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ARS Home » Southeast Area » Fort Pierce, Florida » U.S. Horticultural Research Laboratory » Subtropical Plant Pathology Research » Research » Publications at this Location » Publication #316591
Title: A loop-mediated isothermal amplification assay and sample preparation procedure for sensitive detection of Xanthomonas fragariae in strawberry

Author
item WANG, HEHE - University Of Florida
item Turechek, William

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/29/2015
Publication Date: 1/14/2016
Publication URL: http://handle.nal.usda.gov/10113/62980
Citation: Wang, H., Turechek, W. 2016. A loop-mediated isothermal amplification assay and sample preparation procedure for sensitive detection of Xanthomonas fragariae in strawberry. PLoS ONE. 11(1):e0147122. https://doi.org/10.1371/journal.pone.0147122.
DOI: https://doi.org/10.1371/journal.pone.0147122

Interpretive Summary: Xanthomonas fragariae causes angular leaf spot disease in strawberry. The aim of this study was to develop a loop-mediated isothermal amplification (LAMP) assay with a bacterial enrichment procedure as an efficient method for detection of latent or asymptomatic infections of X. fragariae. The detection limit of the LAMP assay was approximately 2000 CFU/mL for pure bacteria culture and 300 CFU/mL for strawberry leaf and petiole samples processed through enrichment assay. LAMP provided a 2-3 fold lower detection limit than the standard qPCR assay but was much faster, less expensive, and more user-friendly. Therefore, LAMP with bacterial enrichment assay will serve as a rapid, sensitive and cost-effective tool for detecting asymptomatic infections of X. fragariae in strawberry nursery stock and contribute to improved disease management in the industry.

Technical Abstract: Xanthomonas fragariae is a bacterium that causes angular leaf spot of strawberry. Asymptomatic infections are common and contribute to the difficulties in disease management. The aim of this study was to develop a loop-mediated isothermal amplification (LAMP) assay with a bacterial enrichment procedure as a more efficient method for detection of asymptomatic infections of X. fragariae. The enrichment procedure calls for an overnight incubation of strawberry tissues with phosphate-buffered saline (PBS), followed by a quick sample concentration and a boiling step to extract DNA for amplification. The bacterial enrichment step allows sampling of a larger amount of plant tissue, hence increasing the detection rate in real-life samples. The detection limit of the LAMP assay was approximately 2000 CFU/mL for pure bacteria culture and 300 CFU/mL for strawberry leaf and petiole samples processed through enrichment assay. LAMP provided a 2-3 fold lower detection limit than the standard qPCR assay but was much faster, less expensive, and more user-friendly. Therefore, LAMP with bacterial enrichment assay will serve as a rapid, sensitive and cost-effective tool for detecting asymptomatic infections of X. fragariae in strawberry nursery stock and contribute to improved disease management in the industry.