An improved PCR method for detection of HIV-1 proviral DNA of a wide range of subtypes and recombinant forms circulating globally

• Published in: Journal of virological methods Vol. 172; no. 1; pp. 22 - 26
• Main Authors: Weidner, Jürgen, Cassens, Uwe, Göhde, Wolfgang, Sibrowski, Walter, Odaibo, Georgina, Olaleye, David, Reichelt, Doris, Greve, Burkhard
• Format: Journal Article
• Language: English
• Published: Kidlington Elsevier B.V 01.03.2011; Elsevier
• Subjects: antiretroviral agents; Biological and medical sciences; detection limit; DNA; early diagnosis; Electrophoresis, Agar Gel; Flow Cytometry; Fundamental and applied biological sciences. Psychology; gel electrophoresis; HIV infections; HIV Infections - diagnosis; HIV Seropositivity; HIV-1; HIV-1 - genetics; Human immunodeficiency virus 1; Humans; Leukocytes, Mononuclear - virology; Microbiology; mononuclear leukocytes; PCR; people; Polymerase Chain Reaction; Provirus; proviruses; Proviruses - genetics; quantitative polymerase chain reaction; Reproducibility of Results; Sensitivity and Specificity; Subtypes; Techniques used in virology; therapeutics; Virology; Virology - methods; Germany; Africa; HIV-1; Subtypes; DNA; PCR; Provirus; HIV-1 virus; Retroviridae; Method; Lentivirus; Virus; Polymerase chain reaction; Circulating recombinant form; Human immunodeficiency virus; Detection; Subtype
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/j.jviromet.2010.12.008

Proviral DNAs are being measured increasingly as a marker of the efficacy of highly active anti-retroviral therapy (HAART) and is accepted for the early diagnosis of perinatal HIV-1 infections. This requires a standardized test which enables the detection of a wide range of subtypes worldwide including O, N and circulating recombinant forms (CRFs). Based on a previous publication, a PCR – Test for HIV-1 provirus detection in peripheral blood mononuclear cells (PBMCs) was developed. Blood samples from 80 individuals infected with HIV-1 and 20 persons negative for HIV-1&2 from Africa and Germany were tested for the presence of HIV-1 provirus DNA. The primer system used enables the detection of proviral DNA despite the high concentrations of human DNA. The limit of detection was determined to be 5 copies per 10 5 cells. All 20 samples from persons negative for HIV were negative for HIV-1 proviral DNA while provirus DNA was amplified from 76 of the 80 (95%) samples from persons infected with HIV. The amplified products were detected by gel-electrophoresis, flow cytometry and real-time PCR. All three detection systems provided the same results.