Occurrence of a ‘Candidatus Phytoplasma omanense’-related strain in bindweed showing a witches’ broom disease in Iran

In 2014, bindweed witches’ broom and dwarfing symptoms were observed in some bindweed plants grown in alfalfa fields in Bafg (Yazd province, Iran). Direct polymerase chain reaction using P1/P7 and nested PCR using P1/P7 followed by R16mF2/ R16mR2 and R16F2n/R16R2 amplify the 16S ribosomal gene and RFLP analysis of several R16F2n/R16R2 amplicons with AluI, HaeIII, HhaI, HpaI, HpaII, KpnI, RsaI, MseI and Taq1 restriction enzymes showed profiles in which phytoplasma strains belonging to 16SrXXIX group were identified together with 16SrXII-A phytoplasmas by comparison of profiles and confirmed after nested PCR/RFLP analyses with R16(I)F1/R1 primers. Three samples from different fields were directly sequenced using nested PCR products obtained with P1/P7 and R16mF2/R16mR2. Sequence comparison by BLAST analysis showed the highest sequence identity with phytoplasmas in group 16SrXXIX (‘Candidatus Phytoplasma omanense’) and phylogenetic analysis confirmed that the phytoplasma could be considered a ‘Ca. P. omanense’-related strain. While the computer-simulated analysis with the 17 restriction endonucleases in the iPhyClassifier indicated identity of this strain with ‘Ca. P. omanense’, a virtual digestion using BcgI and AatII enzymes allow to enclose it in a new subgroup designed 16SrXXIX-B.