Performance of two HIV-2 primers/probe sets in real-time RT-qPCR assays.

<p>Two HIV-2 primers/probe sets were assessed for amplification of serial dilutions of an HIV-2 isolate, NIH-Z, under standard ampification conditions that were not optimized for each primers/probe set. Primers/probe sets, designated as PD  =  Primer Design LTD HIV-2 PCR Kit and SM  =  Delarue et al <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0096554#pone.0096554-Delarue1" target="_blank">[12]</a>, showed linear amplification profiles that were parallel. PD  =  •. SM  =  ▴.</p

Performance of HIV-1/2 viral stocks in real-time RT-qPCR and HIV-1 p24 antigen tests.

<p>Individual viral stocks identified as HIV-2 were obtained from the NIAID AIDS Reagent and Reference Repository and SeraCare, Inc. The HIV-1 subtype and HIV-2 group identification is based upon data sheets provided by NIAID and from publications. An HIV-1 Subtype B isolate from the United States (91US_4) was used as an HIV-1 control. Viral RNA was extracted and tested in HIV-2 real-time RT-qPCR assays. Viral isolates selected for testing in the Roche COBAS AmpliPrep/COBAS TaqMan HIV-1 v2.0 quantitative RNA assay and the Perkin-Elmer p24 Antigen test included the two viral isolates that were not amplified in the HIV-2 real-time RT-qPCR assays, three HIV-2 viral isolates that amplified well and the HIV-1 91US_4 isolate. Although the HIV-2 RT-qPCR assays were not optimized, the HIV-2 amplifications were robust for the majority of the isolates tested as reflected in the HIV-2 RNA concentrations reported based upon relative values extrapolated from the NIH-Z standard for each primers/probe set. The CDC 310340 and CDC 310342 viral stocks were not amplified in HIV-2 RT-qPCR assays but were quantified and reactive in HIV-1 specific tests. PD  =  Primer Design LTD HIV-2 PCR Kit primers/probe. SM  =  Delarue et al <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0096554#pone.0096554-Delarue1" target="_blank">[12]</a> primers/probe set. TND  =  Target Not Detected. UNK is unknown group. Viral isolate was not tested (−).</p

Phylogenetic Tree Analysis of Protease/Reverse Transcriptase Sequences for CDC 310340 and CDC 310342 Viral Stocks.

<p>HIV-1 sequences were obtained using the Siemens' TRUGENE HIV-1 Genotype Test, analyzed using MegAlign version 9.0.4 (DNASTAR, Inc) and subtyped against the HIV Sequence Database using BLAST (<a href="http://www.hiv.lanl.gov/content/sequence/BASIC_BLAST.html" target="_blank">www.hiv.lanl.gov/content/sequence/BASIC_BLAST.html</a>). Both sequences were found to align with HIV-1 Group M, subtype CRF02_AG. The homology between the two sequences was 98% with 0.3% divergence. Both sequences are designated in the tree with a •. The dotted line indicates a negative branch length, which is a result of averaging.</p