Plant phenotyping methods.

PAT: An Image Analysis Tool for Automated Scoring of Pollen in Alexander-Stained Anthers. →

Quantitative pollen viability analysis is a critical but labor-intensive step in plant reproductive biology. Existing deep-learning Segment Anything Models (SAM) fail to reliably segment viable pollen in Alexander-stained anthers. To address this, we fine-tuned an existing Cellpose-SAM model for pollen segmentation. We integrated it into PAT (Pollen Analysis Tool), a cross-platform desktop application. PAT features instance segmentation with interactive quality control, an in-app model retraining module, and publication-ready statistical outputs. We deployed PAT in an EMS suppressor screen of semi-sterile Arabidopsis smg7-6 mutants, enabling efficient candidate prioritization for whole-genome sequencing and mapping of the candidate mutation. This screen led to the identification of a point mutation in CAP-D2 (capd2-2), a Condensin I subunit, that rescues the smg7-6 meiotic phenotype. Notably, mutation in a Condensin II subunits (CAP-D3 and CAP-H2) does not confer rescue. Further characterization suggests the capd2-2 allele is hypomorphic, showing no defects in vegetative growth, chromocenter compaction, or transposable element silencing. Collectively, we demonstrate that accessible AI tools have the potential to bridge gaps in plant phenotyping and accelerate the pace of biological discovery.

Why it matches plant phenotyping methods花粉生存性を画像から自動推定するセグメンテーション手法とソフトウェアPATの開発が研究の中心であり、植物表現型計測ツールとして明確に該当する。

titlePAT: An Image Analysis Tool for Automated Scoring of Pollen in Alexander-Stained Anthers

Root-TransUNet enables high-throughput phenotyping of Arabidopsis thaliana roots as a parameter in Heterodera schachtii parasitism →

Introduction Plant parasitism by sedentary plant-parasitic nematodes is a dynamic and continuously evolving process, accompanied by profound remodelling of host root system architecture across distinct infection stages. However, the physiology and anisotropic growth of Arabidopsis thaliana roots under Heterodera schachtii infection, together with complex lateral root proliferation and increasingly dense, overlapping morphology, pose substantial challenges for accurate image segmentation. Methods Here, we introduce Root-TransUNet, an optimised segmentation architecture that extends TransUNet by incorporating a composite loss function to improve boundary precision and structural continuity, as well as dual-stage strip-pooling (SP) modules to enhance elongated and directional root features. These adaptations address the unique morphological complexity of the infected root system. Additionally, we integrated Root-TransUNet into a high-throughput phenotyping pipeline and applied it to an existing dataset of ~120,000 images of 362 A. thaliana MAGIC recombinant inbred lines collected over several months of infection. By extracting root system architecture traits, including root surface area and estimated root volume across infection stages, we enabled stage-specific association analyses between host root growth and nematode performance across these genotypes. Results Root-TransUNet achieved strong segmentation performance, demonstrating improved structural continuity and boundary precision compared with widely used CNN- and Transformer-based baselines, including UNet++. Stage-specific analyses revealed that the relationship between host root traits and nematode performance changed as infection progressed. During establishment, nematode number was largely independent of initial root size and varied strongly among genotypes, whereas during the reproductive phase (10-30 dpi), greater root expansion coincided with reduced estimated nematode volume accumulation. Notably, nematode burden was largely independent of host root size before infection, indicating that root quantity was generally not a limiting factor for infection in this experiment. Discussion These results demonstrate that Root-TransUNet can robustly segment infected root systems across a wide range of nematode infection densities, providing a scalable image-analysis framework for studying plant-parasitic nematode parasitism in combination with host root phenotyping.

Why it matches plant phenotyping methods感染根系の画像セグメンテーション手法を開発し、高スループット表現型解析パイプラインに統合して根系形態形質を抽出しており、表現型取得・抽出法が中心的である。

abstractHere, we introduce Root-TransUNet, an optimised segmentation architecture that extends TransUNet by incorporating a composite loss function to improve boundary precision and structural continuity, as well as dual-stage strip-pooling (SP) modules to enhance elongated and directional root features.

The state of plant photosystem II reaction centres affects the rate of non-photochemical quenching →

Abstract Plants employ non-photochemical quenching (NPQ) to protect their photosynthetic apparatus from photodamage. The response latency of NPQ following changes in light intensity is thought to significantly decrease photosynthetic efficiency. The amount of NPQ is commonly quantified from chlorophyll-fluorescence techniques using the Stern–Volmer equation, which requires fully closed reaction centres (RCs) of photosystem II, yielding NPQ in the absence of photochemical quenching ( $${\rm{NPQ}}^{\rm{Closed}}$$ NPQ Closed ). However, in nature, NPQ and photochemical quenching are normally present simultaneously. Therefore, to obtain a full understanding of this process, NPQ should also be explored when the RCs are open. Here we developed two methodologies to obtain NPQ in the presence of photochemistry ( $${\rm{NPQ}}^{\rm{Open}}$$ NPQ Open ) using both fluorescence lifetime and fluorescence yield measurements. A detailed comparison in Arabidopsis thaliana plants reveals that the value of $${\mathrm{NPQ}}^{\mathrm{Open}}$$ NPQ Open is ~35% lower than that of $${\rm{NPQ}}^{\rm{Closed}}$$ NPQ Closed . This difference is consistently observed across all measurements and is seen both upon closing ( $${\rm{NPQ}}^{\rm{Open}}\to {\rm{NPQ}}^{\rm{Closed}}$$ NPQ Open → NPQ Closed ) and upon reopening ( $${\mathrm{NPQ}}^{\mathrm{Closed}}\to {\mathrm{NPQ}}^{\mathrm{Open}}$$ NPQ Closed → NPQ Open ) of the RCs. We show that this difference can be explained by the presence of RC-induced ‘instantaneous’ switching of the NPQ quenching rate. This means that, in plants, NPQ is much more economical than is widely believed, it is large when its presence is needed, and it decreases instantaneously when the need disappears.

Why it matches plant phenotyping methods植物の光合成状態（NPQ）を測定するための蛍光寿命・蛍光収率に基づく2つの方法を開発し、比較検証しているため、方法開発が中心である。

abstractHere we developed two methodologies to obtain NPQ in the presence of photochemistry ( $${\rm{NPQ}}^{\rm{Open}}$$ NPQ Open ) using both fluorescence lifetime and fluorescence yield measurements.

From phenoscope to GreenLab model of Arabidopsis to decipher genotype and treatment effects. →

leaf development was tracked dynamically from high-throughput phenotyping image series using the SAM-2 deep learning model, enabling automated segmentation and tracking of individual leaves across hundreds of plants from different accessions grown under different water and nitrogen conditions. From these time series, leaf-level growth dynamics were reconstructed and used to calculate key developmental traits, including phyllochron and relative expansion rates. These measurements were further integrated into the GreenLab model to infer plant-scale parameters such as radiation use efficiency (RUE) and leaf demand parameters. Statistical analyses revealed significant genotype, treatment, and interaction effects on several traits, with stable genotype rankings but contrasted sensitivities to environmental conditions. While whole-rosette growth captured strong and consistent responses, individual leaf contributions to global growth remained limited, highlighting the integrative nature of rosette-scale dynamics. Model-derived parameters provided additional insight into plant strategies, showing that genotypes differ in both the duration and timing of leaf demand, with generally stable patterns across environments. In parallel, QTL mapping was performed using leaf-level measurements and phyllochron traits. This analysis identified shared and trait-specific genetic loci, including loci associated with leaf emergence dynamics that were not detected using traditional rosette-scale integrative traits. Overall, this study demonstrates that combining deep learning-based image analysis with mechanistic modeling enables large-scale and quantitative characterization of plant development, providing biologically interpretable traits and new insights into their genetic and environmental determinism. It highlights the potential of combining high-throughput image-based phenotyping, deep learning, and mechanistic modeling to generate biologically interpretable traits and to assess genetic and environmental influences on these traits.

Why it matches plant phenotyping methods深層学習による葉の自動セグメンテーション・追跡を開発的に適用し、時系列画像から葉レベルおよび植物体レベルの発達形質を定量化しているため、表現型取得・抽出が研究の中心である。

abstractleaf development was tracked dynamically from high-throughput phenotyping image series using the SAM-2 deep learning model, enabling automated segmentation and tracking of individual leaves across hundreds of plants

Non-destructive quantification of shoot apical meristem homeostasis for prediction of plant architecture and biomass using robot-based 3D imaging and photosynthesis measurements →

Shoot apical meristem (SAM) homeostasis integrates environmental and genetic cues to regulate growth dynamics that drive biomass accumulation and crop yield; however, no robust, non-destructive, quantitative proxy has been established for modeling or monitoring SAM-homeostasis-associated dynamics. Here, we developed a novel robot-based 3D imaging system and a custom pot-chamber gas exchange system to non-destructively measure plant occupation volume (POV) and whole-plant photosynthetic rate in wild-type Arabidopsis plants and nine mutants with disrupted SAM homeostasis. We demonstrate that POV robustly captures 3D plant architecture, whereas whole-plant photosynthetic rate serves as a superior proxy for optimal growth dynamics and final biomass associated with SAM homeostasis, outperforming conventional traits such as leaf number, leaf size, total leaf area, and rosette diameter. The strong positive correlations among POV, whole plant photosynthesis, and biomass accumulation establish a powerful new framework for quantitative studies of SAM homeostasis and data-driven evaluation of plant architecture.

Why it matches plant phenotyping methodsロボット3D画像とカスタムガス交換による非破壊的な植物形態・光合成表現型測定系を開発し、従来形質との比較検証も行っており、方法が研究の中心である。

abstractwe developed a novel robot-based 3D imaging system and a custom pot-chamber gas exchange system to non-destructively measure plant occupation volume (POV) and whole-plant photosynthetic rate

Apical3DTip: Elliptic Cross-section-based Reconstruction for the Embryo Initial Cell of Arabidopsis →

Background Cell geometry plays a central role in determining division orientation and body axis formation during early embryogenesis in Arabidopsis thaliana . However, quantitative analysis of dynamic three-dimensional (3D) morphology remains challenging because live-imaging studies often rely on two-dimensional (2D) projections, while existing 3D reconstruction approaches, including mesh-based methods, often lose the original orientation information relative to the ovule and require labor-intensive mesh correction. In addition, embryo positional fluctuation caused by floating in liquid medium and continuous growth makes it difficult to analyze temporal morphological changes within a common coordinate system. Results We developed a robust framework for quantitative 3D and four-dimensional (4D; 3D + time) analysis of embryo initial cell (apical cell) morphology. The method first establishes a standardized 3D coordinate system by normalizing cell orientation based on the bottom plane and the optical axis of the observation. Cell morphology is then reconstructed through ellipse-based approximation of serial cross-sections extracted from stacked imaging data, enabling accurate geometric characterization without the need for complex surface mesh reconstruction. To evaluate shape anisotropy, we quantified the apical cell shape in 3D. The framework further supports the characterization of volumetric features of subsequent division, providing a basis for quantifying 3D embryogenesis. Conclusion Our framework provides a simple and noise-reduced approach for quantitative analysis of living cell morphology in 3D. We named the integrated method of combining coordinate normalization with elliptical cross-section-based reconstruction Apical3DTip. This method enables consistent comparison of cell shapes without extensive manual corrections. The method overcomes key limitations of 2D projection-based and mesh-dependent analyses and offers a practical platform for quantifying cell shape and daughter cell shapes in 3D. More broadly, it provides a quantitative foundation for exploring the relationship between cell geometry, morphodynamics, and developmental patterning in living plant embryos.

Why it matches plant phenotyping methods植物胚の細胞形態を3D・4D画像から定量化する再構成手法を開発しており、表現型取得・抽出法が研究の中心である。

abstractWe developed a robust framework for quantitative 3D and four-dimensional (4D; 3D + time) analysis of embryo initial cell (apical cell) morphology.

Low-Cost Continuous-Wave Diffusive Microtomography with Fiber-Scanned White-Light Illumination →

Tomographic microscopy enables three-dimensional internal imaging but often requires expensive optical or X-ray instrumentation. Here we present an ultra-low-cost continuous-wave diffusive tomography (CWDT) system for biological samples. The system uses a smartphone microscope, a white LED coupled into an optical fiber, 3D-printed micropositioners, and a physics-based forward model optimized with machine learning. We demonstrate full-color volumetric reconstructions from a tartrazine-cleared poplar section, a scattering phantom, fungal mycelium near an Arabidopsis root, and thick poplar branch imaging with an inserted side-emitting fiber. The current results are qualitative and exploratory, but they show that scanned fiber illumination and inexpensive hardware can produce useful three-dimensional reconstruction outputs for low-cost microscopy experiments.

Why it matches plant phenotyping methods低コスト三次元断層イメージング法そのものを開発し、ポプラ組織・枝やシロイヌナズナ根近傍を対象に植物の内部構造を可視化しているため、植物形態の取得法として中心的です。

abstractHere we present an ultra-low-cost continuous-wave diffusive tomography (CWDT) system for biological samples.

Hyperbolic topological data analysis mapper reveals dynamic trait–environment patterns in plant phenomics →

Modern plant phenotyping faces the challenge of interpreting complex, high-dimensional data. Traditional analytical tools often fail to capture the non-linear, hierarchical, and temporal relationships that define plant responses under multifactorial conditions. We present the Hyperbolic Topological Data Analysis Mapper (HTDA-Mapper), a novel algorithm designed to overcome these limitations by embedding data in Poincaré ball space. Unlike conventional Euclidean approaches, HTDA-Mapper preserves the hierarchical structure of phenotypic traits, improves cluster resolution, and reveals hidden growth trajectories across treatments and time, offering a powerful means to explore latent phenoms. The pipeline supports both quantitative data and images. When integrated with unsupervised contrastive learning, HTDA-Mapper identifies similarities and differences in raw image data without requiring manual labelling or post hoc processing. We applied this framework to a high-throughput phenotyping (HTP) dataset of over 27,000 images of Arabidopsis thaliana seedlings exposed to varying nutrient levels and priming agents at different concentrations over seven days. Using cubical complexes, HTDA-Mapper mapped relationships between treatment variables, compound concentrations, and phenotypic outcomes. Furthermore, it reliably detected compound-specific effects, uncovered dynamic trait–environment interactions, revealed phenotypic trajectories not captured by conventional methods, and facilitated biologically meaningful interpretation of the complex dataset. By preserving the geometry and temporal evolution of plant development, HTDA-Mapper sets a new standard for HTP analysis. Beyond phenomics, it is a versatile tool for other omics, such as transcriptomics and metabolomics, where structured, high-dimensional data is prevalent. HTDA-Mapper can accelerate data-driven crop improvement by uncovering effective compounds, robust genotypes, and adaptive growth strategies that enhance plant resilience.

Why it matches plant phenotyping methods植物フェノミクスの高次元画像・形質データを解析するHTDA-Mapperアルゴリズムを開発し、27,000枚超の植物画像データで適用・評価しているため、解析手法が中心的である。

abstractWe present the Hyperbolic Topological Data Analysis Mapper (HTDA-Mapper), a novel algorithm designed to overcome these limitations by embedding data in Poincaré ball space.

Coupling of high-resolution mass spectrometer and photosynthesis system for comprehensive leaf volatile metabolite profiling. →

Background Leaf-level biogenic volatile organic compounds (BVOCs) emissions represent a major source of organic gases in the atmosphere, influencing both climate and air quality. These emissions are strongly driven by environmental perturbations, which affect individual plant- to ecosystem-level processes. Uncovering all the BVOCs and understanding how their emissions respond to altered environmental conditions provide critical insights into vegetation-driven changes in atmospheric chemistry. We developed a tandem instrumentation setup that integrates a proton transfer reaction time-of-flight mass spectrometer (PTR-ToF-MS) with parts-per-trillion detection limits and a photosynthetic infrared gas exchange system for the untargeted survey of all the BVOCs. This novel system enables simultaneous, real-time monitoring of BVOC emissions and photosynthetic parameters at the leaf level, offering new opportunities to disentangle the physiological and environmental drivers of VOC release. Furthermore, we established the VOC Analysis and Processing Optimization Resource (VAPOR), an open-access software tool designed for rapid data post-processing and the analysis of the variability of hundreds of BVOCs. We assessed the performance of the tandem system under varying background conditions, using standard gas mixtures and a range of environmental factors. Results Blank emissions were substantially lower for major BVOCs (e.g., isoprene) compared to those observed in plant emissions. Despite this, the observation of background-level VOCs highlights the importance of routinely acquiring and accounting for blank measurements in analyses using the coupled instrumentation. Introduction of known VOC concentrations to the system demonstrated a linear response across different compounds with varying molecular compositions, indicating minimal gas loss regardless of chemical moieties within the coupled instrumentation. We applied the optimized system to investigate the physiological mechanisms driving BVOC emissions across different genotypes of poplar and pennycress. The high mass resolution capabilities of the PTR-ToF-MS, coupled with comprehensive VAPOR-driven data analysis, enabled the identification of several important BVOCs, including methanol and methanethiol; these BVOCs displayed substantial variation across pennycress genotypes and showed concentrations ~ 100-350% higher than the blank. Moreover, isoprene emissions varied significantly among poplar genotypes grown in different potting media. Conclusions Tandem instrumentation offers a powerful tool for profiling volatile molecular markers and elucidating their genetic and environmental underpinnings. This approach enhances our ability to predict BVOC emissions in response to genotype by environmental interactions and contributes to a deeper understanding of vegetation responses to environmental changes.

Why it matches plant phenotyping methods葉レベルの植物揮発性物質排出と光合成パラメータを取得するタンデム計測系を開発・検証し、解析ソフトウェアも提供しているため、植物表現型取得法が中心である。

abstractWe developed a tandem instrumentation setup that integrates a proton transfer reaction time-of-flight mass spectrometer (PTR-ToF-MS) with parts-per-trillion detection limits and a photosynthetic infrared gas exchange system for the untargeted survey of all the BVOCs.

Analysis of Cauline Leaf Development in Arabidopsis thaliana Using Time-Lapse Confocal Microscopy. →

Understanding cellular growth dynamics in plants requires precise, long-term imaging of developing tissues. Cauline leaves are produced during the transition from vegetative to reproductive development and provide a useful system for studying how laminar organs diversify in form and function. While other laminar organs, such as rosette leaves and sepals, have been extensively studied, early cauline leaf development remains technically challenging to capture due to their concealed position, curved morphology, and the presence of dense trichomes. Here, we provide a complete pipeline for the dissection, confocal imaging, 2.5D segmentation, and image analysis of initiating cauline leaves in Arabidopsis thaliana . This method enables reproducible, high-resolution imaging of cauline leaves, supporting robust quantitative analysis of growth across developmental stages at cellular scale resolution. Key features • Fine dissection method for exposing initiating cauline leaves in Arabidopsis thaliana . • Long-term confocal live imaging of cauline leaf development at cellular resolution. • Optimized imaging parameters for high-fidelity 2.5D segmentation and growth analysis in MorphoGraphX.

Why it matches plant phenotyping methodsカウリン葉の成長を細胞レベルで定量化するための解剖、共焦点イメージング、2.5Dセグメンテーション、画像解析パイプラインが中心的に開発・提示されている。

abstractHere, we provide a complete pipeline for the dissection, confocal imaging, 2.5D segmentation, and image analysis of initiating cauline leaves in Arabidopsis thaliana .

GraFT: A robust network-based spatiotemporal analysis of filamentous structures. →

The actin cytoskeleton forms a dynamic network composed of filaments that remain flexible when bundled up, leading to complex filamentous structures in plant cells. Understanding the properties of these filamentous structures under different conditions and in different cell types can provide insight into their function. Yet, despite developments in the study of the plant actin cytoskeleton, it remains challenging to segment and identify actin filamentous structures, preventing quantification of their spatiotemporal properties. To address this problem, we devised a network-based approach termed Graph of Filaments over Time (GraFT) to trace and track filamentous structures in cytoskeleton networks extracted from two-dimensional time series imaging data. Our comparative analyses using both synthetic test cases and real-world actin cytoskeleton networks of Arabidopsis thaliana hypocotyls exposed to different treatments demonstrated that GraFT accurately traces and tracks actin filamentous structures. Moreover, GraFT facilitates automated quantification of properties for filamentous structures, providing fine-grained insights of effects of different treatments on the level of individual structures. Therefore, GraFT offers a substantial step toward an automated framework facilitating robust spatiotemporal studies of the plant actin cytoskeleton.

Why it matches plant phenotyping methods植物細胞の画像時系列からアクチン繊維構造を追跡・セグメント化し、その時空間特性を自動定量する手法の開発と検証が中心であるため。

abstractwe devised a network-based approach termed Graph of Filaments over Time (GraFT) to trace and track filamentous structures in cytoskeleton networks extracted from two-dimensional time series imaging data.

Non-Equilibrium Spatial Encoding of Nanoscale Mechanical Relaxation in Growing Plant Epithelial Cells →

A central problem in soft and biological physics is how molecular-scale activity and remodelling coarse-grain into emergent mechanical laws at larger scales. In growing cell walls (polymeric composite materials that surround 90% of living organisms’ cells) irreversible deformation is not controlled by elastic stress alone. Instead, growth depends on the interplay between energy storage, dissipation, and the local timing of viscoelastic relaxation. Although dynamic atomic force microscopy (AFM) resolves storage and loss moduli ( E′, E″) of living walls at nanometre resolution, these observables have remained phenomenological and disconnected from constitutive field variables. Here we introduce a physics-based inversion framework that converts AFM measurements of epidermal cells of living Arabidopsis plants into spatially resolved fields of stiffness k , viscosity η , and relaxation time τ . By analysing the spatial gradients of E′ and E″, we uncover organized mechanical heterogeneities governed by cellular confinement and stress focusing. We demonstrate that the local relaxation time is encoded directly in the coupling between storage and dissipation, yielding the pointwise relation τ = (1/ ω ) ∂ E ’/∂ E ’’, where ω is the indentation frequency. This relation enables model-independent extraction of mechanical timescales and establishes a general route from nanoscale non-equilibrium rheology to continuum descriptions of growth in living and active soft materials. Significance How molecular-scale activity gives rise to tissue-scale form is a central challenge in biological physics. Although growth is fundamentally a non-equilibrium mechanical process, experimental measurements at the nanoscale have not been directly connected to the constitutive parameters that govern morphogenesis. We introduce a framework that converts dynamic atomic force microscopy maps of storage and loss moduli into spatially resolved fields of stiffness, viscosity, and relaxation time in living cell walls. By revealing that mechanical relaxation is encoded in the local coupling between elastic storage and viscous dissipation, our work provides a route from nanoscale rheology to growth-relevant mechanical timing. This establishes a quantitative bridge between molecular remodeling and continuum mechanics, enabling direct experimental constraints on multiscale theories of morphogenesis.

Why it matches plant phenotyping methods生きたArabidopsis細胞のAFM測定を物理ベースで反転し、剛性・粘性・緩和時間という植物細胞壁の機械的形質を空間的に抽出する新規フレームワークが研究の中心である。

abstractHere we introduce a physics-based inversion framework that converts AFM measurements of epidermal cells of living Arabidopsis plants into spatially resolved fields of stiffness k , viscosity η , and relaxation time τ .

AutoSiQ: a curated haploid Arabidopsis thaliana inflorescence dataset with a fine-grained silique ontology and a deep learning application for haploid fertility quantification. →

Doubled haploid (DH) technology can fast-track crop breeding. Haploid induction yields haploids with only one set of genomes, which are usually sterile. Haploid fertility (HF) is the ability of haploid plants to set seed, and it is a critical bottleneck in DH pipelines. Genetic mechanisms to restore HF hold immense potential in DH crop breeding, yet its phenotyping remains manual, destructive, and inconsistent. While recent advances in imaging and machine learning have improved throughput for general plant traits, no curated image dataset exists for Arabidopsis thaliana that explicitly represents HF. Here, we present AutoSiQ, a dataset and baseline deep learning pipeline for automated HF quantification. AutoSiQ includes high-resolution scanned inflorescences annotated with a seven-class ontology encompassing green siliques, green fertile siliques, mature siliques, fertile siliques, cracked fertile siliques, cracked siliques, and flowers. This multi-class annotation scheme preserves biologically meaningful information beyond binary fertile/non-fertile distinctions, enabling reliable fertility estimation and future phenotyping applications. We release baseline object detection models (YOLOv5), trained using the AutoSiQ dataset, and evaluate their performance across confidence thresholds. Model predictions strongly correlate with manual counts, achieving R² up to 0.94 for total silique number estimation. We further demonstrate AutoSiQ's utility for automated haploid fertility rate (HFR) estimation and genotype discrimination between two contrasting genotypes (WT and bmf2 mutant). A longitudinal analysis identifies ~60 days after sowing (DAS) as the optimal harvest time for maximizing mature silique counts by balancing between the number of immature buds and silique shattering. By releasing both the dataset and baseline code, AutoSiQ provides a reproducible and extensible foundation for high-throughput fertility phenotyping in haploid Arabidopsis .

Why it matches plant phenotyping methodsハプロイド稔性を画像から定量するデータセットと深層学習パイプラインを開発・評価しており、植物フェノタイピング手法が中心である。

abstractHere, we present AutoSiQ, a dataset and baseline deep learning pipeline for automated HF quantification.

Segment Any Plant (SAP): Foundation-Model Segmentation for Plant Time-Series Phenotyping →

Quantitative studies of plant growth and environmental responses increasingly rely on time-series imaging, yet automated segmentation remains challenging due to continuous growth, large non-rigid morphological change, and frequent self-occlusion. Traditional image-processing pipelines and task-specific deep learning models often require extensive annotated datasets and retraining, limiting portability across species, developmental stages, and imaging conditions. Here we present SAP (Segment Any Plant), a plant-focused framework that leverages the pretrained Segment Anything Model 2 (SAM2) to enable few-shot, training-free segmentation of plant time-series imagery. SAP integrates interactive prompting, automated temporal mask propagation, and centerline extraction within a web-based interface, allowing users to move from raw images to quantitative descriptors of organ shape and dynamics without programming expertise. Across multiple systems, including Arabidopsis thaliana rosette development, root growth, sunflower gravitropism, and confocal root microscopy, SAP achieves high segmentation accuracy (mean IoU 0.89–0.93) and sub-pixel centerline precision from single-frame prompting. By reducing the need for task-specific retraining, SAP provides a transferable framework for reproducible time-series phenotyping across diverse experimental contexts.

Why it matches plant phenotyping methods植物の時系列画像から器官形状・動態を抽出するセグメンテーション手法とWeb基盤を開発し、複数系で精度検証しているため、植物フェノタイピング手法が中心である。

abstractHere we present SAP (Segment Any Plant), a plant-focused framework that leverages the pretrained Segment Anything Model 2 (SAM2) to enable few-shot, training-free segmentation of plant time-series imagery.

A workflow for absolute apoplastic pH assessment during live cell imaging in plant roots →

Abstract Background Apoplastic pH is a central regulator of plant growth, development, and environmental adaptation, influencing cell expansion, nutrient uptake, and extracellular signaling. Many studies have successfully used HPTS to monitor relative changes in apoplastic pH in plants. At the same time, research increasingly targets pH-dependent biochemical and biophysical processes. Many enzymatic activities, ion binding events, and receptor–ligand interactions depend on defined proton concentrations. Accordingly, the development of reliable approaches to measure absolute pH in living tissues is gaining importance. Methods A calibration-based workflow was developed to enable quantitative assessment of absolute apoplastic pH using ratiometric HPTS imaging. The approach integrates a simplified two-point normalization strategy with an in-vitro derived sigmoidal calibration model, thereby minimizing the need for extensive in-vivo calibration curves. Confocal imaging was performed using HPTS excited at two wavelengths followed by ratiometric image processing. Data analysis is supported by a custom Fiji plugin, Ratio2pH, which converts ratiometric images into pixel-resolved maps of absolute pH. Results In vitro characterization revealed a robust, non-linear relationship between normalized HPTS ratios and pH, enabling accurate pH estimation within the physiologically relevant range of pH 5.0–7.0. When applied in-vivo to Arabidopsis thaliana roots, the workflow yielded extracellular pH estimates consistent with the pH of the incubation medium and detected reproducible pH shifts in response to pharmacological treatments. Conclusions This workflow enables reproducible, spatially resolved measurement of absolute apoplastic pH in living plant tissues. By combining a simplified calibration strategy with accessible image analysis tools, it facilitates quantitative extracellular pH measurements and their integration into biochemical and biophysical analyses.

Why it matches plant phenotyping methods生きた植物組織の絶対アポプラストpHを画像から定量する校正ワークフローを開発・検証し、Fijiプラグインも提供しているため、植物状態の取得法が中心である。

abstractA calibration-based workflow was developed to enable quantitative assessment of absolute apoplastic pH using ratiometric HPTS imaging.

The Tonoplast Topology Index-a new metric for describing vacuole organization. →

Background The plant vacuole arises by orchestrated interplay of membrane trafficking, cytoskeletal rearrangements and a variety of signaling pathways. In the root, the characteristic large central vacuole develops by endomembrane reorganization occurring mainly in the transition zone. The vacuole's bounding membrane-the tonoplast-can be visualized in vivo using fluorescent protein markers, allowing for quantitative analysis of confocal microscopy images. Tonoplast organization can thus serve as a sensitive indicator of changes to any of the processes involved in vacuole biogenesis. The Vacuolar Morphology Index (VMI) is widely accepted as a quantitative measure of vacuole structure. However, this metric has two drawbacks-it only reflects the size of the largest vacuolar compartment (missing therefore possible differences in the organization of smaller compartments), and its determination is labor intensive, limiting its use on large datasets. Results We developed an alternative metric for describing vacuole organization, named the Tonoplast Topology Index (TTI), which overcomes the above-mentioned shortcomings of the VMI. We compared the performance of our protocol with VMI on a simulated dataset and on real data. To validate the methods´ performance, we used it to confirm the previously reported differences in vacuole shape and size between Arabidopsis thaliana roots grown on the surface of an agar medium compared to those embedded inside the agar. Both VMI and TTI could efficiently detect the relatively subtle changes in vacuole organization depending on the position of the root in the agar, and provided correlated results. However, only TTI produced data with close to normal value distribution, simplifying subsequent statistical evaluation. Conclusions We present the protocol for TTI determination as a two-stage semi-automated procedure involving microscopic image analysis employing an ImageJ macro and subsequent processing of numeric data in the Jupyter Notebook environment, together with benchmarking image data. Since this implementation is freeware-based, platform-independent and (relatively) user-friendly, we hope it will find its use as a high throughput, added value alternative to the VMI metric.

Why it matches plant phenotyping methods植物液胞構造を定量化する新規指標と半自動画像解析プロトコルを開発し、シミュレーションおよび実画像で既存指標と比較・検証しているため、植物フェノタイピング手法が中心である。

abstractWe developed an alternative metric for describing vacuole organization, named the Tonoplast Topology Index (TTI)

Electrochromic polyoxometalates for sensing abiotic stress in plants. →

Introduction Understanding plant responses to abiotic stress requires an insight into plant redox activity. This study proposes a novel and cost-effective method for assessing the redox state of plants. Methods The method utilizes the electrochromic properties of polyoxometalate phosphomolybdic acid hydrate (PMA). PMA is reduced proportionally by glutathione (GSH) and ascorbic acid (AsA), which results in a measurable color change. The validity of this method was confirmed through empirical experimentation in Arabidopsis thaliana under conditions of salinity and UV radiation. Results Salinity treatments revealed a non-significant, two-phase trend in redox activity with an increase at moderate levels followed by a decrease. UVC radiation led to a substantial decrease in redox activity, indicating distress. In contrast, UVA promoted resilience, also known as eustress. Notably, UVB significantly increased redox activity, suggesting the activation of an emergency antioxidant response. Discussion A demonstrable correlation has been identified between the redox activity of plants and various stress types. This correlation facilitates the classification of responses into two distinct categories: adaptive eustress and detrimental distress. This advancement contributes to the enhancement of plant metabolic and stress tolerance evaluation.

Why it matches plant phenotyping methods植物のレドックス状態を測定する新規手法を開発し、シロイヌナズナでストレス条件下の妥当性を検証しており、表現型取得が研究の中心である。

abstractThis study proposes a novel and cost-effective method for assessing the redox state of plants.

ChronoRoot 2.0: An Open AI-Powered Platform for 2D Temporal Plant Phenotyping →

BACKGROUND: Plant developmental plasticity, particularly in root system architecture, is fundamental to understanding adaptability and agricultural sustainability. Existing automated phenotyping solutions face limitations, including binary segmentation approaches, restricted structural analysis capabilities, and text-based interfaces that limit accessibility, with most focusing solely on root structures while overlooking valuable information from simultaneous analysis of multiple plant organs. FINDINGS: ChronoRoot 2.0 builds upon established low-cost hardware while significantly enhancing software capabilities and usability. The system employs nnUNet architecture for multi-class segmentation, demonstrating significant accuracy improvements while simultaneously tracking 6 distinct plant structures encompassing root, shoot, and seed components: main root, lateral roots, seed, hypocotyl, leaves, and petiole. This architecture enables easy retraining and incorporation of additional training data without requiring machine learning expertise. The platform introduces dual specialized graphical interfaces: a Standard Interface for detailed architectural analysis with novel gravitropic response parameters and a Screening Interface enabling high-throughput analysis of multiple plants through automated tracking. Functional principal component analysis integration enables discovery of novel phenotypic parameters through temporal pattern comparison. We demonstrate multi-species analysis, with Arabidopsis thaliana and Solanum lycopersicum, both morphologically distinct plant species. Three use cases in Arabidopsis thaliana and validation with tomato seedlings demonstrate enhanced capabilities: circadian growth pattern characterization, gravitropic response analysis in transgenic plants, and high-throughput etiolation screening across multiple genotypes. CONCLUSIONS: ChronoRoot 2.0 maintains the low-cost, modular hardware advantages of its predecessor while dramatically improving accessibility through intuitive graphical interfaces and expanded analytical capabilities. The open-source platform makes sophisticated temporal plant phenotyping more accessible to researchers without computational expertise. SOFTWARE AVAILABILITY: https://chronoroot.github.io.

Why it matches plant phenotyping methods根・シュート・種子を時系列追跡し、植物形態・成長・重力応答などの表現型を抽出するオープンプラットフォームの開発と検証が中心である。

titleChronoRoot 2.0: An Open AI-Powered Platform for 2D Temporal Plant Phenotyping

Abscisic acid-mediated water stress regulation can mechanistically explain oscillations and water stress memory in stomatal conductance →

Stomatal pores, formed by guard cells, govern the critical trade-off between carbon assimilation and water loss in plants. Their dynamic responses to environmental stresses, such as stomatal oscillations and drought “stress memory” (hysteresis), have lacked a unified mechanistic explanation. While abscisic acid (ABA) is believed to play key roles in water stress responses, no model has linked its core regulatory kinetics to these complex stomatal behaviors. Here, we introduce a coupled hydropassive-hydroactive (HP-HA) model that integrates leaf hydraulics with the biokinetics of guard cell-autonomous ABA regulation and plasma membrane-mediated osmoregulation. We demonstrate that this framework predicts accurate, genotype-specific stomatal regulation across wildtype, ABA-insensitive mutant ( ost1-3 ), and ABA-synthesis mutant ( aao3-2 ) in Arabidopsis thaliana ( At ) and that non-linear feedbacks in ABA autoregulation can drive both stomatal oscillations and hysteresis. This work unifies genetic, signaling, and membrane processes with leaf-scale physiological dynamics, providing a new predictive foundation for understanding and modulating plant management of water use and water stress.

Why it matches plant phenotyping methods葉の水理とABA制御を統合した予測モデルを開発し、遺伝子型別の気孔コンダクタンス制御を検証しており、植物生理表現型の取得・予測手法が中心である。

abstractHere, we introduce a coupled hydropassive-hydroactive (HP-HA) model that integrates leaf hydraulics with the biokinetics of guard cell-autonomous ABA regulation and plasma membrane-mediated osmoregulation.

Advanced Tissue Imprinting With Pneumatic Press for Mass Spectrometry Imaging of Plant Tissues →

ABSTRACT Sample preparation is an important first step to obtain high quality mass spectrometry imaging (MSI) data. Preparing plant tissues is especially challenging for MSI of thin tissues along the lateral dimensions. The unique challenges involved with plant tissues, such as fragile cell walls, hydrophobic barriers, and specific tissue structures, often lead to inefficiency and difficulties in sample preparation. Imprinting plant tissues onto porous polytetrafluoroethylene (pPTFE) sheet has been widely used to extract internal metabolites in leaves and petals while keeping spatial resolution for MSI. However, pressure applications were typically made manually using a vise or pliers leading to low reproducibility and resolution in MS images. In this study, we introduce a home‐built pneumatic press (PNP) that has been designed to precisely control the pressure application parameters during imprinting. To evaluate the performance of the new device, Lemna minor fronds, Arabidopsis thaliana , and Bacopa monnieri leaves were imprinted onto the pPTFE with PNP, vise, or pliers, and matrix‐assisted laser desorption/ionization (MALDI) MSI was obtained on the imprints. The PNP showed dramatic improvements in reproducibility and image quality compared to manual pressure application tools.

Why it matches plant phenotyping methods植物組織の空間的な代謝物情報を再現性よく取得するための空気圧式インプリンティング装置を開発し、手動法と性能比較している。植物表現型取得に関わる試料調製・イメージング手法が中心である。

abstractIn this study, we introduce a home‐built pneumatic press (PNP) that has been designed to precisely control the pressure application parameters during imprinting.

Spatial ploidy inference using quantitative imaging. →

Polyploidy (whole-genome duplication) is a common yet under-surveyed property of tissues across multicellular organisms. Polyploidy plays a critical role during tissue development, following acute stress, and during disease progression. Common methods to reveal polyploidy involve either destroying tissue architecture by cell isolation or tedious identification of individual nuclei in intact tissue. Therefore, there is a critical need for rapid and high-throughput ploidy quantification using images of nuclei in intact tissues. Here, we present iSPy (inferring Spatial Ploidy), an unsupervised learning pipeline that is designed to create a spatial map of nuclear ploidy across a tissue of interest. We demonstrate the use of iSPy in Arabidopsis, Drosophila, and human tissue. iSPy can be adapted for a variety of tissue preparations, including whole mount and sectioned. This high-throughput pipeline will facilitate rapid and sensitive identification of nuclear ploidy in diverse biological contexts and organisms.

Why it matches plant phenotyping methodsiSPyは画像から組織内の核倍数性を空間的・高スループットに推定する教師なし学習パイプラインであり、Arabidopsisで実証されている。植物の状態を抽出する計算フェノタイピング手法が中心である。

abstractHere, we present iSPy (inferring Spatial Ploidy), an unsupervised learning pipeline that is designed to create a spatial map of nuclear ploidy across a tissue of interest.

Engineered glycoside hydrolases as fluorescent probes reveal the spatial distribution of the pectic polysaccharide rhamnogalacturonan II in plant cell walls →

Abstract Plant cell walls are dynamic composites whose architecture determines growth, mechanics, and environmental resilience. Efforts to link pectin structure to function have been limited by the lack of molecular probes with sufficient specificity, a gap that becomes even more pronounced for the intricately branched rhamnogalacturonon-II (RG-II) subclass. Here we report the first fluorescent probes with defined specificity to RG-II, engineered from catalytic site mutants of Bacteroides thetaiotaomicron glycoside hydrolases BT1010 and BT0996. These enzyme-derived probes bind RG-II monomer with high affinity, discriminate against dimeric forms, and localize to cell corners and junctions in Arabidopsis thaliana stems, consistent with RG-II’s unique ability among wall polysaccharides to form borate-mediated, covalent crosslinkages between molecules. Application of these probes revealed spatial partitioning distinct from the homogalacturonan (HG)- and rhamnogalacturonan I (RG-I)-enriched middle lamella, highlighting functional specialization among pectic domains, with RG-II reinforcing cell junctions while HG and RG-I mediate wall flexibility. Our work establishes a generalizable framework for transforming CAZymes into high-precision imaging reagents, enabling molecular-level visualization of structurally complex polysaccharides in the cell wall.

Why it matches plant phenotyping methodsRG-IIを特異的に可視化する蛍光プローブを開発し、植物細胞壁内の空間分布という植物状態を画像で測定する手法を示しているため、フェノタイピング手法が中心的です。

abstractHere we report the first fluorescent probes with defined specificity to RG-II

Rapid acquisition of ionomic and morphological data from plant seeds through fast X-ray fluorescence microscopy and computer vision. →

Plant seeds are one of the most important food sources for humans. As a result, seed morphology and the concentrations of essential and toxic elements in seeds have important implications not only for seed yield and quality, but also for human health. To identify natural variation in the accumulation of various elements in seeds and in seed morphology, high-throughput phenotyping methods are needed. Here, we employed X-ray fluorescence microscopy (μ-XRF) as a method for rapid and high-throughput phenotyping of seed libraries and developed a computer vision-based algorithmic workflow to automatically the extraction of elemental and morphological data from single seeds. This workflow enables rapid segmentation of individual seeds from a genome-wide association study (GWAS) panel with 1163 A. thaliana accessions, and facilitates the extraction of elemental and morphological traits at the individual seed level from the μ-XRF image. A total of 7 and 10 loci, respectively associated with the morphology and elemental concentration of A. thaliana seeds, were identified. The high-throughput and nondestructive method for automated phenotyping of plant seed libraries developed in this study provides a tool for investigating natural genetic variation controlling the seed mineral accumulation and seed morphogenesis.

Why it matches plant phenotyping methods種子の元素濃度・形態をμ-XRFとコンピュータビジョンで高速・自動取得する手法を開発しており、植物表現型取得が研究の中心である。

abstracthigh-throughput phenotyping methods are needed

APTES: a high-throughput deep learning-based Arabidopsis phenotypic trait estimation system for individual leaves and siliques. →

High-throughput phenotyping of growth kinetics and organ size in the model plant Arabidopsis thaliana requires rapid and precise methods for trait estimation. To address this need, we developed the Arabidopsis Phenotypic Trait Estimation System, APTES, an open-access, high-throughput program that uses computer vision and deep learning to extract 64 leaf traits and 64 silique traits from photographs. The enhanced segmentation model Cascade Mask Region-based Convolutional Neural Network (Mask R-CNN) achieved precision (measure of positive prediction accuracy), recall (sensitivity in detection), and F1 score values (harmonic mean of precision and recall) of 0.965, 0.958, and 0.961, respectively, for individual leaf segmentation. These metrics demonstrated a consistent improvement of approximately 1 percentage point over the baseline model. For silique segmentation, our enhanced DetectoRS model for silique segmentation attained precision, recall, and F1 scores of 0.954, 0.930, and 0.942, respectively. Notably, precision increased by 1%, while the F1 score improved by 2 percentage points. Trait parameters were automatically calculated with coefficient of determination values for leaf and silique traits ranging from 0.776 to 0.976 and mean absolute percentage error values from 1.89% to 7.90%. We phenotyped 166 Arabidopsis accessions, using APTES, and subjected the resulting values to a genome-wide association study (GWAS), revealing 1,042 single-nucleotide polymorphisms (SNPs) as being significantly associated with 18 leaf and silique traits, and one significant SNP on chromosome 3 linked to silique number. Furthermore, we validated APTES across other public Arabidopsis databases and other plant species, with segmentation results demonstrating its applicability across diverse datasets. In conclusion, APTES is a valuable automated tool for leaf and silique segmentation and trait estimation, which should offer benefits to the broader plant science community. Supplementary information The online version contains supplementary material available at 10.1007/s42994-025-00239-y.

Why it matches plant phenotyping methods植物の葉・莢の形質を画像から抽出する深層学習システムを開発し、性能検証・他データセットでの妥当性確認まで行っており、フェノタイピング手法が研究の中心である。

abstractwe developed the Arabidopsis Phenotypic Trait Estimation System, APTES, an open-access, high-throughput program that uses computer vision and deep learning to extract 64 leaf traits and 64 silique traits from photographs.

DynG: a dynamic scaling factor for thermographic stomatal conductance estimation under changing environmental conditions. →

Summary Thermal imaging is a key plant phenotyping and monitoring technique but faces major bottlenecks in accurately and efficiently inferring stomatal conductance (gsw) from leaf temperature. The conductance index (Ig) was previously proposed to estimate gsw from thermography by linking temperature differences between real and artificial leaves (ALs) based on the leaf energy balance. However, Ig is highly sensitive to environmental fluctuations, hampering interpretation and reducing reproducibility. We developed a simple and novel correction factor (named DynG) for Ig that accounts for environmental fluctuations when scaling Ig to gsw. This was achieved by capturing temperature variations in a set of ALs with a range of known constant pore conductances. This approach provided the Ig–conductance relationship, using ALs as a reference, to infer gsw of real leaves from their measured Ig. In fluctuating environments, gsw estimated using DynG showed greater accuracy and stability than gsw calculated from Ig alone, and was in good agreement with gsw determined using lysimetric and gas exchange methods. DynG's power was further showcased in distinguishing gsw of Arabidopsis genotypes differing in stomatal traits (Col‐0, epf1epf2, and EPF2OE). We conclude that Ig corrected with DynG can reliably estimate gsw in fluctuating environments without complex modeling, opening new avenues for gsw phenotyping and monitoring.

Why it matches plant phenotyping methods熱画像から気孔コンダクタンスを推定する補正係数を開発し、変動環境下で既存法と比較検証した、植物フェノタイピング手法の中心的研究である。

abstractWe developed a simple and novel correction factor (named DynG) for Ig that accounts for environmental fluctuations when scaling Ig to gsw.

Computer vision‐based recognition and distinction of Arabidopsis thaliana ecotypes using supervised deep learning models →

Abstract Image‐based plant phenotyping has diverse applications, ranging from providing quantitative traits for genetic breeding to enhancing management practices for indoor and outdoor production systems. Misidentification of cell lines or ecotypes/varieties is a major problem across all biological research disciplines. With the 1000 Arabidopsis Genome Project facilitating the use of various ecotypes, it is crucial to verify the identity of ecotypes in discovery‐based genetic screens involving hundreds of ecotypes. To address this issue, an RGB image analysis pipeline was established for the accurate recognition of different Arabidopsis thaliana ecotypes. In the developed pipeline, the most crucial aspects for accurately capturing traits and training deep learning models were identified as follows: (i) assessment of data complexity using spatial‐temporal features of the RGB spectrum and data entropy, the latter defined as the variability within the dataset; (ii) data redefinition in instances of high data complexity; and (iii) data partitioning based on extracted morphological similarity among ecotype replicates. The pipeline includes several supervised deep learning models integrated into an auto‐optimization subsystem. Extensive hyperparameter tuning was performed to identify the best‐performing models for single‐image and image‐sequence ecotype classification. Two external datasets were evaluated to demonstrate the robustness of the pipeline, regardless of how they were collected. A graphical user interface is provided to prepare these images for input into the pipeline in cases of extreme variability. The pipeline can automatically verify ecotypes in large‐scale studies and extract traits for further analysis and correlation, as needed, using datasets from a variety of sources.

Why it matches plant phenotyping methodsRGB画像から植物形態情報を抽出し、深層学習による分類・検証を行うパイプライン自体が中心的な貢献であり、外部データセットで頑健性も評価しているため。

abstractan RGB image analysis pipeline was established for the accurate recognition of different Arabidopsis thaliana ecotypes.

The Tonoplast Topology Index - a new metric for describing vacuole organization →

Background The plant vacuole arises by orchestrated interplay of membrane trafficking, cytoskeletal rearrangements and a variety of signalling pathways. In the root, the characteristic large central vacuole develops by endomembrane reorganization occurring mainly in the transition zone. The vacuole’s bounding membrane - the tonoplast - can be visualized in vivo using fluorescent protein markers, allowing for quantitative analysis of confocal microscopy images. Tonoplast organization can thus serve as a sensitive indicator of changes to any of the processes involved in vacuole biogenesis. The Vacuolar Morphology Index (VMI) is widely accepted as a quantitative measure of vacuole structure. However, this metric has two drawbacks - it only reflects the size of the largest vacuolar compartment (missing therefore possible differences in the organization of smaller compartments), and its determination is labor intensive, limiting its use on large datasets. Results We developed an alternative metric for describing vacuole organization, named the Tonoplast Topology Index (TTI), which overcomes the above-mentioned shortcomings of the VMI. We compared the performance of our protocol with VMI on a simulated dataset and on real data. To validate the methods’ performance, we used it to confirm the previously reported differences in vacuole shape and size between Arabidopsis thaliana roots grown on the surface of an agar medium compared to those embedded inside the agar. Both VMI and TTI could efficiently detect the relatively subtle changes in vacuole organization depending on the position of the root in the agar, and provided correlated results. However, only TTI produced data with close to normal value distribution, simplifying subsequent statistical evaluation. Conclusions We present the protocol for TTI determination as a two-stage semi-automated procedure involving microscopic image analysis employing an ImageJ macro and subsequent processing of numeric data in the Jupyter Notebook environment, together with benchmarking image data. Since this implementation is freeware-based, platform-independent and (relatively) user-friendly, we hope it will find its use as a high throughput, added value alternative to the VMI metric.

Why it matches plant phenotyping methods植物の液胞構造を定量化する新規指標と半自動画像解析プロトコルを開発し、既存指標との比較・実データおよびシミュレーションによる検証、ベンチマークデータを提示しており、表現型取得・抽出法が中心である。

abstractWe developed an alternative metric for describing vacuole organization, named the Tonoplast Topology Index (TTI), which overcomes the above-mentioned shortcomings of the VMI.

Time course measurements of leaf elevation angles during shade avoidance response in Arabidopsis thaliana using Raspberry Pi computers and computer vision technique. →

Shade avoidance response in plants includes a higher leaf elevation angle. A cost-effective and noninvasive high throughput image analysis technique was used to measure the dynamics of leaf elevation angles during shade avoidance response in Arabidopsis . Time-lapse images were taken from the top and the side of a plant using Raspberry Pi computers. The leaf elevation index for each plant is determined from the plant dimensions measured by an image analysis software package PlantCV . This method was used to monitor the dynamics of changing leaf elevation angles in wild-type plants and in shade avoidance mutants pif4-2pif5-3 and pif7-2 plants.

Why it matches plant phenotyping methodsRaspberry Piと画像解析を用いて葉の仰角を定量化する高スループット手法が研究の中心であり、植物形質の時系列取得に実質的に適用されている。

abstractA cost-effective and noninvasive high throughput image analysis technique was used to measure the dynamics of leaf elevation angles during shade avoidance response in Arabidopsis .

Growth Cost and Transport Efficiency Tradeoffs Define Root System Optimization Across Varying Developmental Stages and Environments in Arabidopsis →

ABSTRACT Root system architecture (RSA) is central to plant adaptation and fitness, yet the design principles and regulatory mechanisms connecting RSA to environmental adaptation are not well understood. We developed Ariadne, a semi-automated software for quantifying cost-efficiency tradeoffs of RSA by mapping root networks onto a Pareto-optimality framework, which describes the balance between resource transport efficiency and construction cost. Applying Ariadne to Arabidopsis thaliana , we found that root architectures consistently assume Pareto-optimal forms across developmental stages, genotypes, and environmental conditions. Using the Discovery Engine, an engine that combines machine learning together with interpretability techniques, we found developmental stage, the hy5/chl1-5 genotype, and manganese availability as important determinants of the cost-efficiency tradeoff, with manganese exerting a unique influence not observed for other nutrients. These results reveal that RSA plasticity is genetically constrained to cost-efficiency optimal configurations and that developmental and environmental factors shift RSA on the pareto front, with manganese acting as a strong modulator of the transport efficiency and construction cost balance.

Why it matches plant phenotyping methodsRSAのコスト効率トレードオフを定量化する半自動ソフトウェアを開発し、植物形態形質の解析に適用しており、表現型取得・抽出手法が研究の中心である。

abstractWe developed Ariadne, a semi-automated software for quantifying cost-efficiency tradeoffs of RSA by mapping root networks onto a Pareto-optimality framework

Genome-wide association study reveals influence of cell-specific gene networks on Soybean root system architecture →

Abstract Root system architecture (RSA), the three-dimensional arrangement of roots in soil, is a critical determinant of plant productivity, resource use efficiency, and resilience to environmental stress. Despite its agronomic importance, RSA remains a largely untapped breeding target due to historical technical barriers in root phenotyping. We present RADICYL (Root Architecture 3D Cylinder), a scalable, non-invasive, gel-based platform enabling high-throughput, high-resolution quantification of 15 RSA traits in intact root systems. Applying RADICYL to a genetically diverse panel of 371 soybean accessions, we combined 3D phenotyping with genome-wide association studies (GWAS), single-nucleus RNA sequencing (snRNA-seq), and gene co-expression network (GCN) analysis to identify RCE1 and NPR3 as central regulators of RSA, suggesting auxin and salicylic acid-mediated signaling impacts RSA in specific root tissues. Functional validation in Arabidopsis mutants revealed conserved effects on root width and lateral root development. Our findings position the endodermis and metaphloem as key regulatory cell types and demonstrate how multi-omic frameworks can accelerate the discovery of functional genes underlying complex traits. This study establishes a foundation for cell-type-targeted genome editing and climate-smart crop engineering, offering actionable genetic targets to optimize root systems for improved nutrient acquisition, drought resilience, and deep carbon sequestration. By bridging genotype, cellular context, and phenotype, this work redefines RSA as a tractable and transformative trait for the future of crop improvement.

Why it matches plant phenotyping methodsRADICYLという根系構造を定量化する高スループット3Dフェノタイピング基盤の開発・適用が研究の中心であり、15形質を測定している。

abstractWe present RADICYL (Root Architecture 3D Cylinder), a scalable, non-invasive, gel-based platform enabling high-throughput, high-resolution quantification of 15 RSA traits in intact root systems.

CalciumInsights: An Open-Source, Tissue-Agnostic Graphical Interface for High-Quality Analysis of Calcium Signals →

Fluctuations and propagation of cytosolic calcium levels at both the cellular and tissue levels show complex patterns, referred to as calcium signatures, that regulate growth, organ development, damage responses, and survival. The quantitative analysis of calcium signatures at the cellular level is essential for identifying unique patterns that coordinate biological processes. However, a versatile framework applicable to multiple tissue types, allowing researchers to compare, measure, and validate diverse responses and recognize conserved patterns across model organisms, is missing. Here, we present a post-processing tool, CalciumInsights, which leverages the R packages Shiny and Golem. This tool has a graphical user interface and does not require software programming experience to perform calcium signal analysis. The open-source software has a modular framework with standardized functionalities that can be tailored for various research approaches. CalciumInsights provides descriptive statistical analysis through various metrics extracted from dynamic calcium transients and oscillations, such as peak amplitude, area under the curve, frequency, among others. The tool was evaluated with fluorescence imaging data from three model organisms: Danio rerio , Arabidopsis thaliana , and Drosophila melanogaster , demonstrating its ability to analyze diverse biological responses and models. Finally, the open-source nature of CalciumInsights enables community-driven improvements and developments for enabling new applications. Author Summary This manuscript introduces CalciumInsights, an open-source tool for calcium signature analysis. Designed to be a versatile tool that works with various tissue types and biological systems, CalciumInsights has an easy-to-use graphical user interface. Our program simplifies metrics extraction while maintaining the quality of the analysis by integrating several algorithms. CalciumInsights stands out for its user-friendliness, ease of use, and robust data exploration features, such as tunable filters for improved accuracy. These features promote inclusivity and lower barriers to scientific research by making calcium signature analysis accessible to users of all programming skill levels.

Why it matches plant phenotyping methods植物の蛍光イメージングからカルシウム動態という生理状態を抽出・定量するオープンソース解析ツールが中心であり、植物を含む複数生物種のデータで評価されている。

abstractHere, we present a post-processing tool, CalciumInsights, which leverages the R packages Shiny and Golem.

Seed-to-plant-tracking: automated phenotyping of seeds and corresponding plants of Arabidopsis →

Plants adapt seed traits in response to different environmental triggers, supporting the survival of the next generation. To elucidate the mechanistic understanding of such adaptations it is important to characterize the distributions of seed traits by phenotyping seeds on an individual scale and to correlate these traits with corresponding plant properties. Here we introduce a seed-to-plant-tracking pipeline which enables automated handling and high precision phenotyping of Arabidopsis seeds as well as germination detection and early growth quantification of emerging plants. It includes previously published measurement platforms ( pheno Seeder, Growscreen), which were improved for very small seeds. We demonstrate the performance of the pipeline by comparing seeds from two consecutive generations of elevated temperature during flowering with control seeds. Relative standard deviation of repeated seed mass measurements was reduced to 0.2%. We identified an increase in seed mass, volume, length, width, height, and germination time as well as a darkening of the seeds under the treatment. A correlation analysis revealed relationships between seed and plant traits, e.g., a highly significant negative correlation between seed brightness and germination time, and a positive correlation between seed mass and early growth rate, but no correlation between time of emergence and morphometric seed traits (e.g., mass, volume). Thus, the seed-to-plant tracking provides the basis for investigating the mechanism of seed and plant trait variation and transgenerational inheritance.

Why it matches plant phenotyping methods種子から植物までを追跡し、種子形質の高精度自動計測、発芽検出、初期成長定量を行うパイプラインを開発・改良しており、表現型取得法が研究の中心である。

abstractHere we introduce a seed-to-plant-tracking pipeline which enables automated handling and high precision phenotyping of Arabidopsis seeds as well as germination detection and early growth quantification of emerging plants.

ChronoRoot 2.0: An Open AI-Powered Platform for 2D Temporal Plant Phenotyping →

Plant developmental plasticity, particularly in root system architecture, is fundamental to understanding adaptability and agricultural sustainability. ChronoRoot 2.0 builds upon established low-cost hardware while significantly enhancing software capabilities and usability. The system employs nnUNet architecture for multi-class segmentation, demonstrating significant accuracy improvements while simultaneously tracking six distinct plant structures encompassing root, shoot, and seed components: main root, lateral roots, seed, hypocotyl, leaves, and petiole. This architecture enables easy retraining and incorporation of additional training data without requiring machine learning expertise. The platform introduces dual specialized graphical interfaces: a Standard Interface for detailed architectural analysis with novel gravitropic response parameters, and a Screening Interface enabling high-throughput analysis of multiple plants through automated tracking. Functional Principal Component Analysis integration enables discovery of novel phenotypic parameters through temporal pattern comparison. We demonstrate multi-species analysis, with Arabidopsis thaliana and Solanum lycopersicum, both morphologically distinct plant species. Three use cases in Arabidopsis thaliana and validation with tomato seedlings demonstrate enhanced capabilities: circadian growth pattern characterization, gravitropic response analysis in transgenic plants, and high-throughput etiolation screening across multiple genotypes.ChronoRoot 2.0 maintains the low-cost, modular hardware advantages of its predecessor while dramatically improving accessibility through intuitive graphical interfaces and expanded analytical capabilities. The open-source platform makes sophisticated temporal plant phenotyping more accessible to researchers without computational expertise.

Why it matches plant phenotyping methods植物の時系列画像から根・地上部・種子などの形態形質を抽出・追跡するオープンなAI基盤を開発し、精度向上、再学習、GUI、高スループット解析、検証まで扱っており、フェノタイピング手法が研究の中心です。

titleChronoRoot 2.0: An Open AI-Powered Platform for 2D Temporal Plant Phenotyping

Predicting plant trait dynamics from genetic markers →

Molecular and physiological changes across crop developmental stages shape the plant phenome and render its prediction from genetic markers challenging. Here we present dynamicGP, an efficient computational approach that combines genomic prediction with dynamic mode decomposition to characterize the temporal changes and to predict genotype-specific dynamics for multiple morphometric, geometric and colourimetric traits scored by high-throughput phenotyping. Using genetic markers and data from high-throughput phenotyping of a maize multiparent advanced generation inter-cross population and an Arabidopsis thaliana diversity panel, we show that dynamicGP outperforms a baseline genomic prediction approach for the multiple traits. We demonstrate that the developmental dynamics of traits whose heritability varies less over time can be predicted with higher accuracy. The approach paves the way for interrogating and integrating the dynamical interactions between genotype and environment over plant development to improve the prediction accuracy of agronomically relevant traits.

Why it matches plant phenotyping methods遺伝マーカーと高スループット表現型データを統合し、植物形態・幾何・色彩形質の時系列を予測する計算手法dynamicGPが研究の中心であるため。

abstractHere we present dynamicGP, an efficient computational approach that combines genomic prediction with dynamic mode decomposition to characterize the temporal changes and to predict genotype-specific dynamics for multiple morphometric, geometric and colourimetric traits scored by high-throughput phenotyping.

A Detailed Guide to Recording and Analyzing Arabidopsis thaliana Leaf Surface Potential Dynamics Elicited by Mechanical Wounding. →

Recordings of electric potential changes on plant surfaces have been utilized to identify the components and mechanisms involved in the formation and transmission of systemic signals elicited by stimuli such as herbivory, wounding, or burning. The recorded responses, commonly referred to as slow wave or variation potentials, exhibit striking variability in their waveform. The extent to which this variability is due to differences in experimental procedures or plant biological variability remains unclear. Here, we provide a detailed and robust protocol refined from years of experience in conducting leaf surface potential recordings of Arabidopsis thaliana in response to mechanical wounding. This protocol serves as a comprehensive tutorial covering plant growth, procedures for reproducible mechanical wounding, critical aspects of electrophysiological recordings, and statistical analysis of surface potential recordings. It particularly emphasizes the construction and maintenance of electrodes, placement of the reference or ground electrode, mechanisms for wounding, and data analysis. This protocol aims to promote and facilitate the adoption, standardization, and interoperability of plant surface potential recordings among research groups, thereby increasing the reproducibility and comparability of data within the field. Key features • Recording electric potential changes on the petiole of 5-week-old Arabidopsis plants using noninvasive surface electrodes, improving the wounding procedure, reproducibility, and data processing from [1]. • Genotype-independent method for phenotyping, including parallel recordings from multiple plants. • Guidelines for plant growth conditions, unambiguous leaf assignment by order of emergence, and detailed instructions for electrode fabrication and maintenance. • Instructions for constructing devices for standardized, reproducible mechanical wounding along with a custom script for unbiased and semi-automated data analysis.

Why it matches plant phenotyping methods植物の表面電位を再現性高く記録・解析する電気生理学的フェノタイピング手法の詳細プロトコルであり、電極、標準化創傷、データ解析、再現性・相互運用性が中心的に扱われている。

abstractHere, we provide a detailed and robust protocol refined from years of experience in conducting leaf surface potential recordings of Arabidopsis thaliana in response to mechanical wounding.

A 3D morpho-space of sepal geometry reveals the importance of organ curvature. →

How robust three-dimension (3D) organ shape emerges during morphogenesis is a fundamental question in biology. Addressing this question requires a comprehensive quantification of organ geometry in 3D. To tackle these issues, we considered the sepal of Arabidopsis as a model. Using a unique pipeline allowing to recover 3D sepal morphology, we analysed fifteen mutants affected in different pathways. The results of a Principal Component Analysis reveal sepal curvature as an important parameter accounting for variations in sepal morphology within genotypes. Unexpectedly, despite genetic homogeneity of the wild-type plants and reproducible culture conditions, we found a significant level of variability in sepal morphology. Our data also show that sepal shape from wild-type plants is more robust (less variable) than sepal size, hinting to a possible selective pressure on shape parameters.

Why it matches plant phenotyping methods3D萼片形態を復元・定量する独自パイプラインが研究の中心であり、器官形状・曲率という植物表現型を解析している。

abstractUsing a unique pipeline allowing to recover 3D sepal morphology, we analysed fifteen mutants affected in different pathways.

Spatial ploidy inference using quantitative imaging →

Polyploidy (whole-genome multiplication) is a common yet under-surveyed property of tissues across multicellular organisms. Polyploidy plays a critical role during tissue development, following acute stress, and during disease progression. Common methods to reveal polyploidy involve either destroying tissue architecture by cell isolation or by tedious identification of individual nuclei in intact tissue. Therefore, there is a critical need for rapid and high-throughput ploidy quantification using images of nuclei in intact tissues. Here, we present iSPy (Inferring Spatial Ploidy), a new unsupervised learning pipeline that is designed to create a spatial map of nuclear ploidy across a tissue of interest. We demonstrate the use of iSPy in Arabidopsis, Drosophila, and human tissue. iSPy can be adapted for a variety of tissue preparations, including whole mount and sectioned. This high-throughput pipeline will facilitate rapid and sensitive identification of nuclear ploidy in diverse biological contexts and organisms.

Why it matches plant phenotyping methodsArabidopsisを含む組織の核倍数性を画像から空間的に推定する新規計算パイプラインを開発しており、植物の状態計測手法が研究の中心である。

abstractwe present iSPy (Inferring Spatial Ploidy), a new unsupervised learning pipeline that is designed to create a spatial map of nuclear ploidy across a tissue of interest.

Expansion microscopy reveals thylakoid organisation alterations due to genetic mutations and far-red light acclimation. →

The thylakoid membrane is the site of the light-dependent reactions of photosynthesis. It is a continuous membrane, folded into grana stacks and the interconnecting stroma lamellae. The CURVATURE THYLAKOID1 (CURT1) protein family is involved in the folding of the membrane into the grana stacks. The thylakoid membrane remodels its architecture in response to light conditions, but its 3D organisation and dynamics remain incompletely understood. To resolve these details, an imaging technique is needed that provides high-resolution 3D images in a high-throughput manner. Recently, we have used expansion microscopy, a technique that meets these criteria, to visualise the thylakoid membrane isolated from spinach. Here, we show that this protocol can also be used to visualise enveloped spinach chloroplasts. Additionally, we present an improved protocol for resolving the thylakoid structure of Arabidopsis thaliana. Using this protocol, we show the changes in thylakoid architecture in response to long-term far-red light acclimation and due to knocking out CURT1A. We show that far-red light acclimation results in higher grana stacks that are packed closer together. In addition, the distance between stroma lamellae, which are wrapped around the grana, decreases. In the curt1a mutant, grana have an increased diameter and height, and the distance between grana is increased. Interestingly, in this mutant, the stroma lamellae occasionally approach the grana stacks from the top. These observations show the potential of expansion microscopy to study the thylakoid membrane architecture.

Why it matches plant phenotyping methods植物のチラコイド膜構造を高解像度3D画像で取得する拡大顕微鏡法の改良・適用が中心であり、膜構造という植物形態形質を測定しているため。

abstractTo resolve these details, an imaging technique is needed that provides high-resolution 3D images in a high-throughput manner.

The effectiveness of large language models with RAG for auto-annotating trait and phenotype descriptions. →

Ontologies are highly prevalent in biology and medicine and are always evolving. Annotating biological text, such as observed phenotype descriptions, with ontology terms is a challenging and tedious task. The process of annotation requires a contextual understanding of the input text and of the ontological terms available. While text-mining tools are available to assist, they are largely based on directly matching words and phrases and so lack understanding of the meaning of the query item and of the ontology term labels. Large Language Models (LLMs), however, excel at tasks that require semantic understanding of input text and therefore may provide an improvement for the auto-annotation of text with ontological terms. Here we describe a series of workflows incorporating OpenAI GPT's capabilities to annotate Arabidopsis thaliana and forest tree phenotypic observations with ontology terms, aiming for results that resemble manually curated annotations. These workflows make use of an LLM to intelligently parse phenotypes into short concepts, followed by finding appropriate ontology terms via embedding vector similarity or via Retrieval-Augmented Generation (RAG). The RAG model is a state-of-the-art approach that augments conversational prompts to the LLM with context-specific data to empower it beyond its pre-trained parameter space. We show that the RAG produces the most accurate automated annotations that are often highly similar or identical to expert-curated annotations.

Why it matches plant phenotyping methods植物の表現型観察記述をオントロジー語に自動アノテーションするLLM/RAGワークフローの開発・精度評価が中心であり、再利用可能な計算ツールとして植物表現型データを処理する。

abstractHere we describe a series of workflows incorporating OpenAI GPT's capabilities to annotate Arabidopsis thaliana and forest tree phenotypic observations with ontology terms

Order among chaos: High throughput MYCroplanters can distinguish interacting drivers of host infection in a highly stochastic system. →

The likelihood that a host will be susceptible to infection is influenced by the interaction of diverse biotic and abiotic factors. As a result, substantial experimental replication and scalability are required to identify the contributions of and interactions between the host, the environment, and biotic factors such as the microbiome. For example, pathogen infection success is known to vary by host genotype, bacterial strain identity and dose, and pathogen dose. Elucidating the interactions between these factors in vivo has been challenging because testing combinations of these variables quickly becomes experimentally intractable. Here, we describe a novel high throughput plant growth system (MYCroplanters) to test how multiple host, non-pathogenic bacteria, and pathogen variables predict host health. Using an Arabidopsis-Pseudomonas host-microbe model, we found that host genotype and bacterial strain order of arrival predict host susceptibility to infection, but pathogen and non-pathogenic bacterial dose can overwhelm these effects. Host susceptibility to infection is therefore driven by complex interactions between multiple factors that can both mask and compensate for each other. However, regardless of host or inoculation conditions, the ratio of pathogen to non-pathogen emerged as a consistent correlate of disease. Our results demonstrate that high-throughput tools like MYCroplanters can isolate interacting drivers of host susceptibility to disease. Increasing the scale at which we can screen drivers of disease, such as microbiome community structure, will facilitate both disease predictions and treatments for medicine and agricultural applications.

Why it matches plant phenotyping methodsMYCroplantersという植物の高スループット生育・感染評価システム自体を開発・記述し、宿主の感染感受性（病害状態）を測定する方法が研究の中心であるため。

abstractHere, we describe a novel high throughput plant growth system (MYCroplanters) to test how multiple host, non-pathogenic bacteria, and pathogen variables predict host health.

Non-Invasive, Bioluminescence-Based Visualisation and Quantification of Bacterial Infections in Arabidopsis Over Time. →

Plant-pathogenic bacteria colonise their hosts using various strategies, exploiting both natural openings and wounds in leaves and roots. The vascular pathogen Xanthomonas campestris pv. campestris (Xcc) enters its host through hydathodes, organs at the leaf margin involved in guttation. Subsequently, Xcc breaches the hydathode-xylem barrier and progresses into the xylem vessels causing systemic disease. To elucidate the mechanisms that underpin the different stages of an Xcc infection, a need exists to image bacterial progression in planta in a non-invasive manner. Here, we describe a phenotyping setup and Python image analysis pipeline for capturing 16 independent Xcc infections in Arabidopsis thaliana plants in parallel over time. The setup combines an RGB camera for imaging disease symptoms and an ultrasensitive CCD camera for monitoring bacterial progression inside leaves using bioluminescence. The method reliably quantified bacterial growth in planta for two bacterial species, that is, vascular Xcc and the mesophyll pathogen Pseudomonas syringae pv. tomato (Pst). The camera resolution allowed Xcc imaging already in the hydathodes, yielding reproducible data for the first stages prior to the systemic infection. Data obtained through the image analysis pipeline was robust and validated findings from other bioluminescence imaging methods, while requiring fewer samples. Moreover, bioluminescence was reliably detected within 5 min, offering a significant time advantage over our previously reported method with light-sensitive films. Thus, this method is suitable to quantify the resistance level of a large number of Arabidopsis thaliana accessions and mutant lines to different bacterial strains in a non-invasive manner for phenotypic screenings.

Why it matches plant phenotyping methods植物感染を非侵襲的に画像化・定量するフェノタイピング装置とPython解析パイプラインを開発し、複数の細菌感染で検証しているため、方法が中心的です。

abstractHere, we describe a phenotyping setup and Python image analysis pipeline for capturing 16 independent Xcc infections in Arabidopsis thaliana plants in parallel over time.

DFMA: an improved DeepLabv3+ based on FasterNet, multi-receptive field, and attention mechanism for high-throughput phenotyping of seedlings. →

With the rapid advancement of plant phenotyping research, understanding plant genetic information and growth trends has become crucial. Measuring seedling length is a key criterion for assessing seed viability, but traditional ruler-based methods are time-consuming and labor-intensive. To address these limitations, we propose an efficient deep learning approach to enhance plant seedling phenotyping analysis. We improved the DeepLabv3+ model, naming it DFMA, and introduced a novel ASPP structure, PSPA-ASPP. On our self-constructed rice seedling dataset, the model achieved a mean Intersection over Union (mIoU) of 81.72%. On publicly available datasets, including Arabidopsis thaliana, Brachypodium distachyon, and Sinapis alba, detection scores reached 87.69%, 91.07%, and 66.44%, respectively, outperforming existing models. The model generates detailed segmentation masks, capturing structures such as the embryonic shoot, axis, and root, while a seedling length measurement algorithm provides precise parameters for component development. This approach offers a comprehensive, automated solution, improving phenotyping analysis efficiency and addressing the challenges of traditional methods.

Why it matches plant phenotyping methods幼苗画像から器官を分割し、幼苗長を自動測定する深層学習ベースのフェノタイピング手法を開発・検証しており、方法が中心的です。

abstractwe propose an efficient deep learning approach to enhance plant seedling phenotyping analysis.

Super-resolution expansion microscopy in plant roots →

Abstract Super-resolution methods provide far better spatial resolution than the optical diffraction limit of about half the wavelength of light (∼200–300 nm). Nevertheless, they have yet to attain widespread use in plants, largely due to plants' challenging optical properties. Expansion microscopy (ExM) improves effective resolution by isotropically increasing the physical distances between sample structures while preserving relative spatial arrangements and clearing the sample. However, its application to plants has been hindered by the rigid, mechanically cohesive structure of plant tissues. Here, we report on whole-mount ExM of thale cress (Arabidopsis thaliana) root tissues (PlantEx), achieving a 4-fold resolution increase over conventional microscopy. Our results highlight the microtubule cytoskeleton organization and interaction between molecularly defined cellular constituents. Combining PlantEx with stimulated emission depletion microscopy, we increase nanoscale resolution and visualize the complex organization of subcellular organelles from intact tissues by example of the densely packed COPI-coated vesicles associated with the Golgi apparatus and put these into a cellular structural context. Our results show that ExM can be applied to increase effective imaging resolution in Arabidopsis root specimens.

Why it matches plant phenotyping methods植物組織に適用可能な超解像イメージング手法を開発し、Arabidopsis根で解像度向上を実証しており、画像取得法が研究の中心である。

abstractHere, we report on whole-mount ExM of thale cress (Arabidopsis thaliana) root tissues (PlantEx), achieving a 4-fold resolution increase over conventional microscopy.

Unlocking the power of AI for phenotyping fruit morphology in Arabidopsis →

Abstract Deep learning can revolutionise high-throughput image-based phenotyping by automating the measurement of complex traits, a task that is often labour-intensive, time-consuming, and prone to human error. However, its precision and adaptability in accurately phenotyping organ-level traits, such as fruit morphology, remain to be fully evaluated. Establishing the links between phenotypic and genotypic variation is essential for uncovering the genetic basis of traits and can also provide an orthologous test of pipeline effectiveness. In this study, we assess the efficacy of deep learning for measuring variation in fruit morphology in Arabidopsis using images from a multiparent advanced generation intercross (MAGIC) mapping family. We trained an instance segmentation model and developed a pipeline to phenotype Arabidopsis fruit morphology, based on the model outputs. Our model achieved strong performance with an average precision of 88.0% for detection and 55.9% for segmentation. Quantitative trait locus analysis of the derived phenotypic metrics of the MAGIC population identified significant loci associated with fruit morphology. This analysis, based on automated phenotyping of 332,194 individual fruits, underscores the capability of deep learning as a robust tool for phenotyping large populations. Our pipeline for quantifying pod morphological traits is scalable and provides high-quality phenotype data, facilitating genetic analysis and gene discovery, as well as advancing crop breeding research.

Why it matches plant phenotyping methods深層学習による果実形態の画像ベース表現型抽出パイプラインを開発し、検出・セグメンテーション性能を評価しており、植物フェノタイピング手法が研究の中心である。

abstractWe trained an instance segmentation model and developed a pipeline to phenotype Arabidopsis fruit morphology, based on the model outputs.

Nf-Root: A Best-Practice Pipeline for Deep-Learning-Based Analysis of Apoplastic pH in Microscopy Images of Developmental Zones in Plant Root Tissue. →

Hormonal mechanisms associated with cell elongation play a vital role in the development and growth of plants. Here, we report Nextflow-root (nf-root), a novel best-practice pipeline for deep-learning-based analysis of fluorescence microscopy images of plant root tissue from A. thaliana. This bioinformatics pipeline performs automatic identification of developmental zones in root tissue images. This also includes apoplastic pH measurements, which is useful for modeling hormone signaling and cell physiological responses. We show that this nf-core standard-based pipeline successfully automates tissue zone segmentation and is both high-throughput and highly reproducible. In short, a deep-learning module deploys deterministically trained convolutional neural network models and augments the segmentation predictions with measures of prediction uncertainty and model interpretability, while aiming to facilitate result interpretation and verification by experienced plant biologists. We observed a high statistical similarity between the manually generated results and the output of the nf-root.

Why it matches plant phenotyping methods植物根組織の発達ゾーンを画像から自動抽出し、アポプラストpHを測定する再現可能な深層学習パイプラインを開発・検証しており、植物表現型取得が中心である。

abstractThis bioinformatics pipeline performs automatic identification of developmental zones in root tissue images.

NIRSpredict: a platform for predicting plant traits from near infra-red spectroscopy. →

Near-infrared spectroscopy (NIRS) has become a popular tool for investigating phenotypic variability in plants. We developed the Shiny NIRSpredict application to get predictions of 81 Arabidopsis thaliana phenotypic traits, including classical functional traits as well as a large variety of commonly measured chemical compounds, based from near-infrared spectroscopy values based on deep learning. It is freely accessible at the following URL: https://shiny.cefe.cnrs.fr/NirsPredict/ . NIRSpredict has three main functionalities. First, it allows users to submit their spectrum values to get the predictions of plant traits from models built with the hosted A. thaliana database. Second, users have access to the database of traits used for model calibration. Data can be filtered and extracted on user's choice and visualized in a global context. Third, a user can submit his own dataset to extend the database and get part of the application development. NIRSpredict provides an easy-to-use and efficient method for trait prediction and an access to a large dataset of A. thaliana trait values. In addition to covering many of functional traits it also allows to predict a large variety of commonly measured chemical compounds. As a reliable way of characterizing plant populations across geographical ranges, NIRSpredict can facilitate the adoption of phenomics in functional and evolutionary ecology.

Why it matches plant phenotyping methodsNIRスペクトルから植物形質を予測するソフトウェアおよびデータベースを開発しており、形質取得・推定手法が研究の中心である。

abstractWe developed the Shiny NIRSpredict application to get predictions of 81 Arabidopsis thaliana phenotypic traits

Virtual staining from bright-field microscopy for label-free quantitative analysis of plant cell structures →

The applicability of a deep learning model for the virtual staining of plant cell structures using bright-field microscopy was investigated. The training dataset consisted of microscopy images of tobacco BY-2 cells with the plasma membrane stained with the fluorescent dye PlasMem Bright Green and the cell nucleus labeled with Histone-red fluorescent protein. The trained models successfully detected the expansion of cell nuclei upon aphidicolin treatment and a decrease in the cell aspect ratio upon propyzamide treatment, demonstrating its utility in cell morphometry. The model also accurately documented the shape of Arabidopsis pavement cells in both wild type and the bpp125 triple mutant, which has an altered pavement cell phenotype. Metrics such as cell area, circularity, and solidity obtained from virtual staining analyses were highly correlated with those obtained by manual measurements of cell features from microscopy images. Furthermore, the versatility of virtual staining was highlighted by its application to track chloroplast movement in Egeria densa . The method was also effective for classifying live and dead BY-2 cells using texture-based machine learning, suggesting that virtual staining can be applied beyond typical segmentation tasks. Although this method still has some limitations, its non-invasive nature and efficiency make it highly suitable for label-free, dynamic, and high-throughput analyses in quantitative plant cell biology.

Why it matches plant phenotyping methods植物細胞構造の仮想染色と画像解析モデルを開発・評価し、細胞面積・形状・核拡大・葉緑体運動・生死などの表現型を定量化しているため、フェノタイピング手法が中心である。

abstractThe applicability of a deep learning model for the virtual staining of plant cell structures using bright-field microscopy was investigated.

Metabolic modeling identifies determinants of thermal growth responses in Arabidopsis thaliana →

Temperature is a critical environmental factor affecting nearly all plant processes, including growth, development, and yield. Yet, despite decades of research, we lack the ability to predict plant performance at different temperatures, limiting the development of climate-resilient crops. Further, there is a pressing need to bridge the gap between the prediction of physiological and molecular traits to improve our understanding and manipulation of plant temperature responses. Here, we developed the first enzyme-constrained model of Arabidopsis thaliana ’s metabolism, facilitating predictions of growth-related phenotypes at different temperatures. We showed that the model can be employed for in silico identification of genes that affect plant growth at suboptimal growth temperature. Using mutant lines, we validated the genes predicted to affect plant growth, demonstrating the potential of metabolic modeling in accurately predicting plant thermal responses. The temperature-dependent enzyme-constrained metabolic model provides a template that can be used for developing sophisticated strategies to engineer climate-resilient crops.

Why it matches plant phenotyping methods温度依存性の酵素制約代謝モデルを開発し、成長関連表現型の予測と変異体による検証を行っており、植物表現型推定手法が研究の中心です。

abstractHere, we developed the first enzyme-constrained model of Arabidopsis thaliana ’s metabolism, facilitating predictions of growth-related phenotypes at different temperatures.

An optimized live imaging and multiple cell layer growth analysis approach using Arabidopsis sepals. →

Arabidopsis thaliana sepals are excellent models for analyzing growth of entire organs due to their relatively small size, which can be captured at a cellular resolution under a confocal microscope. To investigate how differential growth of connected cell layers generate unique organ morphologies, it is necessary to live-image deep into the tissue. However, imaging deep cell layers of the sepal (or plant tissues in general) is practically challenging. Image processing is also difficult due to the low signal-to-noise ratio of the deeper tissue layers, an issue mainly associated with live imaging datasets. Addressing some of these challenges, we provide an optimized methodology for live imaging sepals, and subsequent image processing. For live imaging early-stage sepals, we found that the use of a bright fluorescent membrane marker, coupled with increased laser intensity and an enhanced Z- resolution produces high-quality images suitable for downstream image processing. Our optimized parameters allowed us to image the bottommost cell layer of the sepal (inner epidermal layer) without compromising viability. We used a 'voxel removal' technique to visualize the inner epidermal layer in MorphoGraphX image processing software. We also describe the MorphoGraphX parameters for creating a 2.5D mesh surface for the inner epidermis. Our parameters allow for the segmentation and parent tracking of individual cells through multiple time points, despite the weak signal of the inner epidermal cells. While we have used sepals to illustrate our approach, the methodology will be useful for researchers intending to live-image and track growth of deeper cell layers in 2.5D for any plant tissue.

Why it matches plant phenotyping methods植物組織の深部をライブイメージングし、画像処理・細胞セグメンテーション・追跡によって成長を解析する方法自体が中心的に開発・最適化されているため。

abstractwe provide an optimized methodology for live imaging sepals, and subsequent image processing.

Time-course analysis system for leaf feeding marks reveals effects of Arabidopsis trichomes on insect herbivore feeding behavior. →

Bioassay with an insect herbivore is a common approach to studying plant defense. While measuring insect growth rate as a negative indicator of plant defense levels is simple and straightforward, analysing more detailed feeding behavior parameters of insects, such as feeding rates, leaf area consumed per feeding event, intervals between feeding events, and spatio-temporal patterns of feeding sites on leaves, is more informative. However, such observations are generally time consuming and labor-intensive. Here, we provide a semi-automated system for quantifying feeding behavior parameters of insects feeding on plant leaves. Automated photo scanners record the time-course development of feeding marks on leaves. An image analysis pipeline processes the scanned images and extracts leaf area. By analysing changes in leaf area over time, it detects insect feeding events and calculates the leaf area consumed during each feeding event, providing quantitative parameters of the feeding behavior of insects. In addition, it visualizes spatio-temporal changes in feeding sites, providing a measure of the complex behavior of insects on leaves. Using this analysis pipeline, we demonstrate that Arabidopsis trichomes reduce insect feeding rate, but not feeding duration or intervals between feeding events. Our image acquisition system requires only a photo scanner and a laptop computer and does not require any specialized equipment. The analysis software is provided as an ImageJ macro and R package and is available at no cost. Taken together, our work provides a scalable method for quantitative assessment of the feeding behavior of insects on leaves, facilitating understanding of plant defense mechanisms.

Why it matches plant phenotyping methods葉の摂食痕をスキャン画像と解析パイプラインで定量し、摂食イベントごとの消費葉面積や時空間的な摂食部位を抽出する方法が研究の中心であるため、植物の損傷状態を測定するフェノタイピング手法として採用する。

abstractHere, we provide a semi-automated system for quantifying feeding behavior parameters of insects feeding on plant leaves.

Data-driven crop growth simulation on time-varying generated images using multi-conditional generative adversarial networks →

BACKGROUND: Image-based crop growth modeling can substantially contribute to precision agriculture by revealing spatial crop development over time, which allows an early and location-specific estimation of relevant future plant traits, such as leaf area or biomass. A prerequisite for realistic and sharp crop image generation is the integration of multiple growth-influencing conditions in a model, such as an image of an initial growth stage, the associated growth time, and further information about the field treatment. While image-based models provide more flexibility for crop growth modeling than process-based models, there is still a significant research gap in the comprehensive integration of various growth-influencing conditions. Further exploration and investigation are needed to address this gap. METHODS: We present a two-stage framework consisting first of an image generation model and second of a growth estimation model, independently trained. The image generation model is a conditional Wasserstein generative adversarial network (CWGAN). In the generator of this model, conditional batch normalization (CBN) is used to integrate conditions of different types along with the input image. This allows the model to generate time-varying artificial images dependent on multiple influencing factors. These images are used by the second part of the framework for plant phenotyping by deriving plant-specific traits and comparing them with those of non-artificial (real) reference images. In addition, image quality is evaluated using multi-scale structural similarity (MS-SSIM), learned perceptual image patch similarity (LPIPS), and Fréchet inception distance (FID). During inference, the framework allows image generation for any combination of conditions used in training; we call this generation data-driven crop growth simulation. RESULTS: Experiments are performed on three datasets of different complexity. These datasets include the laboratory plant Arabidopsis thaliana (Arabidopsis) and crops grown under real field conditions, namely cauliflower (GrowliFlower) and crop mixtures consisting of faba bean and spring wheat (MixedCrop). In all cases, the framework allows realistic, sharp image generations with a slight loss of quality from short-term to long-term predictions. For MixedCrop grown under varying treatments (different cultivars, sowing densities), the results show that adding these treatment information increases the generation quality and phenotyping accuracy measured by the estimated biomass. Simulations of varying growth-influencing conditions performed with the trained framework provide valuable insights into how such factors relate to crop appearances, which is particularly useful in complex, less explored crop mixture systems. Further results show that adding process-based simulated biomass as a condition increases the accuracy of the derived phenotypic traits from the predicted images. This demonstrates the potential of our framework to serve as an interface between a data-driven and a process-based crop growth model. CONCLUSION: The realistic generation and simulation of future plant appearances is adequately feasible by multi-conditional CWGAN. The presented framework complements process-based models and overcomes their limitations, such as the reliance on assumptions and the low exact field-localization specificity, by realistic visualizations of the spatial crop development that directly lead to a high explainability of the model predictions.

Why it matches plant phenotyping methods植物画像を生成し、そこから植物個体別形質を推定する二段階の画像ベース表現型解析フレームワークを開発・評価しており、表現型取得・推定手法が研究の中心である。

abstractWe present a two-stage framework consisting first of an image generation model and second of a growth estimation model, independently trained.

Automated and high throughput measurement of leaf stomatal traits in canola →

Abstract Background Automating stomatal trait measurement has gained popularity because of their inherent importance for field phenotyping application as stomata are critical for both carbon capture and water use efficiency in plants. Such tool has been reported for rice, wheat, tomato, barley and oil palm. However, none exist yet for canola, which is an important economic and agronomic crop globally. Results We developed a new toolkit called Stomatal Comprehensive Automated Neural Network or SCAN by combining the use of high-resolution portable digital microscopy with machine learning based on You Only Look Once algorithm (YOLOv8). Digital micrographs of leaf surfaces enter the SCAN pipeline, which includes stomata detection, stomata segmentation and stomatal pore segmentation models, to measure stomatal density, stomatal size and stomatal pore area, respectively. In addition to SCAN’s ability to measure leaf stomatal traits in canola at 89 to 94% accuracy, we also showed that SCAN can be used to predict stomatal density even in species not included in the training set such as Arabidopsis, tobacco, rice, wheat, maize and proso millet. SCAN was designed for the biological science community with the premise that users are not required to possess advanced programming capabilities to manage dependency prerequisites, execute the models, and integrate the analysis. This was achieved by packaging the models into a desktop application system that can be accessed offline. Conclusion Overall, SCAN provides a non-destructive, real-time, portable, and high-throughput measurement of leaf stomatal traits in canola. The minimised hardware requirement and user-friendly desktop application system make SCAN suitable for field phenotyping application.

Why it matches plant phenotyping methodsカノーラ葉の気孔形質を画像と機械学習で自動抽出するツールを開発し、精度検証と他種での適用性評価を行っており、フェノタイピング手法が中心である。

abstractWe developed a new toolkit called Stomatal Comprehensive Automated Neural Network or SCAN by combining the use of high-resolution portable digital microscopy with machine learning based on You Only Look Once algorithm (YOLOv8).

A low-cost open-source imaging platform reveals spatiotemporal insight into leaf elongation and movement. →

Plant organs move throughout the diurnal cycle, changing leaf and petiole positions to balance light capture, leaf temperature, and water loss under dynamic environmental conditions. Upward movement of the petiole, called hyponasty, is one of several traits of the shade avoidance syndrome (SAS). SAS traits are elicited upon perception of vegetation shade signals such as far-red light (FR) and improve light capture in dense vegetation. Monitoring plant movement at a high temporal resolution allows studying functionality and molecular regulation of hyponasty. However, high temporal resolution imaging solutions are often very expensive, making this unavailable to many researchers. Here, we present a modular and low-cost imaging setup, based on small Raspberry Pi computers that can track leaf movements and elongation growth with high temporal resolution. We also developed an open-source, semiautomated image analysis pipeline. Using this setup, we followed responses to FR enrichment, light intensity, and their interactions. Tracking both elongation and the angle of the petiole, lamina, and entire leaf in Arabidopsis (Arabidopsis thaliana) revealed insight into R:FR sensitivities of leaf growth and movement dynamics and the interactions of R:FR with background light intensity. The detailed imaging options of this system allowed us to identify spatially separate bending points for petiole and lamina positioning of the leaf.

Why it matches plant phenotyping methods低コスト画像計測プラットフォームとオープンソース解析パイプラインを開発し、葉の伸長・運動・角度を高時間分解能で抽出する方法が研究の中心である。

abstractHere, we present a modular and low-cost imaging setup, based on small Raspberry Pi computers that can track leaf movements and elongation growth with high temporal resolution.

ScAnalyzer: an image processing tool to monitor plant disease symptoms and pathogen spread in Arabidopsis thaliana leaves. →

Background Plants are known to be infected by a wide range of pathogenic microbes. To study plant diseases caused by microbes, it is imperative to be able to monitor disease symptoms and microbial colonization in a quantitative and objective manner. In contrast to more traditional measures that use manual assignments of disease categories, image processing provides a more accurate and objective quantification of plant disease symptoms. Besides monitoring disease symptoms, computational image processing provides additional information on the spatial localization of pathogenic microbes in different plant tissues. Results Here we report on an image analysis tool called ScAnalyzer to monitor disease symptoms and bacterial spread in Arabidopsis thaliana leaves. Thereto, detached leaves are assembled in a grid and scanned, which enables automated separation of individual samples. A pixel color threshold is used to segment healthy (green) from chlorotic (yellow) leaf areas. The spread of luminescence-tagged bacteria is monitored via light-sensitive films, which are processed in a similar manner as the leaf scans. We show that this tool is able to capture previously identified differences in susceptibility of the model plant A. thaliana to the bacterial pathogen Xanthomonas campestris pv. campestris. Moreover, we show that the ScAnalyzer pipeline provides a more detailed assessment of bacterial spread within plant leaves than previously used methods. Finally, by combining the disease symptom values with bacterial spread values from the same leaves, we show that bacterial spread precedes visual disease symptoms. Conclusion Taken together, we present an automated script to monitor plant disease symptoms and microbial spread in A. thaliana leaves. The freely available software ( https://github.com/MolPlantPathology/ScAnalyzer ) has the potential to standardize the analysis of disease assays between different groups.

Why it matches plant phenotyping methods植物葉の病徴と病原体拡散を画像処理で定量化するツールおよび解析パイプラインが研究の中心であり、植物の病害状態を直接推定するため。

abstractimage processing provides a more accurate and objective quantification of plant disease symptoms.

Topological analysis of 3D digital ovules identifies cellular patterns associated with ovule shape diversity. →

Tissue morphogenesis remains poorly understood. In plants, a central problem is how the 3D cellular architecture of a developing organ contributes to its final shape. We address this question through a comparative analysis of ovule morphogenesis, taking advantage of the diversity in ovule shape across angiosperms. Here, we provide a 3D digital atlas of Cardamine hirsuta ovule development at single cell resolution and compare it with an equivalent atlas of Arabidopsis thaliana. We introduce nerve-based topological analysis as a tool for unbiased detection of differences in cellular architectures and corroborate identified topological differences between two homologous tissues by comparative morphometrics and visual inspection. We find that differences in topology, cell volume variation and tissue growth patterns in the sheet-like integuments and the bulbous chalaza are associated with differences in ovule curvature. In contrast, the radialized conical ovule primordia and nucelli exhibit similar shapes, despite differences in internal cellular topology and tissue growth patterns. Our results support the notion that the structural organization of a tissue is associated with its susceptibility to shape changes during evolutionary shifts in 3D cellular architecture.

Why it matches plant phenotyping methods3Dデジタルアトラスと神経ベースのトポロジー解析、形態計測を用いて植物器官の細胞構造と形状を定量化しており、表現型取得・解析手法が研究の中心である。

abstractHere, we provide a 3D digital atlas of Cardamine hirsuta ovule development at single cell resolution and compare it with an equivalent atlas of Arabidopsis thaliana.

Four-dimensional quantitative analysis of cell plate development in Arabidopsis using lattice light sheet microscopy identifies robust transition points between growth phases. →

Cell plate formation during cytokinesis entails multiple stages occurring concurrently and requiring orchestrated vesicle delivery, membrane remodelling, and timely deposition of polysaccharides, such as callose. Understanding such a dynamic process requires dissection in time and space; this has been a major hurdle in studying cytokinesis. Using lattice light sheet microscopy (LLSM), we studied cell plate development in four dimensions, through the behavior of yellow fluorescent protein (YFP)-tagged cytokinesis-specific GTPase RABA2a vesicles. We monitored the entire duration of cell plate development, from its first emergence, with the aid of YFP-RABA2a, in both the presence and absence of cytokinetic callose. By developing a robust cytokinetic vesicle volume analysis pipeline, we identified distinct behavioral patterns, allowing the identification of three easily trackable cell plate developmental phases. Notably, the phase transition between phase I and phase II is striking, indicating a switch from membrane accumulation to the recycling of excess membrane material. We interrogated the role of callose using pharmacological inhibition with LLSM and electron microscopy. Loss of callose inhibited the phase transitions, establishing the critical role and timing of the polysaccharide deposition in cell plate expansion and maturation. This study exemplifies the power of combining LLSM with quantitative analysis to decode and untangle such a complex process.

Why it matches plant phenotyping methodsLLSMによる4次元画像取得と、細胞板の小胞体積を定量化する解析パイプラインの開発が研究の中心であり、植物細胞の形態・発達状態を抽出する方法として substantive です。

abstractUsing lattice light sheet microscopy (LLSM), we studied cell plate development in four dimensions

From Organelle Morphology to Whole-Plant Phenotyping: A Phenotypic Detection Method Based on Deep Learning →

The analysis of plant phenotype parameters is closely related to breeding, so plant phenotype research has strong practical significance. This paper used deep learning to classify Arabidopsis thaliana from the macro (plant) to the micro level (organelle). First, the multi-output model identifies Arabidopsis accession lines and regression to predict Arabidopsis’s 22-day growth status. The experimental results showed that the model had excellent performance in identifying Arabidopsis lines, and the model’s classification accuracy was 99.92%. The model also had good performance in predicting plant growth status, and the regression prediction of the model root mean square error (RMSE) was 1.536. Next, a new dataset was obtained by increasing the time interval of Arabidopsis images, and the model’s performance was verified at different time intervals. Finally, the model was applied to classify Arabidopsis organelles to verify the model’s generalizability. Research suggested that deep learning will broaden plant phenotype detection methods. Furthermore, this method will facilitate the design and development of a high-throughput information collection platform for plant phenotypes.

Why it matches plant phenotyping methods深層学習による植物画像からの系統識別・生育状態推定を開発し、時間間隔データで検証しており、植物表現型取得・推定手法が研究の中心である。

abstractThis paper used deep learning to classify Arabidopsis thaliana from the macro (plant) to the micro level (organelle).

Resource allocation modeling for autonomous prediction of plant cell phenotypes. →

Predicting the plant cell response in complex environmental conditions is a challenge in plant biology. Here we developed a resource allocation model of cellular and molecular scale for the leaf photosynthetic cell of Arabidopsis thaliana, based on the Resource Balance Analysis (RBA) constraint-based modeling framework. The RBA model contains the metabolic network and the major macromolecular processes involved in the plant cell growth and survival and localized in cellular compartments. We simulated the model for varying environmental conditions of temperature, irradiance, partial pressure of CO 2 and O 2 , and compared RBA predictions to known resource distributions and quantitative phenotypic traits such as the relative growth rate, the C:N ratio, and finally to the empirical characteristics of CO 2 fixation given by the well-established Farquhar model. In comparison to other standard constraint-based modeling methods like Flux Balance Analysis, the RBA model makes accurate quantitative predictions without the need for empirical constraints. Altogether, we show that RBA significantly improves the autonomous prediction of plant cell phenotypes in complex environmental conditions, and provides mechanistic links between the genotype and the phenotype of the plant cell.

Why it matches plant phenotyping methodsRBAモデルを用いて植物細胞の生長率やC:N比などの表現型を定量予測する計算手法の開発が中心であり、単なる生物学的実験ではない。

abstractHere we developed a resource allocation model of cellular and molecular scale for the leaf photosynthetic cell of Arabidopsis thaliana

PAT (Periderm Assessment Toolkit): A Quantitative and Large-Scale Screening Method for Periderm Measurements. →