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Open AccessArticle

Identification of Spruce Budworm Natural Enemies Using a qPCR-Based Molecular Sorting Approach

1
Laurentian Forestry Centre, Natural Resources Canada, Quebec City, QC G1V4C7 Canada
2
Great Lakes Forestry Centre, Natural Resources Canada, Sault Ste. Marie, ON P6A2E5, Canada
3
Department of Integrative Biology, University of Guelph, Guelph, ON N1G2W1, Canada
4
Atlantic Forestry Centre, Natural Resources Canada, Fredericton, NB E3B5P7, Canada
5
Institut de Biologie Intégrative et des Systèmes, Université Laval, Quebec City, QC G1V0A6, Canada
*
Author to whom correspondence should be addressed.
These authors contributed equally to the work.
Current address: Centre for Ecological and Evolutionary Synthesis, Department of Biosciences, University of Oslo, 0316 Oslo, Norway
Forests 2020, 11(6), 621; https://doi.org/10.3390/f11060621
Received: 29 April 2020 / Revised: 22 May 2020 / Accepted: 26 May 2020 / Published: 1 June 2020
(This article belongs to the Special Issue Biological and Bio-Based Management of Forest Pests and Pathogens)
Annual monitoring of mortality agents in the course of a spruce budworm (Choristoneura fumiferana (Clemens) (Lepidoptera: Tortricidae)) population cycle is essential to understanding the factors governing the rise and collapse of outbreaks. To date, assessments of causes of budworm mortality have relied on laboratory rearing of field-collected larvae, followed by visual identification of emerging parasitoids and/or microscopic analysis of pathogens in larval carcasses. Although this approach has provided vital information on the abundance and identity of mortality agents, the procedure is labor-intensive and has limits in terms of accuracy. To overcome these shortcomings, we developed a molecular identification tool that makes use of real-time quantitative PCR (qPCR) and TaqMan® technologies. The tool relies on taxon-specific molecular variants (single nucleotide polymorphism [SNP] markers) found in mitochondrial (COI) and nuclear (28S rDNA) genes, for parasitoids, and in the nuclear SSU rDNA gene for microsporidian pathogens; these are then used as molecular signatures targeted by qPCR primers and TaqMan probes. Thus, the design of several sets of primers and probes deployed in multiplex format enables the identification of natural enemies via a molecular sorting process, bypassing barcode sequencing. Crude budworm DNA extracts are processed through a first module that detects dipteran and hymenopteran parasitoids, and microsporidian infections. Positive samples are then processed for species determination using three additional modules, enabling the identification of 20 common natural enemies of the spruce budworm. The tool has been fully validated using DNA samples from all comprised taxa, and both its sensitivity and accuracy compared favorably with the rearing-based method in an analysis of field-collected budworms. Using this tool, sample processing can be completed within two days, does not require larval rearing, provides accurate species identification, and can be conducted by technical staff without extensive molecular biology or insect taxonomy training. View Full-Text
Keywords: spruce budworm; natural enemies; molecular identification; qPCR; TaqMan® spruce budworm; natural enemies; molecular identification; qPCR; TaqMan®
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MDPI and ACS Style

Nisole, A.; Stewart, D.; Kyei-Poku, G.; Nadeau, M.; Trudeau, S.; Huron, P.; Djoumad, A.; Kamenova, S.; Smith, M.A.; Eveleigh, E.; Johns, R.C.; Martel, V.; Cusson, M. Identification of Spruce Budworm Natural Enemies Using a qPCR-Based Molecular Sorting Approach. Forests 2020, 11, 621.

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