Submitted to: Phytopathology
Publication Type: Abstract Only
Publication Acceptance Date: 3/29/2005
Publication Date: 7/30/2005
Citation: Chen, J., Groves, R.L., Civerolo, E.L. 2005. High throughput detection of Xylella fastidiosa from almond and grape tissues. Phytopathology. 95:S19.
Technical Abstract: Xylella fastidiosa is an important bacterial pathogen of almond (almond leaf scorch disease) and grape (Pierce’s disease). Pathogen detection is a critical step for epidemiological studies of the diseases. With the recent improvement in technology and availability of whole genome sequences, PCR-based techniques are increasingly promising for bacterial pathogen detection. The bottleneck, however, lies in the preparation of inhibitor-free template DNA from plant tissue. We have developed two simple sample preparation procedures for PCR amplification of X. fastidiosa DNA from infected almond and grape petioles. 1) Sap from fresh petioles was expressed mechanically and mixed with PW broth. The sap-PW broth mix was used immediately for PCR, or stored frozen for later PCR analyses; and 2) Plant petioles were freeze-dried and stored at room temperature, or used for PCR immediately. For PCR, the dried petioles were pulverized and powder suspension was used. Appropriate dilutions of sap-PW broth mix and powder suspension further minimized the effect of possible DNA polymerase inhibition. The two PCR methods were comparable to, but more rapid and much less labor intensive, than pathogen isolation. By coupling with real-time PCR, the two sample preparation procedures demonstrated their high throughput capacity for in planta detection of X. fastdiosa.