High throughput PCR detection of Xylella fastidiosa directly from almond tissues

Xylella fastidiosa, the causal agent of almond leaf scorch disease (ALSD), is currently re-emerging as a serious concern in California. Efficient pathogen detection is critical for ALSD epidemiological studies, particularly when a large sample size is involved. We here report a PCR procedure to detect X. fastidiosa directly from infected almond tissue without the laborious DNA extraction. Plant samples were prepared by freeze-drying and pulverized. Appropriate dilutions of the pulverized freeze-dried tissue (PFT) were determined to minimize the effect of enzyme inhibitors from plant tissue and retain PCR detection of X. fastdiosa cells at a single digit number level. This PFT-PCR procedure was evaluated by comparing to the in vitro cultivation method using 102 symptomatic samples and resulted in a predictive value of 90.8%. PFT-PCR was further applied to monitor the seasonal occurrence of X. fastidiosa from four selected almond trees in two orchards in 2005. The results matched with those of the cultivation method at 92.3%. Considering the simplicity and reliability, we conclude that PFT-PCR is a valuable option for high throughput rapid detection of X. fastidiosa.