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Demonstrates effective detection of Cronartium ribicola in Pinus armandii, suggesting a new method for early monitoring.
White pine blister rust disease (WPBR), caused by Cronartium ribicola, ranks among the most destructive pathogens of five-needle pines. We developed a hydroxynaphthol blue (HNB)-based Loop-mediated isothermal amplification (LAMP) assay enabling rapid, visual detection of C. ribicola directly following DNA extraction. LAMP primers targeting the internal transcribed spacer (ITS) region were designed and validated through in silico comparison with related Cronartium species and in vitro testing against sympatric forest fungi. The optimized 25 μL reaction contained 8.0 mM Mg2+, 1.0 mM dNTPs, and an inner-to-outer primer ratio of 8:1, with amplification conducted at 62 °C for 40 min. Positive amplification produced a distinctive color transition from purple to sky blue, enabling visual interpretation without instrumentation. Under the tested conditions, the assay achieved a detection limit of 460 ± 3.2 fg/μL genomic DNA—a 10-fold improvement over conventional PCR in concentration-based sensitivity. Assay applicability was evaluated using 211 field-collected Pinus armandii samples sourced from China. Detection efficiency varied significantly across tissue types. Symptomatic bark exhibited a substantially higher positive detection rate (68.97%, 95% CI: 49.2–84.7%) compared to needles from symptomatic trees (18.75%, 95% CI: 4.1–45.7%). Among asymptomatic samples, 3.75% of bark samples tested positive for C. ribicola DNA, whereas all needle samples were negative. Geographically, positive detections clustered at several discrete sampling sites in southwestern China, predominantly at elevated elevations. The established LAMP-HNB assay provides a rapid, visually interpretable diagnostic tool for early detection and quarantine monitoring of WPBR following DNA extraction. Beyond its practical utility, this assay establishes valuable baseline data for targeted disease surveillance in the context of evolving climate conditions.
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Zhang et al. (2026) studied this question.
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