Identification and epitope mapping of anti-p72 single-chain antibody against African swine fever virus based on phage display antibody library

African swine fever virus(ASFV)is a lethal pathogen that causes severe threats to the global swine industry and it has already had catastrophic socio-economic effects.To date,no licensed prophylactic vaccine exists.Limited knowledge exists about the major immunogens of ASFV and the epitope mapping of the key antigens.As such,there is a considerable requirement to understand the functional monoclonal antibodies(mAbs)and the epitope mapping may be of utmost importance in our understanding of immune responses and designing improved vaccines,therapeutics,and diagnostics.In this study,we generated an ASFV antibody phage-display library from ASFV convalescent swine PBMCs,further screened a specific ASFV major capsid protein(p72)single-chain antibody and fused with an IgG Fc fragment(scFv-83-Fc),which is a specific recognition antibody against ASFV Pig/HLJ/2018 strain.Using the scFv-83-Fc mAb,we selected a conserved epitope peptide(221MTGYKH226)of p72 retrieved from a phage-displayed random peptide library.Moreover,flow cytometry and cell uptake experiments demonstrated that the epitope peptide can significantly promote BMDCs maturation in vitro and could be effectively uptaken by DCs,which indicated its potential application in vaccine and diagnostic reagent development.Overall,this study provided a valuable platform for identifying targets for ASFV vaccine development,as well as to facilitate the optimization design of subunit vaccine and diagnostic reagents.

The multi-kingdom microbiome catalog of the chicken gastrointestinal tract

Chicken is an important food animal worldwide and plays an important role in human life by providing meat and eggs.Despite recent significant advances in gut microbiome studies,a comprehensive study of chicken gut bacterial,archaeal,and viral genomes remains unavailable.In this study,we constructed a chicken multi-kingdom microbiome catalog(CMKMC),including 18,201 bacterial,225 archaeal,and 33,411 viral genomes,and annotated over 6,076,006 protein-coding genes by integrating 135 chicken gut metagenomes and publicly available metagenome-assembled genomes(MAGs)from ten countries.We found that 812 and 240 MAGs in our dataset were putative novel species and genera,respectively,far beyond what was previously reported.The newly unclassified MAGs were predominant in Phyla Firmicutes_A(n=263),followed by Firmicutes(n=126),Bacteroidota(n=121),and Proteobacteria(n=87).Most of the classified species-level viral oper-ational taxonomic units belong to Caudovirales.Approximately,63.24%of chicken gut viromes are predicted to infect two or more hosts,including complete circular viruses.Moreover,we found that diverse auxiliary metabolic genes and antibiotic resistance genes were carried by viruses.Together,our CMKMC provides the largest integrated MAGs and viral genomes from the chicken gut to date,functional insights into the chicken gastrointestinal tract microbiota,and paves the way for microbial interventions for better chicken health and productivity.

NPC1-regulated dynamic of clathrin-coated pits is essential for viral entry

Viruses utilize cellular lipids and manipulate host lipid metabolism to ensure their replication and spread.Therefore,the identification of lipids and metabolic pathways that are suitable targets for antiviral development is crucial.Using a library of compounds targeting host lipid metabolic factors and testing them for their ability to block pseudorabies virus(PRV)and vesicular stomatitis virus(VSV)infection,we found that U18666A,a specific inhibitor of Niemann-Pick C1(NPC1),is highly potent in suppressing the entry of diverse viruses including pseudotyped severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).NPC1 deficiency markedly attenuates viral growth by decreasing cholesterol abundance in the plasma mem-brane,thereby inhibiting the dynamics of clathrin-coated pits(CCPs),which are indispensable for clathrin-mediated endocytosis.Significantly,exogenous cholesterol can complement the dynamics of CCPs,leading to efficient viral entry and infectivity.Administration of U18666A improves the survival and pathology of PRV-and influenza A virus-infected mice.Thus,our studies demonstrate a unique mechanism by which NPC1 inhibition achieves broad antiviral activity,indicating a potential new therapeutic strategy against SARS-CoV-2,as well as other emerging viruses.

Pseudorabies virus manipulates mitochondrial tryptophanyl-tRNA synthetase 2 for viral replication

The pseudorabies virus(PRV)is identified as a double-helical DNA virus responsible for causing Aujeszky's disease,which results in considerable economic impacts globally.The enzyme tryptophanyl-tRNA synthetase 2(WARS2),a mitochondrial protein involved in protein synthesis,is recognized for its broad expression and vital role in the translation process.The findings of our study showed an increase in both mRNA and protein levels of WARS2 following PRV infection in both cell cultures and animal models.Suppressing WARS2 expression via RNA interference in PK-15 cells led to a reduction in PRV infection rates,whereas enhancing WARS2 expression resulted in increased infection rates.Furthermore,the activation of WARS2 in response to PRV was found to be reliant on the cGAS/STING/TBK1/IRF3 signaling pathway and the interferon-alpha receptor-1,highlighting its regulation via the type I interferon signaling pathway.Further analysis revealed that reducing WARS2 levels hindered PRV's ability to promote protein and lipid synthesis.Our research provides novel evidence that WARS2 facilitates PRV infection through its management of protein and lipid levels,presenting new avenues for developing preventative and therapeutic measures against PRV infections.