Szojka Zsófia, Karlson Sara, Jansson Marianne, Medstrand Patrik
Department of Translational Medicine, Clinical Virology, Lund University, Malmö, Sweden.
Laboratory of Retroviral Biochemistry, Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary.
Bio Protoc. 2019 Oct 20;9(20):e3404. doi: 10.21769/BioProtoc.3404.
Time to AIDS infection is longer with HIV-2, compared to HIV-1, but without antiretroviral therapy both infections will cause AIDS-related mortality. In HIV-2 infection, monitoring of antiretroviral treatment (ART) efficacy is challenging since a large proportion of HIV-2-infected individuals displays low or undetectable plasma RNA levels. Hence, quantification of cellular DNA load may constitute an alternative method for monitoring ART efficacy. Moreover, sensitive HIV-2 DNA quantification protocols are also important for the characterization of the HIV-2 reservoirs, and ultimately for the development of HIV-2 cure strategies. We have developed a sensitive and robust HIV-2 DNA quantification protocol based on whole blood as DNA source, including normalization of leukocyte cell numbers using parallel quantification of the single copy porphobilinogen deaminase gene. The specificity and sensitivity of the assay was 100%. The limit of detection was 1 copy and limit of quantification was 5 copies. When applying this protocol to HIV-2 infected, it was found that HIV-2 viral DNA was detectable in individuals in whom viral RNA was undetectable or under quantification level. Thus, this method provides a sensitive approach to HIV-2 DNA viral quantification from whole blood of HIV-2 infected patients.
与HIV-1相比，感染HIV-2后发展为艾滋病的时间更长，但如果不进行抗逆转录病毒治疗，两种感染都会导致与艾滋病相关的死亡。在HIV-2感染中，监测抗逆转录病毒治疗（ART）的疗效具有挑战性，因为很大一部分感染HIV-2的个体血浆RNA水平较低或检测不到。因此，定量细胞DNA载量可能构成监测ART疗效的另一种方法。此外，灵敏的HIV-2 DNA定量方案对于表征HIV-2储存库也很重要，最终对于开发HIV-2治愈策略也很重要。我们基于全血作为DNA来源开发了一种灵敏且可靠的HIV-2 DNA定量方案，包括使用单拷贝胆色素原脱氨酶基因的平行定量来对白细胞数量进行标准化。该检测方法的特异性和灵敏度均为100%。检测限为1拷贝，定量限为5拷贝。当将该方案应用于HIV-2感染者时，发现在病毒RNA检测不到或低于定量水平的个体中可检测到HIV-2病毒DNA。因此，该方法为从HIV-2感染患者的全血中进行HIV-2 DNA病毒定量提供了一种灵敏的方法。
