Unprecedented three-dimensional X-ray microscope methodology to image
plants at cellular resolution
Improved imaging technology advances how we measure traits in crop
species

Date:
December 7, 2021
Source:
Donald Danforth Plant Science Center
Summary:
Measuring plant phenotypes, a term used to describe the observable
characteristics of an organism, is a critical aspect of studying
and improving economically important crops. Phenotypes central to
the breeding process include traits like kernel number in corn,
seed size in wheat, or fruit color in grape. These features
are visible to the naked human eye but are in fact driven by
microscopic molecular and cellular processes in the plant. Using
three-dimensional (3D) imaging is a recent innovation in the plant
biology sector to capture phenotypes on the 'whole-plant' scale:
from miniscule cells and organelles in the roots, up to the leaves
and flowers. However, current 3D imaging processes are limited by
time-consuming sample preparation and by imaging depth, usually
reaching only a few layers of cells within a plant tissue.



FULL STORY ========================================================================== Measuring plant phenotypes, a term used to describe the observable characteristics of an organism, is a critical aspect of studying
and improving economically important crops. Phenotypes central to the
breeding process include traits like kernel number in corn, seed size in
wheat, or fruit color in grape. These features are visible to the naked
human eye but are in fact driven by microscopic molecular and cellular processes in the plant. Using three-dimensional (3D) imaging is a recent innovation in the plant biology sector to capture phenotypes on the "whole-plant" scale: from miniscule cells and organelles in the roots,
up to the leaves and flowers. However, current 3D imaging processes
are limited by time-consuming sample preparation and by imaging depth,
usually reaching only a few layers of cells within a plant tissue. New
research led by Christopher Topp, PhD, associate member at the Donald
Danforth Plant Science Center, and Keith Duncan, a research scientist
in his lab, have pioneered X-ray microscope technology to image plant
cells, whole tissues, and even organs at unprecedented depths with
cellular resolution. The work, supported by Valent BioSciences LLC and
Sumitomo Chemical Corporation, was recently published in the scientific
journal Plant Physiology, titled X-ray microscopy enables multiscale high-resolution 3D imaging of plant cells, tissues, and organs. This work
will enable plant scientists globally to study above and below-ground
traits at revolutionary clarity.


========================================================================== "This paper focuses on the multiscale," says corresponding author
Chris Topp, "because plants are multiscale. An ear of corn starts off
as a microscopic group of cells called a meristem. Meristem cells will eventually form all the visible parts of the corn plant through division
and growth." Their improved 3D X-ray microscopy (XRM) technology allows
the researchers to relate the developmental microstructure of the plant,
such as meristem cells, to visible traits as they mature, for example
leaves and flowers. In other words, 3D XRM provides cellular-level
resolution of entire plant organs and tissues.

In addition, their XRM methodology can also image below-ground
structures at exceptional resolution, including roots, fungi, and other microbes. "Plant roots drive a lot of important biological processes;
they feed microbes in the soil, and in return the plants get phosphorus
and nitrogen," explains Topp. "We know the interaction between roots
and microbes is important because it was a primary source of phosphorus
and nitrogen before we invented chemical fertilizers." Our dependency on chemical fertilizers in standard agricultural practices have, in turn,
made major contributions to global climate change.

"Half of all the biologically-available nitrogen was made in a factory in
the last 100 years," Topp continues. "This process has been estimated to
use 3% of all available energy and generate 3% of greenhouse gas emissions
on planet Earth every single year." Therefore, a critical component of
the sustainable agriculture movement includes reducing chemical inputs
and instead fostering natural interactions between roots and microbes
below ground. "We haven't had the tools to understand these interactions
until recently," says Topp. "3D XRM can help unlock the potential of re-establishing these natural alliances in our agriculture systems."
3D XRM methodology is unique compared to other imaging approaches in
plant biology because of its ability to yield essentially perfect 3D
clarity of plant structure. Other common methods, such as photon-based tomography, are limited by shallow imaging depths and are optimized in
a select few species of plants.

In contrast, by using 3D XRM, the team led by Topp and Duncan are able to
image "thick tissues that are recalcitrant to typical, optical methods,"
in a whole host of economically important crops, including corn, foxtail millet, soybean, teff, and grape. "This paper is the first of its kind
to show the breadth of what 3D XRM can do," Topp notes.

A major goal of the paper is to establish a reproducible protocol
for other plant scientists interested in 3D XRM imaging. To do so,
lead author Keith Duncan spent a lot of time -- and trial and error -- preparing samples to optimize the contrast between the plant and its background. X-ray imaging works through differential absorption, where
dense material (like minerals in the soil) absorbs more X-rays and shows
up darker on an image. However, biological matter like plant tissue has
low X-ray absorption, and the team was at risk of completely washing out
the material they were interested in imaging. "Solving that problem for
one kind of sample -- like a root tip -- is one thing," explains Topp,
"but the idea of the paper was to give plant scientists working on a
variety of relevant plant tissues and species the access to these methods.

We want to broadly apply 3D XRM to plant systems above and below
ground." As such, their published methodologies greatly advance the number
of plant species and the types of plant tissues that can be imaged at
nearly perfect resolution.

Keith Duncan continues to lead the partnership of the Topp Lab with
Valent Biosciences and Sumitomo Chemical, focusing on improving 3D XRM capabilities.

He often collaborates with Kirk Czymmek, PhD, director of the Danforth
Center's Advanced Bioimaging Laboratory, who was also an author on
the paper.

Next on the horizon is to image 3D structures of fungal networks in
the soil.

Part of that work includes improving machine learning approaches, such
that a computer is trained to recognize what within an image is a root,
soil, or spore (the reproductive cells of a fungus). Their work will
continue to develop new technological approaches to improve our multiscale understanding of the "whole plant," from the microscopic to the visible.

========================================================================== Story Source: Materials provided by
Donald_Danforth_Plant_Science_Center. Note: Content may be edited for
style and length.


========================================================================== Journal Reference:
1. Keith E Duncan, Kirk J Czymmek, Ni Jiang, August C Thies,
Christopher N
Topp. X-ray microscopy enables multiscale high-resolution 3D imaging
of plant cells, tissues, and organs. Plant Physiology, 2021; DOI:
10.1093/ plphys/kiab405 ==========================================================================

Link to news story: https://www.sciencedaily.com/releases/2021/12/211207102000.htm

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