Characterization of three African swine fever viruses from diferent clinical settings revealed a potential attenuation mechanism

African swine fever(ASF)is an acute and fatal hemorrhagic disease in domestic pigs and wild boars caused by African swine fever virus(ASFV)that currently threatens the pig industry worldwide.Since the 2018 ASF outbreak in China,ASFV has evolved and caused diverse clinical manifestations,such as chronic and asymptomatic infections.Therefore,it is important to understand the molecular mechanisms underlying ASFV attenuation in the feld.Here,we isolated three ASFVs from one diseased and two asymptomatic pigs by using primary porcine alveolar macrophages(PAMs)from both domestic pigs and Bama minipigs.The three ASFVs exhibited similar phenotypes in cell culture,includ‑ing cytopathic efects(CPEs),hemadsorptions(HADs),viral protein expressions and growth curves.Genome sequenc‑ing revealed that all three ASFVs were genotype II strains.Genomic comparisons suggested that the disruption of the viral genes MGF360 and MGF110,rather than EP402R and EP153R,is likely involved in the potential attenuation of ASFV via the upregulation of innate immune responses.

Development and application of a SYBR Green I fuorescent PCR assay for the diferentiation of genotypes I and II African swine fever viruses

African swine fever(ASF)is a highly fatal hemorrhagic disease afecting domestic pigs caused by African swine fever virus(ASFV).Genetic analysis of ASFV isolates to date has identifed 24 geographically related genotypes with various subgroups,but only genotype I and II ASFVs have been reported outside Africa.ASFV genotype II and genotype I viruses were reported in China in 2018 and 2021,respectively.In this study,unique and highly conserved noncoding regions were found between MGF_505-9R and MGF_505-10R in the 188 genomes of ASFV genotypes I and II.A pair of primers was designed on the basis of this region.By optimizing the reaction system and conditions,a SYBR Green I fuorescence PCR assay that can distinguish between ASFV genotypes I and II was established,and the sensitivity,reproducibility and specifcity were evaluated.The detection limit was 1 TCID_(50)/0.1 mL for both genotypes,with no cross-reactivity observed with other common pig pathogens.The intra-and interbatch variation coefcients were both less than 1.2%.Clinical sample detection analysis revealed 47 positive cases out of 100,including 3 for genotype I and 44 for genotype II,aligning with results from the WOAH-recommended and national standard methods.The method developed in this study allows for the diferentiation of ASFV genotypes I and II without the need for genome sequencing,ofering a convenient and rapid approach for ASFV detection and genotype identifcation.

Inhibition of African swine fever virus in liquid and feed by medium-chain fatty acids and glycerol monolaurate

Background:The ongoing African swine fever virus(ASFv)epidemic has had a major impact on pig production globally and biosecurity efforts to curb ASFv infectivity and transmission are a high priority.It has been recently identified that feed and feed ingredients,along with drinking water,can serve as transmission vehicles and might facilitate transboundary spread of ASFv.Thus,it is important to test the antiviral activity of regulatory compatible,antiviral feed additives that might inhibit ASFv infectivity in feed.One promising group of feed additive candidates includes medium-chain fatty acids(MCFA)and monoglyceride derivatives,which are known to disrupt the lipid membrane surrounding certain enveloped viruses and bacteria.Results:The antiviral activities of selected MCFA,namely caprylic,capric,and lauric acids,and a related monoglyceride,glycerol monolaurate(GML),to inhibit ASFv in liquid and feed conditions were investigated and suitable compounds and inclusion rates were identified that might be useful for mitigating ASFv in feed environments.Antiviral assays showed that all tested MCFA and GML inhibit ASFv.GML was more potent than MCFA because it worked at a lower concentration and inhibited ASFv due to direct virucidal activity along with one or more other antiviral mechanisms.Dose-dependent feed experiments further showed that sufficiently high GML doses can significantly reduce ASFv infectivity in feed in a linear manner in periods as short as 30 min,as determined by infectious viral titer measurements.Enzyme-linked immunosorbent assay(ELISA)experiments revealed that GML treatment also hinders antibody recognition of the membrane-associated ASFv p72 structural protein,which likely relates to protein conformational changes arising from viral membrane disruption.Conclusion:Together,the findings in this study indicate that MCFA and GML inhibit ASFv in liquid conditions and that GML is also able to reduce ASFv infectivity in feed,which may help to curb disease transmission.

Insights into African swine fever virus immunoevasion strategies

African swine fever(ASF) is an acute and highly contagious disease that causes severe economic losses to the swine industry. ASF is caused by infection of African swine fever virus(ASFV) in domestic pigs, leading to almost 100% mortality. However, no effective vaccines and pharmacologic treatment against ASF are available. ASF poses a severe threat to the swine industry and the economy. Here we summarize potential virus-host cell interaction mechanisms involving the suppression of innate and adaptive immune responses to ASFV entry and infection. These mechanisms include modulation of apoptosis, inhibition of inflammatory responses, reduction of IFN production, inhibition of autophagy, and suppression of MHC-I expression. Insights into immunoevasion strategies by ASFV may shed light on the development of vaccines, as well as preventive and therapeutic drugs.

A multiplex real-time PCR assay for simultaneous detection of classical swine fever virus,African swine fever virus,and atypical porcine pestivirus

With the implementation of the C-strain vaccine,classical swine fever(CSF) has been under control in China,which is currently in a chronic atypical epidemic situation.African swine fever(ASF) emerged in China in 2018 and spread quickly across the country.It is presently occurring sporadically due to the lack of commercial vaccines and farmers’ increased awareness of biosafety.Atypical porcine pestivirus(APPV) was first detected in Guangdong Province,China,in 2016,which mainly harms piglets and has a local epidemic situation in southern China.These three diseases have similar clinical symptoms in pig herds,which cause considerable losses to the pig industry.They are difficult to be distinguished only by clinical diagnosis.Therefore,developing an early and accurate simultaneous detection and differential diagnosis of the diseases induced by these viruses is essential.In this study,three pairs of specific primers and Taq-man probes were designed from highly conserved genomic regions of CSFV(5’ UTR),African swine fever virus(ASFV)(B646L),and APPV(5’ UTR),followed by the optimization of reaction conditions to establish a multiplex real-time PCR detection assay.The results showed that the method did not cross-react with other swine pathogens(porcine circovirus type 2(PCV2),porcine reproductive and respiratory syndrome virus(PRRSV),foot-and-mouth disease virus(FMDV),pseudorabies virus(PRV),porcine parvovirus(PPV),and bovine viral diarrhea virus BVDV).The sensitivity results showed that CSFV,ASFV,and APPV could be detected as low as 1 copy μL–1;the repeatability results showed that the intra-assay and interassay coefficient of variation of ASFV,CSFV,and APPV was less than 1%.Twenty-two virus samples were detected by the multiplex real-time PCR,compared with national standard diagnostic and patented method assay for CSF(GB/T 27540–2011),ASF(GB/T 18648–2020),and APPV(CN108611442A),respectively.The sensitivity of this triple real-time PCR for CSFV,ASFV,and APPV was almost the same,and the compliance results were the same(100%).A total of 451 clinical samples were detected,and the results showed that the positive rates of CSFV,ASFV,and APPV were 0.22% (1/451),1.3%(6/451),and 0%(0/451),respectively.This assay provides a valuale tool for rapid detection and accurate diagnosis of CSFV,ASFV,and APPV.

Development of a recombinant pB602L-based indirect ELISA assay for detecting antibodies against African swine fever virus in pigs

African swine fever(ASF),caused by the African swine fever virus(ASFV),is a devastating disease of domestic and wild pigs.There is no effective vaccine,and the control of the disease relies mainly on surveillance and early detection of infected pigs.Previously,serological assays,such as ELISA,have been developed mainly based on recombinant structural viral proteins of ASFV,including p72,p54,and p30.However,the antibodies against these proteins do not provide efficient protection against ASFV infection in pigs.Therefore,new serological assays that can be applied for clinical diagnosis and evaluating serological immune response in vaccinated pigs are still required.In this study,we expressed and purified a recombinant p B602 L protein.The purified p B602 L protein was then used as an antigen to develop an indirect ELISA assay.This assay has no cross-reaction with the anti-sera against the 15 most common pig pathogens in China,such as classical swine fever virus,pseudorabies virus,and porcine parvovirus.This assay and a commercial ELISA kit were then used to detect 60 field pig serum samples,including an unknown number of antiASFV sera.The coincidence of the two assays was 95%.Furthermore,the p B602 L-based ELISA was employed to test the antibody responses to the seven-gene-deleted ASFV strain HLJ/18-7 GD in pigs.The results showed that the antibody levels in all vaccinated pigs,starting from the 10 th day post-inoculation,have increased continuously during the observation period of 45 days.Our results indicate that this p B602 L-based indirect ELISA assay can be employed potentially in the field of ASFV diagnosis.

Development of a Recombinase-aided Amplification Combined With Lateral Flow Dipstick Assay for the Rapid Detection of the African Swine Fever Virus

Objective To establish a sensitive,simple and rapid detection method for African swine fever virus(ASFV)B646L gene.Methods A recombinase-aided amplification-lateral flow dipstick(RAA-LFD)assay was developed in this study.Recombinase-aided amplification(RAA)is used to amplify template DNA,and lateral flow dipstick(LFD)is used to interpret the results after the amplification is completed.The lower limits of detection and specificity of the RAA assay were verified using recombinant plasmid and pathogenic nucleic acid.In addition,30 clinical samples were tested to evaluate the performance of the RAA assay.Results The RAA-LFD assay was completed within 15 min at 37°C,including 10 min for nucleic acid amplification and 5 minutes for LFD reading results.The detection limit of this assay was found to be 200 copies per reaction.And there was no cross-reactivity with other swine viruses.Conclusion A highly sensitive,specific,and simple RAA-LFD method was developed for the rapid detection of the ASFV.

African swine fever virus MGF505-3R inhibits cGAS-STING-mediated IFN-βpathway activation by degrading TBK1

African swine fever virus(ASFV)is an important pathogen causing acute infectious disease in domestic pigs and wild boars that seriously endangers the global swine industry.As ASFV is structurally complex and encodes a large number of functional proteins,no effective vaccine has been developed to date.Thus,dissecting the mechanisms of immune escape induced by ASFV proteins is crucial.A previous study showed that the ASFV-encoded protein is an important factor in host immunity.In this study,we identified a negative regulator,MGF505-3R,that significantly downregulated cGAS/STING-and poly(dG:dC)-mediated IFN-βand interferon stimulation response element(ISRE)reporter activity and suppressed IFNB1 and IFIT2 mRNA levels.In addition,TBK1,IRF3 and IκBαphosphorylation levels were also inhibited.Mechanistically,MGF505-3R interacted with cGAS/TBK1/IRF3 and targeted TBK1 for degradation,thereby disrupting the cGAS-STING-mediated IFN-βsignaling pathway,which appears to be highly correlated with autophagy.Knockdown MGF505-3R expression enhanced IFN-βand IL-1βproduction.Taken together,our study revealed a negative regulatory mechanism involving the MGF505-3R-cGAS-STING axis and provided insights into an evasion strategy employed by ASFV that involves autophagy and innate signaling pathways.

Viricidal activity of several disinfectants against African swine fever virus

Prevention of African swine fever,a disease caused by African swine fever virus(ASFV),requires maintenance of high biosecurity standards,which principally relies on disinfection.Finding the perfect disinfectant against ASFV is difficult because of the lack of relevant data.Therefore,we aimed to find the most effective disinfectant and to optimise its concentration as well as contact time to confirm the viricidal effect against ASFV in vitro.We evaluated the viricidal activity of three concentrations each of six common disinfectants against ASFV using immersion disinfection assay(IDA)and spray disinfection assay(SDA);the concentrations of these disinfectants at which complete viral inactivation occurred were almost same as the manufacturer-recommended concentrations,but the exposure times for viral inactivation are different.The following disinfectants(assay:concentration,exposure time)showed complete inactivation:iodine and acid mixed solution(IDA/SDA:0.5%,10 min);compound potassium peroxymonosulfate(IDA:0.25%,30 min;SDA:0.25%,60 min);citric acid(IDA:0.25%,60 min;SDA:0.5%,60 min);sodium dichloroisocyanurate(IDA:0.125%,60 min;SDA:0.25%,60 min);and glutaral ang deciquam(IDA/SDA:0.2%,60 min);and deciquam(IDA/SDA:0.5%,60 min).However,in the presence of organic material contamination,disinfectants did not show a marked inactivation effect.Therefore,disinfection procedures should be performed in two steps:thorough mechanical cleaning followed by application of disinfectant.In conclusion,all the tested disinfectants can inactivate ASFV;these can be used as alternative disinfectants to enhance biosecurity.

The Prediction of B Cell Epitopes for VP73 Protein of African Fever Virus

[Objective] The B cell epitopes for VP73 protein of African swine fever virus was predicted. [ Method] Based on the analysis of the amino acid sequence and the flexible regions of VP73 protein, the B cell epitopes for VP73 protein of African swine fever virus were predicted by method of Kyte-Doolittie, Emini and Jameson-Wolf. [Result] The B cell epitopes were located at or adjacent to the N-terminal No. 11 - 18,26 -48,73 -82,136 - 150,159 - 174,181 - 189,191 - 210,247 - 276,279 - 295,313 - 323 and 382 - 392. [Conclusion] The multi-parameters analytic method was adopted to predict the B cell epitopes for VP73 protein of African swine fever virus, which laid solid foundation for further characterizing the protein of VP73 and researching epitope vaccine.

A Universal Real-Time Fluorescence qPCR Method for Identifying Epidemic Strains of African Swine Fever Virus

Objective Establishing a highly sensitive real-time fluorescence quantitative PCR (qPCR) method for universal testing of epidemic African swine fever virus (ASFV) strains. Methods The ASFV p72 gene was targeted to design primer probes covering 24 p72 genotypes. The optimal amount of dimethylsulphoxide (DMSO) for qPCR amplification was determined, Various sensitivity and limit of detection (LOD) tests were performed, and clinical samples from China and imported goods were tested. Results The optimal primer-probe combination could specifically detect ASFV, 1.5% DMSO was optimal for qPCR, and LOD reached 3.2 copies/μL with good reproducibility (n = 20, p = 0.369). The method was employed to test 142 clinically suspected samples, of which 30 pig blood and 37 pig tissue samples were ASFV-positive. Moreover, the positive testing rate for ASFV was higher than for the standard qPCR method recommended by the Office International Des Epizooties (OIE), and for the commercially available kit. Thus, our method is superior for testing weakly positive samples with low virus titre, and epidemic strains present in imported goods. Conclusion Our method could be employed for universal testing of epidemic ASFV strains worldwide, ensuring wider coverage of hosts and ASFV strains/endemic strains, reducing false negatives, and benefitting early diagnosis.

Nanobodies against African swine fever virus p72 and CD2v proteins as reagents for developing two cELISAs to detect viral antibodies

African swine fever virus(ASFV)poses a significant threat to the global swine industry.Currently,there are no effective vaccines or treatments available to combat ASFV infection in pigs.The primary means of controlling the spread of the disease is through rapid detection and subsequent elimination of infected pig.Recently,a lower virulent ASFV isolate with a deleted EP402R gene(CD2v-deleted)has been reported in China,which further complicates the control of ASFV infection in pig farms.Furthermore,an EP402R-deleted ASFV variant has been developed as a potential live attenuated vaccine candidate strain.Therefore,it is crucial to develop detection methods that can distinguish wild-type and EP402R-deleted ASFV infections.In this study,two recombinant ASFV-p72 and-CD2v proteins were expressed using a prokaryotic system and used to immunize Bactrian camels.Subsequently,eight nanobodies against ASFV-p72 and ten nanobodies against ASFV-CD2v were screened.Following the production of these nanobodies with horse radish peroxidase(HRP)fusion proteins,the ASFV-p72-Nb2-HRP and ASFV-CD2v-Nb22-HRP fusions were selected for the development of two competitive ELISAs(cELISAs)to detect anti-ASFV antibodies.The two cELISAs exhibited high sensitivity,good specificity,repeatability,and stability.The coincidence rate between the two cELISAs and commercial ELISA kits was 98.6%and 97.6%,respectively.Collectively,the two cELISA for detecting antibodies against ASFV demonstrated ease of operation,a low cost,and a simple production process.The two cELISAs could determine whether pigs were infected with wild-type or CD2v-deleted ASFV,and could play an important role in monitoring ASFV infections in pig farms.

Identification of host proteins that interact with African swine fever virus pE301R

African swine fever virus(ASFV)infection poses enormous threats and challenges to the global pig industry;however,no effective vaccine is available against ASFV,attributing to the huge viral genome(approximately189 kb)and numerous encoding products(>150 genes)due to the limited understanding on the molecular mechanisms of viral pathogenesis.Elucidating the host-factor/viral-protein interaction network will reveal new targets for developing novel antiviral therapies.Using proteomic analysis,we identified 255 cellular proteins that interact with the ASFV-encoded pE301R protein when transiently expressed in HEK293T cells.Gene ontology(GO)annotation,Kyoto Encyclopedia of Genes and Genomes(KEGG)database enrichment,and protein-protein interaction(PPI)network analyses revealed that pE301R-interacting host proteins are potentially involved in various biological processes,including protein translation and folding,response to stimulation,and mitochondrial transmembrane transport.The interactions of two putative cellular proteins(apoptosis inducing factor mitochondria associated 1(AIFM1)and vimentin(VIM))with pE301R-apoptosis inducing factor have been verified by co-immunoprecipitation.Our study revealed the inhibitory role of pE301R in interferon(IFN)induction that involves VIM sequestration by pE301R,identified interactions between ASFV pE301R and cellular proteins,and predicted the potential function of pE301R and its associated biological processes,providing valuable information to enhance our understanding of viral protein function,pathogenesis,and potential candidates for the prevention and control of ASFV infection.

Regulation of antiviral immune response by African swine fever virus(ASFV)

African swine fever(ASF)is a highly contagious and acute hemorrhagic viral disease with a high mortality approaching 100%in domestic pigs.ASF is an endemic in countries in sub-Saharan Africa.Now,it has been spreading to many countries,especially in Asia and Europe.Due to the fact that there is no commercial vaccine available for ASF to provide sustainable prevention,the disease has spread rapidly worldwide and caused great economic losses in swine industry.The knowledge gap of ASF virus(ASFV)pathogenesis and immune evasion is the main factor to limit the development of safe and effective ASF vaccines.Here,we will summarize the molecular mechanisms of how ASFV interferes with the host innate and adaptive immune responses.An in-depth understanding of ASFV immune evasion strategies will provide us with rational design of ASF vaccines.

稳定表达猪BRD4-BD1/2蛋白的猪肺泡巨噬细胞传代细胞系的构建及其用于ASFV增殖的效果观察

前期研究发现宿主表观遗传调控蛋白——含溴结构域蛋白质4(bromodomain-containing protein 4,BRD4)有助于非洲猪瘟病毒(African swine fever virus,ASFV)的复制,为了深入研究BRD4对ASFV复制的影响,通过筛选出显著促进ASFV复制的BRD4-BD1/2结构域,利用慢病毒表达系统成功构建稳定表达BRD4-BD1/2结构域的3D4/21细胞系,分析ASFV在3D4/21-BRD4-BD1/2细胞系与WT细胞系之间的复制差异。首先,以家猪基因BRD4-BD1/2为靶标,构建了含有3x Flag标签的重组质粒pLVX-IRES-puro-3x Flag-BRD4-BD1/2,并将其与质粒pMD2.G和pSPAX2共同转染至HEK-293T细胞进行慢病毒包装,获得具有感染能力的慢病毒。使用慢病毒感染3D4/21细胞后通过嘌呤霉素药物筛选,成功获得了稳定表达BRD4-BD1/2结构域的3D4/21细胞系。利用RT-qPCR和Western blot技术分别检测了ASFV感染3D4/21-BRD4-BD1/2细胞后CP204L和B602L基因的转录水平以及相应蛋白表达水平,并通过HAD50测定评价ASFV的复制能力。研究结果表明:成功构建了稳定表达BRD4-BD1/2结构域的3D4/21细胞系。与3D4/21-WT细胞系相比,BRD4-BD1/2稳定表达细胞系能够显著促进ASFV复制。本研究提供了深入研究BRD4蛋白在ASFV复制中功能的生物材料,并为ASFV疫苗候选株的开发提供了理论基础。

Diverse immune cell profles in ASFV-associated lymphopenia

Pathogenic African swine fever virus(ASFV)remains a lethal causative agent in the domestic pig industry,which poses a burden on the swine market and causes substantial socioeconomic losses worldwide.Currently,there are no commercially efcacious vaccines or specifc treatments available for ASF prevention and control.Unfortunately,little is known about the swine immune response upon ASFV infection.Here,we investigated the host immune response discrepancy induced by the feld moderately virulent strain ASFV HB-2208 among healthy,diseased and asymptomatic pigs.In the peripheral blood of diseased swine,lymphopenia is caused by the massive loss of bystander lymphocytes,such asγδT cells,B cells and CD4^(+)T cells.Conversely,ASFV has a strong tropism for the mononuclear phagocyte system(MPS)and partial dendritic cells(DCs),whose antigen-presenting ability is impeded by the downregulation of CD80 and MHC I.However,no signifcant diference in the number of CD8α^(high) T cells was detected,whereas the frequencies of NK cells,NKT cells,and regulatory T cells(Tregs)were signifcantly increased.Additionally,an in vitro model was established with a coculture of primary pulmonary alveolar macrophages(PAMs)and peripheral blood mononuclear cells(PBMCs),which signifcantly reducedγδT cells,B cells and CD4^(+)T cells and increased Tregs.The diferentiated immune response might aid in enhancing the understanding of ASFV pathogenesis in suids and provide insights into the mechanism of ASFV-induced lymphopenia for further studies.

基于非洲猪瘟病毒B646 L和I177L基因双重实时荧光定量PCR鉴别ASFV-G-ΔI177 L疫苗株的检测方法

为建立一种可鉴别非洲猪瘟病毒(ASFV)野毒株与ASFV-G-ΔI177L(I177L基因缺失株)疫苗株的实时荧光定量PCR检测方法,本试验针对非洲猪瘟病毒B646L与I177L两种基因保守性高的区域,分别设计了引物和探针,通过引物、探针最佳工作浓度筛选和退火温度优化,建立了一种双重实时荧光定量PCR核酸检测方法,并验证该方法的敏感性、特异性、重复性。结果显示,该检测方法建立的标准曲线线性关系良好,R2均>0.99;对pUC57-B646L和pUC57-I177L质粒标准品最低检出限均为10 copies/μL;与其他常见猪病病原无交叉反应,特异性强;批内变异系数为0.11%~1.39%,批间变异系数为0.21%~0.96%,均小于1.4%,重复性和稳定性良好。结果表明,本试验成功建立了一种性能良好的非洲猪瘟病毒B646L与I177L基因双重实时荧光定量PCR核酸检测方法,可为临床上非洲猪瘟病毒野毒株和ASFV-G-ΔI177L疫苗株的检测和鉴别提供技术支撑。

Emergence and prevalence of naturally occurring lower virulent African swine fever viruses in domestic pigs in China in 2020

African swine fever virus(ASFV)has been circulating in China for more than two years,and it is not clear whether the biological properties of the virus have changed.Here,we report on our surveillance of ASFVs in seven provinces of China,from June to December,2020.A total of 22 viruses were isolated and characterized as genotype II ASFVs,with mutations,deletions,insertions,or short-fragment replacement occurring in all isolates compared with Pig/HLJ/2018(HLJ/18),the earliest isolate in China.Eleven isolates had four different types of natural mutations or deletion in the EP402R gene and displayed a nonhemadsorbing(non-HAD)phenotype.Four isolates were tested for virulence in pigs;two were found to be as highly lethal as HLJ/18.However,two non-HAD isolates showed lower virulence but were highly transmissible;infection with 106 TCID50 dose was partially lethal and caused acute or sub-acute disease,whereas 103 TCID50 dose caused non-lethal,sub-acute or chronic disease,and persistent infection.The emergence of lower virulent natural mutants brings greater difficulty to the early diagnosis of ASF and creates new challenges for ASFV control.

African Swine Fever Virus MGF360-12L Inhibits Type Ⅰ Interferon Production by Blocking the Interaction of Importin α and NF-κB Signaling Pathway

African swine fever(ASF)is an infectious transboundary disease of domestic pigs and wild boar and spreading throughout Eurasia.There is no vaccine and treatment available.Complex immune escape strategies of African swine fever virus(ASFV)are crucial factors affecting immune prevention and vaccine development.MGF360 genes have been implicated in the modulation of the IFN-Ⅰresponse.The molecular mechanisms contributing to innate immunity are poorly understood.In this study,we demonstrated that ASFV MGF360-12 L(MGF360 families 12 L protein)significantly inhibited the mRNA transcription and promoter activity of IFN-βand NF-κB,accompanied by decreases of IRF3,STING,TBK1,ISG54,ISG56 and AP-1 m RNA transcription.Also,MGF360-12 L might suppress the nuclear localization of p50 and p65 mediated by classical nuclear localization signal(NLS).Additionally,MGF360-12 L could interact with KPNA2,KPNA3,and KPNA4,which interrupted the interaction between p65 and KPNA2,KPNA3,KPNA4.We further found that MGF360-12 L could interfere with the NF-κB nuclear translocation by competitively inhibiting the interaction between NF-κB and nuclear transport proteins.These findings suggested that MGF360-12 L could inhibit the IFN-Ⅰproduction by blocking the interaction of importinαand NF-κB signaling pathway,which might reveal a novel strategy for ASFV to escape the host innate immune response.

African Swine Fever Virus MGF-110-9L-deficient Mutant Has Attenuated Virulence in Pigs

African swine fever virus(ASFV)is the etiological agent of African swine fever(ASF),an often lethal disease in domestic and wild pigs.ASF represents a major threat to the swine industry worldwide.Currently,no commercial vaccine is available because of the complexity of ASFV or biosecurity concerns.Live attenuated viruses that are naturally isolated or genetically manipulated have demonstrated reliable protection against homologous ASFV strain challenge.In the present study,a mutant ASFV strain with the deletion of ASFV MGF-110-9L(ASFV-D9L)was generated from a highly virulent ASFV CN/GS/2018 parental strain,a genotypeⅡASFV.Relative to the parental ASFV isolate,deletion of the MGF-110-9L gene significantly decreased the ability of ASFV-D9L to replicate in vitro in primary swine macrophage cell cultures.The majority of animals inoculated intramuscularly with a low dose of ASFV-D9L(10 HAD50)remained clinically normal during the 21-day observational period.Three of five ASFV-D9L-infected animals displayed low viremia titers and low virus shedding and developed a strong virus-specific antibody response,indicating partial attenuation of the ASFV-D9L strain in pigs.The findings imply the potential usefulness of the ASFV-D9L strain for further development of ASF control measures.