Search bioRxiv⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/253724

Robust colony recognition for high-throughput growth analysis from suboptimal low-magnification brightfield micrographs

Plavskin, Y.·Li, S.·Ziv, N.·Levy, S. F.·Siegal, M. L.
View original source ↗Export metadata ↓

Abstract

New technological advances have enabled high-throughput phenotyping at the single-cell level, yet analyzing the large amount of data generated by high throughput phenotyping experiments automatically and accurately is a considerable challenge. Here we introduce Processing Images Easily (PIE), software that automatically tracks growth of microbial colonies in low-magnification brightfield images by combining adaptive object-center recognition with gradient-based object-outline recognition. PIE recognizes colony outlines very robustly and accurately across a wide range of image brightnesses, focal depths, and organisms. Beyond accurate colony recognition, PIE is designed to easily integrate with complex experiments, allowing colony tracking across multiple experimental phases and classification based on fluorescence intensity. We show that PIE can be used to accurately measure the growth rates of large numbers (>90,000) of bacterial or yeast microcolonies in a single-time-lapse experiment, allowing calculation of population-wide growth properties. Finally, PIE is able to track individual colonies across multiple experimental phases, measuring both growth and fluorescence properties of the microcolonies. Author SummaryHigh-throughput microscopy has enabled automated collection of large amounts of growth and gene-expression data in microbes. Computational methods that can precisely recognize and track organisms in images are essential to performing measurements at scale using automated microscopy. We have developed PIE, software that automatically recognizes microbial colonies in microscopy images, tracks them in imaging time-series, and performs measurements of growth and, potentially, gene expression. PIE is highly effective on low-resolution images, outperforming current state-of-the-art approaches in both speed and accuracy, and works well in microbes of varying shapes and sizes. In addition, PIE allows tracking microcolonies across arbitrary sequences of experimental phases, each collecting data in different modalities. We show that PIE allows measurement of growth and fluorescence properties in tens of thousands of microbial colonies in a single experiment, and that in turn the scale of these measurements can lead to important insights about interindividual differences in growth and stress response. PIE is available as a Python package (https://doi.org/10.5281/zenodo.4987127) with documentation currently at https://pie-image.readthedocs.io/; users can also run analysis on individual images or time-series without the need to install PIE by using our web application, currently available at http://pie.hpc.nyu.edu/.

Explore related subjects

bioinformaticsPreprint

Keep this discovery

BibTeXRIS

Plavskin, Y., Li, S., Ziv, N., Levy, S. F., Siegal, M. L.. 2018-01-25. Robust colony recognition for high-throughput growth analysis from suboptimal low-magnification brightfield micrographs. https://doi.org/10.1101/253724

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

spatialMET: an open and scalable framework for spatial metabolomics analysis

Mass spectrometry imaging (MSI) enables spatially resolved metabolomics in intact tissue sections, but analysis remains challenging at scale. Existing MSI workflows often require users to combine multiple software tools, while others rely on proprietary vendor software that limits interoperability and reproducibility. To address these challenges, we developed spatialMET, an open-source framework that provides an end-to-end workflow for MSI analysis. spatialMET provides a unified platform for preprocessing, spatial domain detection, and visualization. Downstream analyses include differential abundance testing, spatial autocorrelation and gradient analysis, dimensionality reduction, and correlation network analysis. Spatial domain detection uses hcdist, a C-based hierarchical clustering implementation that substantially reduces runtime and memory use relative to existing R-based approaches. spatialMET can be run through an interactive R Shiny application or as a standalone command-line workflow for larger datasets or high-performance computing environments. Applied to mouse small cell lung cancer MALDI-MSI data containing 284,673 pixels, spatialMET identified tumor-associated, stromal, and adjacent lung spatial domains that aligned with matched histology. Differential abundance analysis identified 117 m/z features that differed between tumor and stromal regions, while spatial autocorrelation analyses revealed spatially structured abundance patterns. Applying spatialMET to mouse lung adenocarcinoma data from an entire lung lobe containing 338,477 pixels further demonstrated scalability and captured spatial heterogeneity across tumor and surrounding lung tissue. In summary, spatialMET provides a scalable, open-source framework for end-to-end spatial metabolomics analysis, and it is distributed as a Docker container for reproducible deployment. Source code and installation instructions are available at https://github.com/biodatalab/spatialMET.

bioinformatics↗
bioRxiv2026

Probing the transcriptome response to shivering in skeletal muscle using a multilayered bioinformatics approach

Cold acclimation holds therapeutic potential for improving metabolic health. We previously demonstrated that repeated cold-induced shivering enhances insulin sensitivity in humans. However, the molecular pathways that underlie the skeletal muscle shivering response, and how these relate to beneficial physiological effects, remain poorly understood. In this study, we combined complementary bioinformatics approaches to allow in-depth analysis of the transcriptomic response of human skeletal muscle to repeated shivering. We identified a robust transcriptional signature and show a sex-specific component in the shivering skeletal muscle response, which seemed to diminish following cold adaptation. Our findings provide mechanistic insights into cold-induced muscle adaptations, shed light on potential interesting molecular targets for further investigation, and emphasize the importance of including both sexes in future cold acclimation studies.

bioinformatics↗
bioRxiv2026

An Information Geometry approach to model topological trajectories and Gene Expression Radius from UMAP geometry.

Understanding the relationship between gene expression dynamics and cellular identity remains a central challenge in single cell biology. Here, we introduce a novel computational and mathematical framework that integrates information geometry, fuzzy topology, and UMAP analysis to model gene expression landscapes derived from single cell RNA sequencing data. We formalize gene expression data as a fuzzy topological space, where interactions between expression points are governed by probabilistic distributions inspired by manifold learning approaches such as UMAP. Within this framework, we define an information geometric structure through a Fisher metric induced by these distributions, enabling the computation of geodesic trajectories that capture cellular differentiation processes. A key contribution of this work is the derivation of analytical conditions, expressed as expression radius formulas, that characterize local neighborhoods in gene expression space. These conditions allow for the identification of genes associated with stem cell states and predictions in transitional cell types in future work. Application of the proposed framework to single cell datasets reveals biologically meaningful gene sets enriched in key regulatory pathways and transcription factors, demonstrating the capacity of our approach to uncover latent structure in complex gene expression data. Our results suggest that integrating differential geometry with statistical learning theory offers a powerful paradigm for modeling genotype and phenotype relationships and cellular state transitions, with potential implications for precision medicine and systems biology.

bioinformatics↗