Abstract
Plant–pathogen interactions have been widely studied, but mostly from the site of the plant secondary defense. Less is known about the effects of pathogen infection on plant primary metabolism. The possibility to transform a fluorescing protein into prokaryotes is a promising phenotyping tool to follow a bacterial infection in plants in a noninvasive manner. In the present study, virulent and avirulent Pseudomonas syringae strains were transformed with green fluorescent protein (GFP) to follow the spread of bacteria in vivo by imaging Pulse-Amplitude-Modulation (PAM) fluorescence and conventional binocular microscopy. The combination of various wavelengths and filters allowed simultaneous detection of GFP-transformed bacteria, PAM chlorophyll fluorescence, and phenolic fluorescence from pathogen-infected plant leaves. The results show that fluorescence imaging allows spatiotemporal monitoring of pathogen spread as well as phenolic and chlorophyll fluorescence in situ, thus providing a novel means to study complex plant–pathogen interactions and relate the responses of primary and secondary metabolism to pathogen spread and multiplication. The study establishes a deeper understanding of imaging data and their implementation into disease screening.
Highlights
Plants are regularly attacked by several pathogens, such as bacteria, fungi, viruses, oomycetes, nematodes, and others
To validate the signal detected by imaging PAM, the fluorescence of single P. syringae drops in different concentrations on black, nonfluorescing background was quantified
The fluorescing bacteria were detected with fluorescence binocular, which was equipped with a special GFP3 filter
Summary
Plants are regularly attacked by several pathogens, such as bacteria, fungi, viruses, oomycetes, nematodes, and others. Red Chl-F emitted by photosystem II (PSII) provides information on the photosynthetic performance of plants in terms of activity and indirect information on the CO2 assimilation rate (Murchie and Lawson, 2013) Novel technologies such as hyperspectral imaging and Chl-F imaging offer an elegant, noninvasive means to explore indirectly the bacteria spread within the plant tissue (Rolfe and Scholes, 2010; Großkinsky et al, 2017; West et al, 2017; Bohnenkamp et al, 2019; Kuska et al, 2019). In addition to detecting bacterial cells, to follow a pathogen invasion on a leaf by simultaneously monitoring direct changes in plant primary and secondary metabolisms. Pseudomonas syringae is a hemibiotrophic pathogen (Preston, 2000) that can invade several, economically important plant species
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