Evaluation of an affordable HIV-1 virological failure assay and antiretroviral drug resistance genotyping protocol

A qualitative PCR minipool strategy to screen for virologic failure and antiretroviral drug resistance in South African patients on first-line antiretroviral therapy

“…Qualitative rather than quantitative detection of failure has previously been shown to reduce the cost of virological monitoring [24], and sequencing of the reverse transcriptase coding region of the pol gene alone likewise reduces costs compared to longer sequences [25].…”

Performance and Logistical Challenges of Alternative HIV-1 Virological Monitoring Options in a Clinical Setting of Harare, Zimbabwe

“…The IC comprised of the nonhuman RNA virus, encephalomyocarditis virus (EMC), prepared at the UMCU, Utrecht, The Netherlands. Nucleic acids from plasma were extracted using the NucliSENS easyMAG System (bioMérieux) as per manufacturer's instruction, with an on-board lysis incubation as previously described [23, 24]. …”

“…The assay is designed as an open platform and is based on real-time PCR of the HIV-1 LTR fragment as described elsewhere [23, 24]. The VFA is HIV-1 subtype independent, applicable to lower throughput settings and can be applied to dried blood spots (DBS), thanks to the optimized nucleic acid elution methods.…”

“…So far, the VFA has been evaluated on purposefully selected panels of HIV-1 infected plasma and in laboratory-controlled conditions [23, 24]. In order to further demonstrate its clinical utility, we have set out to evaluate the VFA in a clinical setting, using consecutive clinical samples from patients attending Newlands Clinic, an HIV treatment center in Harare, Zimbabwe.…”

Emerging antiretroviral drug resistance in sub-Saharan Africa

“…In-house GART methods and collective bargaining with suppliers – as the case of the Southern African Treatment and Resistance Network –can make testing more affordable, when performed by laboratories participating in international external quality-assurance programmes. Sequencing reverse transcriptase amino acid positions 41–230 is sufficient for patients with virologic failure during first-line ART, and could reduce costs [74], as could approaches that combine screening for virologic failure with sequencing for reverse transcriptase mutations using pooled specimens [75,76] or next generation sequencing (NGS) [77]. Deep sequencing with NGS platforms to detect minor variant NNRTI probably adds clinical value [78] but is costly.…”