Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

A Deep Sequencing Strategy for Investigation of Virus Variants Within African Swine Fever Virus-infected Pigs

Camille Melissa Johnston
,
Ann Sofie Olesen
ORCID logo ,
Louise Lohse
ORCID logo ,
Agnete le Maire Madsen
,
Anette Bøtner
ORCID logo ,
Graham J. Belsham
ORCID logo ,
Thomas Bruun Rasmussen
*
Version 1 : Received: 15 December 2023 / Approved: 18 December 2023 / Online: 18 December 2023 (09:08:56 CET)

A peer-reviewed article of this Preprint also exists.

Johnston, C.M.; Olesen, A.S.; Lohse, L.; le Maire Madsen, A.; Bøtner, A.; Belsham, G.J.; Rasmussen, T.B. A Deep Sequencing Strategy for Investigation of Virus Variants within African Swine Fever Virus-Infected Pigs. Pathogens 2024, 13, 154. Johnston, C.M.; Olesen, A.S.; Lohse, L.; le Maire Madsen, A.; Bøtner, A.; Belsham, G.J.; Rasmussen, T.B. A Deep Sequencing Strategy for Investigation of Virus Variants within African Swine Fever Virus-Infected Pigs. Pathogens 2024, 13, 154.

Abstract

African swine fever virus (ASFV) is the causative agent of African swine fever, an economically important disease of pigs, often with a high case fatality rate. ASFV has demonstrated low genetic diversity among isolates collected within Eurasia. To explore the influence of viral variants on clinical outcomes and infection-dynamics in pigs experimentally infected with ASFV, we have designed a deep sequencing strategy. Variant analysis revealed 3 distinct SNPs present in the virus inoculum (at 2.4%, 0.7%, and 13% frequency, respectively) that were maintained within all infected pigs (1-6 % frequency, and the latter at 16-21%). Several pigs displayed other unique SNPs at <10% frequency. In addition, a deletion of 10487 bp (resulting in the complete loss of 21 genes) was present at nearly 100% frequency in the ASFV DNA from one pig at position 6362-16849. This deletion was also found to be present at low levels in the virus inoculum and in two other infected pigs. The current methodology can be used for the currently circulating Eurasian ASFVs and also adapted to other ASFV strains and genotypes. Comprehensive deep sequencing is critical for following ASFV molecular evolution, especially for identification of modifications that affect virus virulence.

Keywords

African swine fever virus (ASFV); next-generation sequencing (NGS); deep sequencing; Nanopore sequencing; minority variant analysis

Subject

Biology and Life Sciences, Virology

Copyright: This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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