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Duplex Real-Time Polymerase Chain Reaction Reveals Competition Between Erwinia amylovora and E. pyrifoliae on Pear Blossoms

First, second, and fourth authors: Agriculture and Agri-Food Canada, Southern Crop Protection & Food Research Centre, 4902 Victoria Ave. North, P.O. Box 6000, Vineland Station, ON, Canada L0R 2E0; and first and third authors: Department of Biological Science, Brock University, 500 Glenridge Avenue, St. Catharines, ON, Canada L2S 3A1.

Erwinia amylovora and E. pyrifoliae are the causative agents of fire blight and Asian pear blight, respectively. The pathogens are closely related, with overlapping host ranges. Data are unavailable on the current distribution of E. pyrifoliae and on the interaction between the two species when they are present together on the same host. In this study, a duplex real-time polymerase chain reaction (PCR) protocol was developed to monitor the population dynamics of E. amylovora and E. pyrifoliae on the surface of Bartlett pear blossoms. Bacterial cells washed from blossoms were used directly as the PCR template without DNA extraction. Primers and a probe based on the E. amylovora levansucrase gene detected all E. amylovora strains. All E. pyrifoliae strains, including the Japanese Erwinia strains previously described as E. amylovora, were detected with a primer and probe combination based on the E. pyrifoliae hrpW gene. Disease development and severity were not significantly different in blossoms inoculated with individual Erwinia species or with a mixture of the two species. However, E. amylovora grew to greater population sizes than did E. pyrifoliae in both single species inoculations and in mixtures, suggesting that E. amylovora has a greater competitive fitness on Bartlett pear blossoms than E. pyrifoliae.

Additional keywords:bioassay, microbial population dynamics, multiplex detection, quantitative PCR, virulence.