新疆首次临床分离新生隐球菌菌株分子鉴定、基因分型及体外药物敏感性研究

中国真菌学杂志　 2016, Vol. 11 　Issue (6): 327-331.

新疆首次临床分离新生隐球菌菌株分子鉴定、基因分型及体外药物敏感性研究

尤丽吐孜·许合热提¹, 哈地丽亚·哈斯木¹, 王晓东¹, 邓淑文², 帕丽达·阿布利孜¹

1. 新疆医科大学第一附属医院皮肤科, 乌鲁木齐 830053;
2. 第二军医大学上海长征医院真菌研究所, 上海 200003

收稿日期:2016-09-01 出版日期:2016-12-28 发布日期:2016-12-28
通讯作者: 帕丽达·阿布利孜,E-mail:palidae@aliyun.com E-mail:palidae@aliyun.com
作者简介:尤丽吐孜·许合热提,女(维吾尔族),硕士,住院医师.E-mail:m13899841261@163.com
基金资助:
国家自然科学基金（81360237）

Molecular identification and genotyping and in vitro drug sensitivity study of the first five clinical isolates of Cryptococcus neoformans in Xinjiang

Yultuz·Xohrat¹, Kadirye·Kasim¹, WANG Xiao-dong¹, DENG Shu-wen², Prida·Abliz¹

1. Dermatology of the First Affiliated hospital of Xinjiang Medical University, Urumqi 830053;
2. The Fungus Institute of Shanghai Changzheng hospital, Shanghai 200003

Received:2016-09-01 Online:2016-12-28 Published:2016-12-28

摘要/Abstract

摘要：

目的对新疆首次临床分离的5株新生隐球菌进行分子鉴定、RAPD-PCR基因分型及体外药物敏感性研究。方法 5株新生隐球菌分子鉴定采用核糖体DNA大亚基（LSU rDNA）D1/D2基因区域分子鉴定。分子分型采用引物SEQ-6结合RAPD-PCR法扩增5株新疆临床分离新生隐球菌菌株及5株由上海长征医院提供分离自上海新生隐球菌菌株，根据扩增产物带型进行基因型判定。采用临床实验室标准化协会（CLSI）的酵母微量液基稀释法（M27-A3）测定5株新疆临床分离新生隐球菌菌株对6种抗真菌药（伏立康唑、伊曲康唑、两性霉素B、特比萘芬、氟康唑、5-氟胞嘧啶）的体外敏感性。结果 5株新疆临床分离隐球菌菌株经过D1/D2区域序列分析在基因Bank中比对后鉴定为新生隐球菌。5株新疆临床分离新生隐球菌和5株上海新生隐球菌菌株经RAPD-PCR法扩增，带型显示分为A、B、C、D 4个基因型，其中新疆5株菌株A型1株、其余4株均为B型，上海菌株B型为1株，C型3株、D型1株，5株新疆临床分离新生隐球菌株对伏立康唑、伊曲康唑、两性霉素B、特比萘芬的MIC值（μg/mL）范围依次为：0.0625~0.25、0.25~1、0.125~0.5、1~2，对氟康唑、5-氟胞嘧啶MIC值较高，MIC值范围依次为：4~16、8~32。结论新疆临床分离5株隐球菌株采用D1/D2基因序列鉴定为新生隐球菌。RAPD-PCR分型显示新疆临床分离5株新生隐球菌株以B型基因型为主。A型和B型对伏立康唑、两性霉素B敏感，对伊曲康唑、特比萘芬剂量依赖型敏感，对氟康唑、5-氟胞嘧啶耐药。

关键词: 新生隐球菌, 新疆, RAPD-PCR基因分型, 体外药物敏感性

Abstract:

Objective For the first time to perform the molecular identification,RAPD-PCR genotyping and in vitro drug sensitivity study of 5 clinical strains of Cryptococcus neoformans in Xinjiang.Methods Molecular classification using primer SEQ-6 from big subunit ribosomal DNA (LSU rDNA) D1/D2 gene region and RAPD-PCR method were applied in 5 strains of clinical isolated Xinjiang Cryptococcus neoformans strains and 5 Cryptococcus strains provided by Shanghai Changzheng hospital.Sensitivity in vitro of 5 clinical isolated Xinjiang Cryptococcus neoformans strains of the above 6 kinds of antifungal drugs (Voriconazole,itraconazole,amphotericin B,terbinafine,fluconazole,5-fluorine cytosine)were judged according to clinical laboratory standards institute (CLSI) trace liquid dilution method of yeast (M27-A3).Results The 5 strains of Xinjiang were identified as Cryptococcus neoformans through D1/D2 region sequence analysis.All 10 strains are divided into A,B,C,D 4 genotype by RAPD-PCR.One Xinjiang strain was type A,the rest 4 strains were type B;Among 5 Shanghai strains,1 strain was type B,3 strainswere type C,1 strain was typeD.For the 5 Xinjiang clinical Cryptococcus neoformans strains,to the MIC value ranges of voriconazole,itraconazole,amphotericin B and terbinafine were as following:0.062 5-0.25 μg/mL,0.25-1 μg/mL,0.125-0.5 μg/mL,1-2 μg/mL,respectively;MIC value ranges of fluconazole,5-fluorine cytosine MIC value were much higher as:4-16 μg/mL,8-32 μg/mL,respectively.MIC values of genotype A and B to voriconazole were 0.062 5-0.125 μg/mL.Conclusion The 5 Xinjiang clinical isolates of Cryptococcus were identified as Cryptococcus neoformans using D1/D2 genes analysis.RAPD-PCR revealed these clinical Cryptococcus neoformans strain in Xinjiang with priority given to type B genotypes.Type A and type B isolates weremore sensitive to voriconazole,amphotericin B,itraconazole and terbinafine.

Key words: Cryptococcus neoformans, Xinjiang, RAPD-PCR genotyping, antifungal sensitivity

中图分类号:

R379.5

尤丽吐孜·许合热提, 哈地丽亚·哈斯木, 王晓东, 邓淑文, 帕丽达·阿布利孜. 新疆首次临床分离新生隐球菌菌株分子鉴定、基因分型及体外药物敏感性研究[J]. 中国真菌学杂志, 2016, 11(6): 327-331.

Yultuz·Xohrat, Kadirye·Kasim, WANG Xiao-dong, DENG Shu-wen, Prida·Abliz. Molecular identification and genotyping and in vitro drug sensitivity study of the first five clinical isolates of Cryptococcus neoformans in Xinjiang[J]. Chinese Journal of Mycology, 2016, 11(6): 327-331.

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