Dataset Information

High-throughput method for detection and quantification of lesions on leaf scale based on trypan blue staining and digital image analysis.

ABSTRACT: Background:Field-grown leafy vegetables can be damaged by biotic and abiotic factors, or mechanically damaged by farming practices. Available methods to evaluate leaf tissue damage mainly rely on colour differentiation between healthy and damaged tissues. Alternatively, sophisticated equipment such as microscopy and hyperspectral cameras can be employed. Depending on the causal factor, colour change in the wounded area is not always induced and, by the time symptoms become visible, a plant can already be severely affected. To accurately detect and quantify damage on leaf scale, including microlesions, reliable differentiation between healthy and damaged tissue is essential. We stained whole leaves with trypan blue dye, which traverses compromised cell membranes but is not absorbed in viable cells, followed by automated quantification of damage on leaf scale. Results:We present a robust, fast and sensitive method for leaf-scale visualisation, accurate automated extraction and measurement of damaged area on leaves of leafy vegetables. The image analysis pipeline we developed automatically identifies leaf area and individual stained (lesion) areas down to cell level. As proof of principle, we tested the methodology for damage detection and quantification on two field-grown leafy vegetable species, spinach and Swiss chard. Conclusions:Our novel lesion quantification method can be used for detection of large (macro) or single-cell (micro) lesions on leaf scale, enabling quantification of lesions at any stage and without requiring symptoms to be in the visible spectrum. Quantifying the wounded area on leaf scale is necessary for generating prediction models for economic losses and produce shelf-life. In addition, risk assessments are based on accurate prediction of the relationship between leaf damage and infection rates by opportunistic pathogens and our method helps determine the severity of leaf damage at fine resolution.

SUBMITTER: Mulaosmanovic E

PROVIDER: S-EPMC7197134 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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High-throughput method for detection and quantification of lesions on leaf scale based on trypan blue staining and digital image analysis.

Mulaosmanovic Emina E Lindblom Tobias U T TUT Bengtsson Marie M Windstam Sofia T ST Mogren Lars L Marttila Salla S Stützel Hartmut H Alsanius Beatrix W BW

Plant methods 20200504

<h4>Background</h4>Field-grown leafy vegetables can be damaged by biotic and abiotic factors, or mechanically damaged by farming practices. Available methods to evaluate leaf tissue damage mainly rely on colour differentiation between healthy and damaged tissues. Alternatively, sophisticated equipment such as microscopy and hyperspectral cameras can be employed. Depending on the causal factor, colour change in the wounded area is not always induced and, by the time symptoms become visible, a pla ...[more]

PMID: 32391069

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Project description:BackgroundMultipotent mesenchymal stem cells (MSCs) have the potential to repair and regenerate damaged tissues and are considered as attractive candidates for the development of cell-based regenerative therapies. Currently, there are more than 200 clinical trials involving the use of MSCs for a wide variety of indications. However, variations in their isolation, expansion, and particularly characterization have made the interpretation of study outcomes or the rigorous assessment of therapeutic efficacy difficult. An unbiased characterization of MSCs is of major importance and essential to guaranty that only the most suitable cells will be used. The development of standardized and reproducible assays to predict MSC potency is therefore mandatory. The currently used quantification methodologies for the determination of the trilineage potential of MSCs are usually based on absorbance measurements which are imprecise and prone to errors. We therefore aimed at developing a methodology first offering a standardized way to objectively quantify the trilineage potential of MSC preparations and second allowing to discriminate functional differences between clonally expanded cell populations.MethodMSCs originating from several patients were differentiated into osteoblasts, adipocytes, and chondroblasts for 14, 17, and 21 days. Differentiated cells were then stained with the classical dyes: Alizarin Red S for osteoblasts, Oil Red O for adipocytes, and Alcian Blue 8GX for chondroblasts. Quantification of differentiation was then performed with our newly developed digital image analysis (DIA) tool followed by the classical absorbance measurement. The results from the two techniques were then compared.ResultQuantification based on DIA allowed highly standardized and objective dye quantification with superior sensitivity compared to absorbance measurements. Furthermore, small differences between MSC lines in the differentiation potential were highlighted using DIA whereas no difference was detected using absorbance quantification.ConclusionOur approach represents a novel method that simplifies the laboratory procedures not only for the quantification of histological dyes and the degree of differentiation of MSCs, but also due to its color independence, it can be easily adapted for the quantification of a wide range of staining procedures in histology. The method is easily applicable since it is based on open source software and standard light microscopy.

Dataset Information

High-throughput method for detection and quantification of lesions on leaf scale based on trypan blue staining and digital image analysis.

Publications

High-throughput method for detection and quantification of lesions on leaf scale based on trypan blue staining and digital image analysis.

Similar Datasets

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