Multiplex RT‐PCR to simultaneously detect three viruses that infect peach

Abstract

Significance and Impact of the Study: In this study, we developed a multiplex reverse transcriptase PCR (mRT-PCR) assay for simultaneously detecting three viruses that infect peach: peach-associated luteovirus (PaLV), peach virus D (PeVD) and nectarine stem-pitting-associated virus (NSPaV). This assay is simple, easy to perform, reliable and cost-effective, and can thus be applied for large-scale surveys of PaLV, PeVD and NSPaV. AbstractPeach is a major crop in China, and like any other stone fruit, virus and viruslike diseases can reduce the yield and quality of the fruit. Herein, we developed a multiplex RT-PCR (mRT-PCR) assay for simultaneously detecting three viruses known to infect peach: peach-associated luteovirus (PaLV), peach virus D (PeVD) and nectarine stem-pitting-associated virus (NSPaV). Plant nad5 mRNA was used as the internal control. Field samples that were co-infected with PaLV, PeVD and NSPaV were used; we identified three primer pairs to be the most specific for detecting these viruses, followed by determining the ideal concentration of each primer pair and optimizing the annealing temperature for mRT-PCR. We also assessed the detection limit using serial dilutions of RNA and cDNA. The newly developed mRT-PCR assay could simultaneously detect PaLV, PeVD and NSPaV. To validate the reliability of mRT-PCR for virus detection, mRT-PCR was used to detect viruses in the leaves of 21 peach plants collected in Liaoning Province, China. The obtained results revealed the presence of single and co-infections. To conclude, the mRT-PCR assay developed herein is sensitive, reliable and economical, and we believe that it can thus be used for large-scale surveys of PaLV, PeVD and NSPaV.Letters in Applied Microbiology ISSN 0266-8254

Is There a “Biological Desert” With the Discovery of New Plant Viruses? A Retrospective Analysis for New Fruit Tree Viruses

“…Realtime RT-PCR assays were developed for two viruses (ASGV and AGCaV) infecting pome fruit (Beaver-Kanuya et al, 2019). Multiplex RT-PCR was developed to simultaneously detect three new viruses (NSPaV, PaLV, and PeVD) that infect peach (Xu et al, 2019). A loop-mediated isothermal amplification assay was established for CCDaV .…”

Detection and elimination of shallot latent virus and onion yellow dwarf virus for potato onion (Allium cepa L. var. aggregatum Don)

“…RT‐PCR has been developed to detect OYDV in garlic and onions (Da Silva et al, 2019; Kumar et al, 2010). Multiplex RT‐PCR has been widely used for the detection of related viruses in plants, including garlic (Majumder & Baranwal, 2014; Nam et al, 2015), potato (Rashid et al, 2020; Wei et al, 2009), cassava (Aloyce et al, 2013), pear (Yao et al, 2014), peach (Xu et al, 2019) and papaya (Maurastoni et al, 2020). Shallot latent virus (SLV) and onion yellow dwarf virus (OYDV) are the most common viruses in potato onions.…”