Back to basics with VirDisc: integrating High Throughput Sequencing in the framework of plant virus diagnostics

In regulatory plant health, diagnostic outcomes form the basis of phytosanitary decision making, highlighting the importance of accurate and reliable detection and identification of (regulated) pests. Because viruses and viroids cannot be cultured or directly observed, their detection and identification rely on indirect evidence obtained by diagnostic tests. The increasing use of high throughput sequencing has had a major impact on plant virus diagnostics by enabling broad detection of known and unknown viruses and overcoming several limitations associated with conventional diagnostic methods. Despite the increased use of HTS in plant virology, true standardized test descriptions with well-described performance characteristics remained limited so far. Here we present VirDisc, a validated Illumina sequencing-based test for the detection and identification of viruses and viroids in plant material. The test includes seven steps, from sample preparation to reporting, and was validated according to EPPO PM 7/98(6). In addition, we provide guidance for the interpretation of VirDisc results in a plant health context by integrating contextual information such as plant species, sample origin and symptomatology, to enable well-supported conclusions. To substantiate the added value of HTS for plant virus diagnostics, diagnostic cases are presented comparing conventional diagnostic workflows with those including VirDisc results. Furthermore, the broad application of VirDisc is demonstrated by showing the results of tests performed over a decade of use under ISO/IEC 17025 accreditation. It is illustrated how known concepts such as standard operating procedures, test scope, validation and integration of a test in a diagnostic workflow can be fulfilled, thereby placing HTS into the framework of regulatory plant health diagnostics.