Detection and identification of 'Candidatus Phytoplasma prunorum' in Prunus germplasm

Abstract

The molecular characterization of the causal agent of diseases associated with several symptoms such as decline, yellowing, leaf roll and off-season growth in stone fruits made it possible to determine a common etiology, and the name ́European stone fruit yellows phytoplasma was proposed. Recently, the new taxonomical species description within the genus is 'Candidatus Phytoplasma prunorum'. A 2-yr survey was carried out in two different Prunus collections of the Centro de Investigación y Tecnología Agroalimentaria de Aragón (CITA) including European and Japanese plum genotypes of various species and several interspecific hybrids used as rootstocks. Both off-season growth in winter and decline were observed. In order to identify the phytoplasma suspected as a causal agent, two different PCR methods were applied to all inspected trees. The first method was a nested PCR with 16Sr X group-specific primers followed by RFLP analysis. The second method was a direct PCR with specific primers for 'Ca P. prunorum' (Eca1/Eca2). In the most symptomatic trees the presence of this phytoplasma was confirmed by at least one of the methods; negative results were obtained in asymp-tomatic trees. The nested PCR-RFLPs analysis was confirmed as a reliable method for routinary diagnosis rather than direct PCR.