ARS | Publication request: Development of a real-time, quantitative PCR for detection of potato purple top phytoplasma in plants and beet leafhoppers

Submitted to: Phytopathology
Publication Type: Abstract Only
Publication Acceptance Date: June 1, 2006
Publication Date: June 1, 2007
Citation: Crosslin, J., Vandemark, G.J., Munyaneza, J.E. 2007. Development of a real-time, quantitative PCR for detection of potato purple top phytoplasma in plants and beet leafhoppers. Phytopathology. 97:S166. ARIS 204245

Technical Abstract: A quantitative, real-time “TaqMan®” polymerase chain reaction assay (qPCR) was developed which was capable of detecting and quantifying a group 16SrVI phytoplasma in DNA extracts prepared from infected tomatoes, potatoes, and beet leafhoppers (Circulifer tenellus Baker). Primers and probe were designed from the 16S-23S rRNA genes of the Columbia Basin potato purple top phytoplasma, which is closely related to the beet leafhopper transmitted virescence agent. The detection limit in phytoplasma-infected tomato DNA was approximately 50 pg. The detection limit of the assay with a plasmid clone of clover proliferation phytoplasma 16S-23S rDNA was approximately 500 ag. The concentration of phytoplasma varied considerably among potato plants showing symptoms of purple top. The pathogen was readily detected in extracts from single or groups of five beet leafhopper vectors. As with infected potatoes, the concentration of phytoplasma in individual leafhoppers was variable. The assay also detected aster yellows (group 16SrI) and pigeon pea witches’-broom (group 16SrIX) phytoplasmas. The qPCR was at least as sensitive as the commonly used and more labor-intensive nested PCR for detection of the pathogen. To our knowledge this is the first report of qPCR of phytoplasma in potatoes and beet leafhoppers, and only the second report of qPCR of a group 16SrVI phytoplasma.