Our findings show heterogeneous virological failure and illustrate that, in addition to routine access to viral load, good management of ART programs is even more critical to improve treatment outcomes in resource-limited countries.

The development and validation of dried sample spots as a method of specimen collection are urgently needed in developing countries for monitoring of human immunodeficiency virus (HIV) infection. Our aim was to test some crucial steps in the use of dried spots, i.e., viral recovery and storage over time. Moreover, we investigated whether dried plasma and blood spots (DPS and DBS, respectively) give comparable viral load (VL) results. Four manual RNA extraction methods from commercial HIV type 1 (HIV-1) VL assays-a QIAamp minikit (Qiagen), the Abbott Molecular sample preparation system, the Nuclisens assay (bioMarieux), and High Pure viral nucleic acid kit (Roche Applied Science)-were compared for VL quantification and PCR amplification for genotypic drug resistance testing on dried spots from spiked plasma and residual samples from HIV-1 patients (n ‫؍‬ 47; median VL, 4.13 log 10 copies/ml). RNA recovery from DPS was efficient using Nuclisens extraction (median difference, 0.03 log 10 copies/ml) and slightly underestimated using the Abbott Molecular sample preparation system (median difference, 0.35 log 10 copies/ml). PCR amplification results were in concordance. Measurements from DBS overestimated VL for plasma, with VL results showing <3.7 log 10 copies/ml. VL was stable for up to 3 months in spiked DPS stored at 20°C but for only 1 month at 37°C. A faster decline was observed in PCR efficiency: DPS could be stored for 1 week at 37°C and for 1 month at 20°C. In conclusion, the RNA extraction method is an important factor in obtaining reliable RNA quantification and PCR amplification of HIV-1 on DPS/DBS. DBS could be used as an alternative for DPS depending on HIV RNA cutoffs for virological failure. VL measurements remain stable over a longer period than do PCR amplification results.Due to the efforts of national programs and the support of a wide range of international partners, the number of people receiving antiretroviral therapy (ART) in resource-limited countries has increased significantly over recent years, most notably in sub-Saharan Africa (24). In order to limit the emergence of resistance to antiretroviral drugs, human immunodeficiency virus (HIV) treatment should ideally be accompanied by periodic virological monitoring, such as viral load and resistance testing. However, in resource-limited countries, viral load monitoring and drug resistance tests are not yet available at an affordable cost, and infrastructure requirements limit the scale-up in access to these tests. Treatment initiation and modifications are thus guided mainly by clinical disease progression and by CD4 cell counts, when possible (13, 24). As a consequence, many people with virological failure stay on inadequate drug regimens for long periods. Rapid or uncontrolled emergence of HIV drug resistance is thus feared as a potential consequence of ART scale-up in resource-limited countries. Therefore, ART resistance monitoring is not recommended at the individual patient level in low-income countries, and the WHO has established a sys...

Virological monitoring is still feasible for DBS after 3 months of storage at 37 degrees C when VL is >5000 copies/mL, but DNA contributes largely to the final results.