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A Multicenter Study To Evaluate the Performance of High-Throughput Sequencing for Virus Detection.
Arifa S Khan1, Siemon H S Ng2, Olivier Vandeputte3
1Division of Viral Products, Office of Vaccines Research and Review, Center for Biologics Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, Maryland, USA.
Msphere
|September 22, 2017
Summary
High-throughput sequencing (HTS) offers sensitive detection of adventitious viruses in biological products. This study demonstrated comparable virus detection across different labs, platforms, and methods, supporting HTS standardization for biologics manufacturing.
Area of Science:
- Virology
- Genomics
- Biotechnology
Background:
- High-throughput sequencing (HTS) is a powerful tool for detecting known and novel viruses.
- Regulatory applications of HTS require method standardization for reliable detection of viral adventitious agents.
- Novel cell substrates for biological product development necessitate robust viral safety testing.
Purpose of the Study:
- To evaluate the performance of HTS for detecting viral adventitious agents in biological product manufacturing.
- To assess the impact of different sample processing, sequencing platforms, and bioinformatic analyses on virus detection.
- To investigate the potential for HTS standardization in viral safety testing.
Main Methods:
- Four model viruses (RSV, EBV, FeLV, REO) and porcine circovirus (PCV) were used.
- Viruses were spiked into HeLa cell matrices (whole cells, lysate, RNA) to mimic manufacturing materials.
- Samples were processed and sequenced using different HTS platforms (Roche 454, Illumina HiSeq), with independent bioinformatic analyses.
Main Results:
- Comparable virus detection was achieved across three independent laboratories.
- Sensitivity ranged from 0.1 to 3 viral genome copies per cell for all model viruses.
- Detection was consistent regardless of sample preparation, library preparation, sequencing platform, or bioinformatic pipeline.
Conclusions:
- HTS is a sensitive method for detecting adventitious viruses in complex biological samples.
- The study demonstrated the feasibility of comparable HTS virus detection across different methodologies.
- Results support the development of reference materials for HTS standardization and validation in biologics.