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Establishment of a novel virus-induced virulence effector assay for the identification of virulence effectors of plant pathogens using a PVX-based expression vector
Authors:Jinxia Shi  Yuanhong Zhu  Ming Li  Yuqing Ma  Huarong Liu  Peng Zhang  Di Fang  Yushuang Guo  Ping Xu  Yongli Qiao
Institution:1. Shanghai Key Laboratory of Plant Molecular Sciences, College of Life Sciences, Shanghai Normal University, Shanghai, China;2. Shanghai Key Laboratory of Plant Molecular Sciences, College of Life Sciences, Shanghai Normal University, Shanghai, China

College of Agriculture, Yangtze University, Jingzhou, China;3. Laboratory of Molecular Genetics, China National Tobacco Corporation, Guizhou Institute of Tobacco Science, Guiyang, China

Abstract:Plant pathogens deliver virulence effectors into plant cells to modulate plant immunity and facilitate infection. Although species-specific virulence effector screening approaches have been developed for several pathogens, these assays do not apply to pathogens that cannot be cultured and/or transformed outside of their hosts. Here, we established a rapid and parallel screening assay, called the virus-induced virulence effector (VIVE) assay, to identify putative effectors in various plant pathogens, including unculturable pathogens, using a virus-based expression vector. The VIVE assay uses the potato virus X (PVX) vector to transiently express candidate effector genes of various bacterial and fungal pathogens into Nicotiana benthamiana leaves. Using the VIVE assay, we successfully identified Avh148 as a potential virulence effector of Phytophthora sojae. Plants infected with PVX carrying Avh148 showed strong viral symptoms and high-level Avh148 and viral RNA accumulation. Analysis of P. sojae Avh148 deletion mutants and soybean hairy roots overexpressing Avh148 revealed that Avh148 is required for full pathogen virulence. In addition, the VIVE assay was optimized in N. benthamiana plants at different developmental stages across a range of Agrobacterium cell densities. Overall, we identified six novel virulence effectors from seven pathogens, thus demonstrating the broad effectiveness of the VIVE assay in plant pathology research.
Keywords:Nicotiana benthamiana  plant pathogens  PVX  screening assay  virulence effector
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