ABSTRACT Fusarium species cause yield losses in wheat production through fusarium head blight (FHB) and the associated contamination of regulated mycotoxins, such as trichothecenes and zearalenone. Despite the widespread use of PCR‐based molecular approaches for Fusarium detection, quantification, and chemotyping, most primers were developed prior to both modern phylogenetic reclassification and the availability of high‐quality genome assemblies, leaving their specificity and robustness largely untested. Existing PCR‐ and qPCR‐based assays for Fusarium detection in wheat were reviewed and re‐evaluated in silico using a curated genome panel. Of 53 species‐specific primer pairs, 14 (26.4%) achieved high‐specificity grades (A–B), whereas 25 (47.2%) were lower performing (D–E), mainly due to cross‐reactivity or inconsistent target amplification. Chemotype assays targeting TRI and ZEN genes showed stronger agreement with reported chemotypes, especially for informative TRI loci such as Tri3, Tri7, and Tri12. To support improved qPCR assay design and reporting, we propose FusaMIQE, a Fusarium‐adapted framework based on MIQE 2.0 guidelines, tailored to the specific challenges of Fusarium diagnostics in wheat. Together, this manuscript provides the first in silico assessment of PCR/qPCR primers as diagnostic tools for FHB pathogens and associated recommendations for good practice (FusaMIQE) of Fusarium diagnostics in wheat.
Kavitha Vijeandran, Alexey Larionov, C. Cervini et al.· Comprehensive Reviews in Foo...· 0 citations
Verticillium stripe, caused by Verticillium longisporum, is an emerging threat to Brassica napus production in North America, with increasing field incidence and yield loss reported in affected regions. Although visual ratings are useful for rapid field assessment, resistance breeding is constrained by the lack of scalable, objective phenotyping methods that can quantify pathogen colonization across large genotype panels under field conditions. Here, we adapted and validated an established OLG70/OLG71-based SYBR Green qPCR assay within a high-throughput workflow combining freeze-dried hypocotyl sampling and magnetic bead-based DNA extraction to quantify V. longisporum DNA in field-grown B. napus. A panel of 215 B. napus genotypes was evaluated at two naturally infested field sites in Manitoba, Canada. The assay produced robust standard curves across plates, with R² values >0.99, amplification efficiencies of 95-105%, and low intra-plate variability, supporting reliable quantification over a broad dynamic range. Log₁₀-transformed fungal DNA quantities showed strong cross-environment consistency, with genotype rankings largely conserved between sites (Spearman ρ = 0.82). At the Glenlea site, qPCR-derived fungal load was strongly associated with visual Verticillium stripe severity (Spearman ρ = 0.68; R² ≈ 0.44; Kruskal-Wallis P = 1.59 × 10⁻¹⁷), indicating that hypocotyl colonization measured by qPCR captures biologically meaningful variation in disease expression. This scalable qPCR-based platform provides a repeatable, high-resolution phenotype for large-scale germplasm screening and future genetic analyses aimed at improving Verticillium stripe resistance in canola.
M. S. Youssef, S. Walkowiak, C. Obermeier et al.· Plant Disease· 0 citations
This new durum reference genome, enhanced with advanced annotation and an expression atlas linked to QTLome data, is the most comprehensive tool available for durum wheat genomics.
E. Mazzucotelli, C. Forestan, Gina Zastrow-Hayes et al.· Plant Biotechnology Journal· 0 citations
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