Russo Cristiano Teodoro, Alkmim Wagner, Munerato Patricia, Zukurov Jean, Maricato Juliana T, Sucupira M Cecília, Diaz Ricardo S, Janini Luiz Mário
Discipline of Immunology, Medicine Course, Pontifical Catholic University of Paraná, Londrina Paraná, Brazil.
Intervirology. 2014;57(5):277-88. doi: 10.1159/000362415. Epub 2014 Jun 28.
Human immunodeficiency virus type 1 (HIV-1) genetic diversity is one of the most important features of HIV-1 infections and the result of error accumulation during reverse transcription and of high viral turnover. HIV-1 reverse transcription is influenced by factors such as the level of nucleotides and/or the cellular activation state. HIV-1 diversity was investigated after 48 h of viral propagation in peripheral blood mononuclear cells (PBMCs) obtained from healthy donors in three different cell culture conditions: (1) resting PBMCs, (2) simultaneous infection and PBMC activation, and (3) PBMC activation 72 h before infection. Cellular DNA was extracted and proviruses of each culture condition were amplified. Single-genome PCR clones were obtained and the protease and reverse transcriptase of the pol gene were sequenced. An elevated number of nucleotide substitutions in all three culture conditions were observed. In condition 1, the mutational rate observed ranged from 1.0 × 10(-3) to 2.1 × 10(-2), the genetic diversity was 0.6%, and hypermutation was observed in 7.1% of sequenced clones. In condition 2, the mutational rate ranged from 1.0 × 10(-3) to 1.0 × 10(-2), the genetic diversity was 0.8%, and hypermutation affected 6.7% of clones. In condition 3, the mutational rate ranged from 2.8 × 10(-3) to 1.1 × 10(-2), the genetic diversity was 1%, and 5.9% of clones were hypermutated. Substitutions occurred more frequently in some specific nucleotide stretches, and a common pattern for substitutions in all the different conditions was identified. There was a significant accumulation of mutations during the initial periods of in vitro HIV-1 propagation irrespective of culture conditions. The rapid accumulation of virus diversity might represent a viral strategy when colonizing new hosts. Complementary studies are necessary to allow for a better understanding of the initial periods of infection, which represent a crucial event related to disease progression.
1型人类免疫缺陷病毒（HIV-1）的基因多样性是HIV-1感染最重要的特征之一，是逆转录过程中错误积累以及病毒高周转率的结果。HIV-1逆转录受核苷酸水平和/或细胞活化状态等因素影响。在三种不同细胞培养条件下，对从健康供体获得的外周血单核细胞（PBMC）进行48小时病毒增殖后，研究了HIV-1的多样性：（1）静息PBMC，（2）同时感染和PBMC活化，以及（3）感染前72小时PBMC活化。提取细胞DNA并扩增每种培养条件下的前病毒。获得单基因组PCR克隆并对pol基因的蛋白酶和逆转录酶进行测序。在所有三种培养条件下均观察到核苷酸替换数量增加。在条件1中，观察到的突变率范围为1.0×10^(-3)至2.1×10^(-2)，基因多样性为0.6%，在7.1%的测序克隆中观察到超突变。在条件2中，突变率范围为1.0×10^(-3)至1.0×10^(-2)，基因多样性为0.8%，超突变影响6.7%的克隆。在条件3中，突变率范围为2.8×10^(-3)至1.1×10^(-2)，基因多样性为1%，5.9%的克隆发生超突变。替换在某些特定核苷酸片段中更频繁发生，并确定了所有不同条件下替换的共同模式。无论培养条件如何，在体外HIV-1增殖的初始阶段都会有显著的突变积累。病毒多样性的快速积累可能是病毒在定植新宿主时的一种策略。需要进行补充研究以更好地了解感染的初始阶段，这是与疾病进展相关的关键事件。
