Molecular identification of a phytoplasma associated with Elm witches'-broom in China

Elm samples with and without witches'-broom symptoms (EWB) were collected from Tai'an and Zhaoyuan, Shandong Province, China. Phytoplasmal cells were observed in the phloem cells of symptomatic plants under electron microscope. Specific fragments of about 1.2 kb in length were amplified with nested-PCR from symptomatic samples, while no fragment was obtained from healthy plants. The 16S rRNA gene sequences of the phytoplasmas associated with elm witches'-broom in Tai'an (EWB-TA) and Zhaoyuan (EWB-ZHY) had high similarities, and formed a sublineage in phylogenetic tree, with members of subgroup B or D of aster yellows group (16SrI). Computer simulated restriction fragment length polymorphism analysis of 16S rRNA gene revealed that EWB-TA and EWB-ZHY patterns had similarity coefficients of 1.00 with the pattern from the representative strains of subgroup 16SrI-B, and had a similarity coefficient of lower than 0.97 with representatives of other subgroups. These results indicated that the phytoplasma strain associated with elm witches'-broom in China was very closely related to 'Candidatus Phytoplasma asteris' OAY, belonging to subgroup-B of aster yellows group (16SrI-B). This is the first report of a phytoplasma associated with elm witches'-broom disease in China.