Summaryobjective To evaluate treatment results of the paying antiretroviral therapy (ART) clinic of Queen Elizabeth Central Hospital, a large public and teaching hospital in Blantyre, Malawi. The only ART was a fixed drug combination of stavudine, lamivudine and nevirapine.methods Cross sectional study with interviews, laboratory tests (CD4 count, viral load, nevirapine plasma levels, transaminases) and data extraction from files.results A total of 422 (59%) of the patients who started ART since 2000 were lost to follow-up. The 176 patients enrolled in the study had good virological and excellent clinical treatment results. The most common side effect was peripheral neuropathy. Nevirapine plasma levels were remarkably high and associated with successful virological treatment results. Two simple adherence questions pertaining to the use of medication in the previous 8 days corresponded well with nevirapine levels. The most important reasons for non-adherence were shortage of drugs in the hospital pharmacy and personal financial constraints.conclusions (1) Many patients were lost to follow-up. (2) High nevirapine levels contributed to good therapy results in those studied. (3) Simple adherence questions predicted subtherapeutic nevirapine levels. (4) Antiretroviral drug supply needs to be uninterrupted and free of charge, to prevent avoidable non-adherence.

We studied sequence differences in regulatory elements of the long terminal repeat (LTR) and primer-binding site (PBS) among various human immunodeficiency virus type 1 (HIV-1) subtypes. Phylogenetic sequence analysis of a fragment of 729 base pairs (bp) covering the Gag-coding region for half of p24 and all of p17 revealed the gag subtype of all 60 viruses included in the study: A (n = 20), B (n = 12), C (n = 7), D (n = 10), E (n = 3), F (n = 4), G (n = 3), and H (n = 1). The subtype was also determined by analysis of a 689-bp fragment comprising the LTR and the PBS motif. Comparison of the LTR versus gag sequences showed a mosaic genome for seven isolates. After analysis of all sequences, we could describe subtype-specific differences in sequences encompassing the regulatory elements of the LTR and the PBS motif.

Cross-sectional studies have suggested that infection with human immunodeficiency virus (HIV) type 1 could reduce the mitochondrial DNA (mtDNA) content of blood cells. We investigated mtDNA content in peripheral blood mononuclear cells (PBMCs) obtained from 36 antiretroviral therapy-naive documented HIV-1 seroconverters, before and after seroconversion. mtDNA content statistically significantly decreased 1 year after seroconversion and showed a nonsignificant decrease during the subsequent 4 years. These findings confirm that infection with HIV-1 may, itself, reduce mtDNA content, at least within PBMCs. This could have implications for the subsequent development of mitochondrial toxicities associated with the use of nucleoside analogue reverse-transcriptase inhibitors.

We studied the use of dried spots of bodily fluids (plasma, whole blood, and mother's milk) on filter paper as a means of sample collection and storage for human immunodeficiency virus type 1 (HIV-1) viral load testing under stringent field conditions. Plasma placed directly in lysis buffer, which is customarily used for viral load assays, was used for comparison in all our experiments. Utilizing reconstruction experiments, we demonstrate no statistical differences between viral loads determined for plasma and mother's milk spotted on filter paper and those for the same fluids placed directly in lysis buffer. We found that the addition of whole blood directly to lysis buffer was unreliable and could not be considered a feasible option. However, viral load measurements for whole blood spotted onto filter paper correlated with plasma viral load values for both filter spots and lysis buffer (Pearson correlation coefficients, 0.7706 and 0.8155, respectively). In conclusion, dried spots of plasma, whole blood, or mother's milk provide a feasible means for the collection, storage, and shipment of samples for subsequent viral load measurement and monitoring. Virus material spotted and dried on filter paper is a good inexpensive alternative for collecting patient material to monitor the HIV-1 viral load. Measuring the HIV-1 burden from whole blood dried on filter paper provides a suitable alternative for low-technology settings with limited access to refrigeration, as can be found in sub-Saharan Africa.

Because human immunodeficiency virus type 1 (HIV-1) subtypes and circulating recombinant forms (CRFs) are spreading rapidly worldwide and are becoming less confined to a geographical area, RNA assays that can detect and quantify all HIV-1 isolates reliably are in demand. We have developed a fast, real-time monitored RNA assay based on an isothermal nucleic acid sequence-based amplification technology that amplifies a part of the long terminal repeat region of the HIV-1 genome. Real-time detection was possible due to the addition of molecular beacons to the amplification reaction that was monitored in a fluorimeter with a thermostat. The lower level of detection of the assay was 10 HIV-1 RNA molecules per reaction, and the lower level of quantification was 100 copies of HIV-1 RNA with a dynamic range of linear quantification between 10 2 and 10 7 RNA molecules. All HIV-1 groups, subtypes, and CRFs could be detected and quantified with equal efficiency, including the group N isolate YBF30 and the group O isolate ANT70. To test the clinical utility of the assay, a series of 62 serum samples containing viruses that encompassed subtypes A through G and CRFs AE and AG of HIV-1 group M were analyzed, and these results were compared to the results of a commercially available assay. This comparison showed that the quantification results correlated highly (R 2 ‫؍‬ 0.735) for those subtypes that could be well quantified by both assays (subtypes B, C, D, and F), whereas improved quantification was obtained for subtypes A and G and CRFs AE and AG. A retrospective study with six individuals infected with either a subtype A, B, C, or D or an AG isolate of HIV-1 group M, who were treated with highly active antiretroviral therapy, revealed that the assay was well suited to the monitoring of therapy effects. In conclusion, the newly developed real-time monitored HIV-1 assay is a fast and sensitive assay with a large dynamic range of quantification and is suitable for quantification of most if not all subtypes and groups of HIV-1.The viral RNA level in the plasma or serum of human immunodeficiency virus type 1 (HIV-1) infected individuals has become the most important marker for several aspects of an HIV-1 infection. The HIV-1 RNA level is predictive for disease progression in untreated infected individuals (7,13,17,18,25), and it is the best marker for monitoring therapy effects (14,20,31). In addition, the presence of viral RNA in the serum of newborn children is used as evidence for mother-tochild transmission, since maternal antibodies present in infant serum hamper antibody-screening assays. In this decade the number of newborn infants screened will increase as soon as cheaper and easier assays become available for use in developing countries, where they are needed most. The presence of viral RNA will also be one of the most important markers for monitoring breakthroughs in vaccine studies due to the presence of viral antibodies in sera as a result of vaccination. In addition, vaccines that do not have a strict preventive eff...