Incidence and molecular characterization of a 16SrI-B phytoplasma strain associated with Eruca sativa phyllody in Iran

During 2012-2014 surveys for phytoplasma diseases, a phyllody symptomatology was observed in Eruca sativa in Abarkooh (Yazd province, Iran). The highest disease incidence was 32% in Ali Abad-e Shams (Esfandabad), and infected plants showed crown proliferation, witches’ broom, little leaf, flower virescence, phyllody, sterility, and plant stunting. Total DNA extracted from symptomatic and asymptomatic plants was tested for phytoplasma presence verification by direct PCR using P1/P7 primers and nested PCR using P1/P7 and R16F2n/R16R2 primers pairs. PCR amplicons of about 1.8 and 1.25 kb respectively, were obtained from all symptomatic E. sativa plants, but not from the symptomless ones. Restriction fragment length polymorphism analysis of R16F2n/R2 amplicons using AluI, HhaI, HinfI, HpaII, MseI, RsaI and TaqI restriction enzymes showed profile identical to each other and also to those of 16SrI phytoplasmas indicating that the phytoplasmas, associated with E. sativa phyllody are members of 16SrI group. The consensus sequence of Ali Abad-e Shams E. sativa phyllody strain showed 100% identity with those of the ‘Candidatus Phytoplasma asteris’-related strains. Phylogenetic analysis confirmed that this phytoplasma clustered within the 16SrI phytoplasma clade closer to the onion yellows mild strain (OY-M) a 16SrI-B phytoplasma. Restriction analysis using iPhy Classifier confirmed that virtual RFLP patterns of the phytoplasma were identical (similarity coefficient 1.00) to OY-M strain. A 16SrI-B related phytoplasma has been reported associated with rapeseed (Brassica napus) phyllody in Iran and Eruca sativa may be a secondary host for the 16SrI-B phytoplasma associated with rapeseed phyllody.