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采用斑点热群立克次体特异引物扩增福建南部来源蜱标本和动物标本中的DNA ,以发现我国可能存在的新种斑点热群立克次体。以立氏立克次体rOmpA基因序列中的相关序列为引物 ,扩增标本中rOmpA基因部分片段 ,用pGEM -T载体及其宿主菌进行阳性克隆筛选 ,阳性克隆经鉴定之后提取质粒DNA ,进行序列测定。测定序列用Blast软件进行序列同源性比较并采集相关序列 ,用Bootstrip方法随机取样 10 0次 ,用PHYLIP软件包进行聚类分析。共检测采自福建宁化地区的越原血蜱 10 0只和野鼠类血标本 38份。其中从越原血蜱(不同来源 )扩增出斑点热立克次体基因片段 ,阳性率为15 0 4% ( 17/ 113) ;并从社鼠、黄胸鼠、褐家鼠扩增出阳性片段 ,以鼠类为单位计算其阳性率为 44 73%。对寄生于野鼠的越原血蜱扩增的阳性片段序列分析表明新测序列与其它斑点热群立克次体的同一基因片段不同 ,其与日本立克次体同源性最高 ,核苷酸序列同源性为 94% ,推定的氨基酸序列同源性为89 %。从病原学角度首次证实我国福建省存在日本立克次体近缘种立克次体 ,这对斑点热群立克次体的深入研究有重要意义
The speckle heat group Rickettsia specific primers were used to amplify the DNA from ticks and specimens from southern Fujian province to find out the new speckle hot group rickettsia in our country. A fragment of rOmpA gene was amplified from the rOmpA gene sequence of Rickettsia rickettsi by PCR. Positive clones were screened by pGEM-T vector and its host strain. Plasmid DNA was extracted after positive clones were identified. Sequencing. The sequences were sequenced and compared using Blast software and the related sequences were collected. The samples were randomly sampled using the Bootstrip method for 10 times, and the phylogenetic analysis was performed using the PHYLIP software package. A total of 10 phthisic blood smear collected from Ninghua area of Fujian Province and 38 voles were collected. In this study, the genomic DNA fragment of Rickettsia speciosa was amplified from different sources (from different sources), and the positive rate was 15 0 4% (17/113). Positive fragments, in rodents to calculate the positive rate of 44 73%. Sequence analysis of the positive fragments amplified from T. vaporans parasitized in wild mice showed that the new sequence was different from other Rickettsia groups in the same gene segment and had the highest homology with Rickettsia japonica. Nucleotide The sequence homology was 94%, and the putative amino acid sequence homology was 89%. From etiology point of view, it is the first time to confirm the existence of Rickettsia rickettsii in Fujian Province, which is of great significance to the further study of Rickettsia