Between 2016 and 2017, symptoms of witches’ broom, proliferation, drying of twigs and ball-like structures formation were observed in Pinus brutia in urban green areas and parks in Isfahan (Iran). All samples from affected P. brutia trees resulted positive in nested-PCR using primer pairs targeting the phytoplasma 16S rRNA gene. Amplicons were obtained only from all the symptomatic pine tree samples and RFLP analyses of R16F2n/R16R2 amplicons with KpnI, AluI, HaeIII, HhaI, HpaII, MseI, RsaI,ThaI, BfaI and TaqI restriction enzymes showed the presence of phytoplasmas enclosed in the 16SrVI group. Four amplicons were sequenced and showed 100% identity to each other. One of these sequences was deposited in the GenBank database (accession number MK158098) and the virtual RFLP pattern was identical (similarity coefficient 1.00) to the reference pattern of 16Sr group VI, subgroup A confirming that the phytoplasma detected in pine is a member of 16SrVI-A subgroup. This is the first report of a 16SrVI-A phytoplasma strain associated with pine tree witches’ broom (P. brutia) disease.

中文翻译：

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伊朗伊斯法罕黑松病相关植原体的发生与鉴定

2016 年至 2017 年间，在伊斯法罕（伊朗）的城市绿地和公园中，在黑松中观察到了女巫扫帚、增殖、树枝干燥和球状结构形成的症状。使用针对植原体 16S rRNA 基因的引物对，来自受影响的 P. brutia 树的所有样本在巢式 PCR 中均呈阳性。扩增子仅从所有有症状的松树样本中获得，并且 R16F2n/R16R2 扩增子与 KpnI、AluI、HaeIII、HhaI、HpaII、MseI、RsaI、ThaI、BfaI 和 TaqI 限制性内切酶的 RFLP 分析表明，在 16Sr 中存在植原体团体。对四个扩增子进行测序并显示彼此 100% 的同一性。这些序列之一存放在 GenBank 数据库中（登录号 MK158098）并且虚拟 RFLP 模式是相同的（相似系数 1. 00) 参考 16Sr 组 VI，亚组 A，确认在松树中检测到的植原体是 16SrVI-A 亚组的成员。这是与松树女巫扫帚 (P. brutia) 病相关的 16SrVI-A 植原体菌株的第一份报告。