VIEW ARTICLE

Techniques

Identification and Detection of Erwinia amylovora with Monoclonal Antibodies. C. P. Lin, Postdoctoral fellow, Department of Plant Pathology; T. A. Chen(2), J. M. Wells(3), and T. van der Zwet(4). (2)Professor, Department of Plant Pathology; (3)Research plant pathologist, USDA, ARS, Cook College, Rutgers University, New Brunswick, NJ 08903; (4)Research plant pathologist, USDA, ARS, Appalachian Fruit Research Station, Kearneysville, WV 25430. Phytopathology 77:376-380. Accepted for publication 6 August 1986. Copyright 1987 The American Phytopathological Society. DOI: 10.1094/Phyto-77-376.

Forty-eight hybridoma clones secreting monoclonal antibodies (MAs) specific to Erwinia amylovora were produced to identify and detect epiphytic and endophytic E. amylovora in pome fruit tissues by indirect enzyme-linked immunosorbent assay (ELISA) and immunofluorescent staining. These MAs reacted positively in indirect ELISA against pure cultures of 75 E. amylovora isolates from different countries. Serological specificity of 37 of the MAs was further tested against 24 strains of bacteria in six genera and against 56 unidentified epiphytes from pome fruit trees collected in different states. Ten clones of MAs reacted specifically with E. amylovora and did not cross-react with any epiphytes or known bacteria. Nine of the 10 MAs produced strong fluorescent reactions in vitro to E. amylovora in indirect immunofluorescent staining and were further used for in situ detection in diseased apple tissues with an epifluorescent microscope. The MAs were also used to detect E. amylovora in vitro by a modified immunofluorescent direct-counting technique. Bacterial suspensions collected from infected pome fruit blossoms and apple fruits were concentrated in a microcentrifuge tube during staining, then collected on polycarbonate membranes by filtration. This technique conveniently detected 5 × 10³ cells per membrane and was useful in detecting and monitoring infection by E. amylovora in pome fruit trees and fruits.