Sensitive and Specific qPCR and Nested RT-PCR Assays for the Detection of Tobacco Streak Virus in Soybean

Abstract

Tobacco streak virus (TSV) is a reemerging and understudied pathogen of soybean (Glycine max). Management of TSV is challenging due to the multiple modes of transmission, widespread susceptibility of commercial soybean, and lack of reliable diagnostic tests for the virus. Soybean plants with TSV-like, virus-like, or no symptoms were collected from commercial and research fields in seven counties in Wisconsin. Two sensitive assays were developed for the detection of TSV: a fluorescent dye-based quantitative reverse-transcription PCR (qPCR) assay and a nested reverse-transcription PCR (nRT-PCR). TSV was detected in 47 and 91% of symptomatic samples using the qPCR and nRT-PCR assays, respectively, suggesting that the nRT-PCR assay has higher sensitivity for detecting TSV. The qPCR assay’s limit of detection was determined at 10 fg and the assay was used to estimate the viral load in TSV-symptomatic samples. The titer of TSV in these samples was determined by absolute quantification and ranged from 15 fg to 0.796 ng. The two assays reported here provide diagnostic tools for the rapid and accurate detection of TSV that can aid in monitoring outbreaks, assessing management strategies, or screening soybean cultivars or accessions for resistance to the virus. [Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .