African swine fever virus (ASFV) causes a fatal hemorrhagic disease in pigs, leading to severe economic losses worldwide. The viral ubiquitin-conjugating enzyme I215L (E2) is critical for ASFV replication and modulation of host antiviral responses. Host deubiquitinating enzymes (DUBs) bind to E2 enzymes to inhibit the transfer of ubiquitin. In this study, we systematically screened 34 swine DUBs using HDOCK docking and AlphaFold3 modeling, identifying FAM105A as the most probable interactor. Co-immunoprecipitation assay validated the interaction between I215L and FAM105A. Structural analyses revealed seven I215L residues (E19, N20, M99, S102, A104, D108, S116) that form defined polar contacts, among which M99, D108, and S116 are critical for binding stability. These findings identify FAM105A as a novel host target of ASFV I215L, establishing a new paradigm for studying viral-host protein interactions.

Lipid droplets (LDs) are highly conserved, evolutionarily dynamic organelles. In addition to their canonical roles in energy homeostasis, membrane biosynthesis, and lipid signaling, accumulating evidence highlights their critical involvement in viral pathogenesis. As obligate intracellular parasites, viruses are entirely dependent on their host’s cellular machinery. To replicate, they must hijack LDs to obtain essential lipids and energy, which are critical for assembling viral replication complexes and producing new virions. However, host cells do not passively accept viral invasion. Recent evidence demonstrates that pathogen infection triggers the innate immune response of host cells. This response, in turn, rewires lipid metabolism to drive the formation of new LDs. These newly formed LDs are characterized by surfaces enriched with immune proteins. These specialized lipid droplets serve as defensive structures that combat pathogen invasion through both direct and indirect mechanisms. This article provides a systematic review of the dual roles of LDs in viral infection: on the one hand, LDs are hijacked by viruses to facilitate their replication; on the other hand, they serve as central hubs in innate immunity, participating in host cell antiviral defense mechanisms. These findings lend substantial support to the hypothesis that “LDs constitute critical components of innate immunity”, while offering novel perspectives for understanding virus-host interactions. Future investigations should focus on elucidating the specific molecular mechanisms by which LDs mediate immune regulation, and explore innovative antiviral therapeutic strategies through targeted modulation of LD metabolism.

Viral transcription strategies vary significantly depending on the nature of the viral genome and the site of transcription within the host cell. A common challenge for all viruses is to circumvent host defense mechanisms while simultaneously exploiting host cellular machinery to facilitate their own transcription. DNA viruses and retroviruses often hijack the host’s RNA polymerase II (Pol II), while RNA viruses typically encode their own RNA-dependent RNA polymerase (RdRp). Through these diverse mechanisms, viruses effectively manipulate host transcription, translation, and immune responses, thereby evading immune surveillance and completing their replication cycle. This article systematically reviews the transcription strategies employed by major DNA and RNA virus families, providing insights that contribute to a deeper understanding of viral transcriptional regulation mechanisms.

African swine fever (ASF), caused by the African swine fever virus (ASFV), poses a catastrophic threat to global swine production due to the absence of effective vaccines or therapeutics. Targeting the virally encoded RNA polymerase (RNAP), which is essential for viral transcription, offers a promising antiviral strategy. Capitalizing on our previously resolved high-resolution structure of ASFV RNAP, we conducted a rigorous structure-based virtual screening of two compound libraries (T001 and LF1000) to identify selective inhibitors. From 100 candidates validated in vitro, nine compounds identified at the genome level by qPCR demonstrated >98% inhibition of ASFV replication in porcine alveolar macrophages (PAMs). The lead compound, K-7174 — a repurposed agent known to modulate proteasome and GATA activity — exhibited potent, dose-dependent antiviral efficacy (IC₅₀ = 2.502 µmol/L), minimal cytotoxicity, and high binding affinity for ASFV RNAP. Molecular docking simulations indicated potential interactions with key catalytic residues, including Glu1051 and Ala456, proposing a mechanism that may involve active-site disruption. However, definitive confirmation of this binding pose requires further experimental validation through high-resolution structural methods such as cryo-EM. Our study not only nominates K-7174 as a promising anti-ASFV candidate but also establishes an integrated computational–experimental framework for accelerating the discovery of antiviral agents targeting viral polymerases.

Brucella is a zoonotic pathogen that causes substantial economic losses in the livestock industry. However, the categorization of functional genes in Brucella requires further refinement. This knowledge gap was addressed by classifying gene essentiality through constructing a transposon mutant library. Mariner transposon mutant libraries were independently constructed for Brucella abortus A19 and S2308. Transposon sequencing was used to visualize the insertion sites within the mutant library, which revealed a uniform transposon distribution in both genomes and high library saturation. Furthermore, 451 and 393 putative essential genes were identified for B. abortus A19 and S2308, respectively. The functional enrichment analysis of these genes revealed that cofactor biosynthesis was the most highly represented, followed by ribosomal biogenesis, carbon metabolism, amino acid biosynthesis, and oxidative phosphorylation. Many of these genes encode known antimicrobial drug targets. This study provides a foundational resource for Brucella research, and the mutant library serves as a tool for screening functional Brucella genes.

Nipah virus (NiV), a highly pathogenic agent causing severe neurologic and respiratory disease, requires Biosafety Level 4 (BSL-4) containment. While existing pseudovirus models (e.g., based on human immunodeficiency virus (HIV) or vesicular stomatitis virus (VSV) backbones) offer alternatives, their utility is often limited by reliance on a single reporter gene. Here, we developed and optimized a robust NiV pseudovirus system utilizing a VSV-ΔG backbone co-expressing dual reporters, Green Fluorescent Protein (GFP) and NanoLuciferase (NLuc). Preliminary characterization showed that VSV-ΔG-GFP/NLuc exhibited structural integrity, achieved infection rates up to 70% in susceptible cells, and demonstrated relative stability within five freeze-thaw cycles. The study on the correlation between the expression of ephrin-B2 (EFNB2) in various cell lines and the infection efficiency of pseudovirus indicates the effectiveness of the pseudovirus. Furthermore, the platform reliably detected dose-dependent neutralization by established monoclonal antibodies targeting the fusion protein and glycoprotein of NiV, yielding trends comparable to authentic virus neutralization. Collectively, these findings establish this dual-reporter NiV pseudovirus as a safe, scalable, and high-throughput tool for antiviral screening and mechanistic studies under BSL-2 conditions. The synergistic combination of GFP (enabling visual quantification and sorting) and NLuc (providing ultra-sensitive, broad dynamic range quantification) enhances both versatility and data reliability. This study not only provides a critical new tool for NiV research but also serves as a valuable reference for developing pseudovirus platforms targeting other enveloped viruses.