'Candidatus Phytoplasma cirsii', a novel taxon from creeping thistle [Cirsium arvense (L.) Scop].

Creeping thistle [Cirsium arvense (L.) Scop.] and dahlia (Dahlia sp.) plants showing typical symptoms of phytoplasma infection including yellowing, stunting, inflorescence and proliferation, were sampled; the presence of phytoplasma was confirmed by standard PCR using universal primers. RFLP analysis allowed classification of the detected phytoplasma strains CirYS, CirYS1 and DahlP within the 16SrXI group, the unique restriction profile F2nR2 fragment obtained in silico by iPhyClassifier indicated that they belong to the new 16SrXI-E subgroup. Genetic analysis of the 16S rRNA gene revealed that the studied strains shared less than 97.5% similarity with all of the previously described 'Candidatus Phytoplasma' species. The closest relatives are 'Candidatus Phytoplasma cynodontis' and 'Candidatus Phytoplasma oryzae' with 96.8% and 96.6% similarity. All strains studied bear three specific regions in the 16S rRNA gene, discriminating them from the other phytoplasma species. Phylogenetic analysis of the 16S rRNA and secA genes confirmed this specificity, as the creeping thistle and dahlia phytoplasma strains clustered in a distinguishable lineage group. The uniqueness of the genetic analysis agrees with the biological characterization of the studied phytoplasma strains, their host range, and geographical distribution. The strains only infect dicotyledonous plants in Europe, contrary to their closest relatives. Based on their unique properties, it could be concluded that the studied phytoplasma strains represent a discrete group that is proposed as a novel taxon 'Candidatus Phytoplasma cirsii', with strain CirYS as a reference strain.

Tuf and secY PCR amplification and genotyping of phytoplasmas.

Tuf and secY genotyping techniques have been developed to distinguish phytoplasma strains. Tuf polymerase chain reaction sequence analyses are available for phytoplasma taxonomic groups 16SrI, 16SrV, 16SrXII-A, and XII-B. In addition to their use to confirm the taxonomic status of phytoplasma strains, they allow the spread of phytoplasma strains in host plants and insect vectors to be traced. SecY is more variable than tuf and is therefore more discriminatory than tuf, but secY and tuf phylogenies show congruence.