Pseudomonas aeruginosa is a human pathogen, whose treatment is complicated by its frequent antibiotic-resistance. Siphoviridae bacteriophage JBD30 infects and kills bacterium P. aeruginosa, which makes it a potential agent for phage therapy. Here we present the structure of bacteriophage JBD30 virion and its replication strategy, revealed by the combination of cryo-electron microscopy analysis techniques and cryo-electron tomography.
The virion of bacteriophage JBD30 is composed of non-enveloped icosahedral capsid, long flexible non-contractile tail and baseplate decorated with tail fibers. The capsid with a diameter of 60 nm is built from major capsid protein organised in T = 7 icosahedral lattice and decorated on three-fold and pseudo-threefold axis with trimers of minor capsid protein. In one vertex of the capsid, the penton of major capsid protein is replaced by dodecameric portal. The portal complex forms an interface between the capsid and 180 nm long tail. The tail is built from 44 hexameric discs of major tail protein. Distal tail protein trimer follows-up the last tail disc and forms an attachment site for the long tail fibers. The baseplate is terminated with a tripod complex of receptor binding protein trimers.
Using cryo-electron tomography we followed the infection process of P. aeruginosa by JBD30 phage from attachment to bacterial cell, to the production of new phage progeny and host cell lysis. Bacteriophage JBD30 uses its long tail fibres for binding to P. aeruginosa pili type IV. After attachment to pili, the virion either diffuses or is pulled towards the cellular surface, where it irreversibly binds by its receptor binding proteins. Afterwards, the phage punctures the outer cellular membrane, degrades the peptidoglycan layer and injects its DNA into host cell. New phage progeny is released approximately after 80 minutes post infection.
The combination of cryo-electron microscopy methods allowed us, to propose the mechanism of key stages of phage infection and describe it at molecular level.
Core Facility Cryo-electron Microscopy and Tomography of CEITEC Masaryk University is gratefully acknowledged for the obtaining of the scientific data presented in this paper