An Optimized in situ Quantification Method of Leaf H2O2 Unveils Interaction Dynamics of Pathogenic and Beneficial Bacteria in Wheat

Abstract

Hydrogen peroxide (H 2 O 2) functions as an important signaling molecule in plants during biotic interactions. However, the extent to which H 2 O 2 accumulates during these interactions and its implications in the development of disease symptoms is unclear. In this work, we provide a step-by-step optimized protocol for in situ quantification of relative H 2 O 2 concentrations in wheat leaves infected with the pathogenic bacterium Pseudomonas syringae pv. atrofaciens (Psa), either alone or in the presence of the beneficial bacterium Herbaspirillum seropedicae (RAM10). This protocol involved the use of 3-3 diaminobenzidine (DAB) staining method combined with image processing to conduct deconvolution and downstream analysis of the digitalized leaf image. The application of a linear regression model allowed to relate the intensity of the pixels resulting from DAB staining with a given concentration of H 2 O 2. Decreasing H 2 O 2 accumulation patterns were detected at increasing distances from the site of pathogen infection, and H 2 O 2 concentrations were different depending on the bacterial combinations tested. Notably, Psa-challenged plants in presence of RAM10 accumulated less H 2 O 2 in the leaf and showed reduced necrotic symptoms, pointing to a potential role of RAM10 in reducing pathogen-triggered H 2 O 2 levels in young wheat plants.

Loss of function of MusaPUB genes in banana can provide enhanced resistance to bacterial wilt disease

“…To investigate hydrogen peroxide production in edited plants following Xcm in ltration, a histochemical staining assay using DAB (3,3'-diaminobenzidine) solution was performed 21,22 . Fully opened green leaves of six-weeks-old in vitro plantlets of the edited events and wild-type control plants were in ltrated with 100 µl of fresh Xcm culture.…”

Assessment of Oxidative Stress by Detection of H2O2 in Rye Samples Using a CuO- and Co3O4-Nanostructure-Based Electrochemical Sensor

“…Biotic stressors, such as pathogen infection and herbivory [17], can also stimulate an increase in ROS production in plants.…”

The Plant Growth-Promoting Potential of Halotolerant Bacteria Is Not Phylogenetically Determined: Evidence from Two Bacillus megaterium Strains Isolated from Saline Soils Used to Grow Wheat

“…One important approach is to make use of plant growth-promoting bacteria (PGPB). PGPB can improve crop production with multiple modes of action such as the synthesis of growth-promoting substances (including phytohormones such as auxins, strigolactones, and nitric oxide), improved plant nutrition, and resistance to biotic and abiotic stresses [ 18 , 20 , 21 , 22 , 23 ], including the stimulation of plant defence [ 24 , 25 ]. Indeed, the benefits of PGPB for a plant may occur along the plant’s life cycle as they may improve plant germination, survival, health, growth, reproduction, and productivity under optimal and adverse conditions.…”