The plasmids of Chlamydia trachomatis and Chlamydophila pneumoniae (N16):: accurate determination of copy number and the paradoxical effect of plasmid-curing agents
The plasmids of Chlamydia trachomatis and Chlamydophila pneumoniae (N16):: accurate determination of copy number and the paradoxical effect of plasmid-curing agents
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DOI：
10.1099/mic.0.27625-0
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发表时间：
2005-03-01
期刊：
影响因子：
2.8
通讯作者：
Clarke, IN
中科院分区：
文献类型：
--
作者：
Pickett, MA;Everson, JS;Clarke, IN
在几乎所有的沙眼衣原体分离株中都发现了一个7-5 kbp的隐蔽质粒。建立了实时PCR检测方法，使用TaqMan化学，以准确定量衣原体质粒和单拷贝，染色体omcB基因在感染性，初级体（EB）的C。440.第四百四十章还测定了其他6个性病性淋巴肉芽肿（LGV）分离株（血清型L1-L3）、10个沙眼分离株（血清型A-C）和9个泌尿生殖分离株（血清型D-J）的EB中的质粒拷贝数。结果表明，平均质粒拷贝数为4（.）0 +/- 0（.）每个染色体8个（平均值+/-95%置信区间）质粒。在衣原体发育周期中，最多可达7（.）每条染色体检测到6个质粒，表明在活跃复制的网状体中质粒拷贝数增加。使用质粒固化剂溴化乙锭、吖啶橙子或丙咪嗪/新生霉素从菌株L1 440中消除质粒的尝试导致质粒拷贝数的矛盾增加。据推测，化学固化剂诱导的应力可能刺激质粒编码的复制（Rep）蛋白的活性。与C.在沙眼衣原体中，只有单个嗜肺炎衣原体分离物携带质粒。C. pneumoniae菌株N16支持7（.）4 kbp质粒，其中编码一种推定Rep蛋白的ORF 1被缺失破坏并分裂成两个较小的ORF。类似的测定技术揭示了1（。）3 +/- 0（.）该菌株EB中每个染色体2个质粒（平均95%置信区间）。这些发现与ORF 1编码的蛋白质参与质粒复制和拷贝数控制但不是必需的假设一致。
A 7-5 kbp cryptic plasmid is found in almost all isolates of Chlamydia trachomatis. Real-time PCR assays, using TaqMan chemistry, were set up to quantify accurately both the chlamydial plasmid and the single copy, chromosomal omcB gene in the infectious, elementary bodies (EBs) of C. trachomatis L1 440. Plasmid copy number was also determined in the EBs of six other lymphogranuloma venereum (LGV) isolates (serovars L1-L3), ten trachoma isolates (serovars A-C) and nine urogenital isolates (serovars D-J). The results indicated an average plasmid copy number of 4(.)0 +/- 0(.)8 (mean +/- 95 % confidence interval) plasmids per chromosome. During the chlamydial developmental cycle, up to 7(.)6 plasmids per chromosome were detected, indicating an increased plasmid copy number in the actively replicating reticulate bodies. Attempts to eliminate the plasmid from strain L1 440 using the plasmid-curing agents ethidium bromide, acridine orange or imipramine/novobiocin led to a paradoxical increase in plasmid copy number. It is speculated that the stress induced by chemical curing agents may stimulate the activity of plasmid-encoded replication (Rep) proteins. In contrast to C. trachomatis, only a single isolate of Chlamydophila pneurnoniae bears a plasmid. C. pneumoniae strain N16 supports a 7(.)4 kbp plasmid in which ORF1, encoding one of the putative Rep proteins, is disrupted by a deletion and split into two smaller ORFs. Similar assay techniques revealed 1(.)3 +/- 0(.)2 plasmids per chromosome (mean 95 % confidence interval) in EBs of this strain. These findings are in agreement with the hypothesis that the ORF1-encoded protein is involved in, but not essential for, plasmid replication and control of copy number.