A Cell Culture-Adapted Vaccine Virus against the Current African Swine Fever Virus Pandemic Strain

Abstract

African swine fever virus (ASFV) causes a virulent, deadly infection in wild and domestic swine, and is currently causing a pandemic covering a contiguous geographical area from Central and Eastern Europe to Asia. No commercial vaccines are available to prevent African swine fever (ASF), resulting in devastating economic losses to the swine industry. The most advanced vaccine candidates are live attenuated strains developed using genetically modified virulent parental virus. Recently we developed a vaccine candidate, ASFV-G-ΔI177L, by deleting the I177L gene from the genome of the highly virulent pandemic ASFV strain Georgia (ASFV-G). ASFV-G-ΔI177L is safe and highly efficacious in challenge studies using parental ASFV-G. Large-scale production of ASFV-G-ΔI177L has been limited because it can only efficiently replicate in primary swine macrophages. Here we present the development of an ASFV-G-ΔI177L derivative strain, ASFV-G-ΔI177L/ΔLVR, that efficiently replicates in a stable porcine cell line. In challenge studies, ASFV-G-ΔI177L/ΔLVR maintained the same level of attenuation, immunogenic characteristics and protective efficacy as ASFV-G-ΔI177L. ASFV-G-ΔI177L/ΔLVR is the first rationally designed ASF vaccine candidate that can be used for large-scale commercial vaccine manufacturing. Importance African swine fever is currently causing a pandemic causing devastating losses to the swine industry. Experimental ASF vaccines rely on the production of vaccine in primary swine macrophages, which are difficult to use to produce a vaccine on a commercial level. Here we report a vaccine for ASFV with a deletion in the Left Variable Region. This deletion allows for growth in stable cell cultures while maintaining the potency and efficacy of the parental vaccine strain. This discovery will allow for the production of an ASF vaccine on a commercial scale.

Rapid CRISPR/Cas9 Editing of Genotype IX African Swine Fever Virus Circulating in Eastern and Central Africa

“…Additionally, the absence of SNPs, deletions, and rearrangements in these sequenced genomes is an indication that WSL may not interfere with genome integrity of the investigated ASFV. This is in contrast to previous reports where progressive adaptation of ASFV to Vero, PIPEC and CV-1 cell lines resulted in large deletions and re-arrangements in ASFV genome (Tabarés et al, 1987;Krug et al, 2015;Rodríguez et al, 2015;Mazloum et al, 2019;Borca et al, 2021).…”

“…We chose the WSL cell because it is suitable for productive replication of ASFV (Portugal et al, 2012;Keil et al, 2014;Hübner et al, 2018), and it does not interfere with genome integrity of the investigated ASFV strain (unpublished result), whereas after adaptation of ASFV to Vero cells, large deletions resulting in loss of virulence and protective capabilities were frequently observed (Tabarés et al, 1987;Krug et al, 2015;Rodríguez et al, 2015). Similar deletions and re-arrangements in ASFV genome have also been observed for viruses passaged in PIPEC (Borca et al, 2021) and CV-1 (Mazloum et al, 2019) cells. Prior to transfection, we determined that 1 µg/ml of puromycin dihydrochloride was the suitable concentration for selecting resistant clones using dose-response evaluation.…”

Adaptation of African swine fever virus to HEK293T cells

“…to replication in the porcine fetal kidney cell line (PIPEC), exhibits the same level of attenuation, immunogenic characteristics, and protective efficacy as ASFV-G-ΔI177L (Borca et al, 2021). This finding is expected to contribute to the development of ASF vaccines on a commercial scale.…”

In-yeast reconstruction of the African swine fever virus genome isolated from clinical samples

“…ASFV is a large DNA virus belonging to the Asfarviridae family with a genome size ranging between 170 and 190 kbp ( Dixon et al., 2013 ), a size problematic to be stably maintained in E. coli -based systems commonly used to generate and mutagenize molecular clones. The generation of defined ASFV mutants still relies on homologous recombination methods mainly involving eukaryotic cells such as swine macrophages ( Borca et al., 2021 ; Gallardo et al., 2018 ; Monteagudo et al., 2017 ; O’Donnell et al., 2015 ), although the use of CRISPR-Cas9 technology was recently implemented ( Borca et al., 2018 ). In addition, ASFV replication in cell culture has been reported to result in adaptive single nucleotide mutations, genome deletions and rearrangements associated with phenotype changes ( Borca et al., 2021 ; Krug et al., 2015 ).…”