Bacteria associated with BV increase genital-tract HIV RNA levels. Quantitative bacterial counts for lactobacilli and M. hominis are better correlates of CVL HIV RNA than are Nugent score or Amsel criteria. Since plasma virus and CD4 cell levels did not differ between the BV and no-BV groups, these data suggest that the bacterial flora associated with BV influence genital-tract HIV shedding.

Eleven laboratories evaluated the use of dried blood and plasma spots for quantitation of human immunodeficiency virus (HIV) RNA by two commercially available RNA assays, the Roche Amplicor HIV-1 Monitor and the bioMerieux NucliSens HIV-1 QT assays. The recovery of HIV RNA was linear over a dynamic range extending from 4,000 to 500,000 HIV type 1 RNA copies/ml. The Monitor assay appeared to have a broader dynamic range and seemed more sensitive at lower concentrations. However, the NucliSens assay gave more consistent results and could be performed without modification of the kit. HIV RNA was stable in dried whole blood or plasma stored at room temperature or at ؊70°C for up to 1 year. Dried blood and dried plasma spots can be used as an easy and inexpensive means for the collection and storage of specimens under field conditions for the diagnosis of HIV infection and the monitoring of antiretroviral therapy.Human immunodeficiency virus (HIV) RNA levels in blood plasma are used to monitor the response to antiretroviral drug therapy in developed countries. Since the International AIDS Conference held in Durban, South Africa, in 2000, there has been a global effort to provide access to antiretroviral drugs to all infected individuals, including those in the resource-poor countries which have been hit the hardest by the AIDS pandemic. Studies have demonstrated that the provision of treatment is not enough, however. In order for the drugs to be most effective, both for an individual and for public health reasons, it will be necessary to monitor the responses to the therapy. HIV RNA assays typically use plasma, which implies access to laboratory equipment that may not be readily available in all field settings. Therefore, alternative specimens must be considered for use in these situations.Whole blood dried on filter paper (dried blood spots [DBSs]) has been used to qualitatively detect HIV antibodies (13, 15), HIV DNA (1, 3, 5-7, 9, 10, 11, 12, 16, 18) and HIV RNA (4, 14, 17; J. L. Gerstel and A. M. Comeau, Proc. 10th Natl. Neonatal Screening Symp., abstr. P-19, p. 64, 1994). In addition, dried plasma spots (DPSs) have also been used to quantitate the HIV load (8). Although the stabilities of antibodies and DNA in DBSs have been determined (2, 6, 13), there has been some question regarding the stability of the HIV RNA when it is dried and stored at room temperature (14, 17).The purpose of this study was to compare two different methods for quantitatively measuring HIV RNA in DBSs and DPSs and to assess their long-term stability at room temperature and at Ϫ70C.(This study was presented in part at the 12th World AIDS Conference, Geneva, Switzerland, 28 June to 3 July 1998 [S. Cassol, A. Comeau, S. Fiscus, G. Aldrovandi, J. Sullivan, J. Bremer, and B. Jackson, Abstr. 12th World AIDS Conf., abstr. 33166, 1998].) MATERIALS AND METHODS Specimens.Three different panels were prepared by the Virus Quality Assurance Laboratory by using filter paper (903; Schleicher & Schuell, Keene, N.H.) spotted with 50 l of plasma or whol...

Dried blood spots (DBS) are simpler to prepare, store, and transport than plasma or serum and may represent a good alternative for drug resistance genotyping, particularly in resource-limited settings. However, the utility of DBS for drug resistance testing is unknown. We investigated the efficiency of amplification of large human immunodeficiency virus type 1 (HIV-1) pol fragments (1,023 bp) from DBS stored at different temperatures, the type of amplified product(s) (RNA and/or DNA), and the similarity between plasma and DBS sequences. We evaluated two matched plasma/DBS panels stored for 5 to 6 years at several temperatures and 40 plasma/DBS specimens collected from untreated persons in Cameroon and stored for 2 to 3 years at ؊20°C. The amplification of HIV-1 pol was done using an in-house reverse transcriptase-nested PCR assay. Reactions were done with and without reverse transcription to evaluate the contribution of HIV DNA to pol sequences from DBS. Amplification was successful for the DBS samples stored for 5 to 6 years at ؊20°C or at ؊70°C but not for those stored at room temperature. Thirty-seven of the 40 (92.5%) DBS from Cameroon were amplifiable, including 8/11 (72.7%) with plasma virus loads of <10,000 RNA copies/ml and all 29 with plasma virus loads of >10,000. Proviral DNA contributed significantly to DBS sequences in 24 of the 37 (65%) specimens from Cameroon. The overall similarity between plasma and DBS sequences was 98.1%. Our results demonstrate the feasibility of DBS for drug resistance testing and indicate that ؊20°C is a suitable temperature for long-term storage of DBS. The amplification of proviral DNA from DBS highlights the need for a wider evaluation of the concordance of resistance genotypes between plasma and DBS.The introduction of highly active antiretroviral therapy and the demonstration of dramatic improvements in human immunodeficiency virus (HIV)-and AIDS-related mortality and morbidity in North America and Europe have fueled international efforts to expand access to care and treatment in lessdeveloped countries. Several major initiatives to provide treatment in resource-limited settings, including the U.S. President's Emergency Plan for AIDS Relief and the Global Fund against AIDS, TB and Malaria, are now in progress (16). The implementation of these programs requires the development of appropriate and effective patient-monitoring systems, including surveillance for antiretroviral drug resistance. Sentinel drug resistance surveillance systems are important public health tools that can provide information on trends in the prevalence of resistance at the population level and can be used to modify treatment guidelines.Plasma and serum are considered the preferred specimen types for HIV type 1 (HIV-1) drug resistance testing. However, these types of specimens are not optimal in resource-limited settings where the equipment necessary for PCR amplification and sequencing may not be available at collection sites and resistance testing requires transportation of the samples to a refer...

For an individual patient, a change in plasma HIV-1 RNA level of 2.5-fold or more probably indicates a true biological change. Monitoring HIV-1 RNA levels and CD4+ lymphocytes before a change in antiretroviral treatment and monitoring HIV-1 RNA levels shortly thereafter improves prediction of disease progression and decline in CD4+ counts for 1 year compared with monitoring CD4+ counts of HIV-1 RNA levels alone. Additional monitoring of infectious HIV-1 titers in mononuclear cells of peripheral blood is not useful.

Worldwide, 90% of HIV-1 infections are transmitted heterosexually. Because the genital mucosa are the sites of initial contact with HIV-1 for most exposed individuals, study of the virus from the genital tract is critical for the development of vaccines and therapeutics. Previous analyses of HIV-1 in various tissues have documented compartmentalization of viral genomes. Whether compartmentalization was associated with viral phenotypic differences or immune status, however, was not well understood. We compared HIV-1 gp120 env sequences from the genital tract and plasma of 12 women. Eight women displayed compartmentalized HIV-1 RNA genomes, with viral sequences from each site that were clearly discrete, yet phylogenetically related. The remaining four exhibited env sequences that were intermingled between the two sites. Women with compartmentalized HIV-1 genomes had higher CD4 ؉ cell counts than those displaying intermingled strains (P ‫؍‬ 0.02). Intrapatient HIV-1 recombinants comprising sequences that were characteristic of both sites were identified. We next compared viral phenotypes in each compartment. HIV-1 coreceptor usage was often compartmentalized (P < 0.01). The number of N-linked glycosylation sites, associated with neutralization resistance, also differed between compartments (P < 0.01). Furthermore, disparities between the density of gp120 glycosylations in each compartment correlated with higher CD4 ؉ counts (P ‫؍‬ 0.03). These data demonstrate that the genital tract and plasma can harbor populations of replicating HIV-1 with different phenotypes. The association of higher CD4 ؉ cell counts with compartmentalization of viral genomes and density of gp120 glycosylations suggests that the immune response influences the development of viral genotypes in each compartment. These findings are relevant to the prevention and control of HIV-1 infection.