Journal of Phytopathology (2019) 167, 174-184

Ayaka Hieno, Mingzhu Li, Auliana Afandi, Kayoko Otsubo, Haruhisa Suga and Koji Kageyama (2019)
Rapid detection of Phytophthora nicotianae by simple DNA extraction and real-time loop-mediated isothermal amplification assay
Journal of Phytopathology 167 (3), 174-184
Abstract: Phytophthora nicotianae is a phytopathogenic oomycete with a wide host range and worldwide distribution. Rapid detection and diagnosis at the early stages of disease development are important for the effective control of P. nicotianae. In this study, we designed a simple and rapid loop-mediated isothermal amplification (LAMP)-based detection method for P. nicotianae. We tested three DNA extraction methods and selected the Kaneka Easy DNA Extraction Kit version 2, which is rapid and robust for LAMP-based detection. The designed primers were tested using mycelial DNA from 35 species (81 isolates) of Phytophthora, 12 species (12 isolates) of Pythium, one isolate of Phytopythium and one isolate each from seven other soil-borne pathogens. All of the 42 P. nicotianae isolates were detected by these primers, and no other isolates gave positive results. Three isolates were tested for the sensitivity of the reaction, and the lowest amounts of template DNA that could be detected were 10 fg for two isolates and 1 fg for the third. The target was detected within 25 min in all tested samples, including DNA extracted from both inoculated and naturally infected plants. In contrast, PCR assays with P. nicotianae-specific primers failed or showed weakened detection in several samples. Thus, we found that the rapid DNA extraction and LAMP assay methods developed in this study can be used to detect P. nicotianae with high sensitivity, specificity and stability.
(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): Koji Kageyama

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.
	Phytophthora nicotianae