IDENTIFICATION OF ERWINIA-AMYLOVORA BY GROWTH-MORPHOLOGY ON AGAR CONTAINING COPPER-SULFATE AND BY CAPSULE STAINING WITH LECTIN

Erwinia amylovora strains formed yellow colonies on minimal agar mediu m MM2 containing asparagine and copper sulfate (MM2Cu), in contrast to a white morphology on minimal agar without copper salt. Additionally, the colonies were mucoid to various extents. The yellow color was cha racteristic for the fire blight pathogen, including strains from raspb erry and from other unusual host plants, and was used to establish a n ovel plating technique for identification of E. amylovora. The new ide ntification method was especially superior to semi-selective media wit h sucrose when natural levan-deficient strains were assayed. No growth of E. amylovora was observed for the similar medium MM1 containing 2 mM CuSO4, due to its low content of asparagine. Identification by colo ny morphology on MM2 agar with copper was confirmed by staining the ba cterial capsules with FITC-labeled lectin from Abrus precatorious, a c ompound which has a high affinity for galactose residues, the main sug ar in the capsular exopolysaccharide amylovoran of E. amylovora. Other plant-associated bacteria usually did not produce the typical colony morphology of E. amylovora on MM2 agar with copper. Furthermore, those cells were not stained with the Abrus lectin. Capsule staining was al so observed for weakly mucoid strains of E. amylovora, but not for str ains with mutations affecting amylovoran synthesis. The secretion of f luorescent compounds by Pseudomonas syringae pathovars and even growth of any other bacterial colonies adjacent to E. amylovora could interf ere with the formation of typical yellow colonies on MM2Cu, which coul d be white in case of dense plating. After screening for white colonie s on LB agar, E. amylovora was identified in extracts from Cotoneaster leaves and in bark from apple trees with fire blight symptoms by its yellow growth pattern on MM2Cu agar and by capsule staining. The propo sed selective medium gives a clear signal, is easy to prepare, does no t contain dyes or any compounds toxic to humans, and can also detect E . amylovora strains deficient in levan synthesis.