Magnetic-silica based nucleic acid extraction for Human Immunodeficiency Virus Type-1 drug-resistance testing in low viremic patients

Abstract 

Human Immunodeficiency Virus Type-1 (HIV-1) drug-resistance testing is challenging for viral loads below 1,000copies/mL, but, according to HIV-1 guidelines, it should be considered for improving patient management and treatment options. High-recovery and high-purity extraction methods can enhance standard performances of HIV-1 genotyping assays based on direct full-population sequencing.

Aim of the present study was to evaluate performances of the NucliSENS easyMAG (NeM) (BioMerieux, Marcy l’Etoile, F) semi-automated nucleic acid extraction system combined with the direct full-population sequencing ViroSeq HIV-1 genotyping (Abbott, IL, US), for detecting drug resistance in samples with HIV-1 RNA<1,000copies/mL (n=62). Data were compared with those from the ViroSeq manual extraction in 86 samples with HIV-1 RNA<1,000copies/mL and studied on HIV-1 reference standards.

HIV-1 genotyping was successful in 98% of samples extracted with NeM (61/62) and in 84% of those extracted with ViroSeq (72/86) (X²=8.508, p=0.004). HIV-1 RNA levels in samples successfully processed with NeM were significantly lower than those in manually processed ones (mean±SD, respectively, 285±222copies/mL vs. 403±269copies/mL) (p=0.004). For HIV-1 RNA levels <300 and 500copies/mL, performances of HIV-1 genotyping with NeM were significantly high (97% and 98%, vs. 68% and 78% for manual extraction). As assessed on HIV-1 RNA reference standards, the detection rate at 200copies/mL for HIV-1 genotyping with NeM extraction was 100%.

In conclusion, these data support that HIV-1 direct full-population sequencing combined with NeM is associated with a significantly high success rate, thus improving the management of HIV-1 drug resistance in low viremic patients.

Keywords: HIV-1, Human Immunodeficiency Virus Type-1, NeM, NucliSENS easyMAG, PCR, polymerase chain reaction, P, protease, RT, reverse transcriptase, PI, protease inhibitor, NRTI, nucleoside reverse transcriptase inhibitor, NNRTI, non-nucleoside reverse transcriptase inhibitor

Keywords: HIV-1, Drug resistance, Genotypic test, Direct full-population sequencing, Nucleic acid extraction