse stakeholders in multiple regions, (3) hire a Pollinator Extension Educator for Massachusetts, and (4) develop a wide range of regional and national outreach activities.

Project Methods
Obj. 1A: Bombus (Year 1; Adler, Irwin). We will use commercial colonies of B. impatiens, and colonies of B. impatiens, B. griseocollis, and B. bimaculatus produced from wild-caught queens. For commercial B. impatiens, we will use 20 colonies per treatment (3 pollen treatments x 2 parasite treatments), resulting in 120 colonies. Because of the challenges of raising colonies from wild bees, we will reduce sample size to 5-10 colonies per treatment per species in wild bees.Parasite treatment. When colonies have 10 workers, five workers per colony will be removed, inoculated with Crithidia, and returned to their colonies. We will perform a sham inoculation for colonies in the uninfected treatment.Pollen diet treatment. We will start pollen treatments after inoculation. We will give the colonies up to 20 g of pollen every two days, and will record the weight of pollen used/consumed. The experiment will be run for up to 80 days or until the colonies only produce sexuals (new queens and males).Response variables: Parasite infection, food consumption and colony performance. We will assess the time course of within-colony infection every week. At termination, we will assess Crithidia infection using gut samples from 30% of the living workers using microscopy. We will record consumption of sucrose solution weekly and pollen every two days. To assess colony performance, we will record losses (worker mortality, discarded larvae, and oophagy) and weekly colony weight. Any sexuals produced will be counted and removed. When the experiment is terminated, we will freeze the colonies, count the workers, additional sexuals and queen brood clumps, and number of larvae and eggs.Obj 1B: Apis (May-Nov 2017; Evans). We will test the effect of four pollen treatments on adult worker bees from each of three healthy colonies of A. mellifera in which both Deformed wing virus and the trypanosomatid parasite Lotmaria passim have been identified. Newly emerged bees from each colony will be separated into 16 cups, with 4 cups per treatment and 40 bees per cup (3 colonies x 16 cups/colony = 48 total cups, or 12 cups/treatment). All cups will receive ad lib sugar water via a plastic feeder (Evans et al. 2009). Cups will be maintained for three days, at which point half the cups (two of the four cups in each colony x treatment combination) will be used to assess mortality and infection in a field colony, and the other half will be maintained in the lab, with daily measures of mortality, at which point surviving bees will be frozen for genetic analyses.Obj 2. Assess the degree to which sunflower pollen supplements protects against parasite infection in field colonies (Year 2). Obj 2A: Bombus (Summer Year 2, Adler, Irwin, McFrederick). We will deploy 40 colonies each (n=20/treatment/location) of commercial bumble bees (Bombus impatiens) in the Northeastern US (South Deerfield Station, 250 acres, Amherst, MA) and southeastern US (Lake Wheeler Road Field Laboratory, 1500 acres, Raleigh, NC). Colonies will be initially controlled for number of workers and brood. Colonies will be randomly assigned to sunflower or wildflower pollen supplement treatments.Pollen diet treatments. We expect to feed Bombus colonies up to 150 g of pollen weekly. Colonies will not be given supplemental sugar solution, to motivate foraging.Parasite screening. Five workers will be sampled per colony every other week for 12 weeks to assess infection by pathogens, including Crithidia, Nosema spp., Apicystis bombi, iflaviruses and dicistroviruses.Colony foraging. We will use radio frequency ID tags to quantify colony foraging rates. Once per week for 12 weeks, we will observe colonies for entering workers, recording whether they are carrying corbicular pollen loads. For up to 10 bees per colony per collection period, we will collect one corbicular pollen load and identify pollen to species.Colony health. We will estimate productivity weekly by weighing colonies, and will visually inspect the colonies and count and remove any males or queens under red light at night. We will terminate the colonies at 12 weeks. We will freeze the colonies and count final worker production, the production of queen cells and brood in each colony.Obj 2B: Apis (Years 1 and 2; Adler and Evans with BIP, Inc.). We will take advantage of the BIP network of commercial beekeepers and Tech Team standardized disease assessments to test whether pollen supplements increase disease resistance and colony health in commercial honey bee operations.In Year 1, we will conduct a dose trial using 32 colonies within a single commercial apiary; there will be four treatment levels (0, 25%, 50% or 75% sunflower mixed with MegaBee (megabee.com), a standard commercial honey bee protein supplement. Treatments will be assigned randomly, with two pallets assigned to each treatment. Treatments will be applied in early fall, after the last summer honey is removed. Standard BIP colony assessments will be performed 4 times over the study. These measurements include colony size (frames of bees), brood quality (quantitative ranking), overt bee disease (present/absence of Chalkbrood, sac brood, European foulbrood, and American foulbrood), queen status, and lab assessments of Varroa mites (mites/100 bees) and Nosema spores (million spores/bee).In Year 2, we will conduct a similar but expanded field trial comparing the selected optimal supplement dose, a negative control of no supplement, and a positive control of 100% MegaBee. This full trial will be conducted at three commercial apiaries, with three treatments and three pallets (12 colonies)/treatment/apiary, for a total of 36 colonies in each apiary. Sampling for colony performance (colony size and brood quality), visible parasites and Nosema will be conducted as in the dose study.Obj 3. Document whether growing sunflower on farms affects pathogen acquisition and colony productivity.Obj 3A: Apis (Year 1; Baylis). We propose to use the multi-year national survey of honey bee health collected by BIP for USDA-APHIS from 2009-2016, combined with crop data from National Agricultural Statistics Service, and rainfall and temperature forecast survey from the National Oceanic and Atmospheric Administration. Geocoded data on honey bee health outcomes will be matched with data on the acreage of sunflowers grown within the honey bee foraging radius of these apiaries. We will also extract temperature and precipitation data associated with the month prior to the date the sample was collected.Obj 3B: Bombus and Apis (Summer Year 3; Adler, Irwin, McFrederick). We will deploy commercial colonies of bumble bees (B. impatiens) and honey bees at commercial farms (MA) and agricultural stations (NC) to test how varying acreage of sunflower affects pathogen acquisition and colony health. In MA, we will use 22 commercial farms in Hampshire County that are a mix of conventional and organic small-scale farms. In NC we will experimentally plant sunflower along an acreage gradient, using 17 experimental agricultural stations.At each farm in each region, we will deploy three B. impatiens and three Apis colonies (n = 66 colonies in MA and 51 colonies in NC per species). We will place colonies into the field in early to mid June, when workers are naturally starting to forage. Colonies will be initially controlled for number of workers and brood and will be tested to confirm disease absence using PCR, as in Obj 2.Floral resources. We will estimate floral resources at each farm every other week. We will also use GIS to quantify landscape types (farm, suburban, forest, old field, other) around each farm up to 10 km from the center point, and will estimate floral resources in each landscape type.We will assess Colony foraging, Parasitism status and Colony health every other week as in the methods for Obj 2, using molecular analysis for Bombus and visual assessments for Apis.

Progress 04/15/17 to 04/14/22

Outputs
Target Audience:During the entirety of this grant, the primary audiences reached by our efforts have been academic scientists, industry professionals, growers, beekeepers, seventh grade students and the general public. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Over the entire funding period, this NIFA grant provided training opportunities for 10 graduate students, of whom one is first-generation college student, two are under-represented minority students, and one is a non-traditional returning student. One postdoctoral researcher was also trained on this grant, Rosemary Malfi, with a small involvement of an additional postdoc, Jacquelyn Fitzgerald. Hannah Whitehead, our Extension Educator, has developed greatly as a professional in the extension field thanks to the opportunity provided by this grant, and with the ending of this funding, has transitioned to a Vegetable Crops Extension position at UMass and is still spending about half her time serving grower's needs around pollinators and pollinator habitat. This research also engaged 27 undergraduates in research opportunities. We have 6 papers published with graduate student first authors, and one with an undergraduate first author and graduate student co-author who conducted independent research related to this grant. Not all these students were directly paid by the USDA grant (some were funded by other sources), but all were involved in working on the USDA objectives. How have the results been disseminated to communities of interest?We reached a large audience through a wide range of extension accomplishments detailed in the Extension section of the "What was accomplished?" section, reaching over 1700 people directly and over 15,00 through our website. The PIs also gave 10 talks at public or extension events. Our annual Advisory Board meetings were also an excellent venue to disseminate our results to a range of stakeholders and extension educators. We provided an outreach event to 15-20 seventh grade girls per year (except during COVID) from under-represented groups in science through the Eureka! Program. We also presented results at multiple invited university seminars, and scientific meetings. Over the entire grant we presented our research at 13 university seminars and 10 conferences, including 3 international invited symposia or keynote talks. Our work has been disseminated to the science community via 8 journal publications (plus one in review), with 4 more anticipated to submit in the next 2-3 months. Our first results were published in Scientific Reports (see 'products') to substantial and continued media attention. The paper was selected as one of the top 100 read ecologypapers for Scientific Reports in 2018. Media coverage included: · https://www.newsweek.com/save-bees-sunflower-pollen-could-boost-insects-health-1139409 · https://www.upi.com/Science_News/2018/09/26/Sunflower-pollen-protects-bees-from-disease-study-finds/3101537966687/ · https://www.sciencedaily.com/releases/2018/09/180926082721.htm · https://www.livescience.tech/2018/09/26/sunflower-pollen-has-medicinal-protective-effects-on-bees/ · https://www.newsllive.com/nfl/sunflower-pollen-medicinal-protective-effects-bees-48280408 · https://www.reddit.com/r/science/comments/9j3e8j/ sunflower_pollen_has_medicinal_protective_effects/ · https://news.science360.gov/obj/pic-day/bf17114b-fac4-45dc-9c24-38aa87ec41f6/sunflower-pollen-medicinal-protective-effects-bees · https://phys.org/news/2018-09-sunflower-pollen-medicinal-effects-bees.html · https://montgomerycountybeekeepers.com/2017/06/sunflower-pollen-may-boost-honey-bee-health/ In addition, this research and new funding that stemmed from our findings is currently being promoted by UMass media (https://www.umass.edu/news/article/can-sunflowers-help-defeat-insect-apocalypse). What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Research Accomplishments Objective 1. Evaluate the benefits and costs of supplementing bumble and honey bee colonies with sunflower pollen on bee disease and health. Obj 1A. Bombus. - We tested the effect of varying ratios of sunflower pollen on infection in individuals and colonies, and on colony performance. A 1:1 mixture of sunflower combined with wildflower pollen reducedC. bombiinfection prevalence and intensity within individualB. impatiensworkers relative to wildflower pollen. At the colony level, a 1:1 mixture of sunflower and wildflower pollen reducedC. bombiinfection prevalence by 11% and infection intensity by 30% relative to wildflower pollen. Colony performance was similar between pollen diets and infection treatments (Giacomini et al 2021). - We wild-collected queens of Bombus impatiens, Bombus griseocollis, Bombus vagans and Bombus bimaculatus. Queens were allowed to establish nests in the lab and then experimentally inoculated with Crithidia and fed pollen diets of: 100% sunflower, 50-50 sunflower-wildflower, or 100% wildflower. For wild B. impatiens, 100% sunflower pollen and 50-50 sunflower-wildflower providing medicinal benefits relative to wildflower pollen. However, we found no effect of sunflower pollen on Crithidia infection for B. griseocollis, and relatively weak effects for B. bimaculatus and B. vagans (Fowler et al, 2022). - We tested the effect of a one-time exposure to sunflower pollen on the likelihood of Crithidia transmission and found no effect. However, in a separate experiment we found that both duration of dose and earlier timing affect Crithidia growth after infection (LoCascio et al 2019, Ecol Ent). - We found that a wide variety of H. annuus cultivars, wild populations, other Helianthus congeners, and Solidago species all are effective at reducing Crithidia counts, broadening the potential impact of our research (LoCascio et al 2019, Royal Society Open Science). - We tested the effect of sunflower pollen on B. impatiens drones and queens. Interestingly, we found that sunflower reduced Crithidia counts in queens but not drones (Fowler et al 2020). - We tested the effect of several sunflower pollen secondary compounds and fatty acids did not reduced infection, suggesting that it may not be a chemical mechanism (Adler et al 2020). Objective 2. Assess the degree to which sunflower pollen supplements protect against parasite infection in field colonies. Obj 2A. Bombus. We finished sample processing and analyses for this field experiment at NCSU, and have almost completed a full draft of the manuscript. Opposite to our predictions, we found that sunflower pollen supplements did not significantly reduce Crithidia infection or the acquisition of other bee pathogens in field colonies relative to wildflower pollen supplements. We also collaborated with Jamie Strange (Ohio State University) and his colleagues to combine both our groups' data comparing methods for sterilizing pollen (ethylene oxide, gamma radiation and ozone) on viruses, microbes, and pollen palatability. This work arose from our need to effectively sterilize sunflower and buckwheat pollen sourced from China. This manuscript is in preparation with a target submission by the end of the spring semester. Obj 3B. Deploying Bombus colonies in farms with sunflower to assess effects on pathogen acquisition and colony performance. We placed Bombus colonies at 20 farms that varied in sunflower area, from none to approximately 2 acres, and assessed effects on bee infection and colony reproduction. We submitted this work to PNAS and Ecology Letters, where it was rejected (sound science but not broad enough). We are now reformatting for PRSB, where we anticipate submitting very soon. Abstract: Community diversity can reduce the prevalence and spread of disease, but certain species may play a disproportionate role in diluting or amplifying pathogens. Flowers act both as sources of nutrition and sites of pathogen transmission, but the effects of specific plant species in shaping bee disease dynamics is not well understood. We evaluated whether plantings of sunflower (Helianthus annuus), whose pollen reduces infection by some pathogens when fed to bees in captivity, lowered pathogen levels and increased reproduction in free-foraging bumble bee colonies (Bombus impatiens). Sunflower abundance reduced the prevalence of a common gut pathogen, Crithidia bombi, and reduced infection intensity, with an order of magnitude lower infection intensity at high sunflower sites compared to sites with little to no sunflower. Sunflower abundance was also positively associated with greater queen production in colonies. Sunflower did not affect prevalence of other detected pathogens. This work demonstrates that a single plant species can drive disease dynamics in foraging bee populations, and that sunflower plantings can be used as a tool for mitigating a prevalent pathogen while also increasing reproduction of an agriculturally important bee species. We requested a change where instead of repeating this experiment at NCSU, instead we conducted a whole-colony experiment in which all colonies were infected with Crithidia and fed sunflower mixed with wildflower pollen at a 50-50 ratio and then given one box for all colony activities or two boxes (one for the brood and the other for foraging and defecation). Data analyses are in process, but preliminary results suggest that providing an additional box with sunflower pollen can reduce infection further but still cannot fully eliminate transmission. Obj 1B, 2B, and 3A all involved testing effects of sunflower pollen on honey bee health at the individual, colony and landscape scale. These were combined into a single manuscript now in review at the Journal of Economic Entomology. In brief, we tested the effects of sunflower cropland and pollen supplementation on honey bee resistance to macro- and microparasites. Although we observed no significant impacts of sunflower on microparasite levels, our results suggest effects on Varroa mites. At the landscape level, each doubling of sunflower crop area was associated with a 28% reduction in mite infestation. In a field experiment, late-summer supplementation of colonies with sunflower pollen reduced mite infestation by 2.75-fold relative to an artificial pollen. Our findings suggest the potential for sunflower plantings or pollen supplements to counteract a major driver of honey bee losses worldwide. Extension Accomplishments - We hosted three Advisory Board meetings, in which we solicited input about our research and extension plans and conveyed our research and extension activities. We received very positive feedback on our body of work, with potential continuing connections with Biobest to use sunflower pollen to manage Crithidia infection in commercial rearing facilities. Overall, reached 1771 people directly, across 40 speaking engagements or events Presented 27 talks reaching 1362 beekeepers Conducted 8 workshops reaching 159 beekeepers Participated in 5 other events (school groups, panel discussions, etc) reaching 250 participants Created the UMass Pollinator Resources website (https://ag.umass.edu/resources/pollinators), which has had nearly 15,000 unique page views - Created and disseminated 6 fact sheets Created Instructional Video on mite sampling, viewed nearly 160 times Designed and wrote quarterly newsletter about bee research, "The Research Buzz," published in the Mass Bee newsletter Created digital and print resources for veterinarians to conduct honey bee hive inspections Leveraged an additional $133,602 in grants and contracts, allowing us to extend Hannah Whitehead's position for an additional year.

Publications

• Type: Journal Articles Status: Published Year Published: 2019 Citation: **LoCascio GM, **Aguirre L, Irwin RE and LS Adler. 2019. Pollen from multiple sunflower cultivars and species reduces a common bumble bee gut pathogen. Royal Society Open Science 6: 190279. http://dx.doi.org/10.1098/rsos.190279.
• Type: Journal Articles Status: Published Year Published: 2019 Citation: **LoCascio GM, �Pasquale R, ***Amponsah E, Irwin RE and LS Adler. 2019. Effect of timing and exposure of sunflower pollen on a common gut pathogen of bumble bees. Ecological Entomology. DOI: 10.1111/een.12751
• Type: Journal Articles Status: Published Year Published: 2020 Citation: Adler LS, **Fowler AE, *Malfi RL, ***Anderson PR, ***Coppinger LM, ***Deneen PM, ***Lopez S, Irwin RE, Farrell IW and PC Stevenson. 2020. Assessing chemical mechanisms underlying the effects of sunflower pollen on a gut pathogen in bumble bees. Journal of Chemical Ecology. DOI: 10.1007/s10886-020-01168-4
• Type: Journal Articles Status: Published Year Published: 2020 Citation: **Fowler AE, ***Stone EC, Irwin RE and LS Adler. 2020. Sunflower pollen reduces a gut pathogen in worker and queen, but not male bumble bees. Ecological Entomology 45(6): 1318-26. doi: 10.1111/een.12915
• Type: Journal Articles Status: Published Year Published: 2021 Citation: Roch JCU, Mu�oz Agudelo DCG, Adler LS and J Milam. 2021. First records of Perdita bequaerti (Hymenoptera: Andrenidae) from New England. Journal of the Kansas Entomological Society 94(3): 348-354. https://doi.org/10.2317/0022-8567-93.4.354
• Type: Journal Articles Status: Published Year Published: 2021 Citation: Giacomini, J. J., S. J. Connon, D. Marulanda, L. S. Adler, and R. E. Irwin. 2021. The costs and benefits of sunflower pollen on bumble bee colony disease and health. Ecosphere 12:e03663. Doi: 10.1002/ecs2.3663
• Type: Journal Articles Status: Published Year Published: 2022 Citation: Fowler AEG, Giacomini JJ, Connon SJ, Irwin RE and LS Adler. 2022. Sunflower pollen reduces a gut pathogen in the model bee species, Bombus impatiens, but has weaker effects in three wild congeners. Proceedings of the Royal Society of London B 289: 20211909. https://doi.org/10.1098/rspb.2021.1909
• Type: Journal Articles Status: Under Review Year Published: 2022 Citation: Palmer-Young EC, Malfi RP, Zhou Y, Joyce BU, Whitehead H, Van Wyk JIP, Baylis K, Grubbs K, Lopez D, Evans JD, Irwin RE and LS Adler LS (in review). Sunflower crop area and pollen supplementation reduce Varroa mite infestation in honey bee colonies. Journal of Economic Entomology.
• Type: Journal Articles Status: Published Year Published: 2018 Citation: ***Giacomini JJ, ***Leslie J, Tarpy DR, **Palmer-Young EC, Irwin RE and LS Adler. 2018. Medicinal value of sunflower pollen against bee pathogens. Scientific Reports 8: 14394. DOI: 10.1038/s41598-018-32681-y

Progress 04/15/21 to 04/14/22

Outputs
Target Audience:During the past year, the primary audience reached by our efforts has been academic scientists, industry professionals, growers, beekeepers, seventh grade students and the general public. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?In this final no-cost extension year, this NIFA grant provided training opportunities for 5 graduate students, of whom one is first-generation college student, one is an under-represented minority student, and one is a non-traditional returning student These were Carolina Muñoz Agudelo, Jonathan Giacomini, Justin Roch, April Sharp and Erin Eichenberger. One postdoctoral researcher was also trained on this grant, Rosemary Malfi. Hannah Whitehead, our Extension Educator, has developed greatly as a professional in the extension field thanks to the opportunity provided by this grant, and with the ending of this funding, has transitioned to a Vegetable Crops Extension position at UMass and is still spending about half her time serving grower's needs around pollinators and pollinator habitat. This research also engaged 2 undergraduates in research opportunities. During this reporting period we have 2 papers published with graduate student first authors, and one with an undergraduate first author and graduate student co-author who conducted independent research related to this grant. Not all these students were directly paid by the USDA grant (some were funded by other sources), but all were involved in working on the USDA objectives. How have the results been disseminated to communities of interest?Our work has been disseminated to the science community via 3 journal publications (plus one in review) and multiple talks at research universities and conferences. Lynn Adler presented this research at 3 invited university seminars (Cornell, U Wisconsin-Madison, U Maryland - College Park), one invited international symposium (Pollinator Health Conference, London UK). Becky Irwin talked about this research at invited talks at Duke University (Program in Ecology), University of Georgia (Dept of Entomology), and University of Colorado (EPOB Department). In extension and outreach, our fact sheet about sunflower pollen and bee health was posted to the UMass Pollinator Resources website. Lynn Adler presented a UVM extension seminar with 21 attendees, who were mostly growers interested in promoting pollinator health via plantings on farms. Becky Irwin gave a public talk at BugFest that was online live in September 2021 that included some of this work. Quinn McFrederick gave a related talk at the Annual Orchard Bee Association meeting in Corvalis Oregon on December 11, 2021. In addition, this research and new funding that stemmed from our findings is currently being promoted by UMass media (https://www.umass.edu/news/article/can-sunflowers-help-defeat-insect-apocalypse). What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Research Accomplishments Objective 1. Evaluate the benefits and costs of supplementing bumble and honey bee colonies with sunflower pollen on bee disease and health. Obj 1A. Bombus. At NCSU, we focused on one add-on experiment, sample processing, and analyses. We ran a whole-colony experiment in which all colonies were infected with Crithidia and fed sunflower mixed with wildflower pollen at a 50-50 ratio (which has been shown in our prior research to reduce infection), and then colonies were either given one box for all colony activities or two boxes (one for the brood and the other for foraging and defecation). We measured prevalence and intensity of infection in the colonies biweekly and final colony weight and number of workers and new queens. Data analyses are in process, but preliminary results suggest that providing an additional box with sunflower pollen can reduce infection further but still cannot fully eliminate transmission. These results suggest that once Crithidia is in a facility, prevalence and infection intensity can be reduced but not eliminated with diet manipulation. We are finishing analyses and then will write this work up for publication. A manuscript detailing how sunflower pollen supplements affect Crithidia infection in single boxes of commercial Bombus impatiens was published in Ecosphere (accepted during Year 3 reporting and published officially during Year 4 reporting). The work was headed by NCSU graduate student Jonathan Giacomini. We found that a 1:1 mixture of sunflower combined with wildflower pollen reducedC. bombiinfection prevalence and intensity within individualB. impatiensworkers by nearly 4-fold and 12-fold, respectively, relative to wildflower pollen. At the colony level, a 1:1 mixture of sunflower and wildflower pollen reducedC. bombiinfection prevalence by 11% averaged over a 10-week period and infection intensity by 30% relative to wildflower pollen. Colony performance was similar between pollen diets and infection treatments. Infection significantly reduced the probability of queen production in colonies fed a pure wildflower pollen diet, but not colonies fed a mixed sunflower pollen diet, suggesting that the medicinal benefits of a mixed sunflower pollen diet can reverse the negative effects of infection on reproductive success. This study provides evidence that sunflower pollen as part of a mixed pollen diet can reduce infection in individual bees and whole colonies with no significant nutritional trade-offs for colony worker production and most aspects of colony reproduction. Obj 1B, 2B, and 3A all involved honey bee research that was completed in previous reporting periods. Former graduate student Evan Palmer-Young, now a research associate with Jay Evans, took over writing up all the honey bee-related research from this grant. This work is now in review at the Journal of Economic Entomology. In brief, we tested the effects of sunflower cropland and pollen supplementation on honey bee resistance to macro- and microparasites. Although we observed no significant impacts of sunflower on microparasite levels, our results suggest effects on Varroa mites. At the landscape level, each doubling of sunflower crop area was associated with a 28% reduction in mite infestation. In a field experiment, late-summer supplementation of colonies with sunflower pollen reduced mite infestation by 2.75-fold relative to an artificial pollen. Our findings suggest the potential for sunflower plantings or pollen supplements to counteract a major driver of honey bee losses worldwide. Objective 2. Assess the degree to which sunflower pollen supplements protect against parasite infection in field colonies. During this reporting period, at NCSU we finished sample processing and analyses for this field experiment and have almost completed a full draft of the manuscript. Opposite to our predictions, we found that sunflower pollen supplements did not significantly reduce Crithidia infection or the acquisition of other bee pathogens in field colonies relative to wildflower pollen supplements. We suspect this is the case because of high pathogen prevalence in nearby colonies, allowing for constant re-infection. We dissected the colonies and stained pollen to understand what sunflower pollen was used for within the hive and found evidence of preferential sunflower pollen feeding to queen larvae in infected colonies, potentially suggesting self-medication behavior, which could have important consequences for colony reproductive success. Obj 3B. Deploying Bombus colonies in farms with sunflower to assess effects on pathogen acquisition and colony performance. During the current reporting period we finished all data analysis, wrote this manuscript, and submitted it for publication. It was rejected from PNAS and Ecology Letters. We are now reformatting for PRSB, where we anticipate submitting very soon. Abstract: Community diversity can reduce the prevalence and spread of disease, but certain species may play a disproportionate role in diluting or amplifying pathogens. Flowers act both as sources of nutrition and sites of pathogen transmission, but the effects of specific plant species in shaping bee disease dynamics is not well understood. We evaluated whether plantings of sunflower (Helianthus annuus), whose pollen reduces infection by some pathogens when fed to bees in captivity, lowered pathogen levels and increased reproduction in free-foraging bumble bee colonies (Bombus impatiens). Sunflower abundance reduced the prevalence of a common gut pathogen, Crithidia bombi, and reduced infection intensity, with an order of magnitude lower infection intensity at high sunflower sites compared to sites with little to no sunflower. Sunflower abundance was also positively associated with greater queen production in colonies. Sunflower did not affect prevalence of other detected pathogens. This work demonstrates that a single plant species can drive disease dynamics in foraging bee populations, and that sunflower plantings can be used as a tool for mitigating a prevalent pathogen while also increasing reproduction of an agriculturally important bee species. We also collaborated with Jamie Strange (Ohio State University) and his colleagues to combine both our groups' data comparing methods for sterilizing pollen (ethylene oxide, gamma radiation and ozone) on viruses, microbes, and pollen palatability. This work arose from our need to effectively sterilize sunflower and buckwheat pollen sourced from China. This manuscript is in preparation with a target submission by the end of the spring semester.

Publications

• Type: Journal Articles Status: Published Year Published: 2021 Citation: Roch JCU, Mu�oz Agudelo DCG, Adler LS and J Milam. 2021. First records of Perdita bequaerti (Hymenoptera: Andrenidae) from New England. Journal of the Kansas Entomological Society 94(3): 348-354. https://doi.org/10.2317/0022-8567-93.4.354
• Type: Journal Articles Status: Published Year Published: 2021 Citation: Giacomini, J. J., S. J. Connon, D. Marulanda, L. S. Adler, and R. E. Irwin. 2021. The costs and benefits of sunflower pollen on bumble bee colony disease and health. Ecosphere 12:e03663. Doi: 10.1002/ecs2.3663
• Type: Journal Articles Status: Published Year Published: 2022 Citation: Fowler AEG, Giacomini JJ, Connon SJ, Irwin RE and LS Adler. 2022. Sunflower pollen reduces a gut pathogen in the model bee species, Bombus impatiens, but has weaker effects in three wild congeners. Proceedings of the Royal Society of London B 289: 20211909. https://doi.org/10.1098/rspb.2021.1909
• Type: Journal Articles Status: Under Review Year Published: 2022 Citation: Palmer-Young EC, Malfi RP, Zhou Y, Joyce BU, Whitehead H, Van Wyk JIP, Baylis K, Grubbs K, Lopez D, Evans JD, Irwin RE and LS Adler LS (in review). Sunflower crop area and pollen supplementation reduce Varroa mite infestation in honey bee colonies. Journal of Economic Entomology.

Progress 04/15/20 to 04/14/21

Outputs
Target Audience:During the past year, the primary audience reached by our efforts has been academic scientists, industry professionals, growers, beekeepers, seventh grade students and the general public. Changes/Problems:Productivity has been substantially slowed this year due to COVID. Research activities were restricted at all universities, slowing the pace of data collection. In addition, several PIs and people supported by this grant had substantial child-care duties due to the lack of in-person school and child care that reduced their available time for data analysis and writing. We requested and received a second no-cost extension to give us an additional year to complete and publish this research. We describe our revised timeframe for goals in the 'What do you plan to do in the next year?' section. What opportunities for training and professional development has the project provided?This year, this NIFA grant provided training opportunities for 3 graduate students, one of whom is a first-generation college student and one of whom is an under-represented minority student. These were Alison Fowler, Carolina Muñoz, and Jonathan Giacomini. One postdoctoral researcher was also trained on this grant, Rosemary Malfi. Hannah Whitehead, our Extension Educator, has developed greatly as a professional in the extension field thanks to the opportunity provided by this grant, and has transitioned to writing some of her own grants and contracts (mentored by the PI Lynn Adler) in the hopes of continuing to fund her position after the current grant expires. This research also provided independent research opportunities for one non-traditional returning undergraduate, Justin Roch, and engaged 9 other undergraduates in research opportunities. During this reporting period we have two papers published or accepted with graduate student first authors, and one with an undergraduate first author and graduate student co-author who conducted independent research related to this grant. Not all these students were directly paid by the USDA grant (some were funded by other sources), but all were involved in working on the USDA objectives. How have the results been disseminated to communities of interest?Our dissemination activities have been curtailed this year due to COVID. That said, we still reached a large audience virtually through a wide range of extension accomplishments detailed in the Extension section of the "What was accomplished?" section, reaching over 800 people directly and many more through our website. We are in the process of writing fact sheets to share our sunflower results more effectively with growers and beekeepers. What do you plan to do during the next reporting period to accomplish the goals?Research Objectives: Objective 1A: We plan to write and submit this manuscript for publication in the next year. Objective 1B, 2B, 3A: We plan publish these data as a combined manuscript in the next year, now led by Evan Palmer-Young. Objective 2A: This manuscript is in preparation and we intend to submit in the next 2-3 months. Objective 3B: We are currently writing up the Massachusetts portion of this objective and intend to submit in the next 3 months. We made a change to the North Carolina portion of this objective and conducted a two-box colony sunflower supplementation experiment in spring 2020 that was cut short by COVID. We plan to repeat this experiment in the next reporting period. Extension Objectives: We have our final Advisory Board meetings during this final year. We hope to be able to do this in person, but if safety issues continue to make this inadvisable we will conduct it virtually. All PIs will continue to engage collaborators, graduate students, and undergraduates in all aspects of the research. Research results will continue to be disseminated via presentations at regional and national conferences, publication in peer-reviewed journals, and outreach through connections with extension professionals.

Impacts
What was accomplished under these goals? Research Accomplishments Obj 1A: Benefits and costs of sunflower pollen supplements in Bombus. In the current reporting period, graduate student Alison Fowler prepared a manuscript that will be submitted for publication within the next month. At NCSU, we extended to the managed pollinator Megachile rotundata to test how sunflower pollen affects Crithidia bombi infection in this alternate host. We experimentally infected M. rotundata with C. bombi and assigned bees randomly to sunflower or wildflower pollen diet treatments. Our preliminary analyses show no effect of sunflower pollen on the probability of infection or infection intensity in males or females of this alternate host; however, we need to add more females to bolster sample size. At NCSU, we also finished data analyses, wrote and submitted research assessing the costs and benefits of sunflower pollen on whole-colony bumble bee disease and performance using commercial Bombus impatiens. We found that a 1:1 mixture of sunflower combined with wildflower pollen reduced C. bombi infection in individual bee and whole colonies compared to wildflower pollen. Colony performance was similar between pollen diets and infection treatments. Infection significantly reduced the probability of queen production in colonies fed wildflower pollen but a mixed sunflower pollen diet, suggesting that the medicinal benefits of a mixed sunflower pollen diet can reverse the negative effects of infection. This study provides evidence that sunflower pollen as part of a mixed pollen diet can reduce infection in individual bees and whole colonies with few nutritional tradeoffs. This manuscript was accepted for publication in Ecosphere. Obj 1B, 2B, and 3A all involve honey bee research that was completed in previous reporting periods. Because results were largely inconclusive, we are writing them all up as a single manuscript. Writing has been slow this year due to COVID issues and family care constraints for our postdoc. We recently invited a former Adler lab member and current Jay Evans postdoc, Evan Palmer-Young, to take over writing this manuscript, which he has agreed to do in the coming year. Obj 2A. Sunflower pollen supplements in field Bombus colonies. At NCSU, we focused on sample processing of pollen from the colonies, data analyses and writing. At UCR, in October 2020 we completed molecular screening of 432 bumble bee. We screened by first amplifying each bee for host DNA, then qualitatively screening for neogregarines. microsporidians, and two viruses. We additionally performed quantitativePCR for Crithidia bombi. Our preliminary analyses show that sunflower pollen supplements had no effect on field colony C. bombi infection relative to colonies provided with wildflower pollen supplements. We hypothesized that this may be because colonies are using the pollen supplements for building colony structures and feeding larvae as opposed to adult consumption. We dissected all colonies as well as guts of larvae, workers, and reproductives and stained the pollen. During this past reporting period, we have been processing those slides to mechanistically understand the outcome of this field experiment. We are now writing up this manuscript to submit for publication in the coming year. Obj 3B. Deploying Bombus colonies in farms with sunflower to assess effects on pathogen acquisition and colony performance. During 2020 we processed slides collected from foraging bumble bees from this field study to determine relationships between sunflower acreage, foraging on sunflower, and pathogen acquisition. At UCR, in May of 2020 we finished molecular screening of 759 bumble bees from this study for pathogens. We conducted quantitativePCR on these samples for Crithidia bombi and additionally qualitatively screened for microsporidian, neogregarine, and viral (on a subset of samples) pathogens. We additionally amplified host genes for each sample as a positive control for DNA extractions. We found exciting results that sunflower abundance strongly reduces Crithidia prevalence and also increases queen production in colonies. At UMass, we have been analyzing and writing up these results and plan to submit a manuscript to a high-profile journal by early summer. In a previous reporting period requested the change to instead conduct a new experiment that will directly benefit commercial bumble bee companies, rather than replicating a field study in two states. The new experiment was started in the previous reporting period and terminated early (at 4 weeks instead of 8) due to COVID. In the current reporting period at NCSU, we processed samples from this experiment, in which B. impatiens colonies were infected with C. bombi and then assigned to factorial treatments of pollen diet (sunflower or wildflower pollen) crossed by box-type (1-box or 2-box chamber). To assess whether providing colonies a second box in combination with sunflower pollen supplements could be a simple management solution to eliminate C. bombi in managed rearing settings. Preliminary findings show that sunflower pollen supplements significantly reduced C. bombi infection by 65% relative to wildflower pollen. While a second box reduced C. bombi infection by 20%; this difference was not statistically significant, and there was no significant interaction between pollen diet treatment and box-type. Because this experiment was reduced in length from 8 to 4 weeks, during the new reporting period, we will re-run the experiment to assess the consistency of the results over a longer colony time course. We suspect that infection status will start to diverge in the 2-box treatment as the colonies get larger, with infection relief provided by the 2-box design in colonies supplemented with sunflower pollen. Extension Accomplishments: - At UCR, Our largest event (the Riverside Insect Fair) was cancelled due to the pandemic, but McFrederick was a panelist at a well-attended webinar for UCANR (April 8, 2020,https://ucanr.edu/blogs/blogcore/postdetail.cfm?postnum=40983, which counted for continuing education units for IPM practitioners and pesticideapplicators) and was a panelist for the California Citrus Nursery Society'swebinar on October 29, 2020 (https://lists.ucr.edu/pipermail/cafe/2020-October/000014.html). - At UMass, Adler presented this research in a 1-hour virtual talk to 69 beekeepers at the Norfolk County Beekeeper's Association. - We have not yet hosted our final Advisory Board meeting. We decided to postpone this meeting until more of our results are finalized, especially for honey bees, to have maximum impact. - Extension Educator Hannah Whitehead has been engaged in several outreach activities: Overall, reached 801 people directly, across 15 speaking engagements or events Presented at the fall and spring Mass Bee state-wide meetings reaching 232 beekeepers Presented at the fall and spring MDAR Apiary State of the State events, reaching 171 beekeepers Participated as a panelist in the fall and spring New England Honey Bee Updates, reaching 198 beekeepers Spoke at 7 bee club meetings across the state, presenting SARE partnership research and Hobbyist health survey pesticide results, reaching 160 beekeepers. Live-streamed a USDA APHIS inspection for the Barnstable County Beekeepers Association, reaching 30 beekeepers Online Outreach Continued maintaining the UMass Pollinator Resources section of the UMass Extension website (ag.umass.edu/resources/pollinators - launched March 2019) Continued to update the "UMass Extension - Pollinators" Facebook page (@UMassExtPollinators) Print Outreach Published three fact sheets and a tri-fold brochure (in collaboration with MDAR and the Northeast IPM Center). The brochure was published May 22, 2020, and the fact sheets were published June 3, 2020. They can be viewed here: ag.umass.edu/resources/pollinators/varroa. Began drafting a fact sheet summarizing research on sunflower and bee health, to be completed in May 2021

Publications

• Type: Journal Articles Status: Accepted Year Published: 2021 Citation: Giacomini JJ, Connon SJ, Marulanda D, Adler LS and RE Irwin (in press). The costs and benefits of sunflower pollen on bumble bee colony disease and health. Ecosphere.
• Type: Journal Articles Status: Accepted Year Published: 2021 Citation: ***Roch JC, **Mu�oz Agudelo DC, Adler LS and J Milam (in press). First records of Perdita bequaerti (Hymenoptera: Andrenidae) from New England. Journal of the Kansas Entomological Society.
• Type: Journal Articles Status: Published Year Published: 2020 Citation: **Fowler AE, ***Stone EC, Irwin RE and LS Adler. 2020. Sunflower pollen reduces a gut pathogen in worker and queen, but not male bumble bees. Ecological Entomology 45(6): 1318-26. doi: 10.1111/een.12915

Progress 04/15/19 to 04/14/20

Outputs
Target Audience:During the past year, the primary audience reached by our efforts has been academic scientists, industry professionals, growers, beekeepers, seventh grade students and the general public. Changes/Problems:We originally intended to conduct Obj 3B concurrently in Massachusetts and North Carolina. However, the timing has been challenging in North Carolina because sunflower is not as high a priority crop as soybean and so could only be planted as a double crop, which would mean late blooming such that deployed bumble bee colonies may not survive due to heat. We reported these issues in last year's annual report in the 'Changes' section, and at that time were exploring other options. In this reporting period we decided that it would be a better use of resources to conduct a new experiment that will directly benefit commercial bumble bee companies, rather than replicating a field study in two states. In our previous experiments we found that feeding whole commercial colonies of sunflower dramatically reduced but never entirely eradicated infection, and we wondered whether this was because the single box of a commercial colony does not allow the bees to defecate elsewhere as they would under more natural conditions. The Irwin lab therefore conducted an additional factorial experiment testing how colony box design (1-chamber vs 2-chamber) and pollen diet (wildflower vs. 80% sunflower) affect bee disease and health. All colonies were infected with Crithidia and we measured disease incidence and severity every other week, along with final colony size. We intended to run this experiment for 8 weeks, but it only ran for 5 weeks and then got cut short by COVID. Once the Irwin lab reopens they will assess infections in their bees and analyze results. What opportunities for training and professional development has the project provided?In this third year, this NIFA grant provided training opportunities for 5 graduate students, one of whom is a first-generation college student and one of whom is an under-represented minority student. These were Alison Fowler, Carolina Muñoz, Jonathan Giacomini, Yujun Zhou, and Ivan Flores. One postdoctoral researcher was also trained on this grant, Rosemary Malfi. Hannah Whitehead, our Extension Educator, has developed greatly as a professional in the extension field in the past year thanks to the opportunity provided by this grant, and has transitioned to writing some of her own grants and contracts (mentored by the PI Lynn Adler) in the hopes of continuing to fund her position after the current grant expires. This research also provided independent research opportunities for one non-traditional returning undergraduate, Justin Roch, and engaged 12 other undergraduates and 2 high school students in research opportunities. During this reporting period we published a paper that includes 4 undergraduate co-authors who conducted independent research related to this grant. Not all these students were directly paid by the USDA grant (some were funded by other sources), but all were involved in working on the USDA objectives. How have the results been disseminated to communities of interest?We reached a large audience through a wide range of extension accomplishments detailed in the Extension section of the "What was accomplished?" section, reaching over 700 people directly and over 4700 through our website. We also presented results at multiple invited university seminars, and scientific meetings. Specifically: Rebecca Irwin gave two invited seminars at which she talked about research findings from this grant (UC Davis, Department of Entomology, October 2019 and University of Maryland-College Park, Department of Entomology, October 2019). Lynn Adler incorporated results from this grant into invited research seminars at Smith College and New England Botanical Club of Harvard University Conference presentations: NCSU OUR Undergraduate Research Summer Symposium, Raleigh NC, Aug 1 2019 Sam Johnson, Jonathan Giacomini, Lynn Adler and Rebecca Irwin Poster: The Sunflower Effect 2019Gordon Research Conference in Animal-Microbe Symbioses, West Dover VT, June 15-16 2019 Alison Fowler, Rebecca Irwin and Lynn Adler Poster: The Effect of Sunflower Pollen on Bumble Bee Parasitism and Reproduction What do you plan to do during the next reporting period to accomplish the goals?Research Objectives: Objective 1A: We plan to write and submit this manuscript for publication by the end of summer 2020. Objective 1B: We plan publish these data in the next year, in combination with other honey bee objectives. Objective 2A: This experiment was conducted in spring 2018; processing colonies and molecular analysis of pathogens was only recently completed. We intend to submit this manuscript for publication by the end of the final reporting period. Objective 2B: The BIP trials were fall/winter 2019-20. We intend to publish these results with the rest of the honey bee experiments in the final reporting period. Objective 2 add-on: We conducted this experiment in early summer 2019 spring and will include these results in a publication with the other honey bee experiments in the upcoming year). Objective 3A: We will finalize these analyses and publish with the rest of the honey bee data. Objective 3B: The Massachusetts portion of this objective was conducted in summer 2019. Due to COVID, both the pollen counts and the molecular pathogen screening have been delayed. We hope these will be completed by Sept 2020 and a manuscript will be submitted by the end of the reporting period. As described in the 'Changes' section, due to timing and logistical constraints we made a change to the North Carolina portion of this objective and conducted a two-box colony sunflower supplementation experiment in spring 2020 that was cut short by COVID. We hope we will see patterns in the 5-week experiment (intended to run for 8 weeks) and if so, hope to publish in the next reporting period. Extension Objectives: Our honeybee Extension Educator will continue to network with beekeepers across the state, assess beekeeper needs, develop educational materials and website content, attend beekeeper meetings and appropriate conferences, organize Master Beekeeper workshops, and present results of our research as appropriate. We have our final Advisory Board meetings during this final year. We hope to be able to do this in person, but if safety issues continue to make this inadvisable we will conduct it virtually. I will continue to engage collaborators, graduate students, and undergraduates in all aspects of the research. Research results will continue to be disseminated via presentations at regional and national conferences, publication in peer-reviewed journals, and outreach through connections with extension professionals.

Impacts
What was accomplished under these goals? Objective 1A (Adler). These experiments are complete. We plan to submit a manuscript by the end of the summer. Objective 1B (Evans). Honey bee experiments are complete but results did not give a consistent answer. Due to the mixed results, we plan to publish these experiments with our other honey bee work in a single larger publication in 2020. Objective 2A (Irwin, McFrederick). The field experiment was completed in the previous reporting period. In the current reporting period, we processed pollen slides from this experiment to determine sunflower pollen use; this is still ongoing. The McFrederick lab completed quantification of Crithidia infection for 759 individual bees, and screening of iflaviruses, dicistrovirruses, microsporidians, apicomplexa, and trypanosomatids of 279 bees. At present we find no evidence that field-colony supplementation with sunflower pollen reduced Crithidia or other infection, but we are continuing to analyze these data. Objective 2B (Adler) assesses the effect of sunflower pollen supplementation on honey bee pathogens and health. During the current reporting period we worked with BIP using fall supplementation with sunflower or wildflower pollen and assessed impacts on honey bee pathogens and health. The experiment completed in February 2020. We received the final data in March and are beginning data analysis. In addition, the Evans lab conducted viral assays of bees that had previously been fed sunflower pollen versus wildflower in cup assays. These viral measurements helped confirm a slight decrease in disease loads in bees fed pollen from sunflowers, pointing to the potential of this pollen in bee health. Obj 3A (Baylis) uses a long-term BIP data set to analyze relationships between sunflower plantings and honey bee pathogens (mites and Nosema) across the US. In this reporting period the graduate students compiled the landscape and bee morbidity data and put together the data documentation, extending the time frame and using different spatial scales, which was not part of the original data set. Obj 3B (Adler, Irwin, McFrederick), documenting whether sunflower on farms affects pathogen loads and performance of bee colonies, was conducted in Massachusetts in summer 2019. This was an ambitious project that involved 20 farms and extensive field work and data collection. We deployed commercial bumble bee colonies at all 20 farms and opportunistically sampled honey bee apiaries that were located at 15 of the farms. All fall was spent in microscope assessments of bee pathogen infection. Our preliminary findings are that there is a dramatic drop in Crithidia infection prevalence when farms have at least ¾ acre of sunflower. In the spring we began scoring pollen loads from foraging bees, but this was cut short in March due to COVID. We hope to resume this summer. The McFrederick lab surface sterilized and dissected another 795 bees for that willbe screened for parasites using PCR once research ramps up again following COVID-19 shutdown. We also request a change to our proposal. In North Carolina, we originally proposed to conduct a field experiment parallel to the one on Massachusetts. However, the seasonality of sunflower and bumble bee work in NC has made this challenging; when sunflowers are grown in summer it is too hot to successfully deploy bumble bee colonies. We reported these issues in last year's 'Changes' section. In this reporting period we decided that it would be a better use of resources to conduct a new experiment that will directly benefit commercial bumble bee companies, rather than replicating a field study in two states. In our previous experiments we found that feeding whole commercial colonies of sunflower dramatically reduced but never entirely eradicated infection, and we wondered whether this was because the single box of a commercial colony does not allow the bees to defecate outside the colony. The Irwin lab therefore conducted a factorial experiment testing how colony box design (1-chamber vs 2-chamber) and pollen diet (wildflower vs. 80% sunflower) affect bee disease and health. All colonies were infected with Crithidia and we measured disease incidence and severity every other week, along with final colony size. We intended to run this experiment for 8 weeks, but it only ran for 5 weeks and then got cut short by COVID. Once the Irwin lab reopens they will assess infections in their bees and analyze results. Extension Accomplishments: McFrederick presented research and natural history related to the funded research as part of a UC ANR extension webinar offered at the beginning of the pandemic that was attended by ~250 participants. - Adler presented her work at a "Pollinators in Our Landscape" presentation for 200 stakeholders, through a Farm Day for 40 growers, and a BeeFest presentation to the general public. We planned to have our final annual advisory board stakeholder meeting in Massachusetts in April 2019. However, this was postponed due to COVID. We will wait to see if we can have an in-person meeting in fall or winter 2020, or need to move to a virtual format. Some of our beekeeper stakeholders may be less inclined to attend a virtual meeting, but safety precautions may mean this is the only option. Our honey bee Extension Educator, Hannah Whitehead has conducted a wide range of outreach and extension activities, described below. Workshops and Talks Overall, reached 388 people directly, across 13 speaking engagements or events In collaboration with chief apiary inspector Kim Skyrm, designed and led two innovative full-day workshops ("Fight the Mite" held twice and "Honey Bees Under the Microscope" held twice; total of 84 beekeepers). Hosted two workshops at UMass Amherst, led by external speakers. These events reached 27 people total. Gave two talks at the Mass Bee summer field day (6/15/19), reaching 40 beekeepers Spoke at 5 bee club meetings across the state, reaching 225 beekeepers total Gave a tour of the state apiary to a homeschool class from the Hitchcock center (5/21/19), reaching 12 students Online Outreach Continued maintaining the UMass Pollinator Resources section of the UMass Extension website (ag.umass.edu/resources/pollinators - launched March 2019) There were 4,719 visits to this resource section from April 15, 2019-April 14, 2020 Created a "UMass Extension - Pollinators" Facebook page, and used it to advertise events and share relevant information (@UMassExtPollinators) Print Outreach Drafted three fact sheets and a tri-fold brochure (in collaboration with MDAR and the Northeast IPM Center). The brochure was published May 22, 2020, and the fact sheets were published June 3, 2020. They can be viewed here: ag.umass.edu/resources/pollinators/varroa. Research Finished a full report of the MDAR hobbyist health survey (12/20/19), which was posted to the website (https://ag.umass.edu/resources/pollinators/research-projects-at-umass/2018-massachusetts-hobbyist-health-survey). Created a presentation on the results and shared with bee clubs (see bee club visits above). Coordinated the 2019 USDA APHIS survey and collected data for Western MA. Assembled a working group of beekeepers and veterinarians as part of the 2019 MDAR grant. Grants Wrote and received 2 grants to continue extension work (>$55k total)

Publications

• Type: Journal Articles Status: Under Review Year Published: 2020 Citation: **Fowler AE, ***Stone EC, Irwin RE and LS Adler (in review). Sunflower pollen reduces a gut pathogen in worker and queen, but not male bumble bees. Ecological Entomology.
• Type: Journal Articles Status: Published Year Published: 2020 Citation: Adler LS, **Fowler AE, *Malfi RL, ***Anderson PR, ***Coppinger LM, ***Deneen PM, ***Lopez S, Irwin RE, Farrell IW and PC Stevenson. 2020. Assessing chemical mechanisms underlying the effects of sunflower pollen on a gut pathogen in bumble bees. Journal of Chemical Ecology. DOI: 10.1007/s10886-020-01168-4

Progress 04/15/18 to 04/14/19

Outputs
Target Audience:During the past year, the primary audience reached by our efforts has been academic scientists, industry professionals, growers, beekeepers, seventh grade students and the general public. Changes/Problems:As mentioned above in 'Accomplishments,' we have had difficulty collected enough domestic sunflower pollen for use in commercial apiary trials. We now have a permit to deploy sterilized Chinese sunflower pollen in apiary trials this upcoming fall/winter at the University of Maryland under Dennis Van Engelsdorp, where colonies are contained and can be destroyed as necessary, instead of in a California commercial apiary as originally planned. We originally intended to conduct Obj 3B concurrently in Massachusetts and North Carolina. However, the timing has been challenging in North Carolina because sunflower is not as high a priority crop as soybean and so could only be planted as a double crop, which would mean late blooming such that deployed bumble bee colonies may not survive due to heat. This summer we are exploring options to conduct this experiment instead in North Dakota on commercial farms, where sunflower is typically planted at very large acreages, or planning ahead to coordinate sunflower plantings in NC as originally planned. In addition, we realized that the cost of deploying honey bees at each farm was prohibitively high and was not included in our budget. Instead, we are working with local beekeepers in Massachusetts to sample from existing honey bee colonies; these are present at 15 of the 24 farms we will use. What opportunities for training and professional development has the project provided?In this second year, this NIFA grant provided opportunities for 5 graduate students, one of whom is a first-generation college student and one of whom is an under-represented minority. These were Alison Fowler, Jonathan Giacomini, Simon Pinilla, Yujun Zhou, and Laura Leger. Two postdoctoral researchers were also fully or partly involved in research opportunities from this grant; these are Rosemary Malfi and Jacquelyn Fitzgerald. Hannah Whitehead, our Extension Educator, has developed greatly as a professional in the extension field in the past year thanks to the opportunity provided by this grant. This research also provided independent research opportunities for three undergraduates including one low-income student. These were Bryanna Joyce, Lily Coppinger and Stephanie Lopez. Not all these students were directly paid by the USDA grant (some were funded by other sources), but all were involved in working on the USDA objectives. How have the results been disseminated to communities of interest?Through our second Advisory Board committee meeting, a Farm Day for growers at the S Deerfield research station, BeeFest presentations to the general public, an outreach presentation to 7th grade students, multiple invited university seminars, and presentations at scientific meetings. Specifically: We provided an outreach event to approximately 17 seventh grade girls from under-represented groups in science through the Eureka! Program. I spoke at the Bee Fest in Greenfield to about 50 members of the public. I participated in Agricultural Field Day, S Deerfield Farm, 10-4. "The objective of this event is to open the farm to the public for a guided tour of our current research projects." I incorporated these results into invited research seminars at Penn State University, the UMass Stockbridge School of Agriculture, as the invited Keynote Speaker at the International Symposium on the Ecology and Evolution of Flowers, in Zurich, Switzerland, and as an invited speaker at the Gordon Research Conference in Plant-Herbivore Interactions. Conference presentations: 25th Annual Massachusetts Undergraduate Research Conference Amherst MA, April 26 2019 Bryanna Joyce1, Hannah Whitehead1, Jennifer VanWyk1 , Rosemary Malfi1 and Lynn Adler1 Poster: Testing Pollen Diet as a Novel Management Tool in Honey Bees 25th Annual Massachusetts Undergraduate Research Conference Amherst MA, April 26 2019 Lily Coppinger, Alison Fowler and Lynn Adler1 Poster: The Role of Fatty Acids in the Observed Medicinal Properties of Sunflower Pollen on B. impatiens Infected with C. bombi Gordon Research Conference in Plant-Herbivore Interactions, Ventura CA, Feb 25-Mar 1 2019 Lynn S. Adler Invited talk: Floral traits mediating bee-pathogen interactions What do you plan to do during the next reporting period to accomplish the goals?Research Objectives: Objective 1A: We plan to analyze them and write these experiments for publication in the next year. Objective 1B: These experiments have been conducted and we plan to publish them in the next year, in combination with other honey bee objectives. Objective 2A: This experiment was conducted in spring 2018; processing colonies and molecular analysis of pathogens will continue into the next reporting period. When this is complete, we will analyze data and submit this for publication. Objective 2B: The BIP trials will be conducted in fall/winter 2019-20. We intend to publish these result with the rest of the honey bee experiments in spring/summer 2020. Objective 2 add-on: We conducted this experiment in early summer 2019 spring and will include these results in a publication with the other honey bee experiments in the upcoming year (spring/summer 2020). Objective 3A: We will finalize these analyses and publish with the rest of the honey bee data (spring/summer 2020). Objective 3B: The Massachusetts portion of this objective will be conducted in summer 2019. Due to timing and logistical constraints (see 'Changes/Problems'), we will request a no-cost extension to conduct the North Carolina portion in summer 2020. Extension Objectives: Our honeybee Extension Educator will continue to network with beekeepers across the state, assess beekeeper needs, develop educational materials and website content, attend beekeeper meetings and appropriate conferences, organize Master Beekeeper workshops, and present results of our research as appropriate. We will continue to have Advisory Board meetings, with at least one more in MA during this final year. I will continue to engage collaborators, graduate students, and undergraduates in all aspects of the research. Research results will continue to be disseminated via presentations at regional and national conferences, publication in peer-reviewed journals, and outreach through connections with extension professionals.

Impacts
What was accomplished under these goals? Objective 1A (Adler). We assessed effects of sunflower pollen on pathogen load and performance of wild B. impatiens and other Bombus species using microcolonies. These experiments were completed this year and we anticipate manuscript submission in fall or winter 2019. Objective 1B (Evans). Effect of sunflower pollen on honey bee parasites: in the previous reporting period, a small-scale cup assay showed significant benefits to honey bee disease levels when bees were provided with pure sunflower pollen or sunflower pollen mixed 2:1 with wildflower pollen. However, conducting a variation of this study in the current period found limited benefits of sunflower pollen on honey bee pathogen loads or mortality. Due to the mixed results, we plan to publish these experiments with our other honey bee work in a single larger publication in 2020. Objective 2A (Irwin, McFrederick). In the current reporting period, we supplemented field colonies of Bombus impatiens with wildflower or sunflower pollen.Colonies were infected with Crithidia (yes/no) x pollen supplementation (sunflower/wildflower), with10 colonies per treatment for 40 colonies total. We collected workers weekly for pathogen and viral screening, which is still in progress. In the field, we measured colony size weekly, and two times per week we estimated colony foraging.When colonies went sexual, we pulled the colonies in and processed them for the production of workers, males, and queens, and other measures of colony health and pollen use, which is still in progress. Objective 2B (Adler) will assess the effect of sunflower pollen supplementation on honey bee pathogens and health. During the current reporting period we tested the effect of sunflower pollen, wildlflower pollen, mixed sunflower/wildflower pollen, BeePro, or no supplement on 55 apiaries in a commercial beekeeping operation in Massachusetts. We found that pollen (sunflower and wildflower) reduced the incidence of some viruses, but sunflower pollen alone had no notable effects on any pathogen or parasite assessed (varroa, Nosema, and 8 viruses). Due to challenges obtaining pollen, the proposed work with BIP using fall supplementation will be conducted in the next reporting period; we now have the permits, pollen and sterilization procedure in place. Obj 3A (Baylis) uses a long-term BIP data set to analyze relationships between sunflower plantings and honey bee pathogens (mites and Nosema) across the US. The bulk of these analyses were conducted during the reporting period and suggest relationships between sunflower area and both Nosema and mites, but these are preliminary. Obj 3B (Adler, Irwin, McFrederick), documenting whether sunflower on farms affects pathogen loads and performance of bee colonies, will be conducted in Massachusetts in summer 2019. For North Carolina, due to timing and logistical constraints we plan to request a no-cost extension to conduct this work in summer 2020. Extension Accomplishments: We had our second annual advisory board stakeholder meeting in Massachusetts in March 2019, soliciting input from Board Members (including representatives of beekeeping associations, industry, state government and academia) on our research and extension plans and results so far. Feedback was helpful in shaping our planned activities and goals. Our honey bee Extension Educator, Hannah Whitehead, began her position in May 2018 and has conducted a wide range of outreach and extension activities. During the summer of 2018, the extension educator coordinated the USDA pest and disease survey for Massachusetts, and inspected hives as part of the survey. She also coordinated data collection for a beekeeper-initiated research project, in which she and another lab member gathered disease and pesticide samples from 40 hobbyist apiaries at the beginning and end of the summer. During the past year, the extension educator also created and maintains a pollinator sub-section on the UMass Extension website, including information about bees and beekeeping, as well as updates on UMass resources and events (ag.umass.edu/resources/pollinators). She spoke at eight bee meetings to disseminate information about extension and research activities (reaching 440 beekeepers), led a half-day workshop on Integrated Pest Management for Beekeepers at the Berkshire Botanical Gardens (reaching 25 beekeepers), and organized two full-day workshops at UMass Amherst, in collaboration with the Massachusetts Chief Apiary Inspector: Honey Bees Under the Microscope (April 6, 27 beekeepers attended including 4 state bee inspectors), and the Fight the Mite! (planned for May 4, 32 people are currently registered). Several other half-day workshops are in the works for this upcoming summer, including Planting for Pollinators, and Non-chemical Mite Management Tools. The extension educator also writes a quarterly article in the Massachusetts Beekeeping Association Newsletter which summarizes the latest honey bee research papers. Additional accomplishments: Our first results were published in Scientific Reports (see 'products') to substantial and continued media attention. The paper was selected as one of the top 100 read ecologypapers for Scientific Reports in 2018. Media coverage included: · https://www.newsweek.com/save-bees-sunflower-pollen-could-boost-insects-health-1139409 · https://www.upi.com/Science_News/2018/09/26/Sunflower-pollen-protects-bees-from-disease-study-finds/3101537966687/ · https://www.sciencedaily.com/releases/2018/09/180926082721.htm · https://www.livescience.tech/2018/09/26/sunflower-pollen-has-medicinal-protective-effects-on-bees/ · https://www.newsllive.com/nfl/sunflower-pollen-medicinal-protective-effects-bees-48280408 · https://www.reddit.com/r/science/comments/9j3e8j/ sunflower_pollen_has_medicinal_protective_effects/ · https://news.science360.gov/obj/pic-day/bf17114b-fac4-45dc-9c24-38aa87ec41f6/sunflower-pollen-medicinal-protective-effects-bees · https://phys.org/news/2018-09-sunflower-pollen-medicinal-effects-bees.html · https://montgomerycountybeekeepers.com/2017/06/sunflower-pollen-may-boost-honey-bee-health/

Publications

• Type: Journal Articles Status: Published Year Published: 2018 Citation: Giacomini JJ, Leslie J, Tarpy DR, Palmer-Young EC, Irwin RE and LS Adler. 2018. Medicinal value of sunflower pollen against bee pathogens. Scientific Reports 8: 14394. DOI: 10.1038/s41598-018-32681-y equally contributing senior authors
• Type: Journal Articles Status: Published Year Published: 2019 Citation: LoCascio GM, Aguirre L, Irwin RE and LS Adler. 2019. Pollen from multiple sunflower cultivars and species reduces a common bumble bee gut pathogen. Royal Society Open Science 6: 190279. http://dx.doi.org/10.1098/rsos.190279.
• Type: Journal Articles Status: Published Year Published: 2019 Citation: LoCascio GM, Pasquale R, Amponsah E, Irwin RE and LS Adler. 2019. Effect of timing and exposure of sunflower pollen on a common gut pathogen of bumble bees. Ecological Entomology. DOI: 10.1111/een.12751

Progress 04/15/17 to 04/14/18

Outputs
Target Audience:During the past year, the primary audience reached by our efforts has been academic scientists, growers, beekeepers, seventh grade students and the general public. Changes/Problems:As mentioned above in 'Accomplishments,' we have had difficulty collected enough domestic sunflower pollen for use in commercial apiary trials. Instead, we are researching the effects of various sterilization techniques. We have applied for a permit to deploy sterilized Chinese sunflower pollen in apiary trials at the University of Maryland under Dennis Van Engelsdorp, where colonies are contained and can be destroyed as necessary, instead of in a California commercial apiary as originally planned. Due to this challenge and also because our grant did not start until April 15 instead of the requested January start date, we were unable to complete BIP trials in Year 1 as planned. Instead, these trials will take place in Years 2 and 3. We have decided that it is not be the best use of our resources to conduct the same colony deployment of bumble bees in MA and NC that is proposed for Obj 2A. Instead, we are focusing more energy on testing the best dose and timing of sunflower pollen to be most effective, and costs and benefits of sunflower pollen on bee performance. What opportunities for training and professional development has the project provided?In this first year, this NIFA grant provided opportunities for a first-generation college student who is pursuing a Master's in my lab, and independent research opportunities for five undergraduates including two low-income students, one who is an under-represented minority from Ghana. Not all these students were directly paid by the USDA grant (some were funded by other sources), but all were involved in working on the USDA objectives. These were George LoCascio, Eugene Amponsah,, Bryanna Joyce, Elyse Stone, Ariana McFarland, and Toby Shaya. How have the results been disseminated to communities of interest?Through our first Advisory Board committee meeting, a Farm Day for growers at the S Deerfield research station, BeeFest presentations to the general public, an outreach presentation to 7th grade students, multiple invited university seminars, and presentations at scientific meetings. Specifically: We provided an outreach event to approximately 15 seventh grade girls from under-represented groups in science through the Eureka! Program. I spoke at the Bee Fest in Greenfield to about 50 members of the public. I participated in Agricultural Field Day, S Deerfield Farm, 10-4. "The objective of this event is to open the farm to the public for a guided tour of our current research projects." I incorporated these results into invited research seminars at Bates College and Iowa State University. We presented this research at the annual AFRI PD meeting and the Ecological Society of America conference. Conference presentations: AFRI Annual PD meeting Oct 29-31 2017, Washington DC Rebecca Irwin, Lynn Adler, Jay Evans, Quinn McFrederick, Kathy Bayliss, Deborah Delaney Poster: Sunflowers as treatment and preventative for bumble and honey bee pathogens Ecological Society of America Aug 6-11 2017, Portland, OR Jonathan J. Giacomini, Lynn S. Adler, and Rebecca E. Irwin Paper: The role of sunflower pollen in defending bees against parasites What do you plan to do during the next reporting period to accomplish the goals?Research Objectives: Objective 1A: We are conducting these experiments now and plan to analyze them and write them for publication in the next year. Objective 1B: These experiments have been conducted and we plan to publish them in the next year. Objective 2A: This experiment was conducted this spring and molecular analysis of pathogens will take place in the upcoming year. Objective 2B: The BIP trials will begin this year. Objective 2 add-on: We conducted this experiment this spring and will write it up for publication in the upcoming year. Objective 3A: This will be conducted in the upcoming year. Objective 3B: This will not be conducted until Year 3. Extension Objectives: Our honeybee Extension Educator began her position April 29. In the upcoming year she will network with beekeepers across the state, assess beekeeper needs, develop educational materials and website content, attend beekeeper meetings and appropriate conferences, organize Master Beekeeper workshops, and present results of our research as appropriate. We will continue to have Advisory Board meetings, with at least one more in MA during this year. I will continue to engage collaborators, graduate students, and undergraduates in all aspects of the research. Research results will continue to be disseminated via presentations at regional and national conferences, publication in peer-reviewed journals, and outreach through connections with extension professionals.

Impacts
What was accomplished under these goals? a) (Irwin) We used a single-bee experiment to test how various ratios of sunflower pollen affected Crithidia bombi cell counts. We experimentally infected bees with Crithidia and then they were fed a pollen diet of either: 100% sunflower, 25-75 sunflower-wildflower, 50-50 sunflower-wildflower, 100% wildflower. We found that a diet of 50-50 sunflower-wildflower provided as strong a medicinal effect in terms of reducing Crithidia as a 100% sunflower diet. b) (Irwin and Adler) We wild-collected queens of Bombus impatiens, Bombus griseocollis, and Bombus bimaculatus. Queens were allowed to establish nests in the lab and then we used the workers to experimentally inoculate with Crithidia and then fed them pollen diet of: 100% sunflower, 50-50 sunflower-wildflower, or 100% wildflower. For wild B. impatiens, our results from wild colonies matched those of commercial colonies with 100% sunflower pollen and 50-50 sunflower-wildflower providing medicinal benefits relative to wildflower pollen. Our results for B. griseocollis showed no significant effect of pollen diet on parasitism. The results for B. bimaculatus depended on the parasite strain we used. When we used a parasite strain collected from MA, we found no effect of pollen diet on infection. However, when we used a parasite strain from NC, we found 100% sunflower diet significantly reduced infection relative to 50-50 sunflower-wildflower and 100% wildflower pollen diets. C) (Irwin) We performed a whole-colony lab experimental where we manipulated parasitism (yes/no) x pollen diet (50-50 sunflower-wildflower diet or 100% wildflower diet). We measured parasitism weekly and estimates of colony health. The analyses of colony health are in progress, but the infection results revealed that the 50-50 sunflower-wildflower pollen diet significantly reduced whole-colony parasite infection. D) (Irwin) We used a field experiment to assess how pollen supplementation with sunflower affected Crithidia bombi infection, the acquisition of other parasites and viruses, and colony health. Colonies were supplemented with sunflower or wildflower pollen and were either infected or uninfected with a local strain of Crithidia bombi. We measured colony foraging behavior twice weekly, collected workers weekly for parasite analyses using qPCR, and weighed colonies weekly to estimate colony buildup. Finally, when colonies went sexual, we ended the experiment and measured aspects of colony productivity and health. Laboratory analyses of parasitism and colony health are in-progress. E) (Evans, Irwin, Adler) We attempted to collect sufficient domestic sunflower pollen for use in future field studies. We collected enough to conduct field experiments with Bombus (D) but nowhere near enough to conduct the planned commercial experiments in Apis apiaries. We therefore began researching different methods to sterilize pollen, and found that ethylene oxide is as or more effective than gamma radiation, does not reduce the medicinal effect of sunflower pollen, and does not reduce palatability for Bombus (while gamma radiation does). We are collaborating with Dr. Jamie Strange and colleagues to write this work up for publication. F) (Evans) We tested the effect of sunflower and wildflower pollen on honeybees in cup trials. Results are still being analyzed, but we found a surprisingly dramatic effect of sunflower pollen in reducing Deformed Wing Virus. G) (Adler) We tested the effect of a one-time exposure to sunflower pollen on the likelihood of Crithidia transmission and found no effect. However, in a separate experiment we found that both duration of dose and earlier timing affect Crithidia growth after infection. Seven days of exposure to sunflower was more effective than 3.5 days, and 3.5 days of sunflower immediately after infection was more effective than waiting 3.5 days to administer sunflower pollen. This manuscript is in preparation to submit to Ecological Entomology later this summer. H) (Adler) We are writing up pilot data demonstrating that a wide variety of H. annuus cultivars, wild populations, other Helianthus congeners, and Solidago species all are effective at reducing Crithidia counts. This manuscript is in preparation to submit in the next month. I) (Adler) Undergraduate projects: - tested the effect of sunflower pollen on B. impatiens drones and queens. Interestingly, we found that sunflower reduced Crithidia counts in queens but not drones. We hope to submit this manuscript this fall. - compared the medicinal effects of lotus, tea, and poppy pollen to the positive control of sunflower pollen and negative control of buckwheat pollen. We found no evidence that these new pollen species reduced Crithidia relative to buckwheat pollen. - we prepared to test the effect of sunflower pollen, wildlflower pollen, mixed sunflower/wildflower pollen, BeePro, or no supplement on 55 apiaries in a commercial beekeeping operation in Massachusetts. This experiment began after this reporting period ended. J) (Adler) We held our first Advisory Board meeting in Massachusetts, soliciting input from Board Members (including representatives of beekeeping associations, industry, state government and academia) on our research and extension plans and results so far. Feedback was helpful in shaping our planned activities and goals. K) (Adler) We conducted a search and hired a Honeybee Extension Educator, Hannah Whitehead, whose position began just after the reporting period ended.

Publications

• Type: Journal Articles Status: Under Review Year Published: 2018 Citation: Giacomini JJ, ***Leslie J, Tarpy DR, **Palmer-Young EC, Irwin RE and LS Adler (in review). Medicinal value of sunflower pollen against bee pathogens. Scientific Reports.