Histones (the main protein components of chromatin in eukaryotes) have been shown to be bactericidal in vitro, so it had been proposed that they have a physiological role in antimicrobial defences, in particular against intracellular pathogens. This study identifies lipid droplet-bound histones as a new antibacterial defence system. Histones were released from lipid droplets (purified from Drosophila melanogaster embryos) in the presence of bacterial cell envelope components such as lipopolysaccharide (LPS) and lipoteichoic acid, and displayed bactericidal activity in vitro. Importantly, such bactericidal activity was also observed in vivo: D. melanogaster embryos lacking histone deposits on lipid droplets showed reduced survival compared with wild-type embryos when injected with either Gram-negative (Escherichia coli str. DH5a) or Gram-positive (Staphylococcus epidermidis) bacteria. Moreover, the levels of lipid droplet-bound histones in mice increased following LPS injection, suggesting that this antibacterial defence system might be conserved.