一例输入性皮肤利什曼病病原体的鉴定

Pathogen Identification in an Imported Case of Cutaneous Leishmaniasis

对广州一例未知病原体感染引起的皮肤病患者血样进行免疫学和分子生物学鉴定,以确定其病原体。取静脉血,用Leish r K39 dipstick金标试纸检测利什曼原虫抗体情况。取皮损处组织,75%乙醇固定后,提取基因组DNA。用两对利什曼原虫种特异性引物LITSR-L5.8S和NAGTL1-NAGTL4分别PCR扩增利什曼原虫核糖体DNA内转录间隔区1和N-乙酰氨基葡萄糖苷转移酶的基因片段,扩增产物进行测序和BLAST序列分析。Leish r K39 dipstick金标试纸检测结果呈弱阳性。PCR结果显示,引物LITSR-L5.8S和NAGTL1-NAGTL4分别扩增出约404 bp和1 405 bp的片段,两片段序列与硕大利什曼原虫(Leishmania major)相应序列的相似性均为99.7%。其序列Gen Bank登录号分别为KU975160和KX150476。确诊该皮肤病患者为输入性皮肤利什曼病病例,病原体为硕大利什曼原虫。

In this study, immune and molecular biological methods were used to identify the pathogen in a blood sample from a patient with dermatosis. Venous blood was collected and tested with Leish rK39 dipsticks. The lesion sample was collected and fixed in 75% ethanol, and DNA was extracted. The internal transcribed spacer 1 of rDNA and N-acetylglucosamine-1-phosphate transferase of Leishmania were amplified with PCR using primers LITSR-L5.8S and NAGTL1s-NAGTL4, respectively. The amplified products were sequenced and analyzed by BLAST. Weakly positive results were obtained for the gold-labeled Leish rK39 dipstick serological test. PCR resulted in products of 404 bp and 1 405 bp with primers LITSR-L5.8S and NAGTL1-NAGTL4, respectively. Both were 99.7% homologous to the corresponding sequence of Leishmania major. The accession number of the two sequences were KU975160 and KX150476. The case of dermatosis is diagnosed as imported cutaneous leishmaniasis and the pathogen is L. major.