福建省艾滋病患者抗病毒疗效和耐药性检测分析

Evaluation of the effectiveness of anti-retroviral therapy on AIDS patients in Fujian province and the current status of their drug-resistance pattern

目的了解我省艾滋病患者的抗病毒疗效与基因耐药性突变发生情况,为抗病毒治疗提供依据。方法采集我省25例接受抗病毒治疗的艾滋病患者外周静脉抗凝全血进行CD4+T淋巴细胞计数,分离血浆进行HIV病毒载量检测,并提取血浆标本中HIVRNA,用RT-PCR方法扩增HIVpol区蛋白酶基因全序列与部分逆转录酶序列。所得扩增片段进行序列测定后,数据运用ContigExpress编辑校正后上传至斯坦福大学HIV耐药数据库进行比对,同时运用Bio-Edit、Clustal和Mega等软件进行系统发生分析。结果本研究中的17例(68%)患者的病毒载量在1000cp/ml以下,其中40%(10例)降至检测限之下。RT-PCR获得10例患者的扩增片段。对其序列亚型测试结果表明:7例属于HIV-1AE亚型,2例HIV-1B亚型,1例为HIV-1D亚型;耐药性分析表明:10份序列对PI(蛋白酶抑制剂)药物均表现为敏感,8份序列对NRTI(核苷类逆转录酶抑制剂)及NNRTI(非核苷类逆转录酶抑制剂)表现出至少对3种药物高度耐药,最常见的突变位点为G190A、K103N和M184V,造成对NVP、3TC的高度耐药。结论目前我省艾滋病患者的抗病毒治疗取得预期效果。随着治疗时间延长,患者免疫功能也逐渐恢复,但耐药性突变发生种类和数量有所增加,出现对现有各类一线药物不同程度的耐药。为保证可持续性治疗策略的施行,需加强对抗病毒治疗患者的依从性教育,同时加强耐药性监测,及时评估治疗失败者耐药性情况,必要时提供二线药品,减少我省HIV病毒耐药株的产生与传播,保障抗病毒治疗的有效进行。

To evaluate the effectiveness of anti-retroviral therapy on AIDS patients in Fujian province and the current status of their drug-resistance pattern in order to provide experimental evidences for anti-reteroviral therapy, counts of the CD4＋ T lymphocytes in the peripheral blood of 25 AIDS patients receiving anti-retroviral therapy and the virus load of HIV in plasma of these patients were determined, in which the whole sequence of gene coding the pol protein of HIV and the partial reverse transeriptase gene were amplified by RT-PCR, and then the amplified fragments were edited by using Contig Express software and submitted to the website http//hivdb. Stanford. edu. / to analyze the drug-resistance status. Meanwhile, their phylogenesis was analyzed by Bio-Edi,clustal and Mega softwares. It was found that 68% （17/25） of the patients treated with anti-retroviral therapy showed the level of virus load （VL） below 1000 cp/ml and in 40% （10/25） of the treated patients the VL even dropped below the detectable limit（50 cp/ml）. The amplified fragments were obtained in 10 cases of patients treated by means of RT-PCR and their sequence subtype analysis demonstrated that 7 cases belonged to the HIV-1 AE, 2 cases were HIV- 1 B and one was HIV-1 D. As demonstrated by the results in drug-resistance analysis, the sequences obtained from 10 cases was found to be sensitive to the action of protease inhibitors（PI）, however, those from 8 cases were highly resistant to at least 3 anti-retroviral drugs of nucleotide reverse transcriptase inhibitors（NRTI） and non-nucleotide reverse transeriptase inhibitors （NNRTI）. The commonly observed loci of mutations were G190A, K103N and M184V, thus causing high or medium level of resistance to NVP and 3TC. From these observations,it is evident that the types and amount of developing mutations on drugresistance in AIDS patients during treatment with anti-retroviral therapy are definitely increased, although their immune function would be gradually recovered after treatment. The result of these will be the development of drug-resistanee to certain firstline drugs. Therefore, it is necessary to motivate patients to keep them highly compliance to the first line regimes and to have the second line regimes available besides performing timely evaluation of the drug-resistance of oatients.