Detection of stylet-borne and circulative potato viruses in aphids by duplex reverse transcription polymerase chain reaction.

Abstract

A reverse transcription polymerase chain reaction (RT-PCR) assay was designed to amplify stylet-borne potato virus Yo (PVYo) in aphids using primers located in the viral capsid gene. A 480 bp long product was detected in aphids exposed to PVYo-infected potato plants. Approximately 40% of Myzus persicae and 15% of Aphis nasturtii exposed briefly to PVYo-infected plants acquired the virus. This rate of acquisition by both species of aphids was typical of our earlier observation of the virus transmission tests. No significant difference in virus detection was observed whether the aphids were tested immediately after exposure to virus sources or stored for up to 45 days in ethanol at room temperature. The addition of a second pair to primers located in the capsid gene of circulative potato leafroll virus (PLRV) allowed simultaneous amplification of two viruses (duplex RT-PCR) in single aphids. Acquisition of PVYo by the aphids already viruliferous with PLRV was significantly reduced, compared to aphids not carrying PLRV. Duplex RT-PCR for PVYo and PLRV could be applied to analyze aphids collected from the field to ascertain the relative presence of both viruses in a single test.