Quantitative Wood Anatomy—Practical Guidelines

Abstract

Quantitative wood anatomy analyzes the variability of xylem anatomical features in trees, shrubs, and herbaceous species to address research questions related to plant functioning, growth, and environment. Among the more frequently considered anatomical features are lumen dimensions and wall thickness of conducting cells, fibers, and several ray properties. The structural properties of each xylem anatomical feature are mostly fixed once they are formed, and define to a large extent its functionality, including transport and storage of water, nutrients, sugars, and hormones, and providing mechanical support. The anatomical features can often be localized within an annual growth ring, which allows to establish intra-annual past and present structure-function relationships and its sensitivity to environmental variability. However, there are many methodological challenges to handle when aiming at producing (large) data sets of xylem anatomical data. Here we describe the different steps from wood sample collection to xylem anatomical data, provide guidance and identify pitfalls, and present different image-analysis tools for the quantification of anatomical features, in particular conducting cells. We show that each data production step from sample collection in the field, microslide preparation in the lab, image capturing through an optical microscope and image analysis with specific tools can readily introduce measurement errors between 5 and 30% and more, whereby the magnitude usually increases the smaller the anatomical features. Such measurement errors—if not avoided or corrected—may make it impossible to extract meaningful xylem anatomical data in light of the rather small range of variability in many anatomical features as observed, for example, within time series of individual plants. Following a rigid protocol and quality control as proposed in this paper is thus mandatory to use quantitative data of xylem anatomical features as a powerful source for many research topics.

Scientific Merits and Analytical Challenges of Tree‐Ring Densitometry

“…The general procedure to produce wood anatomical density involves (1) the preparation of thin sections of 10-to 20-μm thickness using a microtome, (2) staining the section with a reagent such as safranin to increase contrast, (3) capturing high-resolution imagery of the section, and (4) measuring the tracheid dimensions in the anatomical images with image analysis techniques (Gärtner & Schweingruber, 2013;von Arx et al, 2016). The tracheids should be cut orthogonally and the section thickness should be kept constant within a data set as deviations in both aspects will change the measured tracheid wall and lumen dimensions (Decoux et al, 2004;Elliott & Brook, 1967;von Arx et al, 2016). Cutting can produce cracks and broken tracheid walls that reduce data quality and efficiency of image analysis, but these issues can be largely avoided when stabilizing the wood before cutting (Schneider & Gärtner, 2013;von Arx et al, 2016).…”

“…The tracheids should be cut orthogonally and the section thickness should be kept constant within a data set as deviations in both aspects will change the measured tracheid wall and lumen dimensions (Decoux et al, 2004;Elliott & Brook, 1967;von Arx et al, 2016). Cutting can produce cracks and broken tracheid walls that reduce data quality and efficiency of image analysis, but these issues can be largely avoided when stabilizing the wood before cutting (Schneider & Gärtner, 2013;von Arx et al, 2016). Images of anatomical samples are then manually captured with a camera mounted on a microscope with a resolution of~1-2 pixels/μm, and multiple overlapping images are stitched to form an overall composite image of the anatomical sample using image-stitching software (von Arx et al, 2016).…”

“…Cutting can produce cracks and broken tracheid walls that reduce data quality and efficiency of image analysis, but these issues can be largely avoided when stabilizing the wood before cutting (Schneider & Gärtner, 2013;von Arx et al, 2016). Images of anatomical samples are then manually captured with a camera mounted on a microscope with a resolution of~1-2 pixels/μm, and multiple overlapping images are stitched to form an overall composite image of the anatomical sample using image-stitching software (von Arx et al, 2016). This time-consuming digitization of entire anatomical sections can be improved in efficiency and quality by using automated microscope systems or thin-section slidescanners that automatically batch-process multiple anatomical samples (Castagneri et al, 2018;Pacheco et al, 2018).…”

Axial xylem architecture ofLarix deciduaexposed to CO2enrichment and soil warming at the tree line

“…In total, we selected ten axially well-distributed discs per tree, six from the stem and four from the roots ( Figure S2). We followed the standard protocol for cutting micro-sections and collecting high-resolution images proposed by von Arx, Crivellaro, Prendin, Čufar, and Carrer (2016). From each disc, we extracted radial wood samples from opposite radii ( Figure S2) and produced 10-15 μm thick cross-sections using a rotary microtome (Leica RM2245, Leica Biosystems, Nussloch, Germany).…”

Cell size and wall dimensions drive distinct variability of earlywood and latewood density in Northern Hemisphere conifers

“…However, the relatively new approach of measuring interannual records of anatomical properties (e.g. Pritzkow et al, 2014;Castagneri et al, 2015) has not yet reached its full potential, mainly because of technical difficulties in producing data (see Prendin et al, 2017 andvon Arx et al, 2016 for recent progress). In the broader context of tree growth, one potential application of this approach is in studying the link between, on the one hand, widespread and well-replicated tree-ring data (ring width and density) and, on the other, the fewer and less well replicated data sets for quantitative wood anatomy (e.g.…”