泡桐丛枝病病树周围几种植物上植原体的分子检测

 【目的】初步明确自然条件下可能感染泡桐丛枝病植原体的寄主植物。【方法】用植原体16S rRNA基因的通用引物,对从泡桐丛枝病病树周围采集的表现黄化、小叶、皱叶、丛枝等症状或无症状的16种植物样品的DNA进行巢式PCR扩增,对所扩增的片段进行序列测定和分析。并利用泡桐从枝病植原体延伸因子的抗血清对部分样品进行间接免疫荧光观察。【结果】巢式PCR结果显示从牛筋草（Eleusine indica）、辣椒（Capsicum annuum）、狗尾草（Setaria viridis）、山药（Dioscorea opposita）、灯笼泡（Physalis angulata）、花生（Arachis hypogaea）及南瓜（Cucurbita moschata）共7种植物样品和阳性对照PaWB菏泽分离物（PaWB-HZ）中均得到了约1.2 kb的特异性片段。序列分析表明这7种植原体分离物和PaWB-HZ属于翠菊黄化组的16SrI- D亚组。对所采集的植原体侵染的寄主植株辣椒、山药、花生、南瓜和泡桐进行间接免疫荧光观察结果显示,仅在PaWB-HZ侵染的泡桐中发现翠绿色特异荧光,在健康泡桐及其它4种分离物侵染的植物中均未检测到明显的植原体特异荧光。【结论】首次从泡桐丛枝病病树周围存在的7种其它植物中检测到植原体,并且测定其植原体16S rDNA核苷酸序列与泡桐丛枝植原体相关基因片段高度同源,初步推测这7种植物可能是泡桐丛枝病植原体自然寄主或是通过昆虫偶尔被感染。

Molecular Detection of Phytoplasma Strains from Several Plants Around Diseased Paulownia Infected with Paulownia Witches'-broom Phytoplasma

Objective] The objective of this study is to investigate and identify the alternative host plants of paulownia witches'-broom phytoplasma (PaWB) in natural condition. Method] Using universal primers for phytoplasmal 16S rRNA gene, nested PCR was conducted to detect phytoplasmas from 16 species of plants with/without symptoms of yellowing, little leaf, winkle leaf, witches'-broom and growing around paulownia plants infected with PaWB, and then sequencing and phylogenetic analyses of the amplified fragment were made. Indirect immunofluorescence was also used to analyze partial plant samples. Result] A specific fragment of ca. 1.2 kb in length was amplified with nested PCR from 7 species of plants, Chinese yam (Dioscorea opposita), groundcherry (Physalis angulata), pepper (Capsicum annuum), peanut (Arachis hypogaea), goosegrass (Eleusine indica), bristle grass (Setaria viridis) and musdky guard (Cucurbita moschata) along with the positive control of PaWB. The sequencing and phylogenetic results showed that all the phytoplasmas detected in the 7 species of plants and the one from diseased paulownias belonged to 16Sr I-D subgroup of aster yellows phytoplasma group. Indirect immunofluorescence analysis using antibody to PaWB Tuf protein indicated that specific fluorescence was easily detected in paulownia plants infected with Beijing and Heze isolates of PaWB, but not either in healthy paulownia plants or in CaWL, DoY, AhY, Cm A phytoplasma-infected plants. Conclusion] Paulownia witches'-broom phytoplasma were detected from 7 species of plants, which were probable natural hosts of PaWB.