1a. Objectives (from AD-416):
The objective of this project is to quantify the prevalence of parasite infections in wild populations of bumble bees, and to determine if elevation has any affect on the incidence of disease in these bees. Bumble bees are important pollinators of wild plants and agricultural crops. The populations of some species of wild bumblebee are in apparent decline in Europe and North America, but the extent of the declines cannot be precisely ascertained in the United States because of a lack of baseline population data. However, at least three (3) species of North American bumblebee are now endangered or extinct. Suspected causes for the decline include parasites and pathogens, habitat fragmentation, and pesticides. In this project, we will address four questions: (1) What is the prevalence and intensity of Crithidia and Nosema infections among bumblebee species living at different elevations in Northern Arizona; (2) Are there differences in patterns of infection among these parasites and between elevation zones; (3) What is the phenology of Crithidia and Nosema at different elevations in Northern Arizona bumble bees; and (4) Does the phenology of Crithidia and Nosema differ at different elevations.
1b. Approach (from AD-416):
The unique geography of Northern Arizona offers an opportunity to study the affect of elevation on the prevalence, phenology, and infection intensity of Bombus parasites because it is home to a variety of bumble bee species living along an elevation gradient. The Colorado Plateau rises abruptly from surrounding terrain in a formation of escarpments known as the Mogollon Rim. Not far from the Mogollon Rim, the San Francisco Peaks, which are the highest mountains in Arizona, attain an elevation of nearly 13,000 feet. Bumble bees will be sampled at bi-weekly intervals at three different elevational zones. Bumblebees of all available species at these sites will be collected, but the elevation zones will be selected such that least three target species can be collected at multiple zones so that comparisons can be made within the same host. To quantify infections by two parasites, Nosema bombi and Crithidia bombi, the digestive tract of these bees will be removed and examined microscopically for the presence of Crithidia bombi and Nosema bombi. Pathogens from positive samples will be quantified microscopically to determine the intensity of infection. Bees will be collected until workers are no longer present in the field during the fall. The following spring, queens will be collected, killed, and examined for the Crithidia and Nosema infections. The prevalence and distribution of both the bees and their parasites will then be quantified over both time and elevation to reveal what patterns are occurring in natural settings.
3. Progress Report:
Bumblebees (1,230) were captured at 3 elevation zones in Northern Arizona during the spring and summer of 2011 and the spring of 2012. In addition to capturing individual bees foraging on flowers, 2 nests were collected, one at elevation Zone 2, and one at elevation Zone 3. The bumblebees from these nests still need to be analyzed. The bumble bees were collected from 15 field sites. Initially, four field sites were sampled for each elevation zone, but some of the field sites yielded few bees, so three other field sites were added. Bees were caught one at a time, either by net or by scooping an individual bee into a vial from a plant, then were killed by freezing them. After being killed and photographed, each bee was stored in ethanol and kept frozen until it could be identified and autopsied. Autopsies were conducted by removing the entire gut of each bee, crushing the gut in 100 ul of sterile water, then examining the gut at 400X magnification with a phase-contrast microscope. If spores were found, they were quantified visually under the microscope. Those with at least one Nosema spore were designated ‘positive’ for Nosema. After observations were made, each prepared slide was air dried and stained with Geimsa stain. Cover slips were affixed to dried stained slides using either Permount or Cytoseal 60 mounting medium. Nosema-like spores were examined carefully using Geimsa stained slides to verify or exclude the pathogen. A random selection of Geimsa stained slides for ‘positive’ specimens was examined at 1000X using the bright field microscope and immersion oil. Before finalizing the results, the Geimsa stained slides will be carefully examined for all ‘positive’ specimens. All autopsies have been completed, but a sample of Nosema-positive samples will be evaluated at the ARS lab in Logan to verify the species of the pathogen. Statistical analyses still need to be conducted.