Histological quantification of maize stem sections from FASGA-stained images

Abstract

BackgroundCrop species are of increasing interest both for cattle feeding and for bioethanol production. The degradability of the plant material largely depends on the lignification of the tissues, but it also depends on histological features such as the cellular morphology or the relative amount of each tissue fraction. There is therefore a need for high-throughput phenotyping systems that quantify the histology of plant sections.ResultsWe developed custom image processing and an analysis procedure for quantifying the histology of maize stem sections coloured with FASGA staining and digitalised with whole microscopy slide scanners. The procedure results in an automated segmentation of the input images into distinct tissue regions. The size and the fraction area of each tissue region can be quantified, as well as the average coloration within each region. The measured features can discriminate contrasted genotypes and identify changes in histology induced by environmental factors such as water deficit.ConclusionsThe simplicity and the availability of the software will facilitate the elucidation of the relationships between the chemical composition of the tissues and changes in plant histology. The tool is expected to be useful for the study of large genetic populations, and to better understand the impact of environmental factors on plant histology.Electronic supplementary materialThe online version of this article (doi:10.1186/s13007-017-0225-z) contains supplementary material, which is available to authorized users.

A rapid and quantitative safranin‐based fluorescent microscopy method to evaluate cell wall lignification

“…Hypodermis cell walls are colored in blue, indicating low lignin levels (safranin ratio = 0.53) in comparison with the more heavily lignified cell walls of the epidermis (safranin ratio = 0.93) and vascular bundle sheath, which appear red (safranin ratio = 0.98) (Figure a). This approach is complementary to an approach based on the bioinformatic treatment of FASGA stained stem sections that allows genotyping based on the surface of lignification (Legland et al , ). The safranin ratio methodology also allows a better resolution as it is independent of the section thickness, which can become a limiting point when the cell density and therefore the opacity to transmitted light are too much important, as it occurs in the vicinity of large metaxylem vessels (Figure b).…”

“…To get around this problem and to obtain information on lignin localization more rapidly, researchers have exploited different physical and histochemical methods, including the natural autofluorescence of lignin (Donaldson and Radotic, ), basic fushin, auramine staining and acriflavine (epifluorescence/confocal observations) (Dharmawardhana et al , ; Donaldson et al , ; Pesquet et al , ), revealing global lignin deposition, as well as more ‘specific’ tests such as phloroglucinol staining (Wiesner reagent), revealing lignin hydroxycinnamaldehyde functions (Vallet et al , ; Liljegren, ), and Maüle staining, providing indications of the S/G ratio (Meshitsuka and Nakano, ). Another approach based on safranin/alcian blue staining, the fucsina, alcian blue, safranina, glicerina, agua (FASGA) method (Tolivia and Tolivia, ), coupled to an adapted image workflow (Legland et al , ) allows for the quantification of the histology of Zea mays (maize) based on lignin‐containing tissue detection, but does not permit a strict quantification of cell wall lignin content.…”

Genotypic covariations of traits underlying sorghum stem biomass production and quality and their regulations by water availability: Insight from studies at organ and tissue levels

“…The eight studied genotypes contrasted for stem biomass accumulation and component traits related to stem growth. This gives evidence that histological studies provide a better understanding of the component traits affecting biomass quality (Legland et al, 2017) and should help to better monitor this complex trait in the course of breeding programs. In particular, plant height was positively correlated with stem structural carbohydrate and lignin contents and negatively correlated with stem soluble sugar content.…”

“…This will also help orienting the development of appropriate phenotyping facilities to support the development of varieties and innovative crop management practices (Cabrera-Bosquet et al, 2016;Legland, El-Hage, Mechin, & Reymond, 2017;M. This will also help orienting the development of appropriate phenotyping facilities to support the development of varieties and innovative crop management practices (Cabrera-Bosquet et al, 2016;Legland, El-Hage, Mechin, & Reymond, 2017;M.…”

Crop Phenomics: Current Status and Perspectives

“…The complexity and diversity in microscopic image data poses greater challenges for developing suitable data analysis workflows in the detection and identification of microscopic phenotypes of stalk tissue. Zhang et al (2013) and Legland et al (2017) presented semiautomated and automated analysis method for the stained microscopic images of stalk sections. Legland et al (2014) and Heckwolf et al (2015) created an image analysis tool that could operate on images of hand-cut stalk transections to measure anatomical features in high throughput.…”