Establishment and application of a SYBR Green-based absolute real-time quantitative PCR assay for chilli yellow ringspot virus

Abstract

Chilli yellow ringspot virus (CYRSV), a new emerging pathogen, was first detected in Yunnan Province, China. This virus is a novel agent that causes severe disease in vegetable crops and has a tendency to spread widely. CYRSV infection manifests as yellowing, ringspot and necrosis of the leaves and fruits of tomato plants. Consequently, the development of a sensitive, rapid, and accurate detection system for virus identification and epidemiological surveillance is imperative. In this study, we developed a quantitative PCR (qPCR) assay using SYBR Green for the detection of CYRSV accumulation. To achieve sensitive and rapid detection of CYRSV in tomato, specific primers were designed based on the coding sequence of the CYRSV nucleocapsid protein (N) gene. These primers demonstrated excellent specificity, sensitivity, and reproducibility. After inoculation with CYRSV in a greenhouse, the amount of virus accumulated in tomato leaves was detected. A total of 62 tomato fruit samples were collected from Yuanmou city, Yunnan Province, China, between May and December 2024. Of these, 75.81 % (47/62) tested positive for CYRSV via this qPCR method. These results indicated that the established SYBR Green-based qPCR assay could accurately detect CYRSV in tomato leaves and fruits. In conclusion, our study provides a valuable diagnostic tool for CYRSV detection and epidemiological investigation.