PCR-based techniques for the detection of Erwinia amylovora.
E. amylovora has been reported to survive epiphytically on symptomless flowers, leaves, twigs and immature fruit on apples and pears. Conventional diagnostic methods cannot detect small populations of E. amylovora, so a sensitive and specific PCR-based method was developed. The method involves amplification of a 187 bp length of DNA specific to E. amylovora. The 187 bp product was obtained using DNA or intact bacteria as a template. All 69 cultures of E. amylovora in an international collection from 10 host species in 5 countries were successfully identified using the primers. PCR products were not amplified from 29 strains of other Erwinia species and 9 isolates of 4 other genera of bacteria. The PCR-based method was used to detect E. amylovora in plant tissue. However, amplification of target DNA can be inhibited by large amounts of contaminating DNA and/or interfering components from plant tissue which may lead to false negative results. This difficulty has been overcome using bacterial growth enhancement and immuno-capture techniques enabling a readily detectable 187 bp product to be amplified from reactions containing low levels of E. amylovora.