Poster: Diseases of Plants: Disease Detection & Diagnosis
Rapid diagnostic assay development for detecting patch diseases on turf
B. KARAKKAT (1), M. Franchett (1), M. Olson (1), C. Mullenberg (1), P. Koch; (1) Department of Plant Pathology, University of Madison, WI, USA, (2) Department of Plant Pathology and Molecular and Environmental Toxicology Center, University of Wisconsin, M
Necrotic Ring Spot, Summer Patch, and Take-all Patch are turf root diseases caused by soil-borne plant pathogenic fungi. The infected turf shows aboveground symptoms that are broadly categorized as ‘patch’ diseases that ruin the aesthetic value of cool-season turfgrass on lawns/fields in North America from late spring to early summer. These fungal pathogens are notoriously difficult to isolate from roots. Therefore, we decided to develop a rapid detection assay that can be performed on suspected root samples. We are testing two assays: Loop mediated isothermal amplification (LAMP) and Recombinase polymerase amplification (RPA). Both LAMP and RPA do not require pure genomic DNA but only a solution of root sample. LAMP and RPA rely on DNA strand displacement activity for its rapid amplification and can be completed at 65oC in an hour and 39oC in 20 minutes incubation, respectively. Initial LAMP screening gave false positives repeatedly in almost all assays indicating that very pure conditions are needed, which is not ideal in a diagnostic laboratory. Hence, we have started to work on RPA methodology available as Agdia’s discovery kit to eliminate any false positives. Once we achieve a consistent and specific amplification with these assays, we will proceed to screen samples that will be arriving in late Spring 2016. Assay development, primer-probe design and samples surveyed will be reported in this study.