Exploring the synergistic potential of fungicides and parasites as stressors of bumble bee health and pollination services in greenhouse tomatoes

Final report for GNC21-335

Project Type: Graduate Student
Funds awarded in 2021: $14,993.00
Projected End Date: 03/31/2024
Grant Recipient: The Ohio State University
Region: North Central
State: Ohio
Graduate Student:
Faculty Advisor:
Dr. Frances Sivakoff
The Ohio State University
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Project Information


Exploring the synergistic potential of fungicides and parasites as stressors of bumble bee health and pollination services in greenhouse tomatoes

Bumble bees (Bombus spp.) are important native pollinators critical for the commercial production of tomatoes. However, bumble bees are experiencing global population declines due to an array of known and unknown factors. Recent studies have highlighted the link between fungicide exposure and bee health, and suggested that fungicide exposure may increase susceptibility to other stressors. Thus, it seems likely that multiple stressors may have synergistic negative effects on bumble bee health and threaten the sustainability of this important natural resource base on which agriculture depends (NCR-SARE broad-based outcome).

Ensuring bumble bee health is essential for Ohio tomato greenhouses, a $32 million dollar industry with over 300 operating locations. Commercial greenhouses are also an ideal system to evaluate the effects of multiple stressors. Tomatoes grown in greenhouses regularly have a range of fungicides applied, many of which are systemic, detectable in pollen, nectar, and in the bees themselves. At the same time, growers purchase commercial bumble bees to pollinate their tomatoes, and these colonies can arrive infected with various pathogens. Sub-lethal effects of chemicals can reduce bumble bee foraging efficiency, and it is possible that greenhouse tomato growers are not reaching maximum yield potential when their bumble bees are impacted by this suite of stressors. The goal of my study is to analyze the combined effects of pathogen infection and fungicide exposure in tomato greenhouses on bumble bee health and pollination efficiency, to provide growers with insights that will improve their use of commercial bumble bee pollination services.

I will place bumble bee colonies established from field-collected queens, confirmed to be parasite free, into commercial greenhouses during tomato pollination. I will collaborate with growers to place colonies in greenhouses exposed to a gradient of fungicide applications. These colonies will be equipped with pollen traps to measure fungicide levels in bumble bee-collected pollen. These greenhouses will also contain commercial bumble bee colonies, which I will sample for the presence of the gut parasite, Crithidia bombi. At the end of the study, I will evaluate the relationships between fungicide levels and a series of bee health metrics and pollination efficiency. The key outcomes of this study will be a functional risk assessment for growers of the impacts that fungicides, and potentially pathogens, have on yield due to altered pollinator foraging behavior and health.

Project Objectives:

Learning outcomes: 

(1) Greenhouse tomato growers will learn how fungicide use impacts the bumble bees they purchase for pollination services. I will provide them with an analysis of pollination efficiency related to fungicide exposure, to increase grower understanding of how fungicides affect bumble bee foraging behavior and ultimately crop yield and quality.

(2) Growers will gain knowledge of the prevalence of pathogens in their commercial bumble bee colonies and if the presence of pathogens affects pollination efficiency. I will provide growers an analysis of the infection rate of pathogens in their commercial bumble bees. This is important for growers to understand the quality of the bumble bees they are paying for. I will create extension fact sheets that describe how fungicides affect important bumble bee colony health metrics.

Action outcomes: 

(1) After learning how fungicides affect their bumble bees, tomato greenhouse growers may modify their use of fungicides to maximize the effectiveness of their bumble bee pollinators. If fungicides are shown to have adverse effects on the pollination efficiency of greenhouse bumble bees, then growers will consider this tradeoff before applying fungicides.

(2) After learning about parasite prevalence in commercial bumble bees, tomato greenhouse growers may encourage the commercial bee-keeping industry to implement additional safeguards to prevent against parasite infection. This will be beneficial not only for the tomato growers, who will receive healthier, potentially more efficient bumble bees, but also the natural environment, because there will be less pathogen spillover into wild bee populations.


Materials and methods:

Study Location

All data collection was completed at a commercial tomato greenhouse facility[1] in northern Ohio. This facility consists of three greenhouses sized 265m x 245m. Each greenhouse contains 33 rows of plants on both the north and south sides, 2m wide, for a total of 66 rows of tomatoes. The tomato variety grown in each greenhouse was tomato on the vine.

Study Species

The Common Eastern Bumble Bee (Bombus impatiens)

The common eastern bumble bee is a stable species traditionally found across the eastern half of the united states [1]. B. impatiens is a generalist species, typically easy to rear, and is an important crop pollinator of tomato greenhouses [2,3]. The majority of commercial bumble bees in the US come from two sources, Koppert Biological Systems Inc. or BioBest Ltd.[4]. Each colony arrives with one queen and anywhere between 20-50 female workers.

Study Design

Colony Placement

Each greenhouse had a total of 70 hives at a time, as decided by our grower collaborator, and we monitored 10 randomly selected hives per greenhouse. We placed these 10 colonies of commercial B. impatiens colonies in each of the three tomato greenhouses between July 20 – October 30 of 2022. Each colony was present in the greenhouse for a total of 11 weeks. We weighed each colony prior to placement inside the greenhouse, and upon removal from the greenhouse. Before being placed inside the greenhouse we also removed ten workers from each colony to be sampled for the presence of Crithidia bombi. Each experimental colony was placed along the walkway spanning the length of the greenhouse, roughly equal distance apart (10m).


During the 11-week period that the bumble bee hives were located in the greenhouse, a trained pollination scorer recorded the level of anther bruising on 6 plants in each of the 66 sections within the greenhouse, for a total of 396 pollination-score samples each week. Bumble bees pollinate through a method called “buzz pollination”, where they grasp the anther of a flower and vibrate to dislodge the pollen [5]. The resulting necrotic tissues can be measured on a scale of bruising intensity to quantify pollination efficiency [6]. Higher bruising levels are correlated with more desirable pollination and higher fruit yields [6]. The scorer recorded anther bruising on a scale of 0-3, with 0 = none, 1 = low, 2 = intermediate, 3 = high pollination, respectively [5].

Colony health

At the conclusion of the 11-week period we collected each of the 10 colonies in each greenhouse. Each colony was placed in a freezer until all bees were dead. We then collected all of the workers, queens, drones, and new gynes from each colony. We also collected a sample of brood cells, including new-queen cells when possible, and honey pots from each colony. We counted the number of each caste, and will measure the radial cell of a sample of workers and drones from each colony to make inferences on body quality [7].

Crithidia bombi infection

We gut-dissected 10 workers collected from each colony prior to placement in the greenhouse, and 10 workers collected from each colony after removal from the greenhouse. We will use PCR methods to compare the presence of Crithidia bombi infection in the bumble bees before and after being placed in the greenhouse.

Chemical application and testing

Our collaborator provided us with complete records of pesticide applications that took place during the duration of our experimental hives presence in the greenhouse. We sent samples of the collected bumble bee colony wax to the University of Guelph Agriculture and Food Laboratory (Guelph, Ontario) for testing of the presence of each of these chemicals. This will allow us to investigate if the chemicals sprayed on the plants are carried back to the colony and fed to brood in detectable quantities by workers.

Statistical analysis

We will correlate the pollination score data, as measured by anther bruising record, with timing of each of the pesticides sprayed throughout the monitoring period. We will analyze these results using a generalized linear model with a negative binomial distribution. We will hold chemical as the explanatory variable, with average greenhouse temperature, and average greenhouse hours of light as covariate, and greenhouse number as a random effect. We will use this same model structure to correlate counts of each caste with the presence or absence of Crithidia in the hive, either before or after placement in the greenhouse.

[1] Exact location kept anonymous at the request of collaborator.



[1]       R. Hatfield, S. Colla, S. Jepsen, L. Richardson, R. Thorp, S. Foltz Jordan, IUCN Assessments for North American Bombus Spp., The Xerces Society for Invertebrate Conservation, Portland, OR, 2015.

[2]       D.R. Artz, B.A. Nault, Jnl. Econ. Entom. 104 (2011) 1153–1161.

[3]       J.P. Strange, Journal of Economic Entomology 108 (2015) 873–879.

[4]       H.H.W. Velthuis, A. van Doorn, Apidologie 37 (2006) 421–451.

[5]       L.A. Morandin, T.M. Laverty, P.G. Kevan, Ec 94 (2001) 462–467.

[6]       L.A. Morandin, T.M. Laverty, P.G. Kevan, Ec 94 (2001) 172–179.

[7]       C.B. Muller, T.M. Blackburn, P. Schmid-Hempel, Ins. Soc 43 (1996) 227–233.



Research results and discussion:


Three bar graphs, in a grid format, from left to right. First, shows effect of greenhouse identity on pollination score, second shows effect of greenhouse identity on parasite detection, and third is 
a stacked bar chart which shows effect of greenhouse identity on count of fungicide and pesticide application. Has the short description "Increased fungicide application may be associated with decreased pollination efficiency and increased parasite prevalence. "
(Left) Pollination efficiency score measured for each greenhouse. Higher scores are more desirable and lead to better fruit yield. (Middle) counts of colonies testing positive for a common gut parasite after 12 weeks of pollinating within the greenhouse. (Right) frequencies of fungicide and insecticide applications within each of the three greenhouses.

We are currently analyzing the data collected for deeper inspection. Of the data we have analyzed so far, we have found interesting connections between fungicide, parasite prevalence, and pollination efficiency. We tested 300 bumble bees across 30 colonies from the three greenhouses for C. bombi. Every colony tested negative for the presence of the parasite prior to placement in the greenhouse, however 19 of the 30 colonies tested positive for the parasite at the end of the 12 weeks. Five of the ten colonies in Greenhouse 1 had at least one bee test positive for the parasite, four out of the ten bees in Greenhouse 2, and ten out of ten colonies in Greenhouse 3. We found that Greenhouse 3 also had the lowest pollination score overall, and the greatest occurrences of fungicide application (Figure 1).

Based on this preliminary data analysis, there are strong indicators that fungicide application is negatively correlated with anther bruising levels of greenhouse tomatoes pollinated by bumble bees. 

Participation Summary
1 Farmers participating in research

Educational & Outreach Activities

2 Curricula, factsheets or educational tools
1 Webinars / talks / presentations

Participation Summary:

2 Ag professionals participated
Education/outreach description:
  1. Consultations (0)
  2. Curricula, fact sheets, educational tools
    1. One fact sheet published
  3. Journal articles (0)
  4. On-farm demonstrations (0)
  5. Online trainings (0)
  6. Published press articles
    1. Blog post about project on Sivakoff Lab Website
  7. Tours (0)
  8. Webinars/talks/presentations
    1. Talk planned for the 2024 Entomological Society of America
  9. Workshops/field days (0)
  10. Other
    1. Joined the Ohio Controlled Environment Agriculture Center (OHCEAC) Consortium
      1. Attended the annual 2023 meeting
      2. Will attend the annual 2024 meeting

Education and outreach description

The fact sheet "Bumble Bee Pollination in Tomato Greenhouses" has been published through The Ohio State Extension Office (https://ohioline.osu.edu/factsheet/ent-0092). My lab has also joined the Ohio Controlled Environment Agriculture Center (OHCEAC) Consortium, which is an industry-academic partnership focused on controlled environmental agriculture. I will be attending the annual OHCEAC meeting in June, where I will meet with industry professionals to discuss innovations in the field. 

Participants - The OHCEAC members, and the OSU Extension Office agents 

Project Outcomes

1 Farmers reporting change in knowledge, attitudes, skills and/or awareness
4 Grants received that built upon this project
2 New working collaborations
Project outcomes:

We found that of the bees tested in this study, bumble bees entering greenhouses directly from the commercial rearing facility were free of gut parasites. After 12 weeks of pollinating in the greenhouse, the majority tested positive for parasite infection. From a sustainability perspective, this is important for several reasons. First, it is possible that the greenhouse is not being adequately sterilized between planting cycles. This has implications for other contaminants and diseases not being properly expunged as desired. It is also possible that the bees are exiting the facility, and returning to their colonies with the parasite. In this way, they might introduce external crop diseases into the system, which is the primary threat our grower collaborators wish to prevent. Our collaborators expressed a strong enthusiasm for trying new methods based on data results, and we are excited to explore possibilities to increase pollination efficiency and overall tomato yield going forward. 

Knowledge Gained:

Completing the work funded by SARE improved our lab's understanding of sustainable agriculture by allowing us to work directly with industry personnel, and learn from their needs and perspectives. Our collaborators provided our lab with a tour of their greenhouse facility, which is not open to the public. During visits to the greenhouse space and in meetings with our collaborators we learned that there is a strong desire to pursue non-chemical pest control in controlled agriculture environments (CAE), and that the biggest threat CAE faces is contamination from external entities. Introduction of disease and pests from external clothing, shipping products, and goods brought into the greenhouse is a constant worry. 


Future work should explore the effects of fungicide on bumble bee parasite infection and pollination efficiency in field settings. Work may also extend to other managed pollinator species. 

Any opinions, findings, conclusions, or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture or SARE.