Edmonson P, Murphey-Corb M, Martin L N, Delahunty C, Heeney J, Kornfeld H, Donahue P R, Learn G H, Hood L, Mullins J I
Department of Pathology, University of Washington, Seattle 98195-7740, USA.
J Virol. 1998 Jan;72(1):405-14. doi: 10.1128/JVI.72.1.405-414.1998.
Analysis of disease induction by simian immunodeficiency viruses (SIV) in macaques was initially hampered by a lack of molecularly defined pathogenic strains. The first molecularly cloned SIV strains inoculated into macaques, SIVmacBK28 and SIVmacBK44 (hereafter designated BK28 and BK44, respectively), were cases in point, since they failed to induce disease within 1 year postinoculation in any inoculated animal. Here we report the natural history of infection with BK28 and BK44 in inoculated rhesus macaques and efforts to increase the pathogenicity of BK28 through genetic manipulation and in vivo passage. BK44 infection resulted in no disease in four animals infected for more than 7 years, whereas BK28 induced disease in less than half of animals monitored for up to 7 years. Elongation of the BK28 transmembrane protein (TM) coding sequence truncated by prior passage in human cells marginally increased pathogenicity, with two of four animals dying in the third year and one dying in the seventh year of infection. Modification of the BK28 long terminal repeat to include four consensus nuclear factor SP1 and two consensus NF-kappaB binding sites enhanced early virus replication without augmenting pathogenicity. In contrast, in vivo passage of BK28 from the first animal to die from immunodeficiency disease (1.5 years after infection) resulted in a consistently pathogenic strain and a 50% survival time of about 1.3 years, thus corresponding to one of the most pathogenic SIV strains identified to date. To determine whether the diverse viral quasispecies that evolved during in vivo passage was required for pathogenicity or whether a more virulent virus variant had evolved, we generated a molecular clone composed of the 3' half of the viral genome derived from the in vivo-passaged virus (H824) fused with the 5' half of the BK28 genome. Kinetics of disease induction with this cloned virus (BK28/H824) were similar to those with the in vivo-passaged virus, with four of five animals surviving less than 1.7 years. Thus, evolution of variants with enhanced pathogenicity can account for the increased pathogenicity of this SIV strain. The genetic changes responsible for this virulent transformation included at most 59 point mutations and 3 length-change mutations. The critical mutations were likely to have been multiple and dispersed, including elongation of the TM and Nef coding sequences; changes in RNA splice donor and acceptor sites, TATA box sites, and Sp1 sites; multiple changes in the V2 region of SU, including a consensus neutralization epitope; and five new N-linked glycosylation sites in SU.
由于缺乏分子定义明确的致病菌株，最初对猕猴中猿猴免疫缺陷病毒（SIV）致病作用的分析受到了阻碍。接种到猕猴体内的首批分子克隆SIV菌株，即SIVmacBK28和SIVmacBK44（以下分别称为BK28和BK44），就是典型例子，因为在接种后的1年内，任何接种动物都未因它们感染而发病。在此，我们报告接种恒河猴感染BK28和BK44的自然病程，以及通过基因操作和体内传代提高BK28致病性的努力。4只感染BK44超过7年的动物均未发病，而在长达7年的监测期内，感染BK28的动物不到一半发病。先前在人细胞中传代后截短的BK28跨膜蛋白（TM）编码序列的延长仅略微增加了致病性，4只动物中有2只在感染的第3年死亡，1只在第7年死亡。将BK28长末端重复序列修饰为包含4个共有核因子SP1和2个共有NF-κB结合位点，可增强早期病毒复制，但并未增强致病性。相反，将BK28从第一只因免疫缺陷疾病死亡的动物（感染后1.5年）进行体内传代，产生了一种致病性持续稳定的菌株，50%动物的存活时间约为1.3年，因此该菌株成为了迄今为止鉴定出的致病性最强的SIV菌株之一。为了确定体内传代过程中进化出的多种病毒准种对于致病性是否必要，或者是否进化出了毒性更强的病毒变体，我们构建了一个分子克隆，其由源自体内传代病毒（H824）的病毒基因组3'端一半与BK28基因组5'端一半融合而成。用这种克隆病毒（BK28/H824）诱导疾病的动力学与体内传代病毒相似，5只动物中有4只存活时间不到1.7年。因此，致病性增强的变体的进化可以解释该SIV菌株致病性的增加。导致这种毒性转变的基因变化最多包括59个点突变和3个长度改变突变。关键突变可能是多个且分散的，包括TM和Nef编码序列的延长；RNA剪接供体和受体位点、TATA盒位点以及Sp1位点的变化；SU的V2区域的多个变化，包括一个共有中和表位；以及SU中5个新的N-连接糖基化位点。
