PCR detection ofclavibacter michiganensis subsp.sepedonicus- infected tuber samples in a plate capture assay

Abstract

The speed and sensitivity of PCR-based assays allow shorter turnaround times for the detection of pathogens for which culture and serological methods are difficult or unavailable. PCR was performed with primer sets Cms50 and Cms72, designed previously by Millset al. (1997) through subtractive hybridization to detectClavibacter michiganensis subspeciessepedonicus (Cms). In bacterial suspensions, fewer than three cells/10 ul reaction were detected after PCR amplicons were hybridized with specific DIG-labeled DNA probes in an enzyme-linked oligonucleosorbent assay (ELOSA). In naturally infected tuber samples representing three cultivars of potato, the diagnostic sensitivity of PCR/ ELOSA was 96%, while the specificity exceeded 99%. PCR/ELOSA detectedCms in infected tuber samples with equal sensitivity regardless of colony morphology, potato cultivar, or primer sets.