Detection and enumeration of Erwinia carotovora subsp. atroseptica using spiral plating and immunofluorescence colony staining

Abstract

Immunofluorescence colony staining (IFC) and a new technique using spiral plating combined with IFC were evaluated for the soft-rot pathogen Erwinia carotovora subsp. atroseptica in witloof chicory. Target bacteria could be detected in platings at various dilutions of plant washings. Brilliance of the stained colonies of E. carotovora subsp. atroseptica was high. Spiral plating, used for both the plating of the bacteria and for the delivery of the conjugated antiserum, had a positive effect on the reduction of the background compared with the staining of the bacteria. The combination of spiral plating and IFC proved to be a functional tool for the quantification of target and nontarget bacteria and the isolation of target bacteria as pure culture from IFC-positive colonies. The method uses less conjugated antiserum than traditional IFC and produces results with very small variation within replications. The recovery of the bacteria in both pure culture and plant washing is significantly higher than the recovery using crystal violet pectate medium.Key words: soft rot, witloof chicory, detection, Erwinia carotovora subsp. atroseptica.