Submitted to: Current Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: October 11, 2012
Publication Date: November 16, 2012
Citation: Guan, W., Shao, J.Y., Singh, R., Davis, R.E., Huang, Q. 2012. A TaqMan-based real time PCR assay for specific detection and quantification of Xylella fastidiosa strains causing bacterial leaf scorch in oleander. Current Microbiology. 92:108-112. Interpretive Summary: Xylella fastidiosa is a slow growing, xylem inhabiting, nutritionally fastidious and insect-transmitted bacterium. The bacterium is associated with bacterial leaf scorch and decline in many economically important landscape trees and shrubs including oak, elm, sycamore, maple, and oleander. Oleander leaf scorch caused by X. fastidiosa was first reported in 1996, and is becoming a problem in California, Texas and Louisiana where oleander is used as a popular landscape plant for residential hedges, highway dividers, beach plantings, gardens and parks. In this study, we describe the development of a fluorescent probe-based real-time polymerase chain reaction that is sensitive, specific and quantitative for strains of X. fastidiosa causing bacterial leaf scorch in oleander. As low as one genome of X. fastidiosa can be detected in bacterial cultures and in planta. Specific and sensitive detection and quantification of oleander strains of X. fastidiosa is useful for disease detection, epidemiological studies, disease management of oleander leaf scorch, and resistance screening for oleander shrubs. Our work will be of value primarily to plant pathologists, entomologists and clinicians interested in diseases caused by X. fastidiosa.
Technical Abstract: A TaqMan-based real-time PCR assay is developed for strains of X. fastidiosa causing oleander leaf scorch. The assay uses primers WG-OLS-F1 and WG-OLS-R1 and the fluorescent probe WG-OLS-P1, designed based on unique sequences present only in genomic sequence of oleander strain Ann1. The assay is specific, allowing detection of only oleander strains, not other strains of X. fastidiosa nor other plant-associated bacteria. The assay is also sensitive, with a detection limit of 3 fg/ml DNA of X. fastidiosa in vitro and in planta. The assay can also be applied to detect low numbers of X. fastidiosa in insect samples, or further developed into a multiplex real-time PCR assay to simultaneously detect and differentiate different strains of X. fastidiosa that may be carried by the same hosts or insect vectors. Specific and sensitive detection and quantification of oleander strains of X. fastidiosa is useful for disease detection, epidemiological studies, disease management of oleander leaf scorch, and resistance screening for oleander shrubs.