In this pilot study, we made significant progress in plant tissue imaging and identification of soybean SDS toxin using IR spectroscopy. In plant tissue imaging, we characterized infrared spectral features of SDS fungus Fusarium virguliforme and compared IR spectral features of soybean plants infected with and without SDS toxin. Efforts had been made to characterize fungus and toxin grown on agar using FT-IR and Raman spectroscopy.
We grew soybean plants infected with and without SDS toxin and collected plants for sectioning at different growth periods. We also optimized microtome procedures (sectioning methods) and sample thicknesses for high resolution transmission and attenuated total reflectance (ATR) FT-IR imaging. Transmission and attenuated total reflectance (ATR) FT-IR imaging data had been collected on soybean plant tissue infected with and without SDS toxin.
Preliminary transmission infrared data with 6.25 µm spatial resolutions illustrated differences in the chemical distributions in root and stem tissues between the infected and control plants. Moreover, attenuated total reflectance (ATR) FT-IR imaging data with 1.25 µm spatial resolutions has shown potential for characterizing different cell types within xylem tissue. The imaging was mainly focused on xylem tissue as it is considered also transport vehicle for toxin from root to leaf. The data clearly shows SDS infected plants have different structural/chemical composition of fatty acids, phospholipid and glycolipid having asymmetric CH2 functional groups in stem xylem tissue compared to control plants. Additionally, it shows that lower pectin concentration in xylem tissue of SDS infected plants.
Several efforts could be pursued for a broader scale study. Protocols could be developed to classify different tissue types (e.g. xylem, phloem, epidermis etc) using transmission and attenuated total reflectance (ATR) FT-IR spectroscopy. Cryogenic sectioning or microtoming of leaf sections might be challenging but we have already shown significant progress in sectioning stem and root sections of soybean for FT-IR imaging. Efforts had been made to characterize SDS toxin (FvTox1 13.5-kDa acidic protein) isolated from Fusarium virguliforme culture filtrate using IR and Raman spectroscopy. It is not yet not identified directly from soybean transport tissue and its transport mechanism is still completely unknown.
Though the investigators recognize the challenges of toxin detection, the IR spectral characteristics of the SDS toxin could be compared with the IR images of the plant sections to identify the temporal and spatial distribution of the toxin in the plant tissue. This will open a path for tracking specific types of toxin and their signature inside the plant sections such as root, stem and leaves as well as seeds or grains.