Development of a rapid screening method for testing of Erwinia amylovora (Burill) Winslow et al. from asymptomatic plant material | Razvoj hitrega presejalnega testa za laboratorijsko dolocanje prikrite okuzbe z Erwinia amylovora (Burill) Winslow et al.
2007
Pirc, M., National Inst. of Biology, Ljubljana (Slovenia) | Dreo, T. | Ravnikar, M.
In Slovenia testing of Erwinia amylovora from latent samples has been conducted since 1998 using methods described in EPPO diagnostic protocols as well as using new approaches. ELISA, enrichment ELISA, and immunofluorescence (IF) present three rapid and validated serological screening tests. These serological methods may not be sensitive enough due to the low concentrations of Erwinia amylovora in asymptomatic plant material. PCR (polymerase chain reaction) presents a new molecular screening method. Different compounds present in the extract (polifenolic compounds, phytopharmaceutical residues,_) can inhibit such PCR reaction and lower the sensitivity of the test. Cross-reactivity with closely related bacteria was also noticed and presents another problem. In the last few years real-time PCR has been introduced. The method can be used as either qualitative or quantitative test where the risk for cross-contamination is lowered due to the direct monitoring of the results. Because of the high throughput of samples, real-time PCR can be used as a screening assay. A real-time PCR method for detection of Erwinia amylovora has already been developed (Salm et al., 2004). The target for the test is located on the pEA29 plasmid that is relatively stable but is not present in all bacterial isolates. Erwinia amylovora without pEA29 can still cause appearance of the disease symptoms. For this reason a new real-time PCR targeting chromosomal DNA is being developed in our laboratory.
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