title: Quantification of genital human immunodeficiency virus type 1 (HIV-1) DNA in specimens from women with low plasma HIV-1 RNA levels typical of HIV-1 nontransmitters.
abstract: Studies of human immunodeficiency virus type 1 (HIV-1) transmission suggest that genital HIV-1 RNA and DNA may both be determinants of HIV-1 infectivity. Despite its potential role in HIV-1 transmission, there are limited quantitative data on genital HIV-1 DNA. Here we validated an in-house real-time PCR method for quantification of HIV-1 DNA in genital specimens. In reactions with 100 genomes to 1 genome isolated from a cell line containing one HIV-1 provirus/cell, this real-time PCR assay is linear and agrees closely with a commercially available real-time PCR assay specific for a cellular housekeeping gene. In mock genital samples spiked with low numbers of HIV-1-infected cells such that the expected HIV-1 DNA copy number/reaction was 100, 10, or 5, the average copy number/reaction was 80.2 (standard deviation [SD], 28.3), 9.1 (SD, 5.4), or 3.1 (SD, 2.1), respectively. We used this method to examine genital HIV-1 DNA levels in specimens from women whose low plasma HIV-1 RNA levels are typical of HIV-1 nontransmitters. The median HIV-1 DNA copy number in endocervical secretions from these women (1.8 HIV-1 DNA copies/10,000 cells) was lower than that for women with higher plasma HIV-1 RNA levels (16.6 HIV-1 DNA copies/10,000 cells) (P=0.04), as was the median HIV-1 DNA copy number in vaginal secretions (undetectable versus 1.0 HIV-1 DNA copies/10,000 cells). These data suggest that women with low plasma HIV-1 RNA and thus a predicted low risk of HIV-1 transmission have low levels of genital HIV-1 cell-associated virus. The assay described here can be utilized in future efforts to examine the role of cell-associated HIV-1 in transmission.
authors.affiliation: Department of Microbiology, University of Washington, Seattle, WA 98109, USA, and Coast Provincial General Hospital, Mombasa, Kenya.
authors.forename: Sarah
authors.initials: S
authors.lastname: Benki
authors.forename: R Scott
authors.initials: RS
authors.lastname: McClelland
authors.forename: Sandra
authors.initials: S
authors.lastname: Emery
authors.forename: Jared M
authors.initials: JM
authors.lastname: Baeten
authors.forename: Barbra A
authors.initials: BA
authors.lastname: Richardson
authors.forename: Ludo
authors.initials: L
authors.lastname: Lavreys
authors.forename: Kishorchandra
authors.initials: K
authors.lastname: Mandaliya
authors.forename: Julie
authors.initials: J
authors.lastname: Overbaugh
mesh_terms: D000328:Adult; D001826:Body Fluids / Q000821:virology; D002584:Cervix Uteri / Q000821:virology; D004279:DNA, Viral / Q000032:analysis*; D005260:Female; D015658:HIV Infections / Q000635:transmission / Q000821:virology*; D015497:HIV-1 / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012367:RNA, Viral / Q000097:blood*; D012680:Sensitivity and Specificity; D013223:Statistics as Topic; D014621:Vagina / Q000821:virology
chemical_list: D004279:DNA, Viral; D012367:RNA, Viral