NGS method by library enrichment for rapid pestivirus purity testing in biologics.

Autor: La Polla R; Boehringer Ingelheim Animal Health, Site Lyon porte des Alpes, 813 cours du 3eme Millénaire, 69800 Saint Priest, France; Laboratoire d'Écologie Microbienne - UMR 5557, Université Claude Bernard Lyon 1, 43 Boulevard du 11 Novembre 1918, 69622 Villeurbanne Cedex, France., Goumaidi A; Viroscan3D, Faculté de Médecine et de Pharmacie, 8 avenue Rockefeller, 69373 Lyon, France., Daniau M; Viroscan3D, Faculté de Médecine et de Pharmacie, 8 avenue Rockefeller, 69373 Lyon, France., Legras-Lachuer C; Laboratoire d'Écologie Microbienne - UMR 5557, Université Claude Bernard Lyon 1, 43 Boulevard du 11 Novembre 1918, 69622 Villeurbanne Cedex, France., De Saint-Vis B; Boehringer Ingelheim Animal Health, Site Lyon porte des Alpes, 813 cours du 3eme Millénaire, 69800 Saint Priest, France.
Jazyk: angličtina
Zdroj: Vaccine [Vaccine] 2023 Jan 16; Vol. 41 (3), pp. 855-861. Date of Electronic Publication: 2022 Dec 21.
DOI: 10.1016/j.vaccine.2022.12.040
Abstrakt: NGS sequencing was evaluated to understand its added value for animal health vaccine candidates. We have previously established the proof of concept for its application in purity testing on several Master Seeds. Here we evaluate the NGS method after enrichment to detect pestiviruses. To achieve this, we conducted a spiking study using 6 viruses, consisting of 3 pestiviruses and 3 other RNA-viruses at different concentrations into cell suspension. A deep Illumina random sequencing of all nucleic acids (DNA and RNA) was performed. The bioinformatics analysis including both assembly into contigs and annotation were processed using viral public databases for the spiked viruses' identification. Here we present the results of spiking experiments for the simultaneous spike of 6 viruses at 100-10 and 1 TCID 50 /ml. Using Illumina sequencing, the 3 pestiviruses were all detected at the highest concentration, and even at the lowest one such as 1 TCID 50 /ml for CSFV. Regarding the other viruses, they were not detected at all. Overall, the study showed consistent results for specific detection of pestiviruses with an increase of sensitivity after enrichment. The sensitivity of NGS evaluated by virus spiking experiments of cells demonstrated that NGS method is a valuable and sensitive tool for specific agent detection required in purity testing during vaccine development. This NGS method should be considered as an alternative tool of current purity testing for the prospective testing of biological products.
Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2022. Published by Elsevier Ltd.)
Databáze: MEDLINE
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