Occurrence and molecular characterization of a 16SrI-R subgroup phytoplasma associated with Aquilegia vulgaris phyllody disease

During 2016-2017 surveys, carried out for phytoplasma diseases in ornamental plants in Chaharmahal and Bakhtiari provinces, Iran, found symptoms of virescence, phyllody, reduced size of leaves and flowers in columbine (Aquilegia vulgaris). Total DNAs extracted from symptomatic and symptomless plants were tested for the presence of phytoplasma using P1/P7 and R16F2n/R16R2 primers in direct and nested PCR producing amplicons of about 1.8 and 1.2 kb, respectively, from all symptomatic A. vulgaris plants, but not from symptomless ones. The consensus sequence of the detected phytoplasma named Aquilegia phyllody (APh) was 100% identical with strains clustering to phytoplasmas enclosed in the 16SrI group as also confirmed by phylogenetic analyses. Both real and virtual restriction fragment length polymorphism analysis of R16F2n/R16R2 amplicons showed profiles that were identical to each other and indicated the affiliation of the APh phytoplasma to the 16SrI-R subgroup. This is the first report of a 16SrI-R phytoplasma associated with this A. vulgaris phyllody disease.