Double-Staining Method to Detect Pectin in Plant-Fungus Interaction

Jo&#227;o Paulo  Rodrigues Marques; Laura Galvan Nuevo

doi:10.3791/63432

Double-Staining Method to Detect Pectin in Plant-Fungus Interaction

JoVE Journal
Biology

A subscription to JoVE is required to view this content. Sign in or start your free trial.

JoVE Journal Biology

Double-Staining Method to Detect Pectin in Plant-Fungus Interaction

Full Text

5,276 Views

06:39 min

February 4, 2022

DOI: 10.3791/63432-v

João Paulo Rodrigues Marques¹, Laura Galvan Nuevo²

¹Faculty of Animal Science and Food Engineering, Department of Basic Sciences,University of São Paulo, ²Laboratory of Nuclear Instrumentation, Center of Nuclear Energy in Agriculture,University of São Paulo

Overview

This protocol describes a light microscopy method for detecting pectin in coffee-fungus interactions. It allows for the identification of fungal structures and defects in pathogen cell wall polymers, providing insights into plant-fungal interactions.

Key Study Components

Research Area

Plant-fungal interactions
Histopathological studies
Fungal pathogen detection

Background

Significance in diagnosing diseases caused by fungi like coffee rust and cercosporiosis
Need for techniques that do not require expensive equipment
Rapid techniques for clear distinction of plant structures

Methods Used

Light microscopy for pectin detection
Utilizes coffee leaf samples infected with fungi
Double staining method with cotton blue and ruthenium red

Main Results

Identified dark blue staining in fungal hyphae and structures
Demonstrated interactions between Hemileia vastatrix and host plant cells
Highlighted changes in cell wall integrity and the presence of reproductive structures

Conclusions

The study illustrates a reliable method for examining plant-fungal interactions
Contributes to understanding biotrophic and necrotrophic fungal diseases

Frequently Asked Questions

What types of fungi can be analyzed using this protocol?

The protocol is applicable for studying biotrophic and necrotrophic fungi, including specific pathogens like Hemileia vastatrix and Cercospora coffeicola.

Do I need specialized equipment for this experiment?

No, this protocol is designed to be performed using standard light microscopy without requiring expensive equipment.

Can this technique be applied to other plant species?

Yes, further studies may explore its applicability to various plant-fungal interactions across different pathosystems.

What is the importance of pectin in plant-pathogen interactions?

Pectin is crucial for maintaining cell wall integrity and can influence the outcome of pathogen invasion and plant defense mechanisms.

How does the double staining method enhance visualization?

Double staining with cotton blue and ruthenium red allows clear differentiation of fungal structures and plant tissue under light microscopy.

What are some observed effects on plant cells during fungal infection?

The protocol reveals cell wall breakdown, loss of integrity in interaction zones, and presence of reproductive fungal structures in the plant tissue.

This protocol describes a microscopical method to detect pectin in coffee-fungus interaction.

This protocol is significant as it make it possible to identify the plant-fungus interaction. In addition to coloring the pectin, it also demonstrates defects on the pathogen cell wall polymer and allows the identification of fungus structures. This simple and uncomplicated technique can be performed using light microscopy and it doesn't require any expensive equipment such as scanning electron microscope.

In addition, it is a fast technique that obtains an excellent distinction of plant structures for histopathological studies. The technique identifies the plant-fungus interaction. Hence, it collaborates to diagnose the diseases caused by biotrophic and necrotrophic fungi such as coffee rust and cercosporiosis.

View the full transcript and gain access to thousands of scientific videos