Gundersen D E, Lee I M, Schaff D A, Harrison N A, Chang C J, Davis R E, Kingsbury D T
Molecular Plant Pathology Laboratory, U.S. Department of Agriculture, Beltsville, Maryland 20705, USA.
Int J Syst Bacteriol. 1996 Jan;46(1):64-75. doi: 10.1099/00207713-46-1-64.
Conserved gene sequences, including 16S rRNA and ribosomal protein gene sequences, were used to evaluate genetic variations in phytoplasma strains belonging to 16S rRNA groups I (aster yellows and related phytoplasmas) and III (X-disease and related phytoplasmas). We used PCR to amplify the sequences of the 16S ribosomal DNA and a segment of the ribosomal protein gene operon (encoding the 3' region of rps19, all of rp122, and rps3) from diverse phytoplasma group I and III strains. Additional chromosomal gene sequences of group I strains were also amplified. The PCR products amplified from members of each group of phytoplasmas were compared by performing restriction fragment length polymorphism (RFLP) analyses. On the basis of the RFLP patterns observed and similarity coefficients derived from combined RFLP analyses, the phytoplasma strains belonging to groups I and III were placed in distinct 16S rRNA, ribosomal protein, and 16S rRNA-ribosomal protein subgroups. Analyses of two or more conserved gene sequences revealed that members of the two groups were more diverse than previously thought. Subgroup differentiation on the basis of our combined analyses of 16S rRNA and ribosomal protein gene sequences seemed to adequately reflect the levels of chromosomal homology determined by DNA-DNA hybridization assays. On the basis of unique RFLP profiles, we identified new, previously unclassified group I phytoplasma strains, including the organisms that are associated with Ipomoea obscura witches'-broom [subgroup 16SrI-F(rr-rp)], maize bushy stunt [subgroup 16SrI-I(rr-rp)], and Mexican periwinkle virescence [subgroup 16SrI-J(rr-rp)], and new, previously unclassified group III phytoplasma strains, including the organism that is associated with pecan bunch [subgroup 16SrIII-H(rr-rp)]. On the basis of the results of our analyses of 16S rRNA and ribosomal protein conserved gene sequences, we recognized 9 group I subgroups and eight group III subgroups. We propose that phytoplasma strains belonging to each group I and III subgroup should be distinguished taxonomically at a level equivalent to the subspecies level.
保守基因序列，包括16S rRNA和核糖体蛋白基因序列，被用于评估属于16S rRNA第I组（翠菊黄化病及相关植原体）和第III组（X病及相关植原体）的植原体菌株的遗传变异。我们使用聚合酶链式反应（PCR）从不同的第I组和第III组植原体菌株中扩增16S核糖体DNA序列和核糖体蛋白基因操纵子的一个片段（编码rps19的3'区域、全部rp122和rps3）。还扩增了第I组菌株的其他染色体基因序列。通过进行限制性片段长度多态性（RFLP）分析，比较了从每组植原体成员中扩增出的PCR产物。根据观察到的RFLP模式和联合RFLP分析得出的相似系数，将属于第I组和第III组的植原体菌株归入不同的16S rRNA、核糖体蛋白和16S rRNA - 核糖体蛋白亚组。对两个或更多保守基因序列的分析表明，这两组的成员比之前认为的更加多样。基于我们对16S rRNA和核糖体蛋白基因序列的联合分析进行的亚组区分，似乎充分反映了通过DNA - DNA杂交试验确定的染色体同源性水平。基于独特的RFLP图谱，我们鉴定出了新的、以前未分类的第I组植原体菌株，包括与暗牵牛丛枝病相关的生物体[亚组16SrI - F(rr - rp)]、玉米矮化病相关的生物体[亚组1�SrI - I(rr - rp)]和长春花褪绿病相关的生物体[亚组16SrI - J(rr - rp)]，以及新的、以前未分类的第III组植原体菌株，包括与山核桃簇生病相关的生物体[亚组16SrIII - H(rr - rp)]。基于我们对16S rRNA和核糖体蛋白保守基因序列的分析结果，我们识别出9个第I组亚组和8个第III组亚组。我们建议，属于第I组和第III组每个亚组的植原体菌株在分类学上应在相当于亚种水平的层面上加以区分。
