Development of a User-Friendly Pipeline for Mutational Analyses of HIV Using Ultra-Accurate Maximum-Depth Sequencing

Area of Science:

• Molecular Virology
• Genomics
• Bioinformatics

Background:

• Human immunodeficiency virus type 2 (HIV-2) exhibits lower mutation rates than HIV type 1 (HIV-1).
• Previous studies of HIV mutagenesis were limited by high error rates in standard next-generation sequencing (NGS).

Purpose of the Study:

• To adapt maximum-depth sequencing (MDS) for ultra-accurate characterization of HIV-1 and HIV-2 mutagenesis.
• To develop a user-friendly Galaxy workflow for analyzing MDS sequencing data, enhancing reproducibility and accessibility for molecular virologists.

Main Methods:

• Adaptation of the maximum-depth sequencing (MDS) technique for HIV-1 and HIV-2 studies.
• Development of a Galaxy-based bioinformatics workflow for MDS data analysis.
• Validation of the MDS technique and pipeline by comparing results with existing HIV-1 and HIV-2 mutation data.

Main Results:

• The adapted MDS technique achieved a 100-fold reduction in background error rate compared to standard Illumina sequencing.
• The MDS pipeline demonstrated a 10-fold lower error rate than traditional unique molecular identifier (UMI)-based sequencing.
• The technique and pipeline were validated against previous HIV-1 and HIV-2 mutation analyses.

Conclusions:

• The adapted MDS technique and analysis pipeline provide ultra-accurate characterization of viral mutagenesis in HIV-1 and HIV-2.
• This advancement facilitates the discovery of novel sources of viral mutagenesis, expanding understanding of retroviral diversity and evolution.
• The user-friendly workflow enhances accessibility and replicability for molecular virology research.