pK205R targets the proximal element of IFN-I signaling pathway to assist African swine fever virus to escape host innate immunity at the early stage of infection.

African swine fever virus (ASFV) is a nuclear cytoplasmic large DNA virus (NCLDV) that causes devastating hemorrhagic diseases in domestic pigs and wild boars, seriously threatening the development of the global pig industry. IFN-I plays an important role in the body's antiviral response. Similar to other DNA viruses, ASFV has evolved a variety of immune escape strategies to antagonize IFN-I signaling and maintain its proliferation. In this study, we showed that the ASFV early protein pK205R strongly inhibited interferon-stimulated genes (ISGs) as well as the promoter activity of IFN-stimulated regulatory elements (ISREs). Mechanistically, pK205R interacted with the intracellular domains of IFNAR1 and IFNAR2, thereby inhibiting the interaction of IFNAR1/2 with JAK1 and TYK2 and hindering the phosphorylation and nuclear translocation of STATs. Subsequently, we generated a recombinant strain of the ASFV-pK205R point mutation, ASFV-pK205R7PM. Notably, we detected higher levels of ISGs in porcine alveolar macrophages (PAMs) than in the parental strain during the early stages of ASFV-pK205R7PM infection. Moreover, ASFV-pK205R7PM attenuated the inhibitory effect on IFN-I signaling. In conclusion, we identified a new ASFV immunosuppressive protein that increases our understanding of ASFV immune escape mechanisms.

African swine fever virus pB475L evades host antiviral innate immunity via targeting STAT2 to inhibit IFN-I signaling.

African swine fever virus (ASFV) causes severe disease in domestic pigs and wild boars, seriously threatening the development of the global pig industry. Type I interferon (IFN-I) is an important component of innate immunity, inducing the transcription and expression of antiviral cytokines by activating Janus-activated kinase-signal transducer and activator of transcription (STAT). However, the underlying molecular mechanisms by which ASFV antagonizes IFN-I signaling have not been fully elucidated. Therefore, using coimmunoprecipitation, confocal microscopy, and dual luciferase reporter assay methods, we investigated these mechanisms and identified a novel ASFV immunosuppressive protein, pB475L, which interacts with the C-terminal domain of STAT2. Consequently, pB475L inhibited IFN-I signaling by inhibiting STAT1 and STAT2 heterodimerization and nuclear translocation. Furthermore, we constructed an ASFV-B475L7PM mutant strain by homologous recombination, finding that ASFV-B475L7PM attenuated the inhibitory effects on IFN-I signaling compared to ASFV-WT. In summary, this study reveals a new mechanism by which ASFV impairs host innate immunity.

African Swine Fever Virus MGF505-7R Interacts with Interferon Regulatory Factor 9 to Evade the Type I Interferon Signaling Pathway and Promote Viral Replication.

African swine fever (ASF) is an acute and severe infectious disease caused by the ASF virus (ASFV). The mortality rate of ASF in pigs can reach 100%, causing huge economic losses to the pig industry. Here, we found that ASFV protein MGF505-7R inhibited the beta interferon (IFN-ß)-mediated Janus-activated kinase-signal transducer and activation of transcription (JAK-STAT) signaling. Our results demonstrate that MGF505-7R inhibited interferon-stimulated gene factor 3 (ISGF3)-mediated IFN-stimulated response element (ISRE) promoter activity. Importantly, we observed that MGF505-7R inhibits ISGF3 heterotrimer formation by interacting with interferon regulatory factor 9 (IRF9) and inhibits the nuclear translocation of ISGF3. Moreover, to demonstrate the role of MGF505-7R in IFN-I signal transduction during ASFV infection, we constructed and evaluated ASFV-ΔMGF505-7R recombinant viruses. ASFV-ΔMGF505-7R restored STAT2 and STAT1 phosphorylation, alleviated the inhibition of ISGF3 nuclear translocation, and showed increased susceptibility to IFN-ß, unlike the parental GZ201801 strain. In conclusion, our study shows that ASFV protein MGF505-7R plays a key role in evading IFN-I-mediated innate immunity, revealing a new mode of evasion for ASFV. IMPORTANCE ASF, caused by ASFV, is currently prevalent in Eurasia, with mortality rates reaching 100% in pigs. At present, there are no safe or effective vaccines against ASFV. In this study, we found that the ASFV protein MGF505-7R hinders IFN-ß signaling by interacting with IRF9 and inhibiting the formation of ISGF3 heterotrimers. Of note, we demonstrated that MGF505-7R plays a role in the immune evasion of ASFV in infected hosts and that recombinant viruses alleviated the effect on type I IFN (IFN-I) signaling and exhibited increased susceptibility to IFN-ß. This study provides a theoretical basis for developing vaccines against ASFV using strains with MGF505-7R gene deletions.

Rapid discrimination of African swine fever virus nucleic acid and virions using BenzoNuclease.

African swine fever (ASF) is an acute and severe infectious disease caused by the African Swine Fever Virus (ASFV). ASFV exhibits significant resistance and stability in the environment, which, coupled with its double-stranded DNA and large genome, predisposes it to contaminate laboratory samples. This characteristic can lead to false-positive results in swine farm settings even days after disinfection, as detectable through polymerase chain reaction (PCR) or real-time fluorescent quantitative PCR (qPCR) assays. To meet the demand for efficient clinical methods capable of discriminating between ASFV nucleic acid and ASFV virions, this study aims to ascertain the efficacy of the nuclease "BenzoNuclease" in distinguishing ASFV nucleic acid (ASFV-DNA) from ASFV virions. BenzoNuclease is a versatile nucleic acid enzyme with the capacity to degrade nearly all forms of DNA and RNA. Initially, this research established a highly sensitive general PCR detection method for ASFV. Subsequently, a positive control was constructed using the M13 bacteriophage to substitute for active ASFV, facilitating the development of an improved qPCR method. It is important to note that common disinfectants have the potential to deactivate BenzoNuclease. However, in an environment simulating actual production applications, residual disinfectants do not interfere with the enzymatic efficacy of BenzoNuclease, thus not affecting the detection capabilities of this method. Positive clinical samples from pig farms, upon testing with the improved method, revealed that three samples were positive, indicating the presence of viral particles, while the remaining samples were negative, indicating the presence of nucleic acids. This provides an additional new option for sample testing in pig farms.