Comparison of Nucleic Acid Extraction Methods for a Viral Metagenomics Analysis of Respiratory Viruses

Microorganisms

Grégory Quéromès¹  Valérie Cheynet²  Karen Brengel-Pesce²  Laurence Josset³  Hadrien Regue³  Bruno Lina³  Marina Sabatier³  Antonin Bal³  Grégory Destras³  Vincent Navratil⁴  Claire Bardel⁵ 

[1] CIRI, Centre International de Recherche en Infectiologie, Team VirPatH, Univ Lyon, Inserm, U1111, Université Claude Bernard Lyon 1, CNRS, UMR5308, ENS de Lyon, F-69007 Lyon, France;Laboratoire Commun de Recherche Hospices Civils de Lyon - bioMérieux, Centre Hospitalier Lyon Sud, F-69310 Pierre-Bénite, France;Laboratoire de Virologie, Institut des Agents Infectieux (IAI), Hospices Civils de Lyon, Groupement Hospitalier Nord, F-69004 Lyon, France;PRABI, Rhône Alpes Bioinformatics Center, UCBL, Université Claude Bernard Lyon 1, F-69000 Lyon, France;Université Lyon 1, Laboratoire de Biométrie et Biologie Evolutive, CNRS UMR5558, F-69100 Villeurbanne, France;

关键词: viral metagenomics;    next-generation sequencing;    acid nucleic extraction;    sample cross-contamination;    kitome;   

DOI  :  10.3390/microorganisms8101539

来源: DOAJ

【 摘 要 】

Viral metagenomics next-generation sequencing (mNGS) is increasingly being used to characterize the human virome. The impact of viral nucleic extraction on virome profiling has been poorly studied. Here, we aimed to compare the sensitivity and sample and reagent contamination of three extraction methods used for viral mNGS: two automated platforms (eMAG; MagNA Pure 24, MP24) and the manual QIAamp Viral RNA Mini Kit (QIAamp). Clinical respiratory samples (positive for Respiratory Syncytial Virus or Herpes Simplex Virus), one mock sample (including five viruses isolated from respiratory samples), and a no-template control (NTC) were extracted and processed through an mNGS workflow. QIAamp yielded a lower proportion of viral reads for both clinical and mock samples. The sample cross-contamination was higher when using MP24, with up to 36.09% of the viral reads mapping to mock viruses in the NTC (vs. 1.53% and 1.45% for eMAG and QIAamp, respectively). The highest number of viral reads mapping to bacteriophages in the NTC was found with QIAamp, suggesting reagent contamination. Our results highlight the importance of the extraction method choice for accurate virome characterization.

【 授权许可】

Unknown