Physiological responses of plants to in vivo XRF radiation damage: insights from elemental, histochemical, anatomical and ultrastructural analyses

Abstract

X-ray fluorescence spectroscopy (XRF) is a powerful technique for the in vivo assessment of plant tissues. However, the potential X-ray exposure damages might affect the structure and elemental composition of living plant tissues leading to artefacts in the recorded data. Herein, we exposed soybean (Glycine max (L.) Merrill) leaves to several X-ray doses through a polychromatic benchtop microprobe X-ray fluorescence spectrometer, modulating the photon flux by adjusting either the beam size, focus, or exposure time. The structure, ultrastructure and physiological responses of the irradiated plant tissues were investigated through light and transmission electron microscopy (TEM). Depending on the dose, the X-ray exposure induced decreased K and X-ray scattering intensities, and increased Ca, P, and Mn signals on soybean leaves. Anatomical analysis indicated necrosis of the epidermal and mesophyll cells on the irradiated spots, where TEM images revealed the collapse of cytoplasm and cell-wall breaking. Furthermore, the histochemical analysis detected the production of reactive oxygen species, as well as inhibition of chlorophyll autofluorescence in these areas. Under certain X-ray exposure conditions, e.g., high photon flux and exposure time, XRF measurements may affect the soybean leaves structures, elemental composition, and cellular ultrastructure, and induce programmed cell death. These results shed light on the characterization of the radiation damage, and thus, help to assess the X-ray radiation limits and strategies for in vivo for XRF analysis. HighlightBy exposing soybean leaves to several X-ray doses, we show that the characteristic X-ray induced elemental changes stem from plants physiological signalling or responses rather than only sample dehydration.

Plant phenotyping relevance

植物組織のin vivo XRF測定における放射線損傷と測定アーティファクトを評価し、適用限界と測定条件を検証する研究であり、フェノタイピング手法の技術的妥当性が中心です。

Code and data availability

The paper's DATA AVAILABILITY section states the raw data (XRF spectra/maps and imaging measurements) are fully available on Figshare at the authors' public DOI, which matches an allowed URL.