spacr-nightly 1.4.0

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Features

• Generate Masks: Generate cellpose masks of cell, nuclei, and pathogen objects.

• Object Measurements: Measurements for each object including scikit-image regionprops, intensity percentiles, shannon-entropy, Pearson’s and Manders’ correlations, homogeneity, and radial distribution. Measurements are saved to a SQL database in object-level tables.

• Crop Images: Save objects (cells, nuclei, pathogen, cytoplasm) as images. Object image paths are saved in a SQL database.

• Train CNNs or Transformers: Train Torch models to classify single object images.

• Manual Annotation: Supports manual annotation of single-cell images and segmentation to refine training datasets for training CNNs/Transformers or cellpose, respectively.

• Finetune Cellpose Models: Adjust pre-existing Cellpose models to your specific dataset for improved performance.

• Timelapse Data Support: Track objects in timelapse image data.

• Simulations: Simulate spatial phenotype screens.

• Sequencing: Map FASTQ reads to barcode and gRNA barcode metadata.

• Misc: Analyze Ca oscillation, recruitment, infection rate, plaque size/count.

Overview and data organization of spaCR.

a. Schematic workflow of the spaCR pipeline for pooled image-based CRISPR screens. Microscopy images (TIFF, LIF, CZI, NDI) and sequencing reads (FASTQ) are used as inputs (black). The main modules (teal) are: (1) Mask: generates object masks for cells, nuclei, pathogens, and cytoplasm; (2) Measure: extracts object-level features and crops object images, storing quantitative data in an SQL database; (3) Classify—applies machine learning (ML, e.g., XGBoost) or deep learning (DL, e.g., PyTorch) models to classify objects, summarizing results as well-level classification scores; (4) Map Barcodes: extracts and maps row, column, and gRNA barcodes from sequencing data to corresponding wells; (5) Regression: estimates gRNA effect sizes and gene scores via multiple linear regression using well-level summary statistics. b. Downstream submodules available for extended analyses at each stage. c. Output folder structure for each module, including locations for raw and processed images, masks, object-level measurements, datasets, and results. d. List of all spaCR package modules.

Installation

Linux recommended. If using Windows, switch to Linux—it’s free, open-source, and better.

macOS prerequisites (before install):

brew install libomp hdf5 cmake openssl

Linux GUI requirement: SpaCr GUI requires Tkinter.

sudo apt-get install python3-tk

Installation:

pip install spacr

Run SpaCr GUI:

spacr

Example Notebooks

The following example Jupyter notebooks illustrate common workflows using spaCR.

• Generate masks Generate cell, nuclei, and pathogen segmentation masks from microscopy images using Cellpose.

• Capture single cell images and measurements Extract object-level measurements and crop single-cell images for downstream analysis.

• Machine learning based object classification Train traditional machine learning models (e.g., XGBoost) to classify cell phenotypes based on extracted features.

• Computer vision based object classification Train and evaluate deep learning models (PyTorch CNNs/Transformers) on cropped object images.

• Map sequencing barcodes Map sequencing reads to row, column, and gRNA barcodes for CRISPR screen genotype-phenotype mapping.

• Finetune cellpose models Finetune Cellpose models using your own annotated training data for improved segmentation accuracy.

Interactive Tutorial (under construction)

Click below to explore the step-by-step GUI and Notebook tutorials for spaCR:

Narrated video tutorials

Every home module ships with a scripted narrated MP4 walkthrough that you can regenerate locally on any machine with ffmpeg. The rendering pipeline (spacr.qt.tutorial) uses Piper for neural TTS narration and captures frames directly from the Qt widget tree — no Xvfb needed.

pip install -e '.[tutorial]'
# Fetch a voice model (one-time):
mkdir -p ~/.spacr/piper && curl -sL \
  https://huggingface.co/rhasspy/piper-voices/resolve/main/en/en_US/lessac/medium/en_US-lessac-medium.onnx \
  -o ~/.spacr/piper/en_US-lessac-medium.onnx
curl -sL \
  https://huggingface.co/rhasspy/piper-voices/resolve/main/en/en_US/lessac/medium/en_US-lessac-medium.onnx.json \
  -o ~/.spacr/piper/en_US-lessac-medium.onnx.json

# Render one tutorial:
spacr-tutorial mask         # → ~/spacr-tutorials/mask.mp4 + mask.srt

# Or every module:
spacr-tutorial all          # → mask, measure, crop, classify,
                             #    timelapse, home under ~/spacr-tutorials

Each MP4 is a 1080p30 clip with a burned-in cursor overlay + AAC narration track and a matching .srt sidecar with the exact narration text. Swap the audio track with your own recording for publish-ready videos.

spaCRPower

Power analasys of pooled perterbation spaCR screens.

spaCRPower

Data Availability

• Full microscopy image dataset: EMBL-EBI BioStudies S-BIAD2135

• Testing dataset: Hugging Face toxo_mito

• Sequencing data: NCBI BioProject PRJNA1261935

License

spaCR is distributed under the terms of the MIT License. See the LICENSE file for details.

How to Cite

If you use spaCR in your research, please cite: Olafsson EB, et al. A pooled image-based CRISPR screen identifies EAF1 as a T. gondii modulator of 2 ESCRT subversion. Manuscript under consideration.

• Preprint: A pooled image-based CRISPR screen identifies EAF1 as a T. gondii modulator of 2 ESCRT subversion

Papers Using spaCR

Below are selected publications that have used or cited spaCR:

• Olafsson EB, et al. SpaCr: Spatial phenotype analysis of CRISPR-Cas9 screens. Manuscript in preparation.

• IRE1α promotes phagosomal calcium flux to enhance macrophage fungicidal activity

• Metabolic adaptability and nutrient scavenging in Toxoplasma gondii: insights from ingestion pathway-deficient mutants