African swine fever virus B66L drives extracellular mitochondrial release to promote systemic inflammation in mice

Systemic inflammation is a hallmark of viral infection, but the upstream signals that initiate it remain poorly defined. Here we show that extracellular mitochondria act as inflammatory mediators during infection by vesicular stomatitis virus, influenza A virus, rabies virus, herpes simplex virus 1 and African swine fever virus (ASFV). In ASFV-infected primary porcine alveolar macrophages, the viral protein B66L promotes the capture of damaged mitochondria by autophagosomes while blocking their fusion with lysosomes, causing mitochondria to accumulate outside cells. Extracellular mitochondria are detected in the serum and bronchoalveolar lavage fluid of mice expressing B66L and in the serum of ASFV-infected pigs. Purified extracellular mitochondria trigger inflammatory cytokine production through cGAS-STING signalling and contribute to lung injury in mice. These findings identify virus-associated extracellular mitochondrial release as a pro-inflammatory mechanism during infection.