Michigan is the second most diverse agricultural state, supported by commercial beekeepers who provide crop pollination. However, following pollination contracts on nutritionally poor monoculture crops, such as blueberry, honey bees often suffer from diseases such as European foulbrood (EFB). EFB is a bacterial disease that affects the larvae, often killing them, which can ultimately lead to colony mortality. EFB can also negatively affect colony size, which has economic implications for beekeepers through depressed pollination contract prices and honey production. While the link between EFB and nutrition has not been well established in a field-realistic setting, understanding it is an essential step in making management recommendations for beekeepers. Therefore, this study will evaluate the relationship between EFB recovery rates and nutrition to determine best management practices for mitigating these effects. To do so, it will analyze the effect of supplemental feeding management, as compared to traditional in-hive management on EFB recovery speed. This will be achieved by tracking the recovery rates of infected hives under six different in-hive treatments, including a control (untreated), antibiotics (traditional), antibiotics and a plant-based supplement, plant-based supplement, pollen-based supplement, and probiotics. This research will be conducted in Michigan during the summer feeding period immediately following blueberry pollination. Results will provide beekeepers with best management practices optimize colony recovery and advice on the necessity of supplemental feeding. To evaluate the adoption potential of the management suggestions made through extension to beekeepers, I will administer surveys to assess change in knowledge, awareness and attitudes. Additionally, I will assess intended change in management as a result of extension. Mitigating EFB through nutrition, rather than relying solely on antibiotics is a more environmentally sustainable and affordable solution for beekeepers. Furthermore, because the FDA began mandating veterinary prescriptions for bee antibiotics in 2017, this is a particularly relevant issue. Additionally, disease pressure on the beekeeping industry will have broader implications on national pollination services and the food system. Not only will it cause social and economic tension between growers and beekeepers, as decreased pollination availability increases demand and price, but it will also affect food quality and price for society as a whole.
I will communicate with beekeepers through meetings, articles, extension, and email newsletters to increase knowledge related to EFB treatment, including A) resources to accurately diagnose EFB, B) EFB management costs (economic and environmental), and C) nutrition and supplemental feeding effects on EFB. This information is expected to facilitate changes in skills and attitudes for nutrition-based management, initiating two main action outcomes. First, beekeepers will more accurately field-diagnose EFB, decreasing unnecessary, costly management. Second, beekeepers will practice more sustainable in-hive management practices. These are potentially more cost effective and environmentally conscientious treatments than antibiotics.
Many specialty crop growers are unaware that crop pollination can stress honey bees, and that stressed colonies require more inputs and are more expensive to maintain. Understanding the economic and social inter-dependence of the beekeeper-grower relationship is essential to maintaining reliable stocks of colonies and stable pollination contract services and prices. Through Eextension presentations and resources, blueberry growers will gain a better appreciation for the economic and environmental consequences of their support of honey bees, the role they play in EFB, and how they can support honey bee health on their farms. Because EFB is a stress-related disease, growers adopting stress-reduction practices can reduced EFB incidence. Supportive practices include night spraying, planting flowering strips, and carefully selecting hive drop sites. As a result of this awareness, growers may alter their practices to support the beekeeping industry on which they rely.
We tested colony recovery speed from EFB in a cohort study under six different in-hive treatments, including (1) antibiotics (Terramycine), (2) antibiotics (Terramycine) with a plant-based pollen patty (Bee-Pro), (3) a plant-based pollen patty alone (Bee-Pro), (4) a 15% pollen-based patty (Global Patties), (5) probiotics (SuperDFM), and (6) a no-treatment control in which the colony was opened but nothing was added each time. Antibiotics are the conventional form of treatment for European foulbrood. However, antibiotics can be difficult to obtain since the 2017 FDA guidelines stipulated the need for a veterinary feed directive for antibiotic use in food producing animals, including honey bees (FDA, HSS 2015). Additionally, there are concerns about antibiotic residue in honey (Bargańska et al. 2011) and antibiotic resistance (Waite et al. 2003). As more sustainable alternatives, we tested protein supplements, including a plant-based and pollen-based patty, which we hypothesized would enhance recovery speed by reducing nutrition stress (Forsgren et al. 2005; Roetschi et al. 2008) and boosting immunity (Alaux Cédric et al. 2010). Based on beekeeper feedback we also tested probiotics. Balanced gut microbiomes promote metabolism and boost immunity (Li et al. 2019).
Each treatment was replicated in 10 colonies. These replicates were split between four different apiaries. Two apiaries which each contained 72 colonies were managed by one beekeeper and had four treatment replicates each. These apiaries were in Allegan and Kalamazoo Counties, Michigan, approximately 11.5 km from each other. Approximately 40 km south in Cass and St. Joseph Counties, Michigan were the other two apiaries which were approximately 38 km from each other. These other two apiaries were managed by a different beekeeper and each contained approximately 30 colonies. Each of these apiaries had a single replicate of each treatment. Both collaborating commercial beekeepers pollinate on high bush blueberry (Vaccinium corymbosum) in May, then moved bees into holding yards from June through July 2019. We worked with these commercial beekeepers because in recent years commercial beekeepers across North America have observed elevated incidence of EFB in colonies which service blueberry pollination contracts (Dufour et al. 2020). In our system in 2019 we observed infection in over half of commercial colonies coming off blueberry pollination.
Upon colonies arriving in holding yards, each colony was inspected for visually detectable signs of EFB. Signs of EFB included 3rd-5th instar larvae which showed corkscrewing, yellowing, melting and exposed trachea (Bailey 1961). Atypical signs of EFB also included holes in the cappings and scaling (Forsgren 2010). Colonies were rated on an ordinal scale based upon number of cells exhibiting signs of disease. Colonies with greater than 100 diseased cells received a 3, 11-100 diseased cells received a 2, 1-10 diseased cells received a 1, and no diseased cells received a 0. This ranking system was chosen to be consistent with research being conducted by other researchers across North America. Diagnosis of infected colonies was confirmed by sending sampled larvae to the USDA-ARS Beltsville Laboratory, where the presence of Melissococcus plutonius was confirmed by with light microscopy. Colonies were also inspected for potentially confounding health effects including the presence of a laying queen and other visually detectable signs of pests and disease, such as chalkbrood, small hive beetle, and Varroa destructor mites. Varroa abundance was estimated using an alcohol wash (Fries et al., 1991). All enrolled colonies had an EFB severity rating of at least 2. Within apiary, colonies were blocked by severity into each of the six treatments. Treatments were applied per label. Antibiotics were applied every four to five days for a total of three treatments. Probiotics were applied once, per label, except for in the single-treatment yards, where they were applied upon enrollment, then again five days later, due to a miscommunication of methods. All patty-supplemented colonies were maintained at 0.9 kg per colony at each visit.
Colony inspections occurred every two weeks from enrollment to six weeks post-enrollment to track recovery speed. At each inspection, EFB infection severity was recorded as well as colony survival and queen status. At enrollment and at the six-week follow up, number of frames of adult bees were also recorded (Nasr et al., 1990). At enrollment before treatment, and at the first post-treatment inspection (four-weeks post enrollment), a sample of six nurse bees was obtained by shaking a frame of brood vigorously and scooping bees which remained on the frame. Nurse bees were immediately put on dry ice, then transferred to -80°C until processing. Nurse bee heads were weighed, and fat body size was quantified using the ether wash method, as described in Wilson-Rich 2008 (David et al., 1975; Doums et al., 2002; Wilson-Rich et al., 2008). Nurse bee condition can serve as early indication of colony health and help to elucidate the mechanism through which different treatments are affecting colonies (Forsgren et al. 2005; Roetschi et al. 2008).
All statistical analyses were completed in R-studio version 3.5.2 (R Core Team, 2018). To determine differences in treatment group infection severity, analysis of variance on general linear mixed models (GLMM) was done using the lme4 package (Bates et al., 2015), followed by Tukey’s pair-wise comparisons using the MultComp package (Hothorn et al., 2008). Within each round, treatment was regressed against EFB status, with apiary, nested within beekeeper as random effects. Treatment differences in colony growth in frames of adult bees was likewise calculated using GLMM. Associations between treatments and other pests/ disease were tested for independence using a chi-squared test. To determine differences between treatments in nurse bee head and fat body weight change pre- and post-treatment, we likewise used ANOVA on GLMM, with apiary, nested within beekeeper as random effects and Tukey’s multiple comparison to determine pair-wise differences.
Initial EFB severity was not different among treatments upon enrollment, due to appropriate blocking (F5, 54=0.84, p=0.53). Two-weeks post enrollment, during the last round of treatment application, colonies which were treated with antibiotics, including both the antibiotic only treatment and the antibiotic with soy-patty treatment, had greater recovery than those which were not treated with antibiotics (F5, 45.61=9.64, p<0.01). The non-antibiotic treated colonies had an average severity rating of approximately 2, which would indicate at least 11 affected cells, while 75% of the antibiotic treated colonies had completely recovered and contained no signs of affected brood. By four weeks post-enrollment, which was two-weeks after treatment had concluded, the average severity across all colonies was 0.5, with 78% of colonies showing no signs of disease. There were treatment differences (F5,52=2.57, p=0.04) but only between the antibiotic treatment and the soy-based protein treatment (z=3.2, p=0.02), with the soy-treated colonies displaying more severe signs of EFB than the antibiotic-treated colonies. One-month post-enrollment there were also treatment differences (F5,48=4.07, p<0.01); the pollen-based patties had a resurgence of the EFB signs. While all other treatment groups had recovered to a rating between 0 and 1, the pollen-based patty group had an average severity rating of 1.4.
Half of the colonies treated with probiotics showed signs of chalkbrood two weeks post-enrollment, and half of the colonies that received pollen-based patties had small hive beetles six weeks post-enrollment. However, presence of chalk brood and small hive beetle were independent of treatment within each sampling round, except for the presence of small hive beetle two weeks post-enrollment (X25=11.5, p=0.04). There was no difference in colony growth between the different treatments (F5, 49=1.86, p=0.12).
Processing and analysis of nurse bee samples is currently ongoing, and we anticipate having results in two weeks.
Post treatment course, there were no differences between antibiotic-treated colonies and the control group in this cohort trial. This finding reinforces the best management practice of judicious antibiotic use and suggests that watchful waiting is as efficacious as antibiotics. Recovery speed of both antibiotics was the same as that of the control shortly after treatment concluded. Additionally, recovery maintenance was the same for antibiotic and control colonies. These are promising findings, due to potentially detrimental effects associated with antibiotics including antibiotic resistance (Waite et al. 2003), antibiotic residues in honey (Bargańska et al. 2011), and difficulty obtaining veterinary feed directives (FDA, HSS 2015). Recovery speed was slightly reduced in colonies fed soy-based patties. Colonies in the soy-patty group took an additional two weeks to be comparable in disease severity to the conventional antibiotic treatment. The mechanism behind this slower recovery speed is unclear. We hypothesized that protein feeding would increase colony recovery speed by rescuing colonies from nutritional stress (Forsgren et al. 2005; Roetschi et al. 2008) and providing protein to boost immune function (Alaux Cédric et al. 2010). However, this was not supported by our data. The patties were often consumed by the colony and no other pests or diseases obviously confounded this group’s recovery. During the course of this experiment, Varroa was sufficiently controlled with no counts exceeding a 2% threshold. Therefore, the slower recovery of soy-fed colonies could be an artifact that could be resolved with increased sample size, as there was large variability among the colonies within treatments. The pollen-patty fed colonies recovered at the same speed as the control colonies but increased in disease severity one-month post treatment. Pollen patty fed colonies recovering at the same speed as the control colony provides more evidence that protein feeding imparts no additional benefit to EFB recovery. EFB is a self-limiting disease (Bailey 1960) that clears on its own and the addition of in-hive products are costly implements that provide little benefit or even negative effects. While not statistically significant, colonies that received pollen-based patties had greater small hive beetle pressure, which could have stressed colonies, leading to disease recurrence (Roetschi et al. 2008). We expect that the nurse bee physiology data will further elucidate the mechanism underlying these trends and indicate the potential for long-term colony health.
Educational & Outreach Activities
To date, an educational curriculum has been developed on European foulbrood disease background, in-hive identification, diagnosis support, prevention and sustainable treatment options, based upon the findings of the NC-SARE grant. This curriculum has been presented to the Nebraska Beekeepers’ Association through a webinar, which included 30 beekeeper attendees. There are plans to develop this curriculum for the Great Plaines Master Beekeepers’ Program, which currently has 250+ registered beekeepers. In addition, this curriculum will be presented to Michigan beekeepers through the Southeast Michigan Beekeepers Association Bee School and the Oakland Beekeepers Club. These club meetings primarily reach hobbyist and beginner beekeepers.
Additionally, findings from this project have been presented to growers and commercial beekeepers at the Great Lakes EXPO in Grand Rapids, MI at the blueberry session. This is a grower-focused show, with 3,600+ attendees. Findings were also presented to bee researchers and beekeepers (commercial, sideliners and hobbyists) at the joint American Bee Research Conference and American Beekeeping Federation Annual Conference in Schaumburg, IL. These data were also presented at an entomology departmental seminar in Pennsylvania State University.
Considerations from this research have been included in the Blueberry Pollinator Stewardship Plan, which provides best management suggestions to blueberry growers on how to support healthy pollinators on their farms. I also plan to write an article for the American Bee Journal on these research findings as well as a journal article for a scientific audience.
My project has found support for and communicated the importance of judicious use of antibiotics for honey bee disease management. Based upon my research findings, I offer tools for accurate diagnosis and advocate for watchful waiting to treat cases of European foulbrood. Judicious antibiotic use contributes to sustainability by preventing antibiotic resistance (Waite et al. 2003), antibiotic residues in hive food products (Bargańska et al. 2011), and negative health consequences to adult bees (Li et al. 2019). Healthier adult bee populations and residue-free honey also provides economic benefits to beekeepers in the form of income. Likewise, watchful waiting saves beekeepers money by removing the need for investment in in-hive products. European foulbrood is a spatially- distributed disease (Belloy et al. 2007), so providing beekeepers with the tools for sustainable treatment can also provide environmental benefits by decreasing inter-colony transmission. Responsible beekeeping in turn provides social benefits between neighboring beekeepers. Additional social benefits include enhancing mutual understanding between beekeepers and the blueberry growers with whom they contract. Because blueberry pollination contracts can be risky to honey bee colonies, beekeepers may choose to charge more for pollination services. This would lead to negative economic consequences for growers. Therefore, both groups are stakeholders in sustainable treatment of European foulbrood. My project supports these sustainable treatment options and communicates these findings to both beekeepers and growers, bolstering future sustainability including economic, environmental and social benefits. Indeed, personal communications and feedback from beekeepers have indicated their new knowledge, skills and intended action changes to more sustainable practices as a result of my outreach for this project.
This project revealed that more sustainable agricultural practices can be as efficacious and more affordable than conventional agricultural practices. Before this experience, I often took for granted that conventional methods were the traditional practice because they were the most effective. However, this experience has made me more interested in sustainable agriculture research in order to find more sustainable alternatives. Growers and beekeepers have been very interested in this research due to its applications. Collaborating with and answering questions that serve beekeepers and growers has given me a sense of fulfillment, knowing my research will have direct implications for people and the environment. These collaborations also offer valuable insights and meaningful feedback, which have enhanced my own knowledge, experience and skills. In particular, through this project I gained better communication skills by having the opportunity to communicate research to a new audience. My awareness of the importance of pairing education with sustainability research has increased as a result of this grant experience. It has made me seek more outreach opportunities and consider implications of my other research. Overall, this experience has enhanced my knowledge of treatment-free beekeeping, reinforced my attitude and awareness of the importance of researching sustainable agriculture practices, and given me new skills in sustainable agriculture research communication.