African swine fever virus (ASFV) is a highly pathogenic, double-stranded DNA virus with a marked tropism for cells of the monocyte-macrophage lineage, affecting swine species and provoking severe economic losses and health threats. In the present study, four established porcine cell lines, IPAM-WT, IPAM-CD163, C∆2+ and WSL, were compared to porcine alveolar macrophage (PAM) in terms of surface marker phenotype, susceptibility to ASFV infection and virus production. The virulent ASFV Armenia/07, E70 or the naturally attenuated NHV/P68 strains were used as viral models. Cells expressed only low levels of specific receptors linked to the monocyte/macrophage lineage, with low levels of infection overall, with the exception of WSL, which showed more efficient production of strain NHV/P68 but not of strains E70 and Armenia/07.
African swine fever virus (ASFV) causes a serious disease in domestic pigs and wild boars and is currently expanding worldwide. No safe and efficacious vaccines against ASFV are available, which threats the swine industry worldwide. African swine fever virus (ASFV) is a complex dsDNA virus that displays multiple mechanisms to counteract the host innate immune response, whose efficacy might determine the different degrees of virulence displayed by attenuated and virulent ASFV strains. Here we report that infection with both virulent Arm/07/CBM/c2 and attenuated NH/P68 strains prevents interferon-stimulated gene (ISG) expression in interferon (IFN)-treated cells by counteracting the JAK/STAT pathway. This inhibition results in an impaired nuclear translocation of the interferon-stimulated gene factor 3 (ISGF3) complex, as well as in the proteasome-dependent STAT2 degradation and caspase 3-dependent STAT1 cleavage. The existence of two independent mechanisms of control of the JAK/STAT pathway, suggests the importance of preventing this pathway for successful viral replication. As ASFV virulence is likely associated with the efficacy of the IFN signaling inhibitory mechanisms, a better understanding of these IFN antagonistic properties may lead to new strategies to control this devastating pig disease.
African swine fever (ASF) is an obligated declaration swine disease, provoking farm isolation measures and the closing of affected country boarders. ASF virus (ASFV) is currently the cause of a pandemic across China and Eurasia. By the end of 2019, ASF was detected in nine EU Member States: Bulgaria, Romania, Slovakia, Estonia, Hungary, Latvia, Lithuania, Poland and Belgium. The affected area of the EU extended progressively, moving mostly in a southwestern direction (EFSA). Inactivated and/or subunit vaccines have proven to fail since certain virus replication is needed for protection. LAVs are thus the most realistic option, which must be safe, effective and industrially scalable. We here generated a vaccine prototype from the Arm/07/CBM/c2 genotype II strain, in which we have deleted the EP402R (CD2v) and A238L genes by CRISPR/Cas9 in COS-1 cells, without detectable further genetic changes. The successful immunization of pigs has proven this vaccine to be safe and fully protective against the circulating Korean Paju genotype II strain, opening the possibility of a new vaccine on the market in the near future.
African swine fever virus (ASFV) is the causative agent of one of the most lethal diseases affecting domestic pig and wild boar, which is endangering the swine industry due to its rapid expansion. ASFV has developed different mechanisms to evade the host immune response, including inhibition of type I IFN (IFN-I) production and signaling, since IFN-I is a key element in the cellular antiviral response. Here, we report a novel mechanism of evasion of the IFN-I signaling pathway carried out by the ASFV ubiquitin-conjugating enzyme pI215L. Our data showed that pI215L inhibited IFN-stimulated response element (ISRE) activity and the consecutive mRNA induction of the IFN-stimulated genes ISG15 and IFIT1 through the ubiquitination and proteasomal degradation of STAT2. Additionally, by immunofluorescence, co-immunoprecipitation and nucleus-cytoplasm fractionation approaches, we have confirmed the interaction and colocalization of STAT2 and pI215L, in ectopic experiments and during ASFV infection. Moreover, expression of the catalytic mutant (I215L-C85A) did not inhibit the induction of ISG15 and IFIT1, nor the activity of ISRE. Furthermore, we confirmed that STAT2 degradation by pI215L is dependent on its catalytic activity, since expression of the pI215L-C85A mutant did not affect STAT2 levels, compared to the wild-type protein. Yet, our data reveal that the interaction of pI215L with STAT2 does not require the integrity of its catalytic domain since the pI215L-C85A mutant co-immunoprecipitates with STAT2. All these findings reveal, for the first time, the involvement of E2-ubiquitin-conjugating enzyme activity of pI215L in the immune response modulation.
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