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IntroductionHIV‐1 near full‐length genome (HIV‐NFLG) sequencing from plasma is an attractive multidimensional tool to apply in large‐scale population‐based molecular epidemiological studies. It also enables genotypic resistance testing (GRT) for all drug target sites allowing effective intervention strategies for control and prevention in high‐risk population groups. Thus, the main objective of this study was to develop a simplified subtype‐independent, cost‐ and labour‐efficient HIV‐NFLG protocol that can be used in clinical management as well as in molecular epidemiological studies.MethodsPlasma samples (n=30) were obtained from HIV‐1B (n=10), HIV‐1C (n=10), CRF01_AE (n=5) and CRF01_AG (n=5) infected individuals with minimum viral load >1120 copies/ml. The amplification was performed with two large amplicons of 5.5 kb and 3.7 kb, sequenced with 17 primers to obtain HIV‐NFLG. GRT was validated against ViroSeq™ HIV‐1 Genotyping System.ResultsAfter excluding four plasma samples with low‐quality RNA, a total of 26 samples were attempted. Among them, NFLG was obtained from 24 (92%) samples with the lowest viral load being 3000 copies/ml. High (>99%) concordance was observed between HIV‐NFLG and ViroSeq™ when determining the drug resistance mutations (DRMs). The N384I connection mutation was additionally detected by NFLG in two samples.ConclusionsOur high efficiency subtype‐independent HIV‐NFLG is a simple and promising approach to be used in large‐scale molecular epidemiological studies. It will facilitate the understanding of the HIV‐1 pandemic population dynamics and outline effective intervention strategies. Furthermore, it can potentially be applicable in clinical management of drug resistance by evaluating DRMs against all available antiretrovirals in a single assay.