Morphology and hydrolytic activity of A7, a typing phage of Pseudomonas syringae pv. morsprunorum

Abstract

Bacteriophage A7 has been employed as an indicator for strains of Pseudomonas syringae pv. morsprunorum race 1. Electron microscopy showed that this phage had a hexagonal head, 59 nm in diameter, and a long, flexible, non-contractile tail, 164 nm long and 8 nm wide, containing approximately 15 evenly spaced transverse striations and terminating in a base-plate equipped with six spikes arranged in a radially symmetrical pattern around a central core. Two short fibres projecting at an angle from the base-plate were also visible on some phage particles. Phage A7 can therefore be placed in group B of the classification system of Bradley. Phage particles bound at apparently random sites over the surface of host cells by their base-plates, and after a short time released DNA from their heads. Phage A7 uses lipopolysaccharide (LPS) as its binding site on the bacterial cell surface, removing the d-rhamnan side-chains by the action of a rhamnan hydrolase. The appearance of purified LPS by electron microscopy was either strand-like or vesicular, according to whether it had been stained with phosphotungstate or uranyl acetate respectively. Strands were of approximately uniform width (approx. 9 nm). Vesicular forms included both circles, 25–45 nm in diameter, and larger oval structures of greater electron transparency. The appearance of the LPS did not alter on addition of the phage. Phage particles were observed attached via their base-plates to LPS vesicles, in particular the larger oval vesicles, but were never seen attached to strands. The heads of phage particles attached to LPS were frequently empty. The rhamnanase of phage A7 released the side-chain residues from the LPS as an equimolar mixture of tri- and hexasaccharide. By using 1 h-NMR spectroscopy and methylation analysis, the site of cleavage has been identified as the 2)-α-d-Rhap(1 → 3) residue.