Rapid Detection of Pectolytic Erwinia sp. in Aglaonema sp.

Abstract

The purpose of this research was to develop a rapid microplate assay for detecting and presumptively identifying pathogenic pectolytic Erwinia sp. in ornamental propagative stock and to readily distinguish them from nonpathogenic bacteria associated with Aglaonema. The assay was developed by modifying an existing crystal violet-sodium polypectate (CVP) medium to visualize the depolymerization of pectate by addition of bromcresol purple (BCP), then acidifying with a dilute hydrochloric acid (HCl) solution. The assay was sufficiently sensitive to detect latent (symptomless) infections in small sections of leaf or stem tissue. Based on inoculum titration assays, 40 to 600 colony-forming units (CFU) of Erwinia chrysanthemi (Burkholder et al.) were required to produce symptoms in Aglaonema stems and leaves, respectively. The microplate assay was able to detect the pathogen at low levels (40 to 60 CFU) in tissue segments of ≈1 cm2. In tests of bacterial strains isolated from 211 samples from Aglaonema `Silver Queen', `Emerald Beauty', and `San Luis' grown in Hawaii, only the pectolytic and pathogenic Erwinia sp. reacted positively in the microplate assay. Other bacteria associated with Aglaonema, including Pseudomonas paucimobilis (Holmes), P. vesicularis (Galarneault and Leifson), and nonpectolytic Erwinia sp., were not detected by the assay. Pectolytic strains of Ralstonia (Pseudomonas) solanacearum (Smith) were differentiated from pectolytic Erwinia sp. by the yellow color that developed in wells of the latter strains after acidification of the medium with dilute HCl. The test is visual and can be performed with minimal equipment and cost.