Updated project metadata. Eukaryotes and prokaryotes encode proteins with the histone-fold domain, but only eukaryotes are known to encode "core histones" from four distinct families that heterodimerize selectively and uniquely to form nucleosome particles. Some large DNA viruses, including those in Marseilleviridae family encode histones. The presence of histone proteins in viral particles suggests their role in compacting viral genomes but it is not known whether these histones can form higher-order structures like those found in eukaryotes. Here we show that fused histone pairs Hβ-Hα and Hδ-Hγ from Marseillevirus are structurally analogous to the eukaryotic histone pairs H2B-H2A and H3-H4. We further show that they form “forced” heterodimers and that a heterotetramer of four such heterodimers assembles DNA to form structures virtually identical to those of canonical eukaryotic nucleosomes. We characterize these viral structures using cryo-EM to reveal their unprecedented conservation with the nucleosome—one of the most important and conserved features of eukaryotic chromosomes. The structure and properties of these viral nucleosomes hold clues to understanding the origins of eukaryotic chromatin.