The application of Next-Generation Sequencing for screening seeds for viruses and viroids

Next-Generation Sequencing (NGS) technologies are being increasingly used in plant pathology for the identification of novel pathogens. However, the application of this technology in routine screening work has yet to be fully explored. Preliminary work to validate NGS was carried out through testing serial dilutions of tomato seed samples infected with Potato spindle tuber viroid (PSTVd), Columnea latent viroid (CLVd) and Pepino mosaic virus (PepMV). Infected seeds were obtained via both natural infection (CLVd) and from artificial inoculations (PSTVd and PepMV). A control sample from a commercial seed lot was also used for the serial dilution of test samples. Testing showed CLVd could be detected down to a 100-fold dilution and PSTVd to a 10-fold dilution. The negative control seed lot contained PepMV, Tobacco mosaic virus, Cucumber mosaic virus, Tomato bushy stunt virus and Potato leafroll virus. The viroids from the test samples and the viruses from the control/dilution seed lot were detected simultaneously, the first time this has been demonstrated for botanical seeds. Work is ongoing to further validate this technology for seed screening.