Detection of Erwinia amylovora in symptomless apple plants by bio-PCR and bio-duplex PCR

Two polymerase chain reaction (PCR) protocols (bio-PCR and bio-duplex PCR) that allow the specific detection of live cells of Erwinia amylovora in plant tissues were used to detect the fire blight pathogen in symptomless apple trees. Both protocols have an enrichment step before amplification. In the bio-PCR the E. amylovora specific primers 187a and 187b were used; in the bio-duplex PCR an additional set of primers which amplified a fragment of the 16S rDNA gene was used as an internal positive control. The bio-PCR protocol was capable of detecting 10 2 cfu of the pathogen per apple bud. E. amylovora was rarely found beyond the point of inoculation in symptomless tissues of apple trees which were resistant to fire blight. In diseased trees that were susceptible to fire blight, E. amylovora was detected in some of the symptomless lateral shoots within five months after inoculation. Two years after inoculation, live cells of E. amylovora could still be detected in the susceptible apple trees using either one of these bio-PCR protocols.