Reitz M S, Hall L, Robert-Guroff M, Lautenberger J, Hahn B M, Shaw G M, Kong L I, Weiss S H, Waters D, Gallo R C
Laboratory of Tumor Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892.
AIDS Res Hum Retroviruses. 1994 Sep;10(9):1143-55. doi: 10.1089/aid.1994.10.1143.
Molecular clones of HIV-1 were obtained from isolates cultured from peripheral blood mononuclear cells (PBMCs) and directly from uncultured PBMCs from a laboratory worker accidentally infected with the HIV-1 laboratory strain, HIV-1(HTLV-IIIB). Envelope sequences corresponding to the first 752 amino acids of HIV-1(HTLV-IIIB) clone BH10 were obtained from clones of cultured virus and sequenced. Three env clones obtained shortly after infection differed among themselves at only seven nucleotide positions, resulting in one amino acid substitution and one frameshift mutation. These envelope sequences were as similar to the envelope sequences of various IIIB clones as the latter were to each other. env divergence increased over the course of infection. However, the overall diversity in env clones obtained two or more years after infection was still comparable to that among IIIB env clones from the original IIIB culture. Multiple clones of partial env gene sequences containing the V3 loop were also obtained directly from uncultured PBMCs by polymerase chain reaction amplification. The env sequences of these clones were generally similar to those of the cultured viruses. Within the V3 region, the earliest isolates retained the sequence of the HXB2 clone from IIIB. Clones obtained later showed a progressive divergence in V3. An A-to-T substitution within the GPGRAF sequence at the tip of the V3 loop was observed within 1 year after infection, and this mutation predominated in all subsequent isolates. Antibodies against the V3 loops of IIIB and divergent 1987 and 1990 LW isolates appeared simultaneously in laboratory worker serum and persisted with no significant differences in titer. Furthermore, neutralization studies with autologous sequential sera suggested selection for the A-to-T change in V3 was not due to V3-directed antibodies. These results demonstrate a surprising homogeneity among env sequences of HIV-1 from an infected laboratory worker, perhaps because the initial infection originated from a relatively homogeneous population of tissue culture-adapted virus.
HIV-1分子克隆是从外周血单核细胞（PBMC）培养的分离株中获得的，也直接从一名意外感染HIV-1实验室毒株HIV-1(HTLV-IIIB)的实验室工作人员的未培养PBMC中获得。从培养病毒的克隆中获取了与HIV-1(HTLV-IIIB)克隆BH10的前752个氨基酸相对应的包膜序列并进行测序。感染后不久获得的三个env克隆彼此之间仅在七个核苷酸位置存在差异，导致一个氨基酸替换和一个移码突变。这些包膜序列与各种IIIB克隆的包膜序列相似程度，与后者之间的相似程度相同。env差异在感染过程中增加。然而，感染两年或更长时间后获得的env克隆的总体多样性仍与原始IIIB培养物中IIIB env克隆之间的多样性相当。还通过聚合酶链反应扩增直接从未培养的PBMC中获得了多个包含V3环的部分env基因序列克隆。这些克隆的env序列通常与培养病毒的env序列相似。在V3区域内，最早的分离株保留了来自IIIB的HXB2克隆的序列。后来获得的克隆在V3区域显示出逐渐的差异。在感染后1年内观察到V3环顶端的GPGRAF序列内发生了A到T的替换，并且该突变在所有后续分离株中占主导地位。针对IIIB以及1987年和1990年不同的LW分离株的V3环的抗体同时出现在实验室工作人员血清中，并且效价没有显著差异地持续存在。此外，用自体连续血清进行的中和研究表明，V3中A到T变化的选择不是由于V3导向的抗体。这些结果表明，来自一名受感染实验室工作人员的HIV-1的env序列之间存在惊人的同质性，这可能是因为最初的感染源自相对同质的组织培养适应病毒群体。
