POSTERS: Pathogen detection, quantification and diagnosis
Robust and highly reliable loop-mediated isothermal amplification (LAMP) assay for specific and sensitive detection of Dickeya solani
Mohammad Arif - University of Hawaii at Manoa. Michael Stulberg- USDA-APHIS-PPQ-S&T-CPHST, Gamze Boluk- University of Hawaii at Manoa, Shefali Dobhal- University of Hawaii at Manoa, Mark Nakhla- USDA-APHIS-PPQ S&T CPHST, John Rascoe- USDA-APHIS-PPQ-S&T-CPHST
Dickeya species cause serious bacterial soft rot diseases of potato, an economically important crop worldwide. D. solani is the most aggressive species within soft rot causing bacterial pathogens, but fortunately is not yet reported in the United States. Symptoms produced by soft rot causing bacterial species are nearly indistinguishable. A rapid, field-deployable tool is required to monitor potato farms and to inspect potato tubers at ports-of entry for D. solani. Comparative genomics approach was used to identify signature target regions within the D. solani genome to develop a robust and field-deployable loop-mediated isothermal amplification (LAMP) assay. The specificity of the developed assay was tested with members of inclusivity (D. solani strains from distinct geographical locations) and exclusivity (strains from near-neighbor species and genus including all species from Dickeya and Pectobacterium) panels. No false-positives or -negatives were detected. A detection limit of 100 fg was obtained using DNA from pure cultures. However, a detection limit of 1000 fg was observed when target DNA was mixed with host genomic DNA. The developed LAMP assay has numerous applications from routine diagnostics at point-of-care, biosecurity, surveillance to disease management.