C亚型SHIV-1157ipd3N4中国恒河猴适应性感染及变异分析
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摘要
目前,以C亚型为主的HIV-1仍然在世界上广泛流行,迫切需要研制出安全有效的艾滋病疫苗,进而需要合适的艾滋病动物模型对疫苗进行评价。C亚型SHIV和相应非人灵长类模型是研究HIV-1感染和发病机制及评价以C亚型env区为靶点的候选疫苗功效的有效工具。
为了建立合适的C亚型SHIV动物模型进行候选疫苗评价,本研究选择了R. M.Ruprecht教授构建的SHIV-1157ipd3N4,这株SHIV编码HIV-1 C亚型env基因,具有高度复制能力和单一CCR5辅助受体嗜性,能够粘膜途径感染恒河猴并诱发AIDS类似症状,而且感染猴缓慢的疾病进程与人类感染HIV-1相似。本研究首先制备了SHIV-1157ipd3N4中国恒河猴细胞适应株,发现其序列与SHIV-1157ipd3N4克隆株的相似度为99%。随后,为了向后续动物模型接种剂量提供参考,又对SHIV-1157ipd3N4中国恒河猴细胞适应株和印度恒河猴细胞适应株进行了细胞水平评价,结果前者病毒浓度为500TCID50/ml,而后者为1000TCID50/ml。
模拟血液传播和性传播方式建立SHIV-1157ipd3N4/中国恒河猴模型能够反映艾滋病的传播方式,本研究分别应用SHIV-1157ipd3N4中国恒河猴细胞适应株静脉和直肠途径感染中国恒河猴,SHIV-1157ipd3N4印度恒河猴细胞适应株阴道途径感染中国恒河猴。同时为了更好的反映HIV在人群中自然传播方式,直肠和阴道途径模型使用了小剂量多次的感染方式。结果SHIV-1157ipd3N4中国恒河猴细胞适应株能够通过静脉和直肠途径成功感染中国恒河猴,而SHIV-1157ipd3N4印度恒河猴细胞适应株也成功地通过阴道途径感染中国恒河猴,建立了三种途径的SHIV-1157ipd3N4/中国恒河猴模型。
随后,本研究比较了不同感染途径动物模型间的生物学特性、易感性和发病规律。结果发现三种途径建立的SHIV-1157ipd3N4/中国恒河猴模型生物学特性存在差异。血浆病毒RNA载量方面,静脉组感染猴在21d时出现峰值后迅速下降进入平台期,而直肠组和阴道组感染猴血浆病毒RNA载量阳转时间和峰值时间各不相同,部分感染猴的峰值不明显,全部小剂量感染结束后逐渐进入平台期；外周血CD4+/CD8+T淋巴细胞比值方面,静脉组感染猴21d时均出现下降,随后迅速恢复正常,而直肠组和阴道组感染猴的比值在小剂量多次感染过程中呈现多次小幅度波动,部分猴出现明显下降过程,三组感染猴的比值随着感染时间延长总体上逐渐下降；静脉组和直肠组在整个感染过程中外周血CD4+T淋巴细胞绝对数逐渐上升而阴道组基本保持稳定；三组感染猴血浆结合抗体滴度稳定上升,后期静脉组明显高于直肠组和阴道组。中国恒河猴三种途径对SHIV-1157ipd3N4的易感性不同,静脉途径接种50 TCID50和5 TCID50SHIV-1157ipd3N4中国恒河猴细胞适应株能够成功感染,而0.5 TCID50和0.05 TCID50未感染。直肠和阴道途径分别小剂量接种13次和11次后使实验猴全部感染。至感染后49w止,所有感染猴都未出现明显类似艾滋病的临床症状。
另外,为了揭示艾滋病病毒的变异规律,本研究又进一步对SHIV-1157ipd3N4经不同途径感染中国恒河猴后的变异趋势进行了探索。发现短期内从感染猴血浆中扩增出的gp120碱基和氨基酸序列只存在少数位点的改变,没有N糖基化位点的变化,也没有出现明显变异趋势。
在制备病毒株的过程中,为了证明HIV-1 p24 ELISA试剂盒检测SIV p27抗原结果的可靠性,本研究比较了HIV-1 p24和SIV p27两种ELISA试剂盒对病毒液和血浆中SIV p27抗原检测结果。结果显示HIV-1p24和SIV p27两种ELISA试剂盒检测SIV p27抗原的灵敏度分别是150pg/mL和62.5 pg/mL;两种试剂盒检测病毒液和血浆中SIV p27抗原的定性结果一致；定量结果的统计分析得出病毒液的直线回归决定系数R2=0.857,直线相关系数r=0.926,P<0.01；血浆的直线回归决定系数R2=0.512,直线相关系数r=0.716,P<0.05。
综上所述,本课题首先制备了SHIV-1157ipd3N4中国恒河猴细胞适应株,然后模拟人类自然感染HIV方式和三种主要感染途径分别建立了静脉、直肠和阴道途径SHIV-1157ipd3N4/中国恒河猴模型,并比较不同感染途径动物模型间的生物学特性、易感染性和发病规律,进一步对病毒经不同途径感染中国恒河猴后的变异趋势进行了探索,另外比较了HIV-1 p24和SIV p27两种ELISA试剂盒检测SIV P27抗原结果。这些研究为揭示HIV-1感染和发病机制提供了参考,并为以C亚型env区为靶点的候选疫苗评价提供了有效工具。
Currently. HIV-1 clade C is the most prevalent clade in the world. The safety and effectiveness of AIDS vaccine need ideal animal model for evaluation. The nonhuman primate models made by using SHIV subtype C are useful tools to investigate HIV infection and pathogenesis and evaluate candidate vaccines targeting HIV-1 clade C env.
In order to estabilish an appropriate animal model for evaluating candidate vaccines, SHIV-1157ipd3N4 that constructed by Dr. R. M. Ruprecht was selected in our study. SHIV-1157ipd3N4 encoding env of HIV-1 clade C shows high replication and unique tropsim of CCR5 coreceptor. this chimeric virus is also able to infect rhesus macaque through mucosa and cause symptoms like AIDS eventually. The model of SHIV-1157ipd3N4/rhesus macaque characterrized a slow disease progression that was similar to human HIV infection. First, in this study the SHIV-1157ipd3N4 adapted stock was prepared with PBMCs from Chinese-original rhesus macaque, and we found that sequence of the stock shared 99% similarity with molecular clone of SHIV-1157ipd3N4. For inoculation dose of following models, the titers of SHIV-1157ipd3N4 and its adapted stock with PBMCs from Chinese-original monkeys were measured and the titers were 1000 TCID50/ml and 500 TCID50/ml. respectively.
Establishing models of SHIV-1157ipd3N4/Chinese-original rhesus macaque by the routes of blood and sexual transmission could simulate modes of AIDS transmission, we infected Chinese-original rhesus macaques intravenously and intrarectally with SHIV-1157ipd3N4 harvested from PBMCs from Chinese-original rhesus macaque, and infected Chinese-original rhesus macaques intravaginally with SHIV-1157ipd3N4 prepared from PBMCs of Indian-original rhesus macaque. Meanwhile, in order to closely reflect HIV natural transmission in human, method of repeate-low-dose was used in intravaginal and intrarectal model. Resultes indicated that Chinese-original rhesus macaques were successfully infected intravenously and intrarectally with SHIV-1157ipd3N4 from PBMCs from Chinese-original rhesus macaque, and intravaginally with original SHIV-1157ipd3N4. so that these three routes for models of SHIV-1157ipd3N4/Chinese-original rhesus macaque were established.
Subsequently, biological characteristics and susceptibility of disease were compared among three models. Resultes suggested that there were differences in biological characteristics. In plasma viral RNA load aspect, infectious monkeys of intravenous group appereced peak value at 21 days post inoculation and then rapidly declined into low level, while those of intrarectal and intraviganal groups had various time points that turned positive and appeared peak value, peak values of some of infectious monkeys were not obvious, viral loads declined into low level after all low dose inoculation; In the aspect of ratio of CD4+/CD8+T cell in blood, infected monkeys of intravenous group reduced at 21 days post inoculation and then rapidly recovered, while those of intrarectal and intraviganal groups in process of repeated low dose inoculation showed small curves, and some of infectious monkeys were obviously reduced, generally, ratios of three groups gradually decreased with prolonged time; In the aspect of absolute CD4+T cells in blood, intravenous and intrarectal groups gradually increased and intraviganal group kept stable. In plasma antidody titer aspect, three groups gradually rised, intravenous group were obviously higher than that of intrarectal and intraviganal groups at later period. Susceptbility of Chinese-original rhesus macaque infected SHIV-1157ipd3N4 by three different ways differed from each other,50 TCID50 and 5 TCID50 SHIV-1157ipd3N4 from PBMCs of Chinese-original rhesus macaque successfully infected intravenously but 0.5 TCID50 and 0.05 TCID50 failed. Low-dose intrarectal for 13 times and intraviganal for 11 times inoculation casued all the monkeys infected. There was no obvious symptom like AIDS in all monkeys in 49w post inoculation.
In order to reveal the variation of HIV. this study futhter discovered the variation trends of Chinese-original rhesus macaque infected with SHIV-1157ipd3N4 through different ways. We found that there were only a small number of site changes in gp120 nucleotide and amino acid sequences. There was no N glycosylation site change and no obvious variation trend.
In the process of preparation virus stock, for the purpose of proving reliability of results of detecting SIV p27 antigen got from HIV-1 p24 ELISA kit. the results of detecting SIV p27 antigen in plasma and virus stocks by HIV-1 p24 and SIV p27 ELISA kit were compared in this research. Results showed that sensitivity of detecting SIV p27 antigen was 150 pg/mL by HIV-1 p24 ELISA kit and 62.5 pg/mL by SIV p27 ELISA kit. respectively. The qualitative results obtained by HIV-1 p24 kit consisted with those of SIV p27 kit on detecting SIV p27 antigen in virus stock and plasma. The statistical analysis of quantitative results indicated linear regression R2=0.857. linear correlation r=0.926. P<0.01 in virus stock, while R:=0.512. r=0.716. P<0.05 in plasma.
In summary, SHIV-1157ipd3N4 adapted stock with PBMCs from Chinese-original rhesus macaque was prepared firstly in this study, and then intravenous, intrarectal and intraviganal models of SHIV-1157ipd3N4/Chinese-original rhesus macaque to simulate the three main HIV transmission ways and natural transmission in human. Moreover, biological characteristics and susceptibility of disease were compared among three models, and tried to find the variation trend of Chinese-original rhesus macaque infected with SHIV-1157ipd3N4 through different ways. In addition, the results of detecting SIV p27 antigen in plasma and virus stocks by HIV-1 p24 and SIV p27 ELISA kit were compared. These studies will provide information for HIV infection and pathogenesis and evaluating effectiveness of candidate vaccines targeting HIV-1 clade C env.
为了建立合适的C亚型SHIV动物模型进行候选疫苗评价,本研究选择了R. M.Ruprecht教授构建的SHIV-1157ipd3N4,这株SHIV编码HIV-1 C亚型env基因,具有高度复制能力和单一CCR5辅助受体嗜性,能够粘膜途径感染恒河猴并诱发AIDS类似症状,而且感染猴缓慢的疾病进程与人类感染HIV-1相似。本研究首先制备了SHIV-1157ipd3N4中国恒河猴细胞适应株,发现其序列与SHIV-1157ipd3N4克隆株的相似度为99%。随后,为了向后续动物模型接种剂量提供参考,又对SHIV-1157ipd3N4中国恒河猴细胞适应株和印度恒河猴细胞适应株进行了细胞水平评价,结果前者病毒浓度为500TCID50/ml,而后者为1000TCID50/ml。
模拟血液传播和性传播方式建立SHIV-1157ipd3N4/中国恒河猴模型能够反映艾滋病的传播方式,本研究分别应用SHIV-1157ipd3N4中国恒河猴细胞适应株静脉和直肠途径感染中国恒河猴,SHIV-1157ipd3N4印度恒河猴细胞适应株阴道途径感染中国恒河猴。同时为了更好的反映HIV在人群中自然传播方式,直肠和阴道途径模型使用了小剂量多次的感染方式。结果SHIV-1157ipd3N4中国恒河猴细胞适应株能够通过静脉和直肠途径成功感染中国恒河猴,而SHIV-1157ipd3N4印度恒河猴细胞适应株也成功地通过阴道途径感染中国恒河猴,建立了三种途径的SHIV-1157ipd3N4/中国恒河猴模型。
随后,本研究比较了不同感染途径动物模型间的生物学特性、易感性和发病规律。结果发现三种途径建立的SHIV-1157ipd3N4/中国恒河猴模型生物学特性存在差异。血浆病毒RNA载量方面,静脉组感染猴在21d时出现峰值后迅速下降进入平台期,而直肠组和阴道组感染猴血浆病毒RNA载量阳转时间和峰值时间各不相同,部分感染猴的峰值不明显,全部小剂量感染结束后逐渐进入平台期；外周血CD4+/CD8+T淋巴细胞比值方面,静脉组感染猴21d时均出现下降,随后迅速恢复正常,而直肠组和阴道组感染猴的比值在小剂量多次感染过程中呈现多次小幅度波动,部分猴出现明显下降过程,三组感染猴的比值随着感染时间延长总体上逐渐下降；静脉组和直肠组在整个感染过程中外周血CD4+T淋巴细胞绝对数逐渐上升而阴道组基本保持稳定；三组感染猴血浆结合抗体滴度稳定上升,后期静脉组明显高于直肠组和阴道组。中国恒河猴三种途径对SHIV-1157ipd3N4的易感性不同,静脉途径接种50 TCID50和5 TCID50SHIV-1157ipd3N4中国恒河猴细胞适应株能够成功感染,而0.5 TCID50和0.05 TCID50未感染。直肠和阴道途径分别小剂量接种13次和11次后使实验猴全部感染。至感染后49w止,所有感染猴都未出现明显类似艾滋病的临床症状。
另外,为了揭示艾滋病病毒的变异规律,本研究又进一步对SHIV-1157ipd3N4经不同途径感染中国恒河猴后的变异趋势进行了探索。发现短期内从感染猴血浆中扩增出的gp120碱基和氨基酸序列只存在少数位点的改变,没有N糖基化位点的变化,也没有出现明显变异趋势。
在制备病毒株的过程中,为了证明HIV-1 p24 ELISA试剂盒检测SIV p27抗原结果的可靠性,本研究比较了HIV-1 p24和SIV p27两种ELISA试剂盒对病毒液和血浆中SIV p27抗原检测结果。结果显示HIV-1p24和SIV p27两种ELISA试剂盒检测SIV p27抗原的灵敏度分别是150pg/mL和62.5 pg/mL;两种试剂盒检测病毒液和血浆中SIV p27抗原的定性结果一致；定量结果的统计分析得出病毒液的直线回归决定系数R2=0.857,直线相关系数r=0.926,P<0.01；血浆的直线回归决定系数R2=0.512,直线相关系数r=0.716,P<0.05。
综上所述,本课题首先制备了SHIV-1157ipd3N4中国恒河猴细胞适应株,然后模拟人类自然感染HIV方式和三种主要感染途径分别建立了静脉、直肠和阴道途径SHIV-1157ipd3N4/中国恒河猴模型,并比较不同感染途径动物模型间的生物学特性、易感染性和发病规律,进一步对病毒经不同途径感染中国恒河猴后的变异趋势进行了探索,另外比较了HIV-1 p24和SIV p27两种ELISA试剂盒检测SIV P27抗原结果。这些研究为揭示HIV-1感染和发病机制提供了参考,并为以C亚型env区为靶点的候选疫苗评价提供了有效工具。
Currently. HIV-1 clade C is the most prevalent clade in the world. The safety and effectiveness of AIDS vaccine need ideal animal model for evaluation. The nonhuman primate models made by using SHIV subtype C are useful tools to investigate HIV infection and pathogenesis and evaluate candidate vaccines targeting HIV-1 clade C env.
In order to estabilish an appropriate animal model for evaluating candidate vaccines, SHIV-1157ipd3N4 that constructed by Dr. R. M. Ruprecht was selected in our study. SHIV-1157ipd3N4 encoding env of HIV-1 clade C shows high replication and unique tropsim of CCR5 coreceptor. this chimeric virus is also able to infect rhesus macaque through mucosa and cause symptoms like AIDS eventually. The model of SHIV-1157ipd3N4/rhesus macaque characterrized a slow disease progression that was similar to human HIV infection. First, in this study the SHIV-1157ipd3N4 adapted stock was prepared with PBMCs from Chinese-original rhesus macaque, and we found that sequence of the stock shared 99% similarity with molecular clone of SHIV-1157ipd3N4. For inoculation dose of following models, the titers of SHIV-1157ipd3N4 and its adapted stock with PBMCs from Chinese-original monkeys were measured and the titers were 1000 TCID50/ml and 500 TCID50/ml. respectively.
Establishing models of SHIV-1157ipd3N4/Chinese-original rhesus macaque by the routes of blood and sexual transmission could simulate modes of AIDS transmission, we infected Chinese-original rhesus macaques intravenously and intrarectally with SHIV-1157ipd3N4 harvested from PBMCs from Chinese-original rhesus macaque, and infected Chinese-original rhesus macaques intravaginally with SHIV-1157ipd3N4 prepared from PBMCs of Indian-original rhesus macaque. Meanwhile, in order to closely reflect HIV natural transmission in human, method of repeate-low-dose was used in intravaginal and intrarectal model. Resultes indicated that Chinese-original rhesus macaques were successfully infected intravenously and intrarectally with SHIV-1157ipd3N4 from PBMCs from Chinese-original rhesus macaque, and intravaginally with original SHIV-1157ipd3N4. so that these three routes for models of SHIV-1157ipd3N4/Chinese-original rhesus macaque were established.
Subsequently, biological characteristics and susceptibility of disease were compared among three models. Resultes suggested that there were differences in biological characteristics. In plasma viral RNA load aspect, infectious monkeys of intravenous group appereced peak value at 21 days post inoculation and then rapidly declined into low level, while those of intrarectal and intraviganal groups had various time points that turned positive and appeared peak value, peak values of some of infectious monkeys were not obvious, viral loads declined into low level after all low dose inoculation; In the aspect of ratio of CD4+/CD8+T cell in blood, infected monkeys of intravenous group reduced at 21 days post inoculation and then rapidly recovered, while those of intrarectal and intraviganal groups in process of repeated low dose inoculation showed small curves, and some of infectious monkeys were obviously reduced, generally, ratios of three groups gradually decreased with prolonged time; In the aspect of absolute CD4+T cells in blood, intravenous and intrarectal groups gradually increased and intraviganal group kept stable. In plasma antidody titer aspect, three groups gradually rised, intravenous group were obviously higher than that of intrarectal and intraviganal groups at later period. Susceptbility of Chinese-original rhesus macaque infected SHIV-1157ipd3N4 by three different ways differed from each other,50 TCID50 and 5 TCID50 SHIV-1157ipd3N4 from PBMCs of Chinese-original rhesus macaque successfully infected intravenously but 0.5 TCID50 and 0.05 TCID50 failed. Low-dose intrarectal for 13 times and intraviganal for 11 times inoculation casued all the monkeys infected. There was no obvious symptom like AIDS in all monkeys in 49w post inoculation.
In order to reveal the variation of HIV. this study futhter discovered the variation trends of Chinese-original rhesus macaque infected with SHIV-1157ipd3N4 through different ways. We found that there were only a small number of site changes in gp120 nucleotide and amino acid sequences. There was no N glycosylation site change and no obvious variation trend.
In the process of preparation virus stock, for the purpose of proving reliability of results of detecting SIV p27 antigen got from HIV-1 p24 ELISA kit. the results of detecting SIV p27 antigen in plasma and virus stocks by HIV-1 p24 and SIV p27 ELISA kit were compared in this research. Results showed that sensitivity of detecting SIV p27 antigen was 150 pg/mL by HIV-1 p24 ELISA kit and 62.5 pg/mL by SIV p27 ELISA kit. respectively. The qualitative results obtained by HIV-1 p24 kit consisted with those of SIV p27 kit on detecting SIV p27 antigen in virus stock and plasma. The statistical analysis of quantitative results indicated linear regression R2=0.857. linear correlation r=0.926. P<0.01 in virus stock, while R:=0.512. r=0.716. P<0.05 in plasma.
In summary, SHIV-1157ipd3N4 adapted stock with PBMCs from Chinese-original rhesus macaque was prepared firstly in this study, and then intravenous, intrarectal and intraviganal models of SHIV-1157ipd3N4/Chinese-original rhesus macaque to simulate the three main HIV transmission ways and natural transmission in human. Moreover, biological characteristics and susceptibility of disease were compared among three models, and tried to find the variation trend of Chinese-original rhesus macaque infected with SHIV-1157ipd3N4 through different ways. In addition, the results of detecting SIV p27 antigen in plasma and virus stocks by HIV-1 p24 and SIV p27 ELISA kit were compared. These studies will provide information for HIV infection and pathogenesis and evaluating effectiveness of candidate vaccines targeting HIV-1 clade C env.
引文
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[10]Siddappa NB, Song R, Kramer VG, Chenine AL, Velu V, Ong H, et al. Neutralization-sensitive R5-tropic simian-human immunodeficiency virus SHIV-2873Nip, which carries env isolated from an infant with a recent HIV clade C infection. J Virol 2009 Feb;83(3):1422-32.
[11]Siddappa NB, Watkins JD, Wassermann KJ, Song R, Wang W, Kramer VG, et al. R5 clade C SHIV strains with tier 1 or 2 neutralization sensitivity:tools to dissect env evolution and to develop AIDS vaccines in primate models. PLoS One;5(7):e11689.
[12]丛喆,刘浩,秦川等.B亚型SHIVSF162p3中国恒河猴小剂量多次阴道黏膜感染[J].中国比较医学杂志.2011,21(2)：44-48.
[13]Chenine AL, Siddappa NB, Kramer VG, Sciaranghella G, Rasmussen RA, Lee SJ, et al. Relative transmissibility of an R5 clade C simian-human immunodeficiency virus across different mucosae in macaques parallels the relative risks of sexual HIV-1 transmission in humans via different routes. J Infect Dis Apr 15;201(8):1155-63.
[14]Chen Z, Zhao X, Huang Y, Gettie A, Ba L, Blanchard J, et al. CD4+lymphocytopenia in acute infection of Asian macaques by a vaginally transmissible subtype-C, CCR5-tropic Simian/Human Immunodeficiency Virus (SHIV). J Acquir Immune Defic Syndr 2002 Jun 1;30(2):133-45.
[15]Humbert M, Rasmussen RA, Song R, Ong H, Sharma P, Chenine AL, et al. SHIV-1157i and passaged progeny viruses encoding R5 HTV-1 clade C env cause AIDS in rhesus monkeys. Retrovirology 2008;5:94.
[16]Polacino P, Larsen K, Galmin L, Suschak J, Kraft Z, Stamatatos L, et al. Differential pathogenicity of SHIV infection in pig-tailed and rhesus macaques. J Med Primatol 2008 Dec;37 Suppl 2:13-23.
[17]Ho O, Larsen K, Polacino P, Li Y, Anderson D, Song R, et al. Pathogenic infection of Macaca nemestrina with a CCR5-tropic subtype-C simian-human immunodeficiency virus. Retrovirology 2009;6:65.
[18]Jay A. Levy著；邵一鸣译.艾滋病病毒与艾滋病的发病机制[M].北京：科学出版社,2010：28-49
[19]Chackerian B, Rudensey LM, Overbaugh J. Specific N-linked and O-linked glycosylation modifications in the envelope V1 domain of simian immunodeficiency virus variants that evolve in the host alter recognition by neutralizing antibodies. J Virol 1997 Oct;71(10):7719-27.
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[21]Blay WM, Gnanakaran S, Foley B, Doria-Rose NA, Korber BT, Haigwood NL. Consistent patterns of change during the divergence of human immunodeficiency virus type 1 envelope from that of the inoculated virus in simian/human immunodeficiency virus-infected macaques. J Virol 2006 Jan;80(2):999-1014.
[22]Tso FY, Hoffmann FG, Tully DC, Lemey P, Rasmussen RA, Zhang H, et al. A comparative study of HIV-1 clade C env evolution in a Zambian infant with an infected rhesus macaque during disease progression. AIDS 2009 Sep 10;23(14):1817-28.
[23]Imamichi T, Murphy MA, Adelsberger JW, Yang J, Watkins CM, Berg SC, et al. Actinomycin D induces high-level resistance to thymidine analogs in replication of human immunodeficiency virus type 1 by interfering with host cell thymidine kinase expression. J Virol 2003 Jan;77(2):1011-20.
[24]Nakayama EE, Miyoshi H, Nagai Y, Shioda T. A specific region of 37 amino acid residues in the SPRY (B30.2) domain of African green monkey TRIMSalpha determines species-specific restriction of simian immunodeficiency virus SIVmac infection. J Virol 2005 Jul;79(14):8870-7.
[25]Wei Q, Stallworth JW, Vance PJ, Hoxie JA, Fultz PN. Simian immunodeficiency virus (SIV)/immunoglobulin G immune complexes in SIV-infected macaques block detection of CD16 but not cytolytic activity of natural killer cells. Clin Vaccine Immunol 2006 Jul;13(7):768-78.
[26]Workman S, Wells SK, Pau CP, Owen SM, Dong XF, LaBorde R, et al. Rapid detection of HIV-1 p24 antigen using magnetic immuno-chromatography (MICT). J Virol Methods 2009 Sep;160(1-2):14-21.
[27]Sutthent R, Gaudart N, Chokpaibulkit K, Tanliang N, Kanoksinsombath C, Chaisilwatana P. p24 Antigen detection assay modified with a booster step for diagnosis and monitoring of human immunodeficiency virus type 1 infection. J Clin Microbiol 2003 Mar;41(3):1016-22.
[28]刘强,杨贵波,邵一鸣等.运用实时荧光定量RT-PCR和ELISA方法定量检测SHIV的对比分析[J].中国艾滋病性病.2006,12(4)：300-310.
[29]Jay A. Levy著；邵一鸣译.艾滋病病毒与艾滋病的发病机制[M].北京：科学出版社,2010：1-21
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[32]Wykrzykowska JJ, Rosenzweig M, Veazey RS, Simon MA, Halvorsen K, Desrosiers RC, et al. Early regeneration of thymic progenitors in rhesus macaques infected with simian immunodeficiency virus. J Exp Med 1998 Jun 1;187(11):1767-78.
[33]Diop OM, Gueye A, Dias-Tavares M, Kornfeld C, Faye A, Ave P, et al. High levels of viral replication during primary simian immunodeficiency virus SIVagm infection are rapidly and strongly controlled in African green monkeys. J Virol 2000 Aug;74(16):7538-47.
[34]Pandrea I, Apetrei C, Dufour J, Dillon N, Barbercheck J, Metzger M, et al. Simian immunodeficiency virus SIVagm.sab infection of Caribbean African green monkeys:a new model for the study of SIV pathogenesis in natural hosts. J Virol 2006 May;80(10):4858-67.
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[8]Reynolds MR, Weiler AM, Piaskowski SM, Kolar HL, Hessell AJ, Weiker M, et al. Macaques vaccinated with simian immunodeficiency virus SIVmac239Delta nef delay acquisition and control replication after repeated low-dose heterologous SIV challenge. J Virol Sep;84(18):9190-9.
[9]Hessell AJ, Poignard P, Hunter M, Hangartner L, Tehrani DM, Bleeker WK, et al. Effective, low-titer antibody protection against low-dose repeated mucosal SHIV challenge in macaques. Nat Med 2009 Aug;15(8):951-4.
[10]Siddappa NB, Song R, Kramer VG, Chenine AL, Velu V, Ong H, et al. Neutralization-sensitive R5-tropic simian-human immunodeficiency virus SHIV-2873Nip, which carries env isolated from an infant with a recent HIV clade C infection. J Virol 2009 Feb;83(3):1422-32.
[11]Siddappa NB, Watkins JD, Wassermann KJ, Song R, Wang W, Kramer VG, et al. R5 clade C SHIV strains with tier 1 or 2 neutralization sensitivity:tools to dissect env evolution and to develop AIDS vaccines in primate models. PLoS One;5(7):e11689.
[12]丛喆,刘浩,秦川等.B亚型SHIVSF162p3中国恒河猴小剂量多次阴道黏膜感染[J].中国比较医学杂志.2011,21(2)：44-48.
[13]Chenine AL, Siddappa NB, Kramer VG, Sciaranghella G, Rasmussen RA, Lee SJ, et al. Relative transmissibility of an R5 clade C simian-human immunodeficiency virus across different mucosae in macaques parallels the relative risks of sexual HIV-1 transmission in humans via different routes. J Infect Dis Apr 15;201(8):1155-63.
[14]Chen Z, Zhao X, Huang Y, Gettie A, Ba L, Blanchard J, et al. CD4+lymphocytopenia in acute infection of Asian macaques by a vaginally transmissible subtype-C, CCR5-tropic Simian/Human Immunodeficiency Virus (SHIV). J Acquir Immune Defic Syndr 2002 Jun 1;30(2):133-45.
[15]Humbert M, Rasmussen RA, Song R, Ong H, Sharma P, Chenine AL, et al. SHIV-1157i and passaged progeny viruses encoding R5 HTV-1 clade C env cause AIDS in rhesus monkeys. Retrovirology 2008;5:94.
[16]Polacino P, Larsen K, Galmin L, Suschak J, Kraft Z, Stamatatos L, et al. Differential pathogenicity of SHIV infection in pig-tailed and rhesus macaques. J Med Primatol 2008 Dec;37 Suppl 2:13-23.
[17]Ho O, Larsen K, Polacino P, Li Y, Anderson D, Song R, et al. Pathogenic infection of Macaca nemestrina with a CCR5-tropic subtype-C simian-human immunodeficiency virus. Retrovirology 2009;6:65.
[18]Jay A. Levy著；邵一鸣译.艾滋病病毒与艾滋病的发病机制[M].北京：科学出版社,2010：28-49
[19]Chackerian B, Rudensey LM, Overbaugh J. Specific N-linked and O-linked glycosylation modifications in the envelope V1 domain of simian immunodeficiency virus variants that evolve in the host alter recognition by neutralizing antibodies. J Virol 1997 Oct;71(10):7719-27.
[20]Sagar M, Wu X, Lee S, Overbaugh J. Human immunodeficiency virus type 1 V1-V2 envelope loop sequences expand and add glycosylation sites over the course of infection, and these modifications affect antibody neutralization sensitivity. J Virol 2006 Oct;80(19):9586-98.
[21]Blay WM, Gnanakaran S, Foley B, Doria-Rose NA, Korber BT, Haigwood NL. Consistent patterns of change during the divergence of human immunodeficiency virus type 1 envelope from that of the inoculated virus in simian/human immunodeficiency virus-infected macaques. J Virol 2006 Jan;80(2):999-1014.
[22]Tso FY, Hoffmann FG, Tully DC, Lemey P, Rasmussen RA, Zhang H, et al. A comparative study of HIV-1 clade C env evolution in a Zambian infant with an infected rhesus macaque during disease progression. AIDS 2009 Sep 10;23(14):1817-28.
[23]Imamichi T, Murphy MA, Adelsberger JW, Yang J, Watkins CM, Berg SC, et al. Actinomycin D induces high-level resistance to thymidine analogs in replication of human immunodeficiency virus type 1 by interfering with host cell thymidine kinase expression. J Virol 2003 Jan;77(2):1011-20.
[24]Nakayama EE, Miyoshi H, Nagai Y, Shioda T. A specific region of 37 amino acid residues in the SPRY (B30.2) domain of African green monkey TRIMSalpha determines species-specific restriction of simian immunodeficiency virus SIVmac infection. J Virol 2005 Jul;79(14):8870-7.
[25]Wei Q, Stallworth JW, Vance PJ, Hoxie JA, Fultz PN. Simian immunodeficiency virus (SIV)/immunoglobulin G immune complexes in SIV-infected macaques block detection of CD16 but not cytolytic activity of natural killer cells. Clin Vaccine Immunol 2006 Jul;13(7):768-78.
[26]Workman S, Wells SK, Pau CP, Owen SM, Dong XF, LaBorde R, et al. Rapid detection of HIV-1 p24 antigen using magnetic immuno-chromatography (MICT). J Virol Methods 2009 Sep;160(1-2):14-21.
[27]Sutthent R, Gaudart N, Chokpaibulkit K, Tanliang N, Kanoksinsombath C, Chaisilwatana P. p24 Antigen detection assay modified with a booster step for diagnosis and monitoring of human immunodeficiency virus type 1 infection. J Clin Microbiol 2003 Mar;41(3):1016-22.
[28]刘强,杨贵波,邵一鸣等.运用实时荧光定量RT-PCR和ELISA方法定量检测SHIV的对比分析[J].中国艾滋病性病.2006,12(4)：300-310.
[29]Jay A. Levy著；邵一鸣译.艾滋病病毒与艾滋病的发病机制[M].北京：科学出版社,2010：1-21
[30]Norley SG, Kraus G, Ennen J, Bonilla J, Konig H, Kurth R. lmmunological studies of the basis for the apathogenicity of simian immunodeficiency virus from African green monkeys. Proc Natl Acad Sci U S A 1990 Nov;87(22):9067-71.
[31]Chakrabarti L, Baptiste V, Khatissian E, Cumont MC, Aubertin AM, Montagnier L, et al. Limited viral spread and rapid immune response in lymph nodes of macaques inoculated with attenuated simian immunodeficiency virus. Virology 1995 Nov 10;213(2):535-48.
[32]Wykrzykowska JJ, Rosenzweig M, Veazey RS, Simon MA, Halvorsen K, Desrosiers RC, et al. Early regeneration of thymic progenitors in rhesus macaques infected with simian immunodeficiency virus. J Exp Med 1998 Jun 1;187(11):1767-78.
[33]Diop OM, Gueye A, Dias-Tavares M, Kornfeld C, Faye A, Ave P, et al. High levels of viral replication during primary simian immunodeficiency virus SIVagm infection are rapidly and strongly controlled in African green monkeys. J Virol 2000 Aug;74(16):7538-47.
[34]Pandrea I, Apetrei C, Dufour J, Dillon N, Barbercheck J, Metzger M, et al. Simian immunodeficiency virus SIVagm.sab infection of Caribbean African green monkeys:a new model for the study of SIV pathogenesis in natural hosts. J Virol 2006 May;80(10):4858-67.