The Canadian Entomologist (2010) 142, 271-283

Karen L. Burgher-MacLellan, Geoffrey R. Williams, Dave Shutler, Kenna MacKenzie and Richard E.L. Rogers (2010)
Optimization of duplex real-time PCR with meltingcurve analysis for detecting the microsporidian parasites Nosema apis and Nosema ceranae in Apis mellifera
The Canadian Entomologist 142 (3), 271-283
Abstract: Honey bees, Apis mellifera (L.) (Hymenoptera: Apidae), are parasitized by the microsporidians Nosema apis (Zander) and Nosema ceranae (Fries). Molecular techniques are commonly used to differentiate between these parasites because light microscopy is inadequate. Our objectives were to (i) adapt the previously published duplex polymerase chain reaction (PCR) targeting the 16S rRNA gene of N. apis (321APIS-FOR, 321APIS-REV) and N. ceranae (218MITOC-FOR, 218MITOC-REV) using qualitative real-time PCR assay with SYBR® Green I dye (R-T PCR) and DNA melting-curve analysis, and (ii) determine whether the two Nosema species can be detected simultaneously in honey bees. Total spore counts and purified total genomic DNA were obtained from 37 bee samples (19 individual workers and 18 pooled samples of 15 workers) collected in Nova Scotia, Prince Edward Island, and Newfoundland, Canada. Overall, the prevalence of Nosema species was 86.5% (32/37 samples of bee DNA), based on conventional PCR and the optimized R-T PCR assay. The melting-curve analysis showed three groups of curve profiles that could determine the prevalence of N. apis, N ceranae, and co-infection (21.9%, 56.2%, and 21.9%, respectively). The duplex R-T PCR assay was efficient, specific, and more sensitive than duplex conventional PCR because co-infection was identified in 5.4% (n = 2) more samples. Sequencing of R-T PCR products confirmed the results of the melting-curve analysis. Duplex R-T PCR with melting-curve analysis is a sensitive and rapid method of detecting N. apis, N. ceranae, and co-infection in honey bees.
(The abstract is excluded from the Creative Commons licence and has been copied with permission by the publisher.)
Link to article at publishers website
Database assignments for author(s): Kenna E. MacKenzie, Dave Shutler

Research topic(s) for pests/diseases/weeds:
identification/taxonomy
surveys/sampling/distribution

---

Pest and/or beneficial records:

Beneficial	Pest/Disease/Weed	Crop/Product	Country	Quarant.
	Vairimorpha apis		Canada (east)
	Vairimorpha ceranae		Canada (east)