A new duplex qPCR assay for the quantification of honey bee (Apis mellifera) parasites Nosema ceranae and Nosema apis tested with low dose experimental exposure

Abstract

Nosema ceranae and Nosema apis are microsporidian parasites that cause disease in the European honey bee. Nosema infection is identified as a potential cause of colony loss by beekeepers. Given the importance of Nosema infection in colony mortality and productivity, developing new and improved ways of detecting and quantifying infection loads is vital. We designed and tested a new duplex qPCR assay for the accurate quantification of both N. ceranae and N. apis utilising TaqMan chemistry and the new gBlock^® method for the standards. The assay showed good linearity with natural Nosema infection, and a strong correlation with microscopic spore counts. This new assay has high sensitivity and repeatability and was used to investigate Nosema infection in hive surveys and following low dose experimental exposure. In local hives, we found relativity low levels of N. ceranae and very little N. apis across three sites in Blacksburg, VA. A survey of two bee yards in West Virginia showed much higher levels of N. ceranae, but consistent low levels of N. apis. For the experiment, caged bees from two different hives were fed 100 Nosema spores (or a control solution). Exposed bees were collected after two or five days, and infection was quantified using the new assay. Given the low dose, infection levels were not 100%, with some exposed bees remaining N. ceranae free, while others only developed low level infection. This low dose exposure and subsequent infection status (low level infection or infection free) could provide a new understanding of Nosema infection establishment.

Keywords:

• Nosema
• Nosema ceranae
• qPCR
• Apis mellifera
• infection

Acknowledgements

The authors thank Richard Reed for the access and use of his apiary at the Hale Community Garden, and the Appalachian Beekeeping Collective for access to two bee yards in West Virginia to sample their hives and collecting bees for Nosema spore extraction.

Disclosure statement

The authors declare that there is no conflict of interest.

Additional information

Funding

This work was supported by the National Science Foundation under grant MCB-1817736.