Electron microscopic analysis of the H 2 O 2 accumulation preceding hypersensitive cell death induced by an incompatible strain of Pseudomonas avenae in cultured rice cells

summary Infection of cultured rice cells with an incompatible strain of Pseudomonas avenae induces a hypersensitive reaction of the host, while compatible strain infection produces no such reaction. The induction of H 2 O 2 generation in cultured rice cells by the incompatible strain of P. avenae precedes cell death. To examine the distribution of H 2 O 2 generation sites, cultured rice cells were incubated following infection with a cerium solution. Detection of the reaction product, Ce(OH) 2 OOH, was performed using energy disperse X‐ray microanalysis (EDX) fitted with a variable‐pressure scanning electron microscope (VP‐SEM). We determined that H 2 O 2 accumulation is a local response, appearing as a circular region on the cell surface of only 10% to 15% of the total infected cells. Observation of cross‐sections localized cerium deposition to the plasma membranes of papillae, in the cell walls of a papilla and around the bacterium. Furthermore, immuno‐gold electron microscopy using antibodies for β‐1,3‐glucan suggested that callose synthesis also occurs at the generation site of H 2 O 2 . Therefore, H 2 O 2 functions as an antibacterial agent, serving as a substrate for cell wall cross‐linking. Our detection system employs an EDX system fitted with SEM; this procedure will be useful to examine the function and mechanism of oxidative bursts in plant–pathogen interactions.