LAUSR

Phytophthora infestans, the causal agent of late blight in potatoes and tomatoes, is the most important and ongoing pathogenic threat to agricultural production worldwide. Rapid and early identification of P. infestans is an essential prerequisite for countering the further spread of infection. In this study, a novel method for visual detection of P. infestans has been developed by integrating universal primer mediated asymmetric PCR with gold nanoparticle (AuNP)-based lateral flow biosensor. We employed asymmetric PCR to generate large amounts of single-stranded DNA (ssDNA) by amplifying a region of P. infestans-specific repetitive DNA sequence. The ssDNA products were then applied to the lateral flow biosensor to perform a visual detection using sandwich-type hybridization assays. In the presence of target DNA, sandwich-type hybridization reactions among the AuNP-probe, target DNA and capture probe were performed on the test line of the biosensor, and then a characteristic red band was produced for the accumulation of AuNPs. Quantitative analysis obtained by recording the optical intensity of the red band demonstrated that this biosensor could detect as little as 0.1 pg μL-1 genomic DNA. Furthermore, the specificity of the biosensor was confirmed by detecting three other Phytophthora species and two pathogenic fungi. We believe this method has potential application in early prediction of potato late blight disease and instigation of management actions to reduce the risk of epidemic development.