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 developed a robust and specific semi-nested reverse transcription—polymerase chain 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 ELISA-based methods and allows detection directly from field trees. We have combined the assay with a simple and rapid tissue extraction protocol to make it amenable to large-scale screening. This method will improve the rapidity and reliability of CTLV screening of trees in the Australian Citrus Budwood Scheme. Nucleotide sequence analysis of the amplified CTLV fragments shows 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.