title: Single real-time reverse transcription-PCR assay for detection and quantification of genetically diverse HIV-1, SIVcpz, and SIVgor strains.
abstract: Although antiretroviral treatment availability has improved, the virological monitoring of patients remains largely uneven across regions. In addition, viral quantification tests are suffering from human immunodeficiency virus type 1 (HIV-1) genetic diversity, fueled by the emergence of new recombinants and of lentiviruses from nonhuman primates. We developed a real-time reverse transcription-PCR (RT-PCR) assay that is relatively inexpensive and able to detect and quantify all circulating forms of HIV-1 and its simian immunodeficiency virus (SIV) precursors, SIVcpz and SIVgor. Primers and a probe were designed to detect all variants of the HIV-1/SIVcpz/SIVgor lineage. HIV-1 M plasma (n = 190; 1.68 to 7.78 log(10) copies/ml) representing eight subtypes, nine circulating recombinant forms, and 21 unique recombinant forms were tested. The mean PCR efficiency was 99%, with low coefficients of intra- and interassay variation (<5%) and a limit of quantification of <2.50 log(10) copies/ml, with a 200-μl plasma volume. On the studied range, the specificity and the analytical sensitivity were 100 and 97.4%, respectively. The viral loads were highly correlated (r = 0.95, P < 0.0001) with the reference method (generic HIV assay; Biocentric) and had no systematic difference, irrespective of genotype. Furthermore, 22 HIV-1 O plasmas were screened and were better quantified compared to the gold-standard RealTime HIV-1 assay (Abbott), including four samples that were only quantified by our assay. Finally, we could quantify SIVcpzPtt and SIVcpzPts from chimpanzee plasma (n = 5) and amplify SIVcpz and SIVgor from feces. Thus, the newly developed real-time RT-PCR assay detects and quantifies strains from the HIV-1/SIVcpz/SIVgor lineage, including a wide diversity of group M strains and HIV-1 O. It can therefore be useful in geographical areas of high HIV diversity and at risk for the emergence of new HIV variants.
authors.affiliation: UMI233, TransVIHMI, Institut de Recherche pour le Développement (IRD) and Université Montpellier 1, Montpellier, France.
authors.forename: Lucie
authors.initials: L
authors.lastname: Etienne
authors.forename: Sabrina
authors.initials: S
authors.lastname: Eymard-Duvernay
authors.forename: Avelin
authors.initials: A
authors.lastname: Aghokeng
authors.forename: Christelle
authors.initials: C
authors.lastname: Butel
authors.forename: Marjorie
authors.initials: M
authors.lastname: Monleau
authors.forename: Martine
authors.initials: M
authors.lastname: Peeters
mesh_terms: D000818:Animals; D017931:DNA Primers / Q000235:genetics; D005243:Feces / Q000821:virology; D015497:HIV-1 / Q000145:classification* / Q000235:genetics*; D006801:Humans; D015345:Oligonucleotide Probes / Q000235:genetics; D002679:Pan troglodytes; D010949:Plasma / Q000821:virology; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D015302:Simian Immunodeficiency Virus / Q000145:classification* / Q000235:genetics*; D019562:Viral Load / Q000379:methods*
chemical_list: D017931:DNA Primers; D015345:Oligonucleotide Probes