LSM-W2: laser scanning microscopy worker for wheat leaf surface morphology

Abstract

Background Microscopic images are widely used in plant biology as an essential source of information on morphometric characteristics of the cells and the topological characteristics of cellular tissue pattern due to modern computer vision algorithms. High-resolution 3D confocal images allow extracting quantitative characteristics describing the cell structure of leaf epidermis. For some issues in the study of cereal leaves development, it is required to apply the staining techniques with fluorescent dyes and to scan rather large fragments consisting of several frames. We aimed to develop a tool for processing multi-frame multi-channel 3D images obtained from confocal laser scanning microscopy and taking into account the peculiarities of the cereal leaves staining. Results We elaborated an ImageJ-plugin LSM-W 2 that allows extracting data on L eaf S urface M orphology from L aser S canning M icroscopy images. The plugin is a crucial link in a workflow for obtaining data on structural properties of leaf epidermis and morphological properties of epidermal cells. It allows converting large lsm-files (laser scanning microscopy) into segmented 2D/3D images or tables with data on cells and/or nuclei sizes. In the article, we also represent some case studies showing the plugin application for solving biological tasks. Namely the plugin is applied in the following cases: defining parameters of jigsaw-puzzle pattern for maize leaf epidermal cells, analysis of the pavement cells morphological parameters for the mature wheat leaf grown under control and water deficit conditions, initiation of cell longitudinal rows, and detection of guard mother cells emergence at the initial stages of the stomatal morphogenesis in the growth zone of a wheat leaf. Conclusion The proposed plugin is efficient for high-throughput analysis of cellular architecture for cereal leaf epidermis. The workflow implies using inexpensive and rapid sample preparation and does not require the applying of transgenesis and reporter genetic structures expanding the range of species and varieties to study. Obtained characteristics of the cell structure and patterns further could act as a basis for the development and verification for spatial models of plant tissues formation mechanisms accounting for structural features of cereal leaves. Availability The implementation of this workflow is available as an ImageJ plugin distributed as a part of the Fiji project (FijiisjustImageJ: https://fiji.sc/ ). The plugin is freely available at https://imagej.net/LSM_Worker , https://github.com/JmanJ/LSM_Worker and http://pixie.bionet.nsc.ru/LSM_WORKER/ . Electroni...

Comparison of Sample Preparation Techniques for Inspection of Leaf Epidermises Using Light Microscopy and Scanning Electronic Microscopy

“…For species that emit little epidermis cell wall autofluorescence, such as Arabidopsis and cucumber, staining leaves with a fluorescence dye such as fluorescein diacetate, propidium iodide, and aniline blueS or using plasma-membrane localized GFP materials can help in the detection of cell wall outlines (Talbot and White, 2013a;Li et al, 2015). Furthermore, progress of recent studies on the workflow and analytical tools for obtaining and processing the 3D confocal images contribute to the measurement of epidermal cell sizes (Wuyts et al, 2010;Kierzkowski et al, 2019;Zubairova et al, 2019).…”

“…LSCMs are often used to detect the fluorescence of the cell surface and cell outlines (Wuyts et al, 2010;Jones et al, 2016;Januszkiewicz et al, 2019;Kierzkowski et al, 2019;Vaahtera et al, 2019;Zubairova et al, 2019). Although the bright-field images of chloral hydrated rice leaf tissues were only partially focused ( Figures 3E and 4A), rice tissue exhibited bright autofluorescence and the cell wall outlines could be clearly visualized ( Figure 4B).…”

ImageJ SurfCut: a user-friendly pipeline for high-throughput extraction of cell contours from 3D image stacks

“…In contrast to MGX or MerryProj, SurfCut is an ImageJ plugin and not a standalone software, and it does not require specific hardware. The SurfaceProject and LSM-W 2 [22, 23] ImageJ plugins also represent very good alternatives. For instance, SurfaceProject can be used to extract a layer of signal independently from the surface of the signal, by manually placing points within the 3D stack.…”

“…However, it requires manual processing of each image. Very recently, the ImageJ plugin LSM-W 2 was also introduced [23]. One of the tools developed within this plugin allows the creation of virtual cuts through 3D stacks.…”

Изучение Топологии Ростковых Трубок Бурой Ржавчины На Поверхности Листа Пшеницы На Основе Анализа Конфокальных 3d-Изображений

“…Подбор области сканирования проводился исходя из необходимости обеспечить видимость всех клеточных стенок растения и границ ростковых трубок гриба. Обработка и анализ 3D-изображений проводилась с помощью разработанного авторами программного модуля для плагина LSM-W 2 [3], позволяющего обрабатывать и анализировать конфокальные изображения бурой ржавчины на листьях пшеницы. Основными этапами алгоритма являются:…”