干血斑用于HIV-1耐药基因型检测的研究  

Feasibility of using dried blood spots to detect HIV drug resistance genotyping

作　　者：马鹏飞[1] 邢辉[1] 廖玲洁[1] 陈彬[1] 赵全壁[1] 全宇[1] 孙峰[2] 杨绍敏 苏斌[4] 陈曦[5] 邵一鸣 

机构地区：[1]中国疾病预防控制中心性病艾滋病预防控制中心传染病国家重点实验室,北京102206 [2]新疆伊犁哈萨克自治州疾病预防控制中心 [3]云南省传染病专科医院 [4]安徽省疾病预防控制中心 [5]湖南省疾病预防控制中心

出　　处：《中华预防医学杂志》2010年第11期993-998,共6页Chinese Journal of Preventive Medicine

基　　金：基金项目：国家科技重大专项（2008ZXl00014）04）;国家重点基础研究发展计划（2005CB523103和2005CB522903）;中国全球基金艾滋病项目第3轮（2007-07--2009-06）和第4轮（2006-01-2008-12）

摘　　要：目的通过干血斑样本与血浆、全血的HIV-1基因型耐药性检测结果比较，探讨十血斑样本应用于我国患者HIV—1耐药性检测及监测的可行性。方法选择来自安徽（10例）、云南（13例）、湖南（6例）和新疆维吾尔自治区（10例）4省（自治区）共39例AIDS患者，这些患者感染的HIV-1流行株覆盖中国主要的流行亚型（B、CRF01_AE、CRF07_BC），同时对同一患者的血浆、全血及干血斑3种类型的样本应用实验室自建的套式PCR方法扩增HIV的pol基因区，通过美国斯坦福大学的HIV耐药数据库进行耐药程度判别并比较三者的耐药性结果。结果干血斑、全血与血浆样本相比总扩增成功率分别为95％（37／39）、92％（36／39）及100％（39／39），各亚型样本基因序列的3种样本的一致性均高于99％，同时干血斑样本的耐药性检测结果与血浆相比一致性为86％（31／36），突变位点不一致的主要原因为混合碱基导致。结论综合PCR扩增效果及序列分析差异等因素，十血斑样本可以反映出耐药的整体流行趋势，用于我国HIV-1患者的耐药检测及监测值得推广。Objective This study aimed at exploring the feasibility of using dried blood spots （DBS） to detect HIV drug resistance genotyping in China by comparing the results of drug resistance from DBS,plasma and whole blood samples. Methods Blood samples were collected from 39 AIDS patients from Anhui （ 10）, Yunnan （ 13 ）, Hunan （6） and Xinjiang （10） provinces and autonomous regions. The HIV strains that infected these patients covered all the major HIV-1 subtypes prevailing in China （ B, CRF01_AE, CRF07_BC）. HIV drug resistance genotyping assay was performed on DBS as well as on the whole blood and plasma samples from the same patients simultaneously by using an in-house nest RT-PCR method. Drug resistance levels were determined based on Stanford University HIV drug resistance database,and the results from these three types of samples were compared. Results The percentages of successful amplification of protease and reverse transcriptase regions in the pol gene were 95% （37/39） from DBS,92% （36/39） from whole blood and 100% （39/39） from plasma samples. The sequences from the three types of samples showed more than 99% identity. 86% （31/36） of the DBS samples had the same set of drug resistance mutations as those which were detected from plasma samples. The differences probably resulted from mixed bases. Conclusions There was no major difference in detecting HIV drug resistance genotyping among DBS,plasma and whole blood samples. Therefore, DBS is useful for detection of HIV drug resistance genotyping and is particularly valuable in developing countries like China,especially in remote rural regions.

关 键 词：HIV-1 干血斑 基因型耐药性检测 聚合酶链反应 

分 类 号：R686[医药卫生—骨科学]