biostudies-literaturePeng YNational Natural Science Foundation of China (National Science Foundation of China)1772https://www.ebi.ac.uk/biostudies/studies/S-EPMC12917162biostudies-literatureUnknown17(1)Siphophages possess a long, flexible, and non-contractile tail that is responsible for host recognition, cell wall perforation, and genome delivery. Although the majority of structurally characterized siphophages target Gram-negative bacteria, those infecting Gram-positive bacteria remain elusive. Moreover, structural information concerning phage infection and genome release events in Gram-positive infecting siphophages is sparse. Here, we present a near-atomic resolution structure of Brochothrix thermosphacta bacteriophage NF5 determined by cryo-electron microscopy (cryo-EM). The structure comprises 11 proteins associated with the head, neck, tail tube, and baseplate, amounting to 643 polypeptides in total. Integration of cellular cryo-electron tomography (cryo-ET) showed the infection prNature communicationsStructure of a Brochothrix thermosphacta bacteriophage reveals cell wall adsorption mechanism in Gram-positive infecting siphophages.PMC12917162322009941203400632401014Liu HPeng YZheng JChen WXiao HPang HYang FZhou JfalseStructure of a Brochothrix thermosphacta bacteriophage reveals cell wall adsorption mechanism in Gram-positive infecting siphophages.Siphophages possess a long, flexible, and non-contractile tail that is responsible for host recognition, cell wall perforation, and genome delivery. Although the majority of structurally characterized siphophages target Gram-negative bacteria, those infecting Gram-positive bacteria remain elusive. Moreover, structural information concerning phage infection and genome release events in Gram-positive infecting siphophages is sparse. Here, we present a near-atomic resolution structure of Brochothrix thermosphacta bacteriophage NF5 determined by cryo-electron microscopy (cryo-EM). The structure comprises 11 proteins associated with the head, neck, tail tube, and baseplate, amounting to 643 polypeptides in total. Integration of cellular cryo-electron tomography (cryo-ET) showed the infection pr2026-01-01T00:00:00Z2026 Jan2026-07-16T10:08:26.94Z2026-07-09T10:48:11.808ZS-EPMC129171624154002710.1038/s41467-026-68477-2