Detection of Citrus tatter leaf virus with reverse transcription-polymerase chain reaction (RT-PCR)

Citrus tatter leaf virus (CTLV) has the potential to cause major losses to the Australian citrus industry if an infected clone is propagated, because the predominant rootstocks are intolerant of CTLV infection. We have develo ped a robust and specific semi-nested reverse transcription-polymerase chai n reaction (RT-PCR) assay which detects CTLV in a range of citrus tissues. The sensitivity of the assay is at least 500 times greater than that of ELI SA-based methods and allows detection directly from field trees. We have co mbined the assay with a simple and rapid tissue extraction protocol to make it amenable to large-scale screening. This method will improve the rapidit y and reliability of CTLV screening of trees in the Australian Citrus Budwo od Scheme. Nucleotide sequence analysis of the amplified CTLV fragments sho ws near identity (99.8%) amongst Australian isolates and between Australian isolates and a Japanese isolate of apple stem grooving virus (98.1%), and a high level of identity (92.0%) to a Japanese isolate of CTLV.