A Multicenter Study To Evaluate the Performance of High-Throughput Sequencing for Virus Detection

ABSTRACT The capability of high-throughput sequencing (HTS) for detection of known and unknown viruses makes it a powerful tool for broad microbial investigations, such as evaluation of novel cell substrates that may be used for the development of new biological products. However, like any new assay...

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Autores principales:	Arifa S. Khan, Siemon H. S. Ng, Olivier Vandeputte, Aisha Aljanahi, Avisek Deyati, Jean-Pol Cassart, Robert L. Charlebois, Lanyn P. Taliaferro
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2017
Materias:	454 Roche Illumina adventitious viruses high-throughput sequencing spiking study virus detection Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/53aed62bf6024f899ba66065853665fc
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spelling	oai:doaj.org-article:53aed62bf6024f899ba66065853665fc2021-11-15T15:22:05ZA Multicenter Study To Evaluate the Performance of High-Throughput Sequencing for Virus Detection10.1128/mSphere.00307-172379-5042https://doaj.org/article/53aed62bf6024f899ba66065853665fc2017-10-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSphere.00307-17https://doaj.org/toc/2379-5042ABSTRACT The capability of high-throughput sequencing (HTS) for detection of known and unknown viruses makes it a powerful tool for broad microbial investigations, such as evaluation of novel cell substrates that may be used for the development of new biological products. However, like any new assay, regulatory applications of HTS need method standardization. Therefore, our three laboratories initiated a study to evaluate performance of HTS for potential detection of viral adventitious agents by spiking model viruses in different cellular matrices to mimic putative materials for manufacturing of biologics. Four model viruses were selected based upon different physical and biochemical properties and commercial availability: human respiratory syncytial virus (RSV), Epstein-Barr virus (EBV), feline leukemia virus (FeLV), and human reovirus (REO). Additionally, porcine circovirus (PCV) was tested by one laboratory. Independent samples were prepared for HTS by spiking intact viruses or extracted viral nucleic acids, singly or mixed, into different HeLa cell matrices (resuspended whole cells, cell lysate, or total cellular RNA). Data were obtained using different sequencing platforms (Roche 454, Illumina HiSeq1500 or HiSeq2500). Bioinformatic analyses were performed independently by each laboratory using available tools, pipelines, and databases. The results showed that comparable virus detection was obtained in the three laboratories regardless of sample processing, library preparation, sequencing platform, and bioinformatic analysis: between 0.1 and 3 viral genome copies per cell were detected for all of the model viruses used. This study highlights the potential for using HTS for sensitive detection of adventitious viruses in complex biological samples containing cellular background. IMPORTANCE Recent high-throughput sequencing (HTS) investigations have resulted in unexpected discoveries of known and novel viruses in a variety of sample types, including research materials, clinical materials, and biological products. Therefore, HTS can be a powerful tool for supplementing current methods for demonstrating the absence of adventitious or unwanted viruses in biological products, particularly when using a new cell line. However, HTS is a complex technology with different platforms, which needs standardization for evaluation of biologics. This collaborative study was undertaken to investigate detection of different virus types using two different HTS platforms. The results of the independently performed studies demonstrated a similar sensitivity of virus detection, regardless of the different sample preparation and processing procedures and bioinformatic analyses done in the three laboratories. Comparable HTS detection of different virus types supports future development of reference virus materials for standardization and validation of different HTS platforms.Arifa S. KhanSiemon H. S. NgOlivier VandeputteAisha AljanahiAvisek DeyatiJean-Pol CassartRobert L. CharleboisLanyn P. TaliaferroAmerican Society for Microbiologyarticle454 RocheIlluminaadventitious viruseshigh-throughput sequencingspiking studyvirus detectionMicrobiologyQR1-502ENmSphere, Vol 2, Iss 5 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	454 Roche Illumina adventitious viruses high-throughput sequencing spiking study virus detection Microbiology QR1-502
spellingShingle	454 Roche Illumina adventitious viruses high-throughput sequencing spiking study virus detection Microbiology QR1-502 Arifa S. Khan Siemon H. S. Ng Olivier Vandeputte Aisha Aljanahi Avisek Deyati Jean-Pol Cassart Robert L. Charlebois Lanyn P. Taliaferro A Multicenter Study To Evaluate the Performance of High-Throughput Sequencing for Virus Detection
description	ABSTRACT The capability of high-throughput sequencing (HTS) for detection of known and unknown viruses makes it a powerful tool for broad microbial investigations, such as evaluation of novel cell substrates that may be used for the development of new biological products. However, like any new assay, regulatory applications of HTS need method standardization. Therefore, our three laboratories initiated a study to evaluate performance of HTS for potential detection of viral adventitious agents by spiking model viruses in different cellular matrices to mimic putative materials for manufacturing of biologics. Four model viruses were selected based upon different physical and biochemical properties and commercial availability: human respiratory syncytial virus (RSV), Epstein-Barr virus (EBV), feline leukemia virus (FeLV), and human reovirus (REO). Additionally, porcine circovirus (PCV) was tested by one laboratory. Independent samples were prepared for HTS by spiking intact viruses or extracted viral nucleic acids, singly or mixed, into different HeLa cell matrices (resuspended whole cells, cell lysate, or total cellular RNA). Data were obtained using different sequencing platforms (Roche 454, Illumina HiSeq1500 or HiSeq2500). Bioinformatic analyses were performed independently by each laboratory using available tools, pipelines, and databases. The results showed that comparable virus detection was obtained in the three laboratories regardless of sample processing, library preparation, sequencing platform, and bioinformatic analysis: between 0.1 and 3 viral genome copies per cell were detected for all of the model viruses used. This study highlights the potential for using HTS for sensitive detection of adventitious viruses in complex biological samples containing cellular background. IMPORTANCE Recent high-throughput sequencing (HTS) investigations have resulted in unexpected discoveries of known and novel viruses in a variety of sample types, including research materials, clinical materials, and biological products. Therefore, HTS can be a powerful tool for supplementing current methods for demonstrating the absence of adventitious or unwanted viruses in biological products, particularly when using a new cell line. However, HTS is a complex technology with different platforms, which needs standardization for evaluation of biologics. This collaborative study was undertaken to investigate detection of different virus types using two different HTS platforms. The results of the independently performed studies demonstrated a similar sensitivity of virus detection, regardless of the different sample preparation and processing procedures and bioinformatic analyses done in the three laboratories. Comparable HTS detection of different virus types supports future development of reference virus materials for standardization and validation of different HTS platforms.
format	article
author	Arifa S. Khan Siemon H. S. Ng Olivier Vandeputte Aisha Aljanahi Avisek Deyati Jean-Pol Cassart Robert L. Charlebois Lanyn P. Taliaferro
author_facet	Arifa S. Khan Siemon H. S. Ng Olivier Vandeputte Aisha Aljanahi Avisek Deyati Jean-Pol Cassart Robert L. Charlebois Lanyn P. Taliaferro
author_sort	Arifa S. Khan
title	A Multicenter Study To Evaluate the Performance of High-Throughput Sequencing for Virus Detection
title_short	A Multicenter Study To Evaluate the Performance of High-Throughput Sequencing for Virus Detection
title_full	A Multicenter Study To Evaluate the Performance of High-Throughput Sequencing for Virus Detection
title_fullStr	A Multicenter Study To Evaluate the Performance of High-Throughput Sequencing for Virus Detection
title_full_unstemmed	A Multicenter Study To Evaluate the Performance of High-Throughput Sequencing for Virus Detection
title_sort	multicenter study to evaluate the performance of high-throughput sequencing for virus detection
publisher	American Society for Microbiology
publishDate	2017
url	https://doaj.org/article/53aed62bf6024f899ba66065853665fc
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A Multicenter Study To Evaluate the Performance of High-Throughput Sequencing for Virus Detection

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