Open AccessDevelopment and Application of a LAMP Assay for the Detection of the Latent Apple Tree Pathogen Valsa mali
Author(s) -
Liangsheng Xu,
Yibo Wang,
Shan Zhu,
Jianyu Li,
Yali Chang,
Lili Huang
Publication year - 2021
Publication title -
plant disease
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.663
H-Index - 108
eISSN - 1943-7692
pISSN - 0191-2917
DOI - 10.1094/pdis-07-20-1449-re
Subject(s) - loop mediated isothermal amplification , biology , canker , horticulture , spore , botany , dna , genetics
Valsa mali, the causal agent of apple Valsa canker, produces cankers, resulting in the death of infected tissues and eventually the entire tree. Because of the long latent period of the disease, it is necessary to develop a rapid, sensitive, and reliable field-based assay to effectively diagnose apple Valsa canker when the plant is still symptomless. Loop-mediated isothermal amplification (LAMP) is a novel detection method that synthesizes a large amount of DNA and produces the visible byproduct (magnesium pyrophosphate) without conventional thermal cycling. Six LAMP primers were designed to target a species-specific region of the elongation factor-1α sequence, which can be completed at 61°C in 60 min. A positive result is indicated by color change after the intercalating dye SYBR Green I is added. The specificity of the LAMP was validated with DNA from 45 representative isolates of V. mali and closely related species V. malicola, V. leucostoma, and V. sordida. The sensitivity of the LAMP was determined to be 1 ng of DNA or as few as 10 spores. Because the assay does not require expensive equipment or specialized techniques, the LAMP-based diagnostic method can be applied under field conditions to more precisely and efficiently access disease pressure in apple orchards.