Project Number: 6066-21000-001-022-I
Project Type: Interagency Reimbursable Agreement
Start Date: Feb 1, 2023
End Date: May 31, 2024
To determine if different macronutrient ratios in honey bee diets can increase the survivorship and expression level of genes important for development, growth, and immunity of bees infected with either N. ceranae or Deformed Wing Virus (DWV) through two objectives: 1. To identify an optimal macronutrient ratio for normal, uninfected honey bees. This will be done in caged, laboratory environments. Hypothesis: Experimental honey bees will regulate their P:L ratio to reach a similar P:L ratio found in Brassicaceae pollen, a highly preferred pollen type, as found in our previous (unpublished) colony-level, field study. 2. To determine if offering different macronutrient ratios in diets can affect the survivorship, physiology, and expression level of genes important for growth, development, and immunity in bees infected with either N. ceranae or DWV. Similar to when they are stressed with temperature or nicotine, honey bees infected with either N. ceranae or DWV will likely have lower fitness and expression level of genes important for growth, development, and immunity.
Methods: Determining the macronutrient intake target in uninfected bees. We will perform a series of choice and no-choice assays to determine nutrient regulation in nurse bees with unmanipulated background levels of disease prevalence. The no-choice assay will confine cohorts of bees to a single artificial diet of varying P:L ratios. These results will provide us with the information necessary to address objective 2. Diets: Artificial diets will be made using proteins composed of isolated soy protein. The final P:L ratios will be 35:15, 30:20, 25:25, and 20:30, respectively. Water and 50% sugar solution (w:v) will be provided in separate gravity feeders ad libitum. No-choice assays: To control for bee age, frames of capped brood will be chosen from randomly selected. Cohorts of 30 newly emerged bees will be collected from the frames and placed in cages with water and 50% sugar syrup. Each cohort will be presented with a single P:L diet, which is typically done to test a diet functional significance. We will measure total macronutrient consumption and fitness parameters. Choice assays: Each cohort will be presented with two different diets to select from concurrently. Providing bees with two suboptimal diets will allow them to choose the ratio of nutrients that will maximize their fitness. This framework assumes that organisms will self-select nutrients to reach an optimal intake target for growth and development. This assay will provide us with an intake target toward which young bees are regulating as they consume the diets. Assessment of fitness parameters: I will measure hypopharyngeal gland acini size and total lipid content as the physiological parameters that are indicative of individual bee fitness. Obj. 2 Methods: Determining the effects of diet on bee pathogen tolerance. We will perform a series of no-choice assays to determine if diet can impact the bees; defense against pathogens. Diets and no-choice assays: The methodology for diet creation, no-choice assays, and fitness quantification will be similar to Obj. 1, with modifications for pathogen inoculation. Within the first 24 hours post emergence, caged bees will be group fed either a 1) 40,000 Nosema spores/bee inoculum diluted into a 50:50 w:v sucrose solution, 2) 108 genomes/bee Deformed Wing Virus (DWV) inoculum diluted into a 50:50 w:v sucrose solution 11, or 3) control treatment of solely a 50:50 w:v sucrose solution. Based on the results from the previous objective, we will use diets with varying P:L ratios to conduct no-choice assays with either untreated control bee cohorts or with cohorts of bees inoculated with either N. Ceranae or DWV. Diet consumption will be weighed daily to quantify the amount of macronutrients consumed over time. Individual assessment: Survivorship and bee physiology will be assessed using similar methods described in the previous objective. We will also sequence bees in each treatment group and compare gene expression between inoculated and control bees fed diets with varying macronutrient ratios. By doing so, we can better determine how nutrition impacts the growth, development, and immunity of workers infected with either N. ceranae or DWV.