Distinguishing integrated sub-viral fragments from infectious Rubus yellow net virus in Scottish commercial red raspberry cultivars

Abstract

Short read sequencing of two field-grown plants of red raspberry cv Glen Dee identified sequences derived from Rubus yellow net virus (RYNV). Surprisingly, PCR primers designed to target these sequences amplified RYNV-specific DNA fragments from some high health nuclear stock raspberry plants, previously tested as free from RYNV infection. The complete sequence of a Scottish isolate of RYNV (RYNV LG) was determined and a panel of primers covering the entire virus genome was designed to demonstrate that nuclear stock plants of cultivars Glen Dee, Glen Ericht, Glen Fyne and Glen Moy all contain partial fragments of the RYNV genome as integrated elements but do not contain the entire RYNV genome. The complete, circular genome of RYNV could be detected in RYNV LG-infected plants by rolling circle amplification but was not detected in Glen Dee plants. Grafting experiments showed that infectious RYNV could be transferred from RYNV LG plants but not from Glen Dee or Glen Moy plants, further demonstrating that these cultivars contain only partial, integrated RYNV elements. Grafting of RYNV LG to the certification-approved indexing species R. occidentalis (black raspberry) and R. macraei, showed that RYNV LG induced recognisable symptoms in both plants but that R. macraei was more susceptible and produced more recognizable symptoms. Effective testing for RYNV in high health certified raspberry plants can be achieved by combining graft testing with multifragment PCR.