绵羊附红细胞体部分16S rRNA基因序列测定和系统进化分析

从自然感染附红细胞体的石河子和杭州两地的绵羊无菌采集血液,分离附红细胞体并提取基因组,根据已公布的绵羊附红细胞体16S rRNA基因序列设计一对引物,进行PCR扩增。结果扩增出约1 100bp目的片段。测序结果表明:目的片段长1 080bp、1 079bp(GenBank收录号EU916726、FJ440328),同源性分析表明该序列与参考序列(AF338268)同源性达99.7%,证实该病原是绵羊附红细胞体。将该序列与5种支原体、14种血营养菌及立克次氏体等相应序列进行同源性比对,系统进化树表明,绵羊附红细胞体和其他血营养菌在进化关系上组成一个大的分支,与支原体科,支原体属病原最为接近,与立克次氏体科的病原较远。分析结果与Neimark等提出的观点一致,将这类血营养菌划归支原体科、支原体属。

Sequencing and phylogenetic analysis of the partial 16S rRNA in Eperythrozoon ovis

The genomic DNA of Eperthrozoon ovis was extracted from blood samples of sheeps naturally infected with eperthrozoons in Shihezi and Hangzhou districts and was amplified with PCR by a pair of primers designed according to the published genomic sequence of 16S rRNA of eperthrozoon gene locating in the conserved region of the corresponding sequence of haematophilie bacteria. In this way, a target fragment With size about 1 100 bps was amplified. The amplified fragments were cloned and sequenced and the sequencing result showed that this fragments comprised of two sequences of 1 080 bps and 1 079 bps respectively （GenBank No. EU916726 and FJ440328）. The result in phylogenetic analysis indicated that these two sequences were 99.7% identical to the published reference sequence AF338268. These sequences were compared with those sequences of 5 mycoplasma, 14 haematophilic bacteria and rickettsia available in GenBank ,thus the phylogenetic tree was estab- lished, in which they formed a large phylogenetic branch with other haematophilic bacteria and were very close to the Mycoplasma genus of Mycoplasmataloes as proposed by Neimark et al. This result of analysis sustains the advice that Erthrozoon and Haemobartonella genera should be reclassified in Mycoplasrna genus of mycoplasmatales.