Abstract
Lily symptomless virus (LSV), Lily mottle virus (LMoV), Cucumber mosaic virus (CMV), Shallot yellow stripe virus (SYSV), and Plantago asiatica mosaic virus (PlAMV) are five of the economically important viruses infecting lilies (Lilium spp.) worldwide. In order to prevent the occurrence and spread of these viruses, it is necessary to develop a rapid, effective, and sensitive detection method for the simultaneous detection and specific quantification of these viruses. In this study, specific primers and probes for multiplex TaqMan real-time PCR assays designed from conserved regions of the coat protein sequence of each virus were used for the simultaneous detection of these viruses in lilies (Lilium spp.). The optimal concentration of primers and probes and reaction annealing temperature were 20 µM and 55.9 °C, respectively. The detection limits of the assay were 1.33 × 102, 1.27 × 101, 1.28 × 101, 2.33 × 102, and 2.01 × 102 copies·μL−1 for LSV, LMoV, CMV, SYSV, and PlAMV, respectively. Specificity was determined using seven viral pathogens of lilies. Variability tests of intra- and inter-assays showed high reproducibility with coefficients of variation <2%. The multiplex TaqMan real-time PCR assay was used to detect these viruses from lily samples in China. In brief, our developed assay showed high specificity, sensitivity, and reproducibility for the simultaneous detection and differentiation of five lily-infecting viruses and can be used for certification and quarantine programs.
Highlights
Academic Editor: Salvatore DavinoLilies (Lilium spp.) are among the most important ornamental plants worldwide and are widely grown for their nutritional and medicinal value in Eastern Asia [1,2,3].Lily cultivars are usually propagated vegetatively, resulting in virus transmission from generation to generation
Fresh leaves from lily samples grown in greenhouses at the Chinese Academy of Agricultural Sciences (Beijing, China), which were confirmed to be infected with Lily symptomless virus (LSV), Lily mottle virus (LMoV), Cucumber mosaic virus (CMV), Shallot yellow stripe virus (SYSV), and Plantago asiatica mosaic virus (PlAMV) by RT-PCR and sequencing, were used to establish and optimize the multiplex TaqMan real-time PCR assay
The results showed the optimum annealing temperatures for LSV, LMoV, and PlAMV
Summary
Academic Editor: Salvatore DavinoLilies (Lilium spp.) are among the most important ornamental plants worldwide and are widely grown for their nutritional and medicinal value in Eastern Asia [1,2,3].Lily cultivars are usually propagated vegetatively, resulting in virus transmission from generation to generation. Symptomless virus (LSV; genus Carlavirus, family Flexiviridae), Cucumber mosaic virus (CMV; genus Cucumovirus, family Bromoviridae), Lily mottle virus (LMoV; genus Potyvirus, family Potyviridae), Shallot yellow stripe virus (SYSV; genus Potyvirus, family Potyviridae), and Plantago asiatica mosaic virus (PlAMV; genus Potexvirus, family Alphaflexiviridae) are five economically important viruses [4,5,6,7,8,9]. LSV-infected plants show mild, pale vein clearing and mottle on leaves, which generally turn yellow fairly quickly [6,7]. LMoV can cause flower-color breaking, leaf mottle, leaf mosaic, chlorotic and yellow streaking, vein clearing, leaf curling, and narrowing [6,7]. CMV-infected lily leaves show chlorotic or yellow spotting, inter-veinal striping or vein-clearing and sometimes malformations [6,7]. Mixed viral infections in lily plants can cause much more severe symptoms, including dwarfism [7]
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