Highly sensitive detection of Pseudomonas savastanoi pv. savastanoi in asymptomatic olive plants by nested-PCR in a single closed tube

A nested-polymerase chain reaction (PCR) has been set up to be performed in a single closed tube for the detection of Pseudomonas savastanoi pv. savastanoi. Nested-PCR coupled with dot-blot hybridization was able to detect up to one cell of the target per ml of olive extract, showing the greatest sensitivity compared with all previously reported detection assays. Validation of the developed procedure for diagnosis and epidemiological purposes was achieved by testing ca. 240 asymptomatic plant samples from olive trees. When performing the other previously reported techniques (bacterial isolation and single PCR), P. savastanoi was detected in 50 of the analyzed samples, while with the new developed nested-PCR assay, the bacterium was detected in 82 samples. (C) 2003 Elsevier Science B.V. All rights reserved.