Poster Session: Diseases of Plants - Disease Detection and Diagnosis
418-P
LAMP and multiplex endpoint PCR-based diagnostics for detection and discrimination among Pseudomonas syringae pv. actinidiae strains.
G. Y. BUSOT (1), M. Arif (1), J. Rascoe (2), M. K. Nakhla (2), J. P. Stack (1)
(1) Department of Plant Pathology, Kansas State University, Manhattan, KS, U.S.A.; (2) USDA-APHIS-PPQ CPHST Beltsville Lab, USA, Beltsville, MD, U.S.A.
Strain-specific detection of the kiwifruit pathogen, P. syringae pv. actinidiae (Psa), is essential to support plant biosecurity decisions regarding trade and production. To detect and discriminate among Psa strains (including the 2008 global outbreak strain, Psa-V), a multiplex assay using conventional endpoint PCR and a Loop Mediated Isothermal Amplification (LAMP) assay were developed for laboratory and field use, respectively. The primer sets were screened against isolates of Psa and closely related P. syringae pathovars. The expected amplicons were generated and no cross-reaction was observed with any primer set. hopZ3 and hopS2 amplicons were obtained in all Psa strains tested. HopS2 amplicon was also observed in PsD (ex Psa-LV; formerly considered Psa low virulent strain). hopO1 and hopZ5 amplicons were specific to PsD and Psa-V strains, respectively. The LAMP protocol targeting three effector genes (hopZ3, hopZ5 and hrpW) was highly specific, discriminating between Psa virulent strains (hopZ5 and hrpW primers) and other Psa strains (hopZ3 primers). The LAMP assay is a valuable tool for in-field pathogen detection to support surveillance and at ports of entry to facilitate interception. Both assays can be used for the accurate detection and discrimination of Psa.