In vivo label-free mapping of the effect of a photosystem II inhibiting herbicide in plants using chlorophyll fluorescence lifetime

Abstract

BackgroundIn order to better understand and improve the mode of action of agrochemicals, it is useful to be able to visualize their uptake and distribution in vivo, non-invasively and, ideally, in the field. Here we explore the potential of plant autofluorescence (specifically chlorophyll fluorescence) to provide a readout of herbicide action across the scales utilising multiphoton-excited fluorescence lifetime imaging, wide-field single-photon excited fluorescence lifetime imaging and single point fluorescence lifetime measurements via a fibre-optic probe.ResultsOur studies indicate that changes in chlorophyll fluorescence lifetime can be utilised as an indirect readout of a photosystem II inhibiting herbicide activity in living plant leaves at three different scales: cellular (~μm), single point (~1 mm2) and macroscopic (~8 × 6 mm2 of a leaf). Multiphoton excited fluorescence lifetime imaging of Triticum aestivum leaves indicated that there is an increase in the spatially averaged chlorophyll fluorescence lifetime of leaves treated with Flagon EC—a photosystem II inhibiting herbicide. The untreated leaf exhibited an average lifetime of 560 ± 30 ps while the leaf imaged 2 h post treatment exhibited an increased lifetime of 2000 ± 440 ps in different fields of view. The results from in vivo wide-field single-photon excited fluorescence lifetime imaging excited at 440 nm indicated an increase in chlorophyll fluorescence lifetime from 521 ps in an untreated leaf to 1000 ps, just 3 min after treating the same leaf with Flagon EC, and to 2150 ps after 27 min. In vivo single point fluorescence lifetime measurements demonstrated a similar increase in chlorophyll fluorescence lifetime. Untreated leaf presented a fluorescence lifetime of 435 ps in the 440 nm excited chlorophyll channel, CH4 (620–710 nm). In the first 5 min after treatment, mean fluorescence lifetime is observed to have increased to 1 ns and then to 1.3 ns after 60 min. For all these in vivo plant autofluorescence lifetime measurements, the plants were not dark-adapted.ConclusionsWe demonstrate that the local impact of a photosystem II herbicide on living plant leaves can be conveniently mapped in space and time via changes in autofluorescence lifetime, which we attribute to changes in chlorophyll fluorescence. Using portable fibre-optic probe instrumentation originally designed for label-free biomedical applications, this capability could be deployed outside the laboratory for monitoring the distribution of herbicides in growing plants.Electronic supplementary materialThe online version of this article (doi:10.1186/s13007-017-0201-7) contains supplementary material, which is available to authorized users.

Assessment of Soil Fertility Using Induced Fluorescence and Machine Learning

“…For plants, pigments (chlorophyll and anthocyanin) act as key responsive parameters for fluorescence emission at a specific wavelength of excitation. Differential sensitivity of the emission spectrum (at far-red and red) at specific excitation wavelengths (green and red) can be observed in wheat leaves [26]. These differences in the emission spectrum help to rationalize the chlorophyll indices (SFR_G and SFR_R) and anthocyanin indices (ANTH_RG and ANTH_RB) for plants that are generated by the Multiplex3 sensor.…”

Can chlorophyll fluorescence imaging make the invisible visible?

“…Other studies, in which dynamic Chl-FI was used, include the analysis of chlorosis induced by plant virus infection (Lei et al 2017), PSII-inhibiting herbicide (Noble et al 2017), reversible UV-induced photosynthetic activity (Kristoffersen et al 2016), discrimination of maize genotypes to drought (de Sousa et al 2017), dynamic photosynthetic fingerprints of salt overly sensitive (sos)mutants of Arabidopsis to drought stress (Sun et al 2019), and sensitivity to pH of Chlorella algae (Marcek Chorvatova et al 2020). A combination of kinetic Chl fluorescence and multicolour fluorescence imaging was used in a study on drought stress in Arabidopsis (Yao et al 2018).…”

A Review of Imaging and Sensing Technologies for Field Phenotyping

“…This technique can also be used to screen multiple plants, for example in agriculture (Betemps et al, 2012;Baluja, Diago, Goovaerts, Tardaguila, 2012;Van Iersel et al, 2016;Miao et al, 2018;Sytar, Bruckova, Plotnitskaya, Zivcak, Brestic, 2019), forestry (Hernández-Clemente, North, Hornero, Zarco-Tejada, 2017;Sonti, Hallett, Griffin, Trammell, Sullivan, 2020), climate change studies (He et al, 2019;Luus et al, 2017), and plant breeding programmes (Watanabe, Fekih, Kasajima, 2019;Kalaji, Guo, 2008). In addition, the chlorophyll fluorescence imaging technique can easily be used at different scales -from the whole plant (Eguchi, Konishi, Hosoi, Omasa, 2008;Miao et al, 2018) through the leaf (Montero et al, 2016;Leipner, Oxborough, Baker, 2001) to the cellular resolution (Noble et al, 2017). Several typical sensors that were implemented in the fluorescence imaging technique and representative products are listed in Table 2.…”

“…There is currently a large number of experiments using the UAV in plant ecology (Nowak, Dziób, Bogawski, 2019;Cruzan et al, 2016), forestry (Klosterman, Richardson, 2017;Weil, Lensky, Resheff, Levin, 2017), and agriculture (Burkart, Hecht, Kraska, Rascher, 2018;Yue et al, 2017). Examples of several chosen platforms that were generally implemented to different aspects of plant phenotyping, are described in Table 6.…”