African swine fever virus remodels cellular host metabolism and promotes lipid droplet formation in macrophages and cell lines.

BACKGROUND: African swine fever virus (ASFV) is a large cytoplasmic DNA virus that causes a highly lethal disease in pigs and poses a major threat to global animal health and food security. ASFV infection is characterized by extensive reprogramming of host cellular pathways to support viral replication, including immune regulation, stress responses, and metabolic processes. Increasing evidence indicates that lipid metabolism is actively modulated during infection; however, the functional contribution of specific lipid-related organelles to the ASFV replicative cycle remains to be fully defined. METHODS: We characterized the host response to ASFV infection in primary porcine alveolar macrophages using Illumina-based RNA sequencing at two representative post-infection time points. Transcriptomic analyses were complemented by functional assays in Vero cells, employing pharmacological inhibitors of lipid droplet (LD) biogenesis, quantitative measurements of viral replication, infectivity, and gene expression, as well as proteomic profiling of purified LDs from infected cells. RESULTS: ASFV infection induced profound changes in host gene expression, with significant enrichment of pathways involved in immune modulation, apoptosis, autophagy, and cellular metabolism. A remarkable finding was the marked upregulation of enzymes driving LD biogenesis. Functional studies showed that LDs were dynamically remodeled during infection, increasing in abundance and redistributing toward viral replication factories. Proteomic analysis of LDs from infected macrophages identified both structural and non-structural ASFV proteins, including subunits of the viral transcription machinery. Accordingly, pharmacological disruption of LD formation pathway prominently reduced viral genome replication, viral protein synthesis, infectivity, and virus production, exceeding 95% in some conditions. CONCLUSIONS: These findings establish lipid droplets as central host organelles that support efficient ASFV replication and reveal a tight physical and functional coupling between viral replication and host lipid metabolism. Targeting LD biogenesis represents a promising host-directed antiviral strategy against ASFV.