Near-Full-Length Single-Genome HIV-1 DNA Sequencing

Area of Science:

• Virology
• Genomics
• Immunology

Background:

• Human Immunodeficiency Virus type 1 (HIV-1) establishes lifelong reservoirs by integrating into host chromosomes.
• Most integrated viral DNA contains defects, hindering viral replication and rebound viremia.
• Accurate quantification of intact HIV-1 DNA is essential for understanding reservoir dynamics and viral evolution.

Purpose of the Study:

• To introduce Full-Length Individual Proviral Sequencing (FLIP-Seq), a novel next-generation sequencing method.
• To enable near full-length, single-genome analysis of HIV-1 DNA in human cells.
• To distinguish between intact and defective HIV-1 DNA sequences.

Main Methods:

• Sequential dilution of proviral DNA to single genomes.
• Amplification of near full-length viral DNA.
• Deep sequencing and biocomputational analysis to identify intact proviruses.

Main Results:

• FLIP-Seq provides absolute quantification of HIV-1 DNA in cell populations.
• The method can be applied to small numbers of purified CD4 T cells.
• Identifies phylogenetic associations and clonal expansion of intact proviruses.

Conclusions:

• FLIP-Seq is a powerful tool for characterizing the HIV-1 reservoir.
• Enables precise measurement of intact viral DNA, crucial for assessing rebound viremia risk.
• Advances understanding of HIV-1 persistence and evolutionary dynamics.