High throughput SARS-CoV-2 variant analysis using molecular barcodes coupled with next generation sequencing.

Area of Science:

• Virology
• Genomics
• Molecular Diagnostics

Background:

• Emerging SARS-CoV-2 variants pose significant challenges to pandemic control, necessitating advanced diagnostic tools.
• Current diagnostic methods struggle with rapid, reliable identification of viral mutations and epidemiological surveillance.

Purpose of the Study:

• To develop and validate a high-throughput diagnostic solution for large-population SARS-CoV-2 screening and variant analysis.
• To integrate DNA barcoding, sample pooling, and Next Generation Sequencing (NGS) for efficient viral mutation mapping.

Main Methods:

• Patient samples were barcoded using RT-PCR primers, followed by multi-step sample pooling.
• Amplified viral and human housekeeping genes were sequenced using an Illumina NGS system.
• Data analysis focused on identifying positive samples and specific SARS-CoV-2 variants at single nucleotide resolution.

Main Results:

• The developed method achieved a sensitivity of 82.5% and a specificity of 97.3%.
• Screening of 960 samples identified 258 positive cases (27%) and detected seven SARS-CoV-2 variants (six known, one unknown).
• The solution demonstrated effective large-population screening and precise variant identification.

Conclusions:

• The integrated DNA barcoding, pooling, and NGS approach offers a sensitive and specific solution for epidemiological screening of SARS-CoV-2.
• This method facilitates rapid identification of positive samples and detailed variant analysis, crucial for pandemic response.
• The diagnostic solution supports high-throughput testing and single nucleotide resolution for viral mutation surveillance.