diff --git a/.github/actions/build-dev-image/action.yml b/.github/actions/build-dev-image/action.yml index 76d6d939..116e9e3e 100644 --- a/.github/actions/build-dev-image/action.yml +++ b/.github/actions/build-dev-image/action.yml @@ -1,9 +1,6 @@ name: "build-dev-image" description: "Build a dev image and push it to the Pixelgen container registries" inputs: - tag: - description: "The tag to use for the image" - required: true cache-from: description: "The cache to use for the build" default: "" @@ -61,7 +58,12 @@ runs: bakeSetCommands.push(...cacheToArray) bakeSetCommands.push(...platformArray) - bakeSetCommandsText = bakeSetCommands.join('\n') + // Override the version in the container if we are run in a version tag push event + if ( context.eventName === "push" && context.ref.startsWith('refs/tags/v') ) { + bakeSetCommands.push("*.args.VERSION_OVERRIDE=" + "${{ steps.meta.outputs.version }}" ) + } + + const bakeSetCommandsText = bakeSetCommands.join('\n') core.setOutput('set', bakeSetCommandsText) - name: Build and push to ghcr and ecr diff --git a/.github/workflows/tests.yml b/.github/workflows/tests.yml index 94d65436..e38d7ece 100644 --- a/.github/workflows/tests.yml +++ b/.github/workflows/tests.yml @@ -132,7 +132,8 @@ jobs: fail-fast: false matrix: case: - - D21 + - mpx_v1 + - mpx_v2 if: needs.pre_job.outputs.should_skip == '{}' || !fromJSON(needs.pre_job.outputs.paths_result).python.should_skip steps: diff --git a/.pre-commit-config.yaml b/.pre-commit-config.yaml index 27b8f9e6..ec20bc93 100644 --- a/.pre-commit-config.yaml +++ b/.pre-commit-config.yaml @@ -18,7 +18,7 @@ repos: - id: ruff-format - repo: https://github.com/pre-commit/mirrors-mypy - rev: "v1.3.0" # Use the sha / tag you want to point at + rev: "v1.10.1" # Use the sha / tag you want to point at hooks: - id: mypy exclude: "(^cue.mod/)|(^docs/)" diff --git a/CHANGELOG.md b/CHANGELOG.md index 56fbba85..03aa59db 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -5,194 +5,222 @@ All notable changes to this project will be documented in this file. The format is based on [Keep a Changelog](https://keepachangelog.com/en/1.0.0/), and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0.html). +## [0.18.0] - 2024-07-11 + +### Added + +- Add minimum marker count `colocalization_min_marker_count` parameter to calculate colocalization score. +- Add `density_scatter_plot` function to make two-marker abundance scatter plots with pseudo-density coloring. +- Add `wpmds` option in `pmds_layout` to compute edge weighted layouts. This is now set as the default layout algorithm. +- Add `dsb_normalization` function for normalization of marker abundance. +- Add a `Fraction of Outlier Cells` metric to the QC report. +- Add a `Panel Version` metadata field to the QC report. +- Add support for datasets generated using the `human-sc-immunology-spatial-proteomics-2` panel. + +### Changed + +- The default value for `normalize_counts` in `local_g` is now `False` instead of `True`. +- The default transformation for the calculation of the colocalization score is now `rate-diff` instead of `log1p`. +- Rename `edge_rank_plot` function to `molecule_rank_plot`. + +### Fixed + +- Fix a bug in `compute_transition_probabilities` when `k>1` where the stochastic matrix was not correctly row-normalized. +- Fix a bug in `local_g` when `use_weights=False` where the adjacency matrix was not correctly expended if `k>1`. +- Fix a bug where `a_pixels_per_b_pixel` summary statistics where equal to the `b_pixels_per_a_pixel` statistics. +- `collapse` will return exit code 137 when one of the child processes is killed by the system (e.g. because it is + to much memory). This allows e.g. Nextflow to retry the process with more memory automatically. +- Hide the `Sample Description` metadata field in the QC report when no value is available. +- Fix an issue where boolean parameters were formatted as integers in the Parameters section of the QC report. +- Fix a bug in aggregating files with precomputed layouts, where the lazy-loading of the layouts was not working correctly. + +### Removed + +- Remove the `Pixel Version` metadata field from the QC report. + ## [0.17.1] - 2024-05-27 ### Fixed -* Poor performance when writing many small layouts to pxl file (~45x speed-up). This should almost only - impact test scenarios, since most real components should be large enough for this not to be an issue. +- Poor performance when writing many small layouts to pxl file (~45x speed-up). This should almost only + impact test scenarios, since most real components should be large enough for this not to be an issue. ## [0.17.0] - 2024-05-23 ### Added -* Add `rate_diff_transformation` function with `rate-diff` alias as an alternative option for transforming marker counts before colocalization calculation. -* Add `local_g` function to compute spatial autocorrelation of marker counts per node. -* Add `compute_transition_probabilities` function to compute transition probabilities for k-step random walks for node pairs in a graph. -* Add QC plot showing UMIs per UPIA vs Tau. -* Add plot functions showing edge rank and cell counts. -* Add 2D and 3D graph plot functions. -* Add heatmap plot functions showing colocalization and differential colocalization. -* Add volcano plot (value difference vs log p-value) function for differential colocalization. -* Add a function to calculate the differential colocalization between two conditions. -* Performance improvements and reduced bundle size in QC report. -* Improved console output in verbose mode. -* Improved logging from multiprocessing jobs. -* Improved runtime for graph creation. -* Added PMDS layout algorithm. -* Add `--sample_name` option to `single-cell amplicon` to overwrite the name derived from the input filename. -* Add `--skip-input-checks` option to `single-cell amplicon` to make input filename checks warnings instead of errors. -* `PixelDataset` instances are now written to disk without creating intermediate files on-disk. -* A nice string representation for the `Graph` class, to let you know how many nodes and edges there are in the current graph object instance. -* Metric to collect molecules (edges) in cells with outlier distributions of antibodies (aggregates). -* Provide typed interfaces for all per-stage report files using pydantic. -* Centralize pixelator intermediate file lookup and access. -* Add a `precomputed_layouts` property to `PixelDataset` to allow for loading precomputed layouts. -* Add `pixelator single-cell layout` stage to pixelator, which allows users to compute layouts for a PXL file that can then be used to visualize the graph in 2D or 3D downstream. -* Add minimum marker count `polarization_min_marker_count` parameter to calculate Polarity Score. -* Add "log1p" as an alternative for `PolarizationNormalizationTypes`. -* Add `convert_indices_to_integers` option when creating graphs. -* Add a feature flag module to aid in the development of new features. +- Add `rate_diff_transformation` function with `rate-diff` alias as an alternative option for transforming marker counts before colocalization calculation. +- Add `local_g` function to compute spatial autocorrelation of marker counts per node. +- Add `compute_transition_probabilities` function to compute transition probabilities for k-step random walks for node pairs in a graph. +- Add QC plot showing UMIs per UPIA vs Tau. +- Add plot functions showing edge rank and cell counts. +- Add 2D and 3D graph plot functions. +- Add heatmap plot functions showing colocalization and differential colocalization. +- Add volcano plot (value difference vs log p-value) function for differential colocalization. +- Add a function to calculate the differential colocalization between two conditions. +- Performance improvements and reduced bundle size in QC report. +- Improved console output in verbose mode. +- Improved logging from multiprocessing jobs. +- Improved runtime for graph creation. +- Added PMDS layout algorithm. +- Add `--sample_name` option to `single-cell amplicon` to overwrite the name derived from the input filename. +- Add `--skip-input-checks` option to `single-cell amplicon` to make input filename checks warnings instead of errors. +- `PixelDataset` instances are now written to disk without creating intermediate files on-disk. +- A nice string representation for the `Graph` class, to let you know how many nodes and edges there are in the current graph object instance. +- Metric to collect molecules (edges) in cells with outlier distributions of antibodies (aggregates). +- Provide typed interfaces for all per-stage report files using pydantic. +- Centralize pixelator intermediate file lookup and access. +- Add a `precomputed_layouts` property to `PixelDataset` to allow for loading precomputed layouts. +- Add `pixelator single-cell layout` stage to pixelator, which allows users to compute layouts for a PXL file that can then be used to visualize the graph in 2D or 3D downstream. +- Add minimum marker count `polarization_min_marker_count` parameter to calculate Polarity Score. +- Add "log1p" as an alternative for `PolarizationNormalizationTypes`. +- Add `convert_indices_to_integers` option when creating graphs. +- Add a feature flag module to aid in the development of new features. ### Changed -* Change name and description of `Avg. Reads per Cell` and `Avg. Reads Usable per Cell` in QC report. -* The output name of the `.pxl` file from the `annotate` step is now `*.annotated.dataset.pxl`. -* The output name of the `.pxl` file from the `analysis` step is now `*.analysis.dataset.pxl`. -* The term `edges` in `metrics` and `adata` is now replaced with `molecules`. -* Renaming of variables in per-stage JSON reports. -* Changed name of TCRb to TCRVb5 antibody in human-immunology-panel file and bumped to version 0.5.0. -* Renaming of component metrics in adata. -* Use MPX graph compatible permutation strategy when calculating Moran's I related statistics. -* Marker filtering is now done after count transformation in polarization score calculation. -* Use the input read count at the annotate stage for the `fraction_antibody_reads_in_outliers` metric denominator instead of the total raw input reads. -* Use common analysis engine to orchestrate running different "per component" analyses, like polarization and colocalization analysis (yielding a roughly 3x speed-up over the previous approach). -* The default transformation for the calculation of the polarity score is now `log1p` instead of `clr`. +- Change name and description of `Avg. Reads per Cell` and `Avg. Reads Usable per Cell` in QC report. +- The output name of the `.pxl` file from the `annotate` step is now `*.annotated.dataset.pxl`. +- The output name of the `.pxl` file from the `analysis` step is now `*.analysis.dataset.pxl`. +- The term `edges` in `metrics` and `adata` is now replaced with `molecules`. +- Renaming of variables in per-stage JSON reports. +- Changed name of TCRb to TCRVb5 antibody in human-immunology-panel file and bumped to version 0.5.0. +- Renaming of component metrics in adata. +- Use MPX graph compatible permutation strategy when calculating Moran's I related statistics. +- Marker filtering is now done after count transformation in polarization score calculation. +- Use the input read count at the annotate stage for the `fraction_antibody_reads_in_outliers` metric denominator instead of the total raw input reads. +- Use common analysis engine to orchestrate running different "per component" analyses, like polarization and colocalization analysis (yielding a roughly 3x speed-up over the previous approach). +- The default transformation for the calculation of the polarity score is now `log1p` instead of `clr`. ### Fixed -* Fix a bug in how discarded UMIs are calculated and reported. -* Fix deflated counts in the edgelist after collapse. -* Fix a bug where an `r1` or `r2` in the directory part of a read file would break file name sanity checks. -* Fix a bug where the wrong `r1` or `r2` in the filename would be removed when multiple matches are present. -* Logging would cause deadlocks in multiprocessing scenarios, this has been resolved by switching to a server/client-based logging system. -* Fix a bug in the amplicon stage where read suffixes were not correctly recognized. -* Ensure deterministic results from `pmds_layout` (given a set seed). -* Fix an issue with the `fraction_antibody_reads_usable_per_cell` metric where the denominator read count was not correctly averaged with the cell count. +- Fix a bug in how discarded UMIs are calculated and reported. +- Fix deflated counts in the edgelist after collapse. +- Fix a bug where an `r1` or `r2` in the directory part of a read file would break file name sanity checks. +- Fix a bug where the wrong `r1` or `r2` in the filename would be removed when multiple matches are present. +- Logging would cause deadlocks in multiprocessing scenarios, this has been resolved by switching to a server/client-based logging system. +- Fix a bug in the amplicon stage where read suffixes were not correctly recognized. +- Ensure deterministic results from `pmds_layout` (given a set seed). +- Fix an issue with the `fraction_antibody_reads_usable_per_cell` metric where the denominator read count was not correctly averaged with the cell count. ### Removed -* Remove multi-sample processing from all `single-cell` subcommands. -* Remove `--input1_pattern` and `--input2_pattern` from `single-cell amplicon` command. -* Self-correlations, e.g. CD8 vs CD8 are no longer part of the colocalization results, as these values will always be undefined. -* Remove `umi_unique_count` and `upi_unique_count` from `edgelist`. -* Remove `umi` and `median_umi_degree` from `component` metrics. -* Remove `normalized_rel` and `denoised` from `obsm` in `anndata`. -* Remove the `denoise` function. -* Remove cell type selector in QC report for UMAP colored by molecule count plots. -* Remove `clr` as a transformation option in `pixelator analysis`. +- Remove multi-sample processing from all `single-cell` subcommands. +- Remove `--input1_pattern` and `--input2_pattern` from `single-cell amplicon` command. +- Self-correlations, e.g. CD8 vs CD8 are no longer part of the colocalization results, as these values will always be undefined. +- Remove `umi_unique_count` and `upi_unique_count` from `edgelist`. +- Remove `umi` and `median_umi_degree` from `component` metrics. +- Remove `normalized_rel` and `denoised` from `obsm` in `anndata`. +- Remove the `denoise` function. +- Remove cell type selector in QC report for UMAP colored by molecule count plots. +- Remove `clr` as a transformation option in `pixelator analysis`. ## [0.16.2] - 2024-03-19 ### Fixed -* Uninitialized value for `--polarization-n-permutations` +- Uninitialized value for `--polarization-n-permutations` ## [0.16.1] - 2024-01-12 ### Fixed -* Bug in README shield formatting - +- Bug in README shield formatting ## [0.16.0] - 2024-01-12 This release introduces two major change in pixelator: - 1) the Graph backend has been switched from using igraph to using networkx - 2) the license has been changed from GLP2.0 to MIT + +1. the Graph backend has been switched from using igraph to using networkx +2. the license has been changed from GLP2.0 to MIT ### Added -* Experimental 3D heatmap plotting feature. -* Optional caching of layouts to speed up computations in some scenarios. -* `experimental` mark that can be added to functions that are not yet production ready. -* The underlying graph instance e.g. a networkx `Graph` instance is exposed as a property called `raw` from the pixelator `Graph` class. -* Monte Carlo permutations supported when calculating Moran's I (`morans_z_sim`) in `polarization_scores`. +- Experimental 3D heatmap plotting feature. +- Optional caching of layouts to speed up computations in some scenarios. +- `experimental` mark that can be added to functions that are not yet production ready. +- The underlying graph instance e.g. a networkx `Graph` instance is exposed as a property called `raw` from the pixelator `Graph` class. +- Monte Carlo permutations supported when calculating Moran's I (`morans_z_sim`) in `polarization_scores`. ### Changed -* The default (and only) graph backend in pixelator is now based on networkx. -* `mean_reads` and `median_reads` in adata.obs to `mean_reads_per_molecule` and `median_reads_per_molecule` respectively. -* Drop support for python 3.8 and 3.9. -* Change output format of `collapse` from csv to parquet. -* Change input and output format of `graph` from csv to parquet. -* Change input format of `annotate` from csv to parquet. -* Rename the report to "qc report" -* Add a Reads per Molecule frequency figure to the sequencing section of the qc report. -* Remove placeholder warning of missing data for not yet implemented features. -* Change "Median antibody molecules per cell" to "Average antibody molecules per cell" in the qc report. -* Refactoring of the graph backend implementations module. -* Speeding up the `amplicon` step by roughly 3x. +- The default (and only) graph backend in pixelator is now based on networkx. +- `mean_reads` and `median_reads` in adata.obs to `mean_reads_per_molecule` and `median_reads_per_molecule` respectively. +- Drop support for python 3.8 and 3.9. +- Change output format of `collapse` from csv to parquet. +- Change input and output format of `graph` from csv to parquet. +- Change input format of `annotate` from csv to parquet. +- Rename the report to "qc report" +- Add a Reads per Molecule frequency figure to the sequencing section of the qc report. +- Remove placeholder warning of missing data for not yet implemented features. +- Change "Median antibody molecules per cell" to "Average antibody molecules per cell" in the qc report. +- Refactoring of the graph backend implementations module. +- Speeding up the `amplicon` step by roughly 3x. ### Fixed -* Nicer error messages when there are no components valid for computing colocalization. -* A bunch of warnings. +- Nicer error messages when there are no components valid for computing colocalization. +- A bunch of warnings. ### Removed -* `graph` no longer outputs the raw edge list. -* igraph has been dropped as a graph backend for pixelator. - +- `graph` no longer outputs the raw edge list. +- igraph has been dropped as a graph backend for pixelator. ## [0.15.2] - 2023-10-23 ### Fixed -* Fixed broken pixeldataset aggregation for more than two samples. -* Fixed a bug in graph generation caused by accidentally writing the index to the parquet file. - For backwards compatibility, if there is a column named `index` in the edgelist, this - will be removed and the user will get a warning indicating that this has happened. - +- Fixed broken pixeldataset aggregation for more than two samples. +- Fixed a bug in graph generation caused by accidentally writing the index to the parquet file. + For backwards compatibility, if there is a column named `index` in the edgelist, this + will be removed and the user will get a warning indicating that this has happened. ## [0.15.1] - 2023-10-18 ### Fixed -* Fixed a bug in filtering pixeldataset causing it to return the wrong types. -* Fixed a bug in graph layout generation due to incorrect data frame concatenation. - +- Fixed a bug in filtering pixeldataset causing it to return the wrong types. +- Fixed a bug in graph layout generation due to incorrect data frame concatenation. ## [0.15.0] - 2023-10-16 ### Added -* Add support for Python 3.11. -* Add early enablement work for a networkx backend for the graph stage. +- Add support for Python 3.11. +- Add early enablement work for a networkx backend for the graph stage. ### Fixed -* Fix report color axis in report figures not updating when selecting markers or cell types. -* Remove placeholder links in report tooltips. -* Fix a bug where aggregating data did not add the correct sample, and unique component columns. - +- Fix report color axis in report figures not updating when selecting markers or cell types. +- Remove placeholder links in report tooltips. +- Fix a bug where aggregating data did not add the correct sample, and unique component columns. ## [0.14.0] - 2023-10-05 ### Added -* Lazy option for edge list loading (`pixeldataset.edgelist_lazy`), which returns a - `polars` `LazyFrame` that can be used to operate on the edge list without reading - all of it into memory. -* Option (`ignore_edgelists`) to skip the edge lists when aggregating files. This defaults - to `False`. +- Lazy option for edge list loading (`pixeldataset.edgelist_lazy`), which returns a + `polars` `LazyFrame` that can be used to operate on the edge list without reading + all of it into memory. +- Option (`ignore_edgelists`) to skip the edge lists when aggregating files. This defaults + to `False`. ### Changed -* Types on the edge list in memory will utilize the `pandas` `category` type for string, and - `uint16` for numeric values to lower the memory consumption when working with the - edge list -* Remove `--pbs1` and `--pbs2` commandline arguments to `pixelator single-cell adapterqc`. -* Restructure report figures. -* Improve metric names and tooltips in the report. -* Synchronize zoom level between the scatter plots in cell annotations section of the report. -* Add report placeholder for missing cell annotation data -* Add `Fraction of discarded UMIs` and `Avg. Reads per Molecule` metrics to the report. +- Types on the edge list in memory will utilize the `pandas` `category` type for string, and + `uint16` for numeric values to lower the memory consumption when working with the + edge list +- Remove `--pbs1` and `--pbs2` commandline arguments to `pixelator single-cell adapterqc`. +- Restructure report figures. +- Improve metric names and tooltips in the report. +- Synchronize zoom level between the scatter plots in cell annotations section of the report. +- Add report placeholder for missing cell annotation data +- Add `Fraction of discarded UMIs` and `Avg. Reads per Molecule` metrics to the report. ### Fixed -* Fix an issue where pixelator --version would return 0.0.0 when installing in editable mode. - +- Fix an issue where pixelator --version would return 0.0.0 when installing in editable mode. ## [0.13.1] - 2023-09-15 @@ -200,21 +228,20 @@ This release introduces two major change in pixelator: ### Changed -* Unpin igraph dependency to allow for newer versions of igraph to be used. -* Cleanup README and point to the external documentation site. -* Change PyPi package name to pixelgen-pixelator. +- Unpin igraph dependency to allow for newer versions of igraph to be used. +- Cleanup README and point to the external documentation site. +- Change PyPi package name to pixelgen-pixelator. ### Fixed -* Fix an issue where `--keep-workdirs` option for pytest was not available when running pytest without - restricting the testdir to `tests/integration`. -* Fix an issue where pixelator --version would return 0.0.0. +- Fix an issue where `--keep-workdirs` option for pytest was not available when running pytest without + restricting the testdir to `tests/integration`. +- Fix an issue where pixelator --version would return 0.0.0. ### Removed -* `clr` and `relative` transformation options for the colocalization computations in `analysis` - +- `clr` and `relative` transformation options for the colocalization computations in `analysis` ## [0.13.0] - 2023-09-13 -* First public release of pixelator. +- First public release of pixelator. diff --git a/CITATIONS.md b/CITATIONS.md index fe12af2d..9f77e8f4 100644 --- a/CITATIONS.md +++ b/CITATIONS.md @@ -25,3 +25,7 @@ - [local G](https://doi.org/10.1007/s11749-018-0599-x) > Bivand, R.S., Wong, D.W.S. Comparing implementations of global and local indicators of spatial association. TEST 27, 716–748 (2018). https://doi.org/10.1007/s11749-018-0599-x + +- [dsb](https://doi.org/10.1038/s41467-022-29356-8) + + > Mulè, M.P., Martins, A.J. & Tsang, J.S. Normalizing and denoising protein expression data from droplet-based single cell profiling. Nat Commun 13, 2099 (2022). https://doi.org/10.1038/s41467-022-29356-8 diff --git a/poetry.lock b/poetry.lock index 1ac78bed..82a905bd 100644 --- a/poetry.lock +++ b/poetry.lock @@ -1,4 +1,4 @@ -# This file is automatically @generated by Poetry 1.8.2 and should not be changed by hand. +# This file is automatically @generated by Poetry 1.7.1 and should not be changed by hand. [[package]] name = "alabaster" @@ -13,13 +13,13 @@ files = [ [[package]] name = "anndata" -version = "0.10.7" +version = "0.10.8" description = "Annotated data." optional = false python-versions = ">=3.9" files = [ - {file = "anndata-0.10.7-py3-none-any.whl", hash = "sha256:8b50af790f9a73414d0b94fd7b8e005f5cc6c53c33eb09206d2bd6720115bffc"}, - {file = "anndata-0.10.7.tar.gz", hash = "sha256:9b186bea18956fefdc35e7464d822a17e9faa456ca2d20c7a0966a4837b822a2"}, + {file = "anndata-0.10.8-py3-none-any.whl", hash = "sha256:1b24934dc2674eaf3072cb7010e187aa2b2f4f0e4cf0a32ffeab5ffebe3b1415"}, + {file = "anndata-0.10.8.tar.gz", hash = "sha256:b728a33225eeaaefddf6bed546d935c0f06881c9166621b24de3b492b2f406bb"}, ] [package.dependencies] @@ -36,17 +36,17 @@ scipy = ">1.8" dev = ["pytest-xdist", "setuptools-scm"] doc = ["awkward (>=2.0.7)", "ipython", "myst-parser", "nbsphinx", "readthedocs-sphinx-search", "scanpydoc[theme,typehints] (>=0.13.4)", "sphinx (>=4.4)", "sphinx-autodoc-typehints (>=1.11.0)", 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a/src/pixelator/analysis/__init__.py +++ b/src/pixelator/analysis/__init__.py @@ -33,6 +33,7 @@ class AnalysisParameters: colocalization_neighbourhood_size: int colocalization_n_permutations: int colocalization_min_region_count: int + colocalization_min_marker_count: int def analyse_pixels( diff --git a/src/pixelator/analysis/colocalization/__init__.py b/src/pixelator/analysis/colocalization/__init__.py index 241427b4..09c59615 100644 --- a/src/pixelator/analysis/colocalization/__init__.py +++ b/src/pixelator/analysis/colocalization/__init__.py @@ -51,6 +51,7 @@ def colocalization_from_component_edgelist( n_permutations: int = 50, use_full_bipartite: bool = True, min_region_count: int = 5, + min_marker_count: int = 5, random_seed: Optional[int] = None, ) -> pd.DataFrame: """Get the colocalization scores for the component in the given `edgelist`. @@ -65,6 +66,8 @@ def colocalization_from_component_edgelist( projection, defaults to True :param min_region_count: minimum number of counts in region to consider, defaults to 5 + :param min_marker_count: the minimum number of counts of a marker to calculate + colocalization :param random_seed: Set the random seed for the permutation tests, defaults to None :return: a dataframe with computed colocalization scores :rtype: pd.DataFrame @@ -84,6 +87,7 @@ def colocalization_from_component_edgelist( neighbourhood_size=neighbourhood_size, n_permutations=n_permutations, min_region_count=min_region_count, + min_marker_count=min_marker_count, random_seed=random_seed, ) @@ -109,6 +113,7 @@ def colocalization_from_component_graph( neighbourhood_size: int = 1, n_permutations: int = 50, min_region_count: int = 5, + min_marker_count: int = 5, random_seed: Optional[int] = None, ) -> pd.DataFrame: """Compute the colocalization scores for this component graph. @@ -121,20 +126,29 @@ def colocalization_from_component_graph( p-values and z-scores, defaults to 50 :param min_region_count: minimum number of counts in region to consider, defaults to 5 + :param min_marker_count: the minimum number of counts of a marker to calculate + colocalization :param random_seed: Set the random seed for the permutation tests, defaults to None :return: a dataframe containing colocalization scores for this component :rtype: pd.DataFrame """ logger.debug("Computing colocalization for component: %s", component_id) logger.debug("Prepare the graph data for computing colocalization") - marker_counts_by_region = prepare_from_graph(graph, n_neighbours=neighbourhood_size) + raw_marker_counts = graph.node_marker_counts + # Record markers to keep + # Remove markers with zero variance and markers below minimum marker count + markers_to_keep = raw_marker_counts.columns[ + (raw_marker_counts != 0).any(axis=0) + & (raw_marker_counts.nunique() > 1) + & (raw_marker_counts.sum() >= min_marker_count) + ] + + marker_counts_by_region = prepare_from_graph(graph, n_neighbours=neighbourhood_size) + marker_counts_by_region = marker_counts_by_region[markers_to_keep] marker_counts_by_region = filter_by_region_counts( marker_counts_by_region, min_region_counts=min_region_count ) - marker_counts_by_region = filter_by_unique_values( - marker_counts_by_region, at_least_n_unique=2 - ) nrow, ncols = marker_counts_by_region.shape if nrow == 0: @@ -187,6 +201,7 @@ def colocalization_scores( neighbourhood_size: int = 1, n_permutations: int = 50, min_region_count: int = 5, + min_marker_count: int = 5, random_seed: Optional[int] = None, ) -> pd.DataFrame: """Compute colocalization scores for antibody pairs. @@ -225,6 +240,8 @@ def colocalization_scores( :param min_region_count: The minimum size of the region (e.g. number of counts in the neighbourhood) required for it to be considered + :param min_marker_count: the minimum number of counts of a marker to calculate + colocalization :param random_seed: Set a random seed for the permutation function :returns: a pd.DataFrame of scores :rtype: pd.DataFrame @@ -273,6 +290,7 @@ def data(): neighbourhood_size=neighbourhood_size, n_permutations=n_permutations, min_region_count=min_region_count, + min_marker_count=min_marker_count, random_seed=random_seed, ) @@ -402,6 +420,7 @@ def __init__( neighbourhood_size: int, n_permutations: int, min_region_count: int, + min_marker_count: int, ): """Initialize the ColocalizationAnalysis. @@ -413,11 +432,14 @@ def __init__( :param min_region_count: The minimum size of the region (e.g. number of counts in the neighbourhood) required for it to be considered for colocalization analysis + :param min_marker_count: the minimum number of counts of a marker to calculate + colocalization """ self.transformation_type = transformation_type self.neighbourhood_size = neighbourhood_size self.n_permutations = n_permutations self.min_region_count = min_region_count + self.min_marker_count = min_marker_count def run_on_component(self, component: Graph, component_id: str) -> pd.DataFrame: """Run colocalization analysis on the component.""" @@ -429,6 +451,7 @@ def run_on_component(self, component: Graph, component_id: str) -> pd.DataFrame: neighbourhood_size=self.neighbourhood_size, n_permutations=self.n_permutations, min_region_count=self.min_region_count, + min_marker_count=self.min_marker_count, ) def post_process_data(self, data: pd.DataFrame) -> pd.DataFrame: diff --git a/src/pixelator/analysis/normalization/__init__.py b/src/pixelator/analysis/normalization/__init__.py new file mode 100644 index 00000000..c4592373 --- /dev/null +++ b/src/pixelator/analysis/normalization/__init__.py @@ -0,0 +1,66 @@ +"""Functions for the colocalization analysis in pixelator. + +Copyright © 2024 Pixelgen Technologies AB. +""" + +from typing import List, Union + +import numpy as np +import pandas as pd +from sklearn.decomposition import PCA +from sklearn.mixture import GaussianMixture +from sklearn.preprocessing import StandardScaler + + +def _regress_out_confounder(pheno, exprs, rcond=1e-8): + """Linear regression to remove confounding factors from abundance data.""" + design_matrix = np.column_stack((np.ones((len(pheno), 1)), pheno)) + coefficients, res, rank, s = np.linalg.lstsq(design_matrix, exprs, rcond=rcond) + beta = coefficients[1:] # remove intercept term + return exprs - design_matrix[:, 1:].dot(beta) + + +def _get_background_abundance(dataframe: pd.DataFrame, axis=0): + """Fit a double gaussian distribution to the abundance data and return the mean of the first gaussian as an estimation of the background level.""" + background = pd.Series(index=dataframe.index if axis == 0 else dataframe.columns) + scores = pd.Series(index=dataframe.index if axis == 0 else dataframe.columns) + gmm = GaussianMixture(n_components=2, max_iter=1000, random_state=0) + if axis not in {0, 1}: + raise ValueError(f"Axis was {axis}. Must be 0 or 1") + ax_iter = dataframe.index if axis == 0 else dataframe.columns + for i in ax_iter: + current_axis = dataframe.loc[i, :] if axis == 0 else dataframe.loc[:, i] + gmm = gmm.fit(current_axis.to_frame()) + background[i] = np.min(gmm.means_) + scores[i] = np.abs(gmm.means_[1] - gmm.means_[0]) / np.sum(gmm.covariances_) + return background, scores + + +def dsb_normalize( + raw_abundance: pd.DataFrame, isotype_controls: Union[List, None] = None +): + """empty-droplet-free method as implemented in Mulè et. al. dsb package. + + The normalization steps are: 1- log1p transformation, 2- remove background + abundance per marker, 3- regularize abundance per component. + + :param raw_abundance: the raw abundance count data. + :param isotype_controls: list of isotype controls. + :return: normalized abundance data. + """ + log_abundance = np.log1p(raw_abundance) + marker_background, _ = _get_background_abundance(log_abundance, axis=1) + log_abundance = log_abundance - marker_background + component_background, _ = _get_background_abundance(log_abundance, axis=0) + + if isotype_controls is not None: + control_signals = log_abundance.loc[:, isotype_controls] + control_signals["component_background"] = component_background + control_signals = StandardScaler().fit_transform(control_signals) + pheno = PCA(n_components=1).fit_transform(control_signals) + else: + raise ValueError(f"At least one isotype control must be provided.") + + normalized_abundance = _regress_out_confounder(pheno, log_abundance) + + return normalized_abundance diff --git a/src/pixelator/cli/analysis.py b/src/pixelator/cli/analysis.py index e607c8de..f8a6b9b2 100644 --- a/src/pixelator/cli/analysis.py +++ b/src/pixelator/cli/analysis.py @@ -92,7 +92,7 @@ ) @click.option( "--colocalization-transformation", - default="log1p", + default="rate-diff", required=False, type=click.Choice(get_args(TransformationTypes)), show_default=True, @@ -134,6 +134,14 @@ "valid for computing colocalization" ), ) +@click.option( + "--colocalization-min-marker-count", + default=5, + required=False, + type=click.IntRange(min=0), + show_default=True, + help=("The minimum number of marker counts in component for colocalization"), +) @output_option @click.pass_context @timer @@ -150,6 +158,7 @@ def analysis( colocalization_neighbourhood_size, colocalization_n_permutations, colocalization_min_region_count, + colocalization_min_marker_count, output, ): """ @@ -171,6 +180,7 @@ def analysis( colocalization_neighbourhood_size=colocalization_neighbourhood_size, colocalization_n_permutations=colocalization_n_permutations, colocalization_min_region_count=colocalization_min_region_count, + colocalization_min_marker_count=colocalization_min_marker_count, ) # some basic sanity check on the input files @@ -214,6 +224,7 @@ def analysis( neighbourhood_size=colocalization_neighbourhood_size, n_permutations=colocalization_n_permutations, min_region_count=colocalization_min_region_count, + min_marker_count=colocalization_min_marker_count, ) ) diff --git a/src/pixelator/cli/collapse.py b/src/pixelator/cli/collapse.py index 1c0138e5..7cd91258 100644 --- a/src/pixelator/cli/collapse.py +++ b/src/pixelator/cli/collapse.py @@ -3,6 +3,7 @@ Copyright © 2022 Pixelgen Technologies AB. """ +import sys from collections import defaultdict from concurrent import futures from pathlib import Path @@ -25,6 +26,38 @@ ) +def _handle_errors(jobs, executor): + for job in jobs: + exception = job.exception() + if exception is None: + continue + + logger.error( + "Found an issue in the process pool. Trying to determine what went wrong and set the correct exit code. Exception was: %s", + exception, + ) + process_map = executor._processes + for pid in process_map.keys(): + exit_code = process_map[pid].exitcode + if exit_code is not None and exit_code != 0: + logger.error( + "The child process in the process pool returned a non-zero exit code: %s.", + exit_code, + ) + # If we have an out of memory exception, make sure we exit with that. + if abs(exit_code) == 9: + logger.error( + "One of the child processes was killed (exit code: 9). " + "Usually this is caused by the out-of-memory killer terminating the process. " + "The parent process will return an exit code of 137 to indicate that it terminated because of a kill signal in the child process." + ) + sys.exit(137) + logger.error( + "Was unable to determine what when wrong in process pool. Will raise original exception." + ) + raise exception + + @click.command( "collapse", short_help=( @@ -238,12 +271,12 @@ def collapse( min_count=min_count, ) ) + jobs = list(futures.as_completed(jobs)) + _handle_errors(jobs, executor) total_input_reads = 0 tmp_files = [] - for job in futures.as_completed(jobs): - if job.exception() is not None: - raise job.exception() + for job in jobs: # the worker returns a path to a file (temp antibody edge list) tmp_file, input_reads_count = job.result() if tmp_file is not None: diff --git a/src/pixelator/cli/layout.py b/src/pixelator/cli/layout.py index c0a4e595..faba6f90 100644 --- a/src/pixelator/cli/layout.py +++ b/src/pixelator/cli/layout.py @@ -48,7 +48,7 @@ "--layout-algorithm", required=False, multiple=True, - default=["pmds_3d"], + default=["wpmds_3d"], help="Select a layout algorithm to use. This can be specified multiple times to compute multiple layouts. Default: pmds_3d", type=click.Choice(get_args(SupportedLayoutAlgorithm)), ) diff --git a/src/pixelator/cli/misc.py b/src/pixelator/cli/misc.py index 09867cea..66a6476c 100644 --- a/src/pixelator/cli/misc.py +++ b/src/pixelator/cli/misc.py @@ -41,7 +41,7 @@ def list_single_cell_panels(ctx: click.Context, param: Any, value: Any) -> None: if not value or ctx.resilient_parsing: return - options = list(config.panels.keys()) + options = config.list_panel_names(include_aliases=True) for option in options: click_echo(option) diff --git a/src/pixelator/config/config_class.py b/src/pixelator/config/config_class.py index 0a7388a9..837c0766 100644 --- a/src/pixelator/config/config_class.py +++ b/src/pixelator/config/config_class.py @@ -7,6 +7,7 @@ import itertools import typing +import warnings from collections import defaultdict from pathlib import Path from typing import Dict, List, Optional, Tuple @@ -16,6 +17,7 @@ from pixelator.config.assay import Assay from pixelator.config.panel import AntibodyPanel +from pixelator.exceptions import PixelatorBaseException from pixelator.types import PathType DNA_CHARS = {"A", "C", "G", "T"} @@ -25,6 +27,12 @@ ) +class PanelException(PixelatorBaseException): + """Exception raised for failures to load a panel into the global configuration.""" + + pass + + class Config: """Class containing the pixelator configuration (assay settings).""" @@ -36,6 +44,7 @@ def __init__( """Initialize the config object.""" self.assays: Dict[str, Assay] = {} self.panels: typing.MutableMapping[str, List[AntibodyPanel]] = defaultdict(list) + self.panel_aliases: Dict[str, str] = {} if assays is not None: self.assays.update({a.name: a for a in assays}) @@ -51,11 +60,29 @@ def load_assay(self, path: PathType) -> None: self.assays[assay.name] = assay def load_panel_file(self, path: PathType) -> None: - """Load the panel file.""" + """Load the panel file. + + :param path: The path to the panel file. + :raises PanelException: If the panel alias already exists in the config. + """ panel = AntibodyPanel.from_csv(path) key = panel.name if panel.name is not None else str(panel.filename) self.panels[key].append(panel) + # Enable panel lookup by aliases + for alias in panel.aliases: + if (alias in self.panel_aliases) and (key != self.panel_aliases[alias]): + raise PanelException( + f'Panel alias "{alias}" already exists in the ' + f'config for panel "{self.panel_aliases[alias]}".' + "If you provided your own panel file, please " + "ensure the panel name an aliases are unique " + "in the header of the file." + ) + continue + + self.panel_aliases[alias] = key + def load_assays(self, path: PathType): """Load all assays from a directory containing yaml files.""" search_path = Path(path) @@ -83,11 +110,40 @@ def get_assay(self, assay_name: str) -> Optional[Assay]: """Get an assay by name.""" return self.assays.get(assay_name) + def list_panel_names(self, include_aliases: bool = False) -> List[str]: + """Return a list of all panel names. + + :param include_aliases: Include panel aliases in the list + :returns: A list of panel names + """ + out = sorted(list(self.panels.keys())) + + if not include_aliases: + return out + + out += sorted(list(self.panel_aliases.keys())) + return out + def get_panel( - self, panel_name: str, version: Optional[str] = None + self, + panel_name: str, + version: Optional[str] = None, + allow_aliases: bool = True, ) -> Optional[AntibodyPanel]: - """Get a panel by name.""" + """Get a panel by name. + + :param panel_name: The name of the panel + :param version: The optional version of a panel to return + :param allow_aliases: Allow panel aliases to be used + """ panels_with_key = self.panels.get(panel_name) + + # Try to load using an alias if no name matches are found + if panels_with_key is None and allow_aliases: + panel_alias = self.panel_aliases.get(panel_name) + if panel_alias is not None: + panels_with_key = self.panels.get(panel_alias) + if panels_with_key is None: return None diff --git a/src/pixelator/config/panel.py b/src/pixelator/config/panel.py index 0382c60b..b8c9cf1c 100644 --- a/src/pixelator/config/panel.py +++ b/src/pixelator/config/panel.py @@ -29,6 +29,7 @@ class AntibodyPanelMetadata(pydantic.BaseModel): version: Optional[str] = None name: Optional[str] = None description: Optional[str] = None + aliases: List[str] = [] class AntibodyPanel: @@ -118,6 +119,11 @@ def description(self) -> Optional[str]: """Return the panel file description.""" return self._metadata.description + @property + def aliases(self) -> list[str]: + """Return the (optional) list of panel file aliases.""" + return self._metadata.aliases + @classmethod def validate_antibody_panel(self, panel_df: pd.DataFrame) -> list[str]: """Perform validation on an antibody panel file. diff --git a/src/pixelator/graph/backends/implementations/_networkx.py b/src/pixelator/graph/backends/implementations/_networkx.py index 430e2889..ce1baa58 100644 --- a/src/pixelator/graph/backends/implementations/_networkx.py +++ b/src/pixelator/graph/backends/implementations/_networkx.py @@ -320,7 +320,7 @@ def clusters(leiden_communities): def _layout_coordinates( self, - layout_algorithm: SupportedLayoutAlgorithm = "pmds_3d", + layout_algorithm: SupportedLayoutAlgorithm = "wpmds_3d", random_seed: Optional[int] = None, **kwargs, ) -> pd.DataFrame: @@ -352,6 +352,10 @@ def _layout_coordinates( layout_inst = pmds_layout(raw, seed=random_seed, **kwargs) if layout_algorithm == "pmds_3d": layout_inst = pmds_layout(raw, dim=3, seed=random_seed, **kwargs) + if layout_algorithm == "wpmds_3d": + layout_inst = pmds_layout( + raw, dim=3, weights="prob_dist", seed=random_seed, **kwargs + ) coordinates = pd.DataFrame.from_dict( layout_inst, @@ -396,7 +400,7 @@ def node_marker_counts(self) -> pd.DataFrame: def layout_coordinates( self, - layout_algorithm: SupportedLayoutAlgorithm = "pmds_3d", + layout_algorithm: SupportedLayoutAlgorithm = "wpmds_3d", only_keep_a_pixels: bool = True, get_node_marker_matrix: bool = True, random_seed: Optional[int] = None, @@ -414,9 +418,14 @@ def layout_coordinates( - fruchterman_reingold_3d - kamada_kawai - kamada_kawai_3d + - wpmds_3d - The `pmds` options are much (~10-100x) faster than the `fruchterman_reingold` and - the `kamada_kawai`. + For most cases the `pmds` options should be about 10-100x faster + than the force directed layout methods, i.e. `fruchterman_reingold` + and `kamada_kawai`. Among the force directed layout methods, + `fruchterman_reingold` is generally faster than `kamada_kawai`. The + `wpmds_3d` method uses edge weights to improve the layout, but is slightly + slower than `pmds_3d`. :param layout_algorithm: the layout algorithm to use to generate the coordinates :param only_keep_a_pixels: If true, only keep the a-pixels @@ -739,33 +748,50 @@ def subgraphs(self) -> Iterable[Graph]: def pmds_layout( - g: nx.Graph, pivots: int = 50, dim: int = 2, seed: Optional[int] = None + g: nx.Graph, + pivots: int = 200, + dim: int = 2, + weights: Optional[Union[np.ndarray, str]] = None, + seed: Optional[int] = None, ) -> pd.DataFrame: """Calculate a pivot MDS layout for a graph as described in [1]_. - The algorithm is similar to classical multidimensional scaling (MDS), but uses only - a smalls set of random pivot nodes. The algorithm is considerably faster than MDS - and therefore scales better to large graphs. The topology of resulting layouts are - deterministic for a given seed, but may be mirrored across different systems due to - variations in floating-point precision. + The algorithm is similar to classical multidimensional scaling (MDS), + but uses only a smalls set of random pivot nodes. The algorithm is + considerably faster than MDS and therefore scales better to large graphs. + The topology of resulting layouts are deterministic for a given seed, + but may be mirrored across different systems due to variations in + floating-point precision. - .. [1] Brandes U, Pich C. Eigensolver Methods for Progressive Multidimensional - Scaling of Large Data. International Symposium on Graph Drawing, 2007. - Lecture Notes in Computer Science, vol 4372. doi: 10.1007/978-3-540-70904-6_6. + .. [1] Brandes U, Pich C. Eigensolver Methods for Progressive + Multidimensional Scaling of Large Data. International Symposium + on Graph Drawing, 2007. Lecture Notes in Computer Science, vol + 4372. doi: 10.1007/978-3-540-70904-6_6. :param g: A networkx graph object :param pivots: The number of pivot nodes to use :param dim: The dimension of the layout + :param weights: Edge weights to use for the layout computation. + Options are: + + * an np.array with non-negative values (same number of elements as edges in g) + * "prob_dist" to use -log(P)^3, where P is the probability of a random walker to traverse the end nodes of and edge (i->j) and back again (j->i) in 5 steps. + * None to use unweighted shortest path lengths :param seed: Set seed for reproducibility :return: A dataframe with layout coordinates :rtype: pd.DataFrame - :raises: ValueError raises if conditions are not met + :raises ValueError: if conditions are not met """ if not nx.is_connected(g): raise ValueError("Only connected graphs are supported.") if pivots >= len(g.nodes): - raise ValueError("'pivots' must be less than the number of nodes in the graph.") + total_nodes = len(g.nodes) + warnings.warn( + f"'pivots' ({pivots}) should be less than the number of " + f"nodes ({total_nodes}) in the graph. Using all nodes as 'pivots'." + ) + pivots = total_nodes if dim not in [2, 3]: raise ValueError("'dim' must be either 2 or 3.") @@ -779,6 +805,28 @@ def pmds_layout( f"'pivots' must be greater than or equal to {pivot_lower_bound}" ) + if isinstance(weights, str): + if weights != "prob_dist": + raise ValueError("If 'weights' is a string, it must be 'prob_dist'.") + weights = -(np.log(_prob_edge_weights(g, k=5)) ** 3) + elif isinstance(weights, np.ndarray): + if len(weights) != len(g.edges): + raise ValueError( + "'weights' must have the same length as the number of edges in the graph." + ) + if np.any(weights < 0): + raise ValueError("All elements in 'weights' must be non-negative.") + elif weights is not None: + raise ValueError("'weights' must be a string or an array.") + + if weights is not None: + edges = list(g.edges) + edge_weight_dict = {edges[i]: weights[i] for i in range(len(edges))} + nx.set_edge_attributes(g, edge_weight_dict, "weight") + weight = "weight" + else: + weight = None + if seed is not None: np.random.seed(seed) @@ -788,7 +836,7 @@ def pmds_layout( pivs = np.random.choice(node_list, pivots, replace=False) # Calculate the shortest path length from the pivots to all other nodes - A = nx.to_scipy_sparse_array(g, weight=None, nodelist=node_list, format="csr") + A = nx.to_scipy_sparse_array(g, weight=weight, nodelist=node_list, format="csr") # This is a workaround for what seems to be a bug in the type of # the indices of the sparse array created above @@ -797,15 +845,16 @@ def pmds_layout( D = sp.sparse.csgraph.shortest_path( A, directed=False, - unweighted=True, + unweighted=weight is None, method="D", indices=np.where(np.isin(g.nodes, pivs))[0], ).T # Center values in rows and columns - cmean = np.mean(D**2, axis=0) - rmean = np.mean(D**2, axis=1) - D_pivs_centered = D**2 - np.add.outer(rmean, cmean) + np.mean(D**2) + D2 = D**2 + cmean = np.mean(D2, axis=0) + rmean = np.mean(D2, axis=1) + D_pivs_centered = D2 - np.add.outer(rmean, cmean) + np.mean(D2) # Compute SVD and use distances to compute coordinates for all nodes # in an abstract cartesian space @@ -818,3 +867,59 @@ def pmds_layout( coordinates = {node_list[i]: coordinates[i, :] for i in range(coordinates.shape[0])} return coordinates + + +def _prob_edge_weights( + g: nx.Graph, + k: int = 5, +) -> np.ndarray: + """Compute edge weights based on k-step transition probabilities. + + The transition probabilities are computed using powers of the + stochastic matrix of the graph with self-loops allowed. + + :param g: A networkx graph object + :param k: The number of steps in the random walk + :return: An array of edge weights + :rtype: np.array + :raises ValueError: if conditions are not met + """ + # Check that k is an integer between 1 and 10 + if not isinstance(k, int) and k < 1 or k > 10: + raise ValueError("'k' must be an integer between 1 and 10.") + + A = nx.to_scipy_sparse_array(g, weight=None, nodelist=list(g.nodes), format="csr") + + # Add 1 to the diagonal to allow self-loops + A = A + sp.sparse.diags([1] * A.shape[0], format="csr") + + # Divide by row sum + D = sp.sparse.diags(1 / A.sum(axis=1), format="csr") + P = D @ A + + # Compute the transition probabilities for a k-step walk + P_step = _mat_pow(P, k) + + # Keep edges from original graph + P_step = P_step.multiply(A) + + # Compute bi-directional transition probabilities which are symmetric. + # Now we get the probability of going from i to j and back again in k steps, + # so it doesn't matter if we start in i or j. + P_step_bidirectional = P_step.multiply(P_step.T) + + # Extract the transition probabilities for edges in g + edges = list(g.edges) + edge_probs = P_step_bidirectional[ + [list(g.nodes).index(u) for u, _ in edges], + [list(g.nodes).index(v) for _, v in edges], + ] + + return np.array(edge_probs)[0] + + +def _mat_pow(mat, power): + mat_power = mat + for _ in range(power - 1): + mat_power = mat @ mat_power + return mat_power diff --git a/src/pixelator/graph/backends/protocol.py b/src/pixelator/graph/backends/protocol.py index 41886998..28e898ca 100644 --- a/src/pixelator/graph/backends/protocol.py +++ b/src/pixelator/graph/backends/protocol.py @@ -34,6 +34,7 @@ "kamada_kawai_3d", "pmds", "pmds_3d", + "wpmds_3d", ] @@ -149,7 +150,7 @@ def node_marker_counts(self) -> pd.DataFrame: def layout_coordinates( self, - layout_algorithm: SupportedLayoutAlgorithm = "pmds_3d", + layout_algorithm: SupportedLayoutAlgorithm = "wpmds_3d", only_keep_a_pixels: bool = True, get_node_marker_matrix: bool = True, random_seed: Optional[int] = None, @@ -167,11 +168,14 @@ def layout_coordinates( - kamada_kawai_3d - pmds - pmds_3d + - wpmds_3d For most cases the `pmds` options should be about 10-100x faster than the force directed layout methods, i.e. `fruchterman_reingold` and `kamada_kawai`. Among the force directed layout methods, - `fruchterman_reingold` is generally faster than `kamada_kawai`. + `fruchterman_reingold` is generally faster than `kamada_kawai`. The + `wpmds_3d` method uses edge weights to improve the layout, but is slightly + slower than `pmds_3d`. :param layout_algorithm: the layout algorithm to use to generate the coordinates :param only_keep_a_pixels: If true, only keep the a-pixels diff --git a/src/pixelator/graph/graph.py b/src/pixelator/graph/graph.py index d93ea0d2..c11aa5d6 100644 --- a/src/pixelator/graph/graph.py +++ b/src/pixelator/graph/graph.py @@ -183,7 +183,7 @@ def community_leiden( def layout_coordinates( self, - layout_algorithm: SupportedLayoutAlgorithm = "pmds_3d", + layout_algorithm: SupportedLayoutAlgorithm = "wpmds_3d", only_keep_a_pixels: bool = True, get_node_marker_matrix: bool = True, cache: bool = False, @@ -202,10 +202,11 @@ def layout_coordinates( - kamada_kawai_3d - pmds - pmds_3d + - wpmds_3d - The `fruchterman_reingold` options are in general faster, but less - accurate than the `kamada_kawai` ones. + The `pmds` options are much faster than the force-directed algorithms fruchterman_reingold + and kamada_kawai. The `wpmds_3d` option is a weighted version of the `pmds_3d` algorithm. :param layout_algorithm: the layout algorithm to use to generate the coordinates :param only_keep_a_pixels: If true, only keep the a-pixels diff --git a/src/pixelator/graph/node_metrics.py b/src/pixelator/graph/node_metrics.py index d6495376..30a6e92c 100644 --- a/src/pixelator/graph/node_metrics.py +++ b/src/pixelator/graph/node_metrics.py @@ -52,7 +52,9 @@ def compute_transition_probabilities( # Set diagonal of W to 0 if k > 1 for gi to avoid self-loops W_out.setdiag(values=0) # Renormalize transition probabilities to sum to 1 - W_out = W_out / W_out.sum(axis=0)[:, None] + row_sums = np.array(W_out.sum(axis=1)).flatten() + inv_row_sums = np.reciprocal(row_sums, where=row_sums != 0) + W_out = W_out.multiply(inv_row_sums[:, np.newaxis]) return W_out @@ -63,7 +65,7 @@ def local_g( counts: pd.DataFrame, k: int = 1, use_weights: bool = True, - normalize_counts: bool = True, + normalize_counts: bool = False, W: sp.sparse.csr_array | None = None, method: Literal["gi", "gstari"] = "gi", ) -> pd.DataFrame: @@ -97,7 +99,7 @@ def local_g( :param k: The number of steps in the k-step random walk. Default is 1. :param use_weights: Whether to use weights in the computation. When turned off, all edge weights will be qeual to 1. Default is True. - :param normalize_counts: Whether to normalize counts to proportions. Default is True. + :param normalize_counts: Whether to normalize counts to proportions. Default is False. :param W: A sparse matrix of custom edge weights. This will override the automated computation of edge weights. `W` must have the same dimensions as A. Note that weights can be defined for any pair of nodes, not only the pairs represented by edges in `A`. Default is None. @@ -156,6 +158,17 @@ def local_g( W = W.T else: W = A + if k > 1: + # Expand local neighborhood using matrix powers + W = sp.sparse.linalg.matrix_power(W, k) + # Set all positive elements to 1 + W.data = np.ones_like(W.data) + if method == "gstari": + # Set diagonal of A to 1 for gstari which expects self-loops + W = W + sp.sparse.eye(n_nodes) + else: + # Set diagonal of W to 0 for gi to avoid self-loops + W.setdiag(values=0) # Compute lag matrix lag_mat = W @ counts diff --git a/src/pixelator/graph/utils.py b/src/pixelator/graph/utils.py index 5117f2ed..af2f989a 100644 --- a/src/pixelator/graph/utils.py +++ b/src/pixelator/graph/utils.py @@ -101,8 +101,8 @@ def components_metrics(edgelist: pd.DataFrame) -> pd.DataFrame: median_b_pixels_per_a_pixel = b_pixels_per_a_pixel_series.median() a_pixels_per_b_pixel = group_df.groupby("upib", observed=True)["upia"].nunique() - mean_a_pixels_per_b_pixel = b_pixels_per_a_pixel_series.mean() - median_a_pixels_per_b_pixel = b_pixels_per_a_pixel_series.median() + mean_a_pixels_per_b_pixel = a_pixels_per_b_pixel.mean() + median_a_pixels_per_b_pixel = a_pixels_per_b_pixel.median() # Same reasoning as above molecule_count_per_a_pixel_series = group_df.groupby("upia", observed=True)[ diff --git a/src/pixelator/logging.py b/src/pixelator/logging.py index cd8ec403..bdea09d3 100644 --- a/src/pixelator/logging.py +++ b/src/pixelator/logging.py @@ -214,6 +214,11 @@ def log_exception(exc_type, exc_value, traceback_obj): # and thus we ignore them here. return False + if issubclass(exc_type, SystemExit): + # SystemExit is raised when the application has been explicitly + # directed to exit, so we don't what a trace dumped for that. + return False + self._root_logger.critical( "Unhandled exception of type: {}".format(exc_type.__name__) ) diff --git a/src/pixelator/pixeldataset/precomputed_layouts.py b/src/pixelator/pixeldataset/precomputed_layouts.py index d512ee68..a9e99875 100644 --- a/src/pixelator/pixeldataset/precomputed_layouts.py +++ b/src/pixelator/pixeldataset/precomputed_layouts.py @@ -212,6 +212,8 @@ def __init__( self._data_provider: _DataProvider = _EmptyDataProvider() elif isinstance(layouts_lazy, pl.LazyFrame): self._data_provider = _SingleFrameDataProvider(layouts_lazy) + elif isinstance(layouts_lazy, pl.DataFrame): + self._data_provider = _SingleFrameDataProvider(layouts_lazy.lazy()) elif isinstance(layouts_lazy, Iterable): self._data_provider = _MultiFrameDataProvider(layouts_lazy) else: @@ -363,7 +365,7 @@ def data(): try: return PreComputedLayouts( - pl.concat(data(), rechunk=False), + pl.concat(data(), rechunk=True).collect(), partitioning=["sample"] + PreComputedLayouts.DEFAULT_PARTITIONING, ) except ValueError: @@ -474,7 +476,7 @@ def generate_precomputed_layouts_for_components( components: set[str] | None = None, add_node_marker_counts: bool = True, layout_algorithms: SupportedLayoutAlgorithm - | list[SupportedLayoutAlgorithm] = "pmds_3d", + | list[SupportedLayoutAlgorithm] = "wpmds_3d", ) -> PreComputedLayouts: """Generate precomputed layouts for the components in the PixelDataset. diff --git a/src/pixelator/plot/__init__.py b/src/pixelator/plot/__init__.py index 44f14e78..ca7f048d 100644 --- a/src/pixelator/plot/__init__.py +++ b/src/pixelator/plot/__init__.py @@ -4,57 +4,32 @@ """ import warnings -from typing import Literal, Optional, Tuple, Union +from typing import Optional, Tuple +import anndata import matplotlib.pyplot as plt -import networkx as nx import numpy as np import pandas as pd -import plotly.graph_objects as go -import polars as pl -import scipy import seaborn as sns -from matplotlib import cm -from matplotlib.colors import Normalize +from matplotlib.colors import LinearSegmentedColormap +from matplotlib.patches import Rectangle +from scipy.stats import gaussian_kde -from pixelator.analysis.colocalization import get_differential_colocalization -from pixelator.graph import Graph -from pixelator.marks import experimental -from pixelator.pixeldataset import PixelDataset from pixelator.plot.constants import Color +from pixelator.plot.layout_plots import ( + plot_2d_graph, + plot_3d_from_coordinates, + plot_3d_graph, + plot_3d_heatmap, +) +from pixelator.plot.spatial_analysis_plots import ( + plot_colocalization_diff_heatmap, + plot_colocalization_diff_volcano, + plot_colocalization_heatmap, +) sns.set_style("whitegrid") - - -def _unit_sphere_surface(horizontal_resolution, vertical_resolution): - horizontal_angles = np.linspace(0, 2 * np.pi, horizontal_resolution) - vertical_angles = np.linspace(0, np.pi, vertical_resolution) - - X = np.outer(np.cos(horizontal_angles), np.sin(vertical_angles)) - Y = np.outer(np.sin(horizontal_angles), np.sin(vertical_angles)) - Z = np.outer(np.ones(np.size(horizontal_angles)), np.cos(vertical_angles)) - return X, Y, Z - - -def _calculate_distance_to_unit_sphere_zones(coordinates, unit_sphere_surface): - X, Y, Z = unit_sphere_surface - zones_on_sphere_surface = np.stack([X, Y, Z], axis=2).reshape(-1, 3) - return scipy.spatial.distance.cdist( - zones_on_sphere_surface, coordinates, "euclidean" - ) - - -def _calculate_densities(coordinates, distance_cutoff, unit_sphere_surface): - dist = _calculate_distance_to_unit_sphere_zones( - coordinates=coordinates, unit_sphere_surface=unit_sphere_surface - ) - raw_sums = np.sum( - 1 - (dist / distance_cutoff), where=(dist < distance_cutoff), axis=1 - ) - raw_sums[raw_sums <= 1] = 0 - densities = np.log(raw_sums, out=raw_sums, where=(raw_sums > 1)) - densities = densities / np.max(densities) - return densities +jet_colormap = LinearSegmentedColormap.from_list("jet_colormap", Color.JETSET) def scatter_umi_per_upia_vs_tau( @@ -120,13 +95,68 @@ def scatter_umi_per_upia_vs_tau( return plt.gcf(), plt.gca() +def molecule_rank_plot( + data: pd.DataFrame, group_by: Optional[str] = None +) -> Tuple[plt.Figure, plt.Axes]: + """Plot the number of molecules per component against its molecule rank. + + :param data: a pandas DataFrame with a column 'molecules' containing edge counts for MPX + components. + :param group_by: a column in the DataFrame to group the plot by. + + :return: a plot showing the number of molecules per component against its edge rank used + for quality control. + :rtype: Tuple[plt.Figure, plt.Axes] + :raises: AssertionError if the required column(s) are not present in the DataFrame + :raises: ValueError if the data types are invalid + """ + if "molecules" not in data.columns and "edges" in data.columns: + data["molecules"] = data["edges"] + assert "molecules" in data.columns, "column 'molecules' is missing from DataFrame" + assert ( + isinstance(data["molecules"], pd.Series) and data["molecules"].dtype == int + ), "'molecules' must be a vector of integer values" + + if group_by is not None: + assert ( + group_by in data.columns + ), f"group variable '{group_by}' not found in DataFrame" + + if data[group_by].dtype not in ["object", "category"]: + raise ValueError( + f"Invalid class '{data[group_by].dtype}' for column '{group_by}'. " + f"Expected a string or categorical value" + ) + else: + edge_rank_df = data[[group_by, "molecules"]].copy() + edge_rank_df["rank"] = edge_rank_df.groupby([group_by])["molecules"].rank( + ascending=False, method="first" + ) + else: + edge_rank_df = data[["molecules"]].copy() + edge_rank_df["rank"] = edge_rank_df["molecules"].rank( + ascending=False, method="first" + ) + + ( + sns.relplot( + data=edge_rank_df, x="rank", y="molecules", hue=group_by, aspect=1.6 + ) + .set(xscale="log", yscale="log") + .set_xlabels("Component rank (by number of molecules)") + .set_ylabels("Number of molecules") + ) + + return plt.gcf(), plt.gca() + + def edge_rank_plot( data: pd.DataFrame, group_by: Optional[str] = None ) -> Tuple[plt.Figure, plt.Axes]: """Plot the number of edges per component against its edge rank. :param data: a pandas DataFrame with a column 'edges' containing edge counts for MPX - momponents. + components. :param group_by: a column in the DataFrame to group the plot by. :return: a plot showing the number of edges per component against its edge rank used @@ -135,6 +165,11 @@ def edge_rank_plot( :raises: AssertionError if the required column(s) are not present in the DataFrame :raises: ValueError if the data types are invalid """ + warnings.warn( + "edge_rank_plot is deprecated and will be removed in a future version. Use molecule_rank_plot instead.", + DeprecationWarning, + stacklevel=2, + ) assert "edges" in data.columns, "column 'edges' is missing from DataFrame" assert ( isinstance(data["edges"], pd.Series) and data["edges"].dtype == int @@ -250,739 +285,172 @@ def cell_count_plot( return plt.gcf(), plt.gca() -def _get_component_graph(pxl_data: PixelDataset, component: str): - component_edges = pxl_data.edgelist_lazy.filter(pl.col("component") == component) - component_graph = Graph.from_edgelist( - component_edges, - add_marker_counts=True, - simplify=True, - use_full_bipartite=True, - ) - return component_graph - - -def _get_coordinates( - component_graph: Graph, - layout_algorithm: str, - cache_layout: bool = False, - show_b_nodes: bool = False, - random_seed: int | None = None, -) -> pd.DataFrame: - coordinates = component_graph.layout_coordinates( - layout_algorithm=layout_algorithm, # type: ignore - cache=cache_layout, - only_keep_a_pixels=not show_b_nodes, - random_seed=random_seed, - ) - filtered_coordinates = coordinates - filtered_coordinates["pixel_type"] = [ - component_graph.raw.nodes[ind]["pixel_type"] - for ind in filtered_coordinates.index - ] - - if not show_b_nodes: - filtered_coordinates = filtered_coordinates[ - filtered_coordinates["pixel_type"] == "A" - ] - - edgelist = pd.DataFrame(component_graph.es) - edgelist = edgelist[edgelist[0].isin(filtered_coordinates.index)] - edgelist = edgelist[edgelist[1].isin(filtered_coordinates.index)] - edgelist = edgelist.loc[:, [0, 1]].to_numpy() - - return filtered_coordinates, edgelist - - -def _plot_for_legend(coordinates: pd.DataFrame, axis, show_b_nodes, cmap, node_size): - a_node_example = coordinates[coordinates["pixel_type"] == "A"].iloc[0, :] - axis.scatter( - a_node_example["x"], - a_node_example["y"], - c=0, - cmap=cmap, - vmin=0, - s=node_size, - label="A-nodes", - ) - if show_b_nodes: - b_node_example = coordinates[coordinates["pixel_type"] == "B"].iloc[0, :] - axis.scatter( - b_node_example["x"], - b_node_example["y"], - c=1, - cmap=cmap, - vmin=0, - s=node_size, - label="B-nodes", - ) - - -def _decorate_plot( - im=None, - include_colorbar: bool = False, - vmax: float = 0, - cmap: str = "cool", +def _plot_joint_distribution(data, x, y, show_marginal, **kargs): + g = sns.JointGrid(data, x=x, y=y) + g.plot_marginals(sns.kdeplot, fill=True) + g.ax_marg_x.axis("off") + g.ax_marg_y.axis("off") + g.plot_joint( + sns.scatterplot, + legend=False, + data=data, + hue="density", + palette=jet_colormap, + size=0.1, + ) + if not show_marginal: + g.ax_marg_x.set_visible(False) + g.ax_marg_y.set_visible(False) + return g + + +def _add_gate_box( + data, + gate: pd.Series | pd.DataFrame, + marker1, + marker2, + facet_row=None, + facet_column=None, ax=None, - fig=None, - legend_ax=None, + **kargs, ): - if include_colorbar: - if isinstance(ax, np.ndarray): - fig.colorbar( - im, - ax=ax.ravel().tolist(), - cmap=cmap, - norm=Normalize(vmin=0, vmax=vmax), - ) - else: - fig.colorbar( - im, - ax=ax, - cmap=cmap, - norm=Normalize(vmin=0, vmax=vmax), - ) - if legend_ax is not None: - if not include_colorbar: - handles, labels = legend_ax.get_legend_handles_labels() - fig.legend(handles, labels, loc="right") - plt.subplots_adjust(right=0.80) - else: - raise AssertionError( - "Plotting pixel_type together with other markers is not supported." - ) - - -def _get_current_axis(ax, i_m, i_c, marker_list, component): - if len(marker_list) == 1: - if len(component) == 1: - crnt_ax = ax - else: - crnt_ax = ax[i_c] + if ax is None: + ax = plt.gca() + + if facet_row is not None and facet_column is not None: + condition = (data.iloc[0][facet_row], data.iloc[0][facet_column]) + elif facet_row is not None: + condition = data.iloc[0][facet_row] + elif facet_column is not None: + condition = data.iloc[0][facet_column] else: - if len(component) == 1: - crnt_ax = ax[i_m] - else: - crnt_ax = ax[i_m, i_c] + condition = None - if i_c == 0: # Set the y-label only for the first column - crnt_ax.set_ylabel(marker_list[i_m], rotation=90, size="large") - if i_m == 0: # Set the title only for the first row - crnt_ax.set_title(component[i_c]) - - return crnt_ax - - -def _get_color_values(mark, filtered_coordinates, log_scale, vmax): - if mark is None: - color_val = Color.NETWORKX_NODE_COLOR - elif mark == "pixel_type": - color_val = filtered_coordinates["pixel_type"] == "B" - elif mark not in filtered_coordinates.columns: - raise AssertionError(f"Marker {mark} not found in the component graph.") - else: - if log_scale: - color_val = np.log1p(filtered_coordinates.loc[:, mark]) + if isinstance(gate, pd.DataFrame): + if condition in gate.index: + gate = gate.loc[condition, :] else: - color_val = filtered_coordinates.loc[:, mark] - vmax = max(vmax, np.max(color_val)) - - return color_val, vmax - - -def plot_2d_graph( - pxl_data: PixelDataset, - component: Union["str", list], - marker: str = "pixel_type", - layout_algorithm: Literal["fruchterman_reingold", "kamada_kawai", "pmds"] = "pmds", - show_edges: bool = False, - log_scale: bool = True, - node_size: float = 10.0, - edge_width: float = 1.0, - show_b_nodes: bool = False, - cmap: str = "cool", - alpha: float = 0.5, - cache_layout: bool = False, - random_seed: int | None = None, -) -> Tuple[plt.Figure, plt.Axes]: - """Plot a (collection of) 2D graph(s) based on the given pixel data. - - The graph can be plotted for one or a list of components. - The graph nodes can be colored by a marker. The marker can be a (list of) marker(s) or "pixel_type". - Example usage: plot_2d_graph(pxl, component=["PXLCMP0000000"], marker=["HLA-ABC", "HLA-RA"]). - - :param pxl_data: The pixel dataset to plot. - :param component: The component(s) to plot. Defaults to None. - :param marker: The marker attribute to use for coloring the nodes. Defaults to "pixel_type". - :param layout_algorithm: The layout algorithm to use. Defaults to "pmds". - :param show_edges: Whether to show the edges in the graph. Defaults to False. - :param log_scale: Whether to use a logarithmic scale for the marker attribute. Defaults to True. - :param node_size: The size of the nodes. Defaults to 10.0. - :param edge_width: The width of the edges. Defaults to 1.0. - :param show_b_nodes: Whether to show the B-nodes. Defaults to False. - :param cmap: The colormap to use for coloring the nodes. Defaults to "cool". - :param alpha: The alpha value for the nodes. Defaults to 0.7. - :param cache_layout: Whether to cache the layout coordinates. Defaults to False. - :param random_seed: The random seed to use for the layout algorithm. Defaults to None. - - :return: The figure and axes objects of the plot. - :rtype: Tuple[plt.Figure, plt.Axes] - - :raises: AssertionError if the marker is not found in the component graph. - :raises: AssertionError if no nodes are found with the specified marker. - :raises: AssertionError if "pixel_type" is in the markers together with other markers. - - """ - if isinstance(component, str): - component = [component] - - if isinstance(marker, str): - marker_list = [marker] - else: - marker_list = marker - - fig, ax = plt.subplots(nrows=len(marker_list), ncols=len(component)) - - include_colorbar = False - vmax = 0 # maximum value for colorbar - for i_c, comp in enumerate(component): - component_graph = _get_component_graph(pxl_data=pxl_data, component=comp) - coordinates, edgelist = _get_coordinates( - component_graph=component_graph, - layout_algorithm=layout_algorithm, - cache_layout=cache_layout, - show_b_nodes=show_b_nodes, - random_seed=random_seed, + return + ax.add_patch( + Rectangle( + xy=(gate["xmin"], gate["ymin"]), + width=gate["xmax"] - gate["xmin"], + height=gate["ymax"] - gate["ymin"], + fill=False, + linestyle="--", + linewidth=1, + edgecolor="black", ) - - for i_m, mark in enumerate(marker_list): - crnt_ax = _get_current_axis(ax, i_m, i_c, marker_list, component) - color_val, vmax = _get_color_values(mark, coordinates, log_scale, vmax) - if mark != "pixel_type": - include_colorbar = True - - im = nx.draw_networkx( - component_graph.raw, - nodelist=coordinates.index, - pos=coordinates.loc[:, ["x", "y"]].T.to_dict("list"), - ax=crnt_ax, - node_size=node_size, - node_color=color_val, - cmap=cmap, - width=edge_width, - with_labels=False, - edgelist=edgelist if show_edges else [], - label="_nolegend_", - alpha=alpha, - ) - - if mark == "pixel_type": - # Re-plot one point from each pixel type to add a legend - _plot_for_legend(coordinates, crnt_ax, show_b_nodes, cmap, node_size) - legend_ax = crnt_ax - else: - legend_ax = None - - crnt_ax.grid(False) - crnt_ax.spines[:].set_visible(False) - crnt_ax.set_xticks([]) - crnt_ax.set_yticks([]) - - _decorate_plot( - im=im, - include_colorbar=include_colorbar, - vmax=vmax, - cmap=cmap, - ax=ax, - fig=fig, - legend_ax=legend_ax, - ) - - return fig, ax - - -def plot_3d_from_coordinates( - coordinates: pd.DataFrame, - node_size: float = 3.0, - opacity: float = 0.4, - cmap: str = "Inferno", - suppress_fig: bool = False, -) -> go.Figure: - """Plot a 3D graph from the given coordinates. - - :param coordinates: The coordinates to plot. - :param node_size: The size of the nodes. Defaults to 3.0. - :param opacity: The opacity of the nodes. Defaults to 0.4. - :param cmap: The colormap to use for coloring the nodes. Defaults to "Inferno". - :param suppress_fig: Whether to suppress (i.e. not plot) the figure. Defaults to False. - :return: The plotted 3D graph. - :rtype: go.Figure - - """ - fig = go.Figure( - data=[ - go.Scatter3d( - x=coordinates["x"], - y=coordinates["y"], - z=coordinates["z"], - mode="markers", - marker=dict( - size=node_size, - color=coordinates["color"], - colorscale=cmap, - opacity=opacity, - colorbar=dict(thickness=20, title="color"), - ), - ) - ] - ) - - if not suppress_fig: - fig.show() - return fig - - -def plot_3d_graph( - pxl_data: PixelDataset, - component: str, - marker: Union[list, None] = None, - layout_algorithm: Literal[ - "fruchterman_reingold_3d", "kamada_kawai_3d", "pmds_3d" - ] = "fruchterman_reingold_3d", - log_scale: bool = True, - normalize: bool = False, - node_size: float = 3.0, - opacity: float = 0.4, - show_b_nodes: bool = False, - cmap: str = "Inferno", - cache_layout: bool = False, - suppress_fig: bool = False, -) -> go.Figure: - """Plot a 3D graph of the specified component in the given PixelDataset. - - :param pxl_data: The PixelDataset containing the data. - :param component: The component to plot. - :param marker: The marker to use for coloring the nodes. Defaults to None. - :param layout_algorithm: The layout algorithm to use for positioning the nodes. Defaults to "fruchterman_reingold_3d". - :param log_scale: Whether to apply logarithmic scaling to the marker values. Defaults to True. - :param normalize: Whether to normalize the coordinates. Defaults to False. - :param node_size: The size of the nodes. Defaults to 3.0. - :param opacity: The opacity of the nodes. Defaults to 0.4. - :param show_b_nodes: Whether to show nodes of type B. Defaults to False. - :param cmap: The colormap to use for coloring the nodes. Defaults to "Inferno". - :param cache_layout: Whether to cache the layout coordinates. Defaults to False. - :param suppress_fig: Whether to suppress (i.e. not plot) the figure. Defaults to False. - :return: The plotted 3D graph. - :rtype: go.Figure - - """ - component_graph = _get_component_graph(pxl_data=pxl_data, component=component) - coordinates, _ = _get_coordinates( - component_graph=component_graph, - layout_algorithm=layout_algorithm, - cache_layout=cache_layout, - show_b_nodes=show_b_nodes, - ) - - if marker is not None: - if log_scale: - coordinates["color"] = np.log1p(coordinates[marker]) - else: - coordinates["color"] = coordinates[marker] - else: - coordinates["color"] = Color.SKYBLUE3 - - if normalize: - coordinates[["x", "y", "z"]] = coordinates[["x_norm", "y_norm", "z_norm"]] - - fig = plot_3d_from_coordinates( - coordinates=coordinates, - node_size=node_size, - opacity=opacity, - cmap=cmap, - suppress_fig=True, ) - fig.update_layout(title=component) - - if not suppress_fig: - fig.show() - - return fig - - -def _pivot_colocalization_data( - colocalization_data: pd.DataFrame, - value_col: str = "pearson", - markers: Union[list, None] = None, + # Counting data points inside the gate box + inside_box = ( + (data[marker1] >= gate["xmin"]) + & (data[marker1] < gate["xmax"]) + & (data[marker2] >= gate["ymin"]) + & (data[marker2] < gate["ymax"]) + ) + inside_percentage = f"{inside_box.mean() * 100:.1f}%" + ax.text( + x=gate["xmin"], + y=gate["ymax"], + s=inside_percentage, + verticalalignment="top", + horizontalalignment="left", + fontsize=10, + color="black", + ) + return + + +def density_scatter_plot( + adata: anndata.AnnData, + marker1: str, + marker2: str, + layer: str | None = None, + facet_row: str | None = None, + facet_column: str | None = None, + gate: pd.Series | pd.DataFrame | None = None, + show_marginal=False, ): - colocalization_data_pivot = pd.pivot_table( - colocalization_data, - index="marker_1", - columns="marker_2", - values=value_col, - fill_value=0, - ) - - if markers is not None: - colocalization_data_pivot = colocalization_data_pivot.loc[markers, markers] - - for m in colocalization_data_pivot.index: - colocalization_data_pivot.loc[m, m] = 0 # remove autocorrelations - - colocalization_data_pivot = colocalization_data_pivot.fillna(0) - - return colocalization_data_pivot - - -def _make_colocalization_symmetric( - colocalization_data: pd.DataFrame, - value_col: str = "pearson", -): - colocalization_data = pd.DataFrame( - np.concatenate( - [ - colocalization_data[["marker_1", "marker_2", value_col]].to_numpy(), - colocalization_data[["marker_2", "marker_1", value_col]].to_numpy(), - ], - ), - columns=["marker_1", "marker_2", value_col], - ) - colocalization_data = ( - colocalization_data.groupby(["marker_1", "marker_2"])[value_col] - .apply(lambda x: np.mean(x)) - .reset_index() - ) - - return colocalization_data - - -def plot_colocalization_heatmap( - colocalization_data: pd.DataFrame, - markers: Union[list, None] = None, - cmap: str = "vlag", - use_z_scores: bool = False, -) -> Tuple[plt.Figure, plt.Axes]: - """Plot a colocalization heatmap based on the provided colocalization data. - - The colocalization_data DataFrame should contain the columns "marker_1", "marker_2", "pearson", "pearson_z". - Example usage: plot_colocalization_heatmap(pxl.colocalization). - - :param colocalization_data: The colocalization data to plot. The colocalization data frame that can be found in a pixel variable "pxl" through pxl.colocalization. The data frame should contain the columns "marker_1", "marker_2", "pearson", "pearson_z", and "component". - :param markers: The markers to include in the heatmap. Defaults to None. - :param cmap: The colormap to use for the heatmap. Defaults to "vlag". - :param use_z_scores: Whether to use z-scores. Defaults to False. - - :return: The figure and axes objects of the plot. - :rtype: Tuple[plt.Figure, plt.Axes] - + """Pseudocolor density scatter plot. + + This function creates a scatter plot of abundance data for two markers colored + based on the density of points: + Example usage: `density_scatter_plot(pxl.adata, "CD3E", "CD4")`. + It is also possible to specify one or two variables for faceting the plot by + columns and rows: + Example usage: `density_scatter_plot(pxl.adata, "CD3E", "CD4", + facet_row = "stimulation", facet_column = "donor")`. + `facet_row` and `facet_column` should be names of categorical columns in `adata.obs`. + In addition, a gate can be specified as a Series with xmin, xmax, ymin, and ymax + to mark a range for components of interest. Alternatively, gate can be specified + as a DataFrame to allow for different gate ranges for various conditions. + When both facet_row and facet_column are specified, the condition becomes a + tuple (facet_row, facet_column), if only one is specified, that parameter + becomes the condition. The condition permuations are used as the index of + the gate. + Example usage: + gate = pd.DataFrame(columns = ["xmin", "ymin", "xmax", "ymax"]) + gate.loc["Resting"] = [2, 2, 5, 4] + gate.loc["PHA"] = [1.5, 1.5, 5, 4] + fig, ax = density_scatter_plot(pixel.adata, "CD3E", "CD4", layer="dsb", + facet_column="sample", gate=gate) + + + :param adata: Anndata object containing the marker abundance data per + component. + :param marker1: The first marker to plot (x-axis). + :param marker2: The second marker to plot (y-axis). + :param layer: The anndata layer (e.g. transformation) to use for the marker + data. Defaults to None. + :param facet_row: The column to use for faceting the plot by rows. + Defaults to None. + :param facet_column: The column to use for faceting the plot by columns. + Defaults to None. + :param gate: The gate to use for marking a range of interest. Defaults to + None. + :param show_marginal: Whether to show marginal distributions. Defaults to False. """ - if use_z_scores: - value_col = "pearson_z" - else: - value_col = "pearson" - - colocalization_data = _make_colocalization_symmetric(colocalization_data, value_col) - - colocalization_data_pivot = _pivot_colocalization_data( - colocalization_data, value_col, markers=markers - ) - sns.clustermap( - colocalization_data_pivot, - yticklabels=True, - xticklabels=True, - linewidths=0.1, - method="complete", - cmap=cmap, - ) - - return plt.gcf(), plt.gca() - - -def _get_top_marker_pairs( - colocalization_data: pd.DataFrame, - n_top_marker_pairs: int, - value_col: str = "pearson", -) -> list: - colocalization_data["abs_val"] = colocalization_data[value_col].abs() - top_marker_pairs = colocalization_data.nlargest(n_top_marker_pairs, "abs_val") - top_markers = list( - set(top_marker_pairs["marker_1"]).union(set(top_marker_pairs["marker_2"])) - ) - - return top_markers - - -def plot_colocalization_diff_heatmap( - colocalization_data: pd.DataFrame, - target: str, - reference: str, - contrast_column: str = "sample", - markers: Union[list, None] = None, - n_top_marker_pairs: Union[int, None] = None, - cmap: str = "vlag", - use_z_score: bool = True, -) -> Tuple[plt.Figure, plt.Axes]: - """Plot the differential colocalization between reference and target components. - - Example usage: plot_colocalization_diff_heatmap(pxl.colocalization, target:"stimulated", reference:"control", contrast_column="sample"). - - :param colocalization_data: The colocalization data frame that can be found in a pixel variable "pxl" through pxl.colocalization. The data frame should contain the columns "marker_1", "marker_2", "pearson", "pearson_z", and the contrast_column. - :param target: The label for target components in the contrast_column. - :param reference: The label for reference components in the contrast_column. - :param contrast_column: The column to use for the contrast. Defaults to "sample". - :param markers: The markers to include in the heatmap. Defaults to None. At most only one of n_top_marker_pairs or markers should be provided. - :param n_top_marker_pairs: The number of top marker pairs to include in the heatmap. Defaults to None. At most only one of n_top_marker_pairs or markers should be provided. - :param cmap: The colormap to use for the heatmap. Defaults to "vlag". - :param use_z_score: Whether to use the z-score. Defaults to True. - - :return: The figure and axes objects of the plot. - :rtype: Tuple[plt.Figure, plt.Axes] - """ - assert ( - markers is None or n_top_marker_pairs is None - ), "Only one of markers or n_top_marker_pairs can be provided." - - if use_z_score: - value_col = "pearson_z" - else: - value_col = "pearson" - - if markers is not None: - filter_mask = (colocalization_data["marker_1"].isin(markers)) & ( - colocalization_data["marker_2"].isin(markers) - ) - colocalization_data = colocalization_data[filter_mask] - - differential_colocalization = get_differential_colocalization( - colocalization_data, - target=target, - reference=reference, - contrast_column=contrast_column, - use_z_score=use_z_score, - ) - - differential_colocalization = differential_colocalization.fillna(0).reset_index() - - if n_top_marker_pairs is not None: - top_markers = _get_top_marker_pairs( - differential_colocalization, n_top_marker_pairs, "median_difference" + layer_data = adata.to_df(layer) + data = layer_data.loc[:, [marker1, marker2]] + data.loc[:, "density"] = gaussian_kde(data.T)(data.T) + if facet_column is not None: + data.loc[:, facet_column] = adata.obs.loc[:, facet_column] + + if facet_row is not None: + data.loc[:, facet_row] = adata.obs.loc[:, facet_row] + + if facet_column is not None or facet_row is not None: + if show_marginal: + warnings.warn("show_marginal is not supported for faceted plots.") + plot_grid = sns.FacetGrid(data=data, col=facet_column, row=facet_row) + plot_grid.map_dataframe( + sns.scatterplot, + x=marker1, + y=marker2, + hue="density", + palette=jet_colormap, + size=0.1, ) + if gate is not None: + plot_grid.map_dataframe( + _add_gate_box, + gate=gate, + marker1=marker1, + marker2=marker2, + facet_row=facet_row, + facet_column=facet_column, + ) + plot_grid.refline(x=0, y=0) else: - top_markers = None - - # Making the differential colocalization symmetric - differential_colocalization = _make_colocalization_symmetric( - differential_colocalization, "median_difference" - ) - - pivoted_differential_colocalization = _pivot_colocalization_data( - differential_colocalization, - "median_difference", - markers=top_markers, - ) - - max_value = np.max(np.abs(pivoted_differential_colocalization.to_numpy().flatten())) - sns.clustermap( - pivoted_differential_colocalization, - yticklabels=True, - xticklabels=True, - method="complete", - linewidths=0.1, - vmin=-max_value, - vmax=max_value, - cmap=cmap, - ) - + plot_grid = _plot_joint_distribution( + data, x=marker1, y=marker2, show_marginal=show_marginal + ) + if gate is not None: + _add_gate_box( + data, marker1=marker1, marker2=marker2, ax=plot_grid.ax_joint, gate=gate + ) + plot_grid.ax_joint.axhline(0, color="black", linewidth=1, linestyle="--") + plot_grid.ax_joint.axvline(0, color="black", linewidth=1, linestyle="--") return plt.gcf(), plt.gca() - - -def _add_top_marker_labels( - differential_colocalization, - ax, - n_top_pairs: int = 5, - min_log_p: float = 5.0, -): - differential_colocalization = differential_colocalization.sort_values( - "median_difference" - ) - differential_colocalization = differential_colocalization.loc[ - -np.log10(differential_colocalization["p_adj"]) > min_log_p, : - ] - - ## Labels for marker pair withs highest negative differential colocalization scores - for _, row in differential_colocalization.head(n_top_pairs).iterrows(): - x, y = row[["median_difference", "p_adj"]] - y = -np.log10(y) - if x > 0: - continue - ax.text(x, y, row["markers"], horizontalalignment="left", fontsize="xx-small") - - ## Labels for marker pair with highest positive differential colocalization scores - for _, row in differential_colocalization.tail(n_top_pairs).iterrows(): - x, y = row[["median_difference", "p_adj"]] - y = -np.log10(y) - if x < 0: - continue - ax.text(x, y, row["markers"], horizontalalignment="left", fontsize="xx-small") - - -def _add_target_mean_colocalizations( - differential_colocalization, - target_coloc, - value_col, -): - differential_colocalization = differential_colocalization.fillna(0).reset_index() - - target_values = ( - target_coloc.groupby(["marker_1", "marker_2"])[value_col].mean().reset_index() - ) - differential_colocalization = pd.merge( - target_values, - differential_colocalization, - on=["marker_1", "marker_2"], - how="inner", - ) - - return differential_colocalization - - -def plot_colocalization_diff_volcano( - colocalization_data: pd.DataFrame, - target: str, - reference: str, - contrast_column: str = "sample", - cmap: str = "vlag", - use_z_score: bool = True, - n_top_pairs: int = 5, - min_log_p: float = 5.0, - ax: Optional[plt.Axes] = None, -) -> Tuple[plt.Figure, plt.Axes]: - """Generate the volcano plot of differential colocalization between reference and target components. - - Example usage: `plot_colocalization_diff_volcano(pxl.colocalization, target:"stimulated", reference:"control", contrast_column="sample")`. - - :param colocalization_data: The colocalization data frame that can be found in a pixel variable - "pxl" through pxl.colocalization. The data frame should contain the - columns "marker_1", "marker_2", "pearson", "pearson_z", and the contrast_column. - :param target: The label for target components in the contrast_column. - :param reference: The label for reference components in the contrast_column. - :param contrast_column: The column to use for the contrast. Defaults to "sample". - :param cmap: The colormap to use for the heatmap. Defaults to "vlag". - :param use_z_score: Whether to use the z-score. Defaults to True. - :param n_top_pairs: Number of high value marker-pairs to label from positive and negative sides. - :param min_log_p: marker-pairs only receive a label if -log10 of their p-value is higher than - this parameter. - - :return: The figure and axes objects of the plot. - :rtype: Tuple[plt.Figure, plt.Axes] - """ - if use_z_score: - value_col = "pearson_z" - else: - value_col = "pearson" - - differential_colocalization = get_differential_colocalization( - colocalization_data, - target=target, - reference=reference, - contrast_column=contrast_column, - use_z_score=use_z_score, - ) - - differential_colocalization = _add_target_mean_colocalizations( - differential_colocalization, - target_coloc=colocalization_data.loc[ - colocalization_data[contrast_column] == target, : - ], - value_col=value_col, - ) - - if ax is None: - fig, ax = plt.subplots() - - p = ax.scatter( - x=differential_colocalization["median_difference"], - y=-np.log10(differential_colocalization["p_adj"]), - c=differential_colocalization[value_col], - s=20, - marker="o", - cmap=cmap, - ) - - ax.set(xlabel="Median difference", ylabel=r"$-\log_{10}$(adj. p-value)") - fig = plt.gcf() - fig.colorbar(p, label="Mean target colocalization score", cmap=cmap) - _add_top_marker_labels( - differential_colocalization, - n_top_pairs=n_top_pairs, - min_log_p=min_log_p, - ax=ax, - ) - - return fig, ax - - -@experimental -def plot_3d_heatmap( - component_graph: Graph, - marker: str, - distance_cutoff: float, - layout_algorithm: Literal[ - "fruchterman_reingold_3d", "kamada_kawai_3d" - ] = "fruchterman_reingold_3d", - cache_layout: bool = False, -) -> Tuple[plt.Figure, plt.Axes]: - """Plot a 3D heatmap for the marker in the provided component. - - :param component_graph: A component graph to plot for. - :param marker: marker to plot this for. - :param distance_cutoff: a distance cutoff to use for determining size of - area to consider as close in the density calculation. - :param layout_algorithm: Layout algorithm to use. Options are: - "fruchterman_reingold_3d" and "kamada_kawai_3d" - :param cache_layout: set this to `True` to cache the layout - or faster computations on subsequent calls. This comes at the - cost of additional memory usage. - :return: A matplotlib 3D heatmap figure, and it's associated Axes instance - :rtype: Tuple[plt.Figure, plt.Axes] - :raises: AssertionError if the provided `layout_algorithm` is not valid, - or there are no with markers for the provided `marker` - """ - coordinates = component_graph.layout_coordinates( - layout_algorithm=layout_algorithm, cache=cache_layout - ) - coordinates = coordinates[coordinates[marker] > 0] - coordinates = np.array(coordinates[["x_norm", "y_norm", "z_norm"]]) - if len(coordinates) < 1: - raise AssertionError(f"No nodes found with {marker}.") - - horizontal_resolution = 120 - vertical_resolution = 80 - X, Y, Z = _unit_sphere_surface( - horizontal_resolution=horizontal_resolution, - vertical_resolution=vertical_resolution, - ) - - densities = _calculate_densities( - coordinates=coordinates, - distance_cutoff=distance_cutoff, - unit_sphere_surface=(X, Y, Z), - ) - - fig = plt.figure(figsize=(6, 6)) - ax = fig.add_subplot(1, 1, 1, projection="3d") - color_scale = cm.inferno( # type: ignore - densities.reshape(horizontal_resolution, -1) - ) - ax.plot_surface( # type: ignore - X, - Y, - Z, - cstride=1, - rstride=1, - facecolors=color_scale, - shade=False, - ) - ax.set_axis_off() - ax.set_box_aspect([1.0, 1.0, 1.0]) # type: ignore - return fig, ax diff --git a/src/pixelator/plot/constants.py b/src/pixelator/plot/constants.py index ceeaf2d0..cd2d94f7 100644 --- a/src/pixelator/plot/constants.py +++ b/src/pixelator/plot/constants.py @@ -1,6 +1,6 @@ """Contains constants used as part of plotting modules. -Copyright (c) 2023 Pixelgen Technologies AB. +Copyright © 2023 Pixelgen Technologies AB. """ @@ -11,3 +11,14 @@ class Color: ORANGERED2 = "#ee4000" SKYBLUE3 = "#6ca6cd" NETWORKX_NODE_COLOR = "#1f78b4" + JETSET = [ + "#00007F", # dark blue + "#0000FF", # blue + "#007FFF", # azure + "#00FFFF", # cyan + "#7FFF7F", # light green + "#FFFF00", # yellow + "#FF7F00", # orange + "#FF0000", # red + "#7F0000", # dark red + ] diff --git a/src/pixelator/plot/layout_plots.py b/src/pixelator/plot/layout_plots.py new file mode 100644 index 00000000..1c94ecf9 --- /dev/null +++ b/src/pixelator/plot/layout_plots.py @@ -0,0 +1,491 @@ +"""Functions for plotting component layouts for MPX data. + +Copyright © 2024 Pixelgen Technologies AB. +""" + +from typing import List, Literal, Tuple, Union + +import matplotlib.pyplot as plt +import networkx as nx +import numpy as np +import pandas as pd +import plotly.graph_objects as go +import polars as pl +import scipy +from matplotlib import cm +from matplotlib.colors import Normalize + +from pixelator.graph import Graph +from pixelator.graph.backends.protocol import SupportedLayoutAlgorithm +from pixelator.marks import experimental +from pixelator.pixeldataset import PixelDataset +from pixelator.plot.constants import Color + + +def _unit_sphere_surface(horizontal_resolution, vertical_resolution): + horizontal_angles = np.linspace(0, 2 * np.pi, horizontal_resolution) + vertical_angles = np.linspace(0, np.pi, vertical_resolution) + + X = np.outer(np.cos(horizontal_angles), np.sin(vertical_angles)) + Y = np.outer(np.sin(horizontal_angles), np.sin(vertical_angles)) + Z = np.outer(np.ones(np.size(horizontal_angles)), np.cos(vertical_angles)) + return X, Y, Z + + +def _calculate_distance_to_unit_sphere_zones(coordinates, unit_sphere_surface): + X, Y, Z = unit_sphere_surface + zones_on_sphere_surface = np.stack([X, Y, Z], axis=2).reshape(-1, 3) + return scipy.spatial.distance.cdist( + zones_on_sphere_surface, coordinates, "euclidean" + ) + + +def _calculate_densities(coordinates, distance_cutoff, unit_sphere_surface): + dist = _calculate_distance_to_unit_sphere_zones( + coordinates=coordinates, unit_sphere_surface=unit_sphere_surface + ) + raw_sums = np.sum( + 1 - (dist / distance_cutoff), where=(dist < distance_cutoff), axis=1 + ) + raw_sums[raw_sums <= 1] = 0 + densities = np.log(raw_sums, out=raw_sums, where=(raw_sums > 1)) + densities = densities / np.max(densities) + return densities + + +def _get_component_graph(pxl_data: PixelDataset, component: str): + component_edges = pxl_data.edgelist_lazy.filter(pl.col("component") == component) + component_graph = Graph.from_edgelist( + component_edges, + add_marker_counts=True, + simplify=True, + use_full_bipartite=True, + ) + return component_graph + + +def _get_coordinates( + pxl_data: PixelDataset, + component: str, + layout_algorithm: SupportedLayoutAlgorithm | None = None, + cache_layout: bool = False, + show_b_nodes: bool = False, + random_seed: int | None = None, +) -> pd.DataFrame: + component_graph = _get_component_graph(pxl_data=pxl_data, component=component) + if ( + layout_algorithm is None + and hasattr(pxl_data, "precomputed_layouts") + and pxl_data.precomputed_layouts is not None + ): + coordinates = pxl_data.precomputed_layouts.filter( + component_ids=component + ).to_df() + else: + if layout_algorithm is None: + layout_algorithm = "pmds_3d" + coordinates = component_graph.layout_coordinates( + layout_algorithm=layout_algorithm, # type: ignore + cache=cache_layout, + only_keep_a_pixels=not show_b_nodes, + random_seed=random_seed, + ) + + filtered_coordinates = coordinates + filtered_coordinates["pixel_type"] = [ + component_graph.raw.nodes[ind]["pixel_type"] + for ind in filtered_coordinates.index + ] + + if not show_b_nodes: + filtered_coordinates = filtered_coordinates[ + filtered_coordinates["pixel_type"] == "A" + ] + + edgelist = pd.DataFrame(component_graph.es) + edgelist = edgelist[edgelist[0].isin(filtered_coordinates.index)] + edgelist = edgelist[edgelist[1].isin(filtered_coordinates.index)] + edgelist = edgelist.loc[:, [0, 1]].to_numpy() + + return filtered_coordinates, edgelist, component_graph + + +def _plot_for_legend(coordinates: pd.DataFrame, axis, show_b_nodes, cmap, node_size): + a_node_example = coordinates[coordinates["pixel_type"] == "A"].iloc[0, :] + axis.scatter( + a_node_example["x"], + a_node_example["y"], + c=0, + cmap=cmap, + vmin=0, + s=node_size, + label="A-nodes", + ) + if show_b_nodes: + b_node_example = coordinates[coordinates["pixel_type"] == "B"].iloc[0, :] + axis.scatter( + b_node_example["x"], + b_node_example["y"], + c=1, + cmap=cmap, + vmin=0, + s=node_size, + label="B-nodes", + ) + + +def _decorate_plot( + im=None, + include_colorbar: bool = False, + vmax: float = 0, + cmap: str = "cool", + ax=None, + fig=None, + legend_ax=None, +): + if include_colorbar: + if isinstance(ax, np.ndarray): + fig.colorbar( + im, + ax=ax.ravel().tolist(), + cmap=cmap, + norm=Normalize(vmin=0, vmax=vmax), + ) + else: + fig.colorbar( + im, + ax=ax, + cmap=cmap, + norm=Normalize(vmin=0, vmax=vmax), + ) + if legend_ax is not None: + if not include_colorbar: + handles, labels = legend_ax.get_legend_handles_labels() + fig.legend(handles, labels, loc="right") + plt.subplots_adjust(right=0.80) + else: + raise AssertionError( + "Plotting pixel_type together with other markers is not supported." + ) + + +def _get_current_axis(ax, i_m, i_c, marker_list, component): + if len(marker_list) == 1: + if len(component) == 1: + crnt_ax = ax + else: + crnt_ax = ax[i_c] + else: + if len(component) == 1: + crnt_ax = ax[i_m] + else: + crnt_ax = ax[i_m, i_c] + + if i_c == 0: # Set the y-label only for the first column + crnt_ax.set_ylabel(marker_list[i_m], rotation=90, size="large") + if i_m == 0: # Set the title only for the first row + crnt_ax.set_title(component[i_c]) + + return crnt_ax + + +def _get_color_values(mark, filtered_coordinates, log_scale, vmax): + if mark is None: + color_val = Color.NETWORKX_NODE_COLOR + elif mark == "pixel_type": + color_val = filtered_coordinates["pixel_type"] == "B" + elif mark not in filtered_coordinates.columns: + raise AssertionError(f"Marker {mark} not found in the component graph.") + else: + if log_scale: + color_val = np.log1p(filtered_coordinates.loc[:, mark]) + else: + color_val = filtered_coordinates.loc[:, mark] + vmax = max(vmax, np.max(color_val)) + + return color_val, vmax + + +def plot_2d_graph( + pxl_data: PixelDataset, + component: str | List[str], + marker: str | List[str] = "pixel_type", + layout_algorithm: SupportedLayoutAlgorithm | None = None, + show_edges: bool = False, + log_scale: bool = True, + node_size: float = 10.0, + edge_width: float = 1.0, + show_b_nodes: bool = False, + cmap: str = "cool", + alpha: float = 0.5, + cache_layout: bool = False, + random_seed: int | None = None, +) -> Tuple[plt.Figure, plt.Axes | np.ndarray]: + """Plot a (collection of) 2D graph(s) based on the given pixel data. + + The graph can be plotted for one or a list of components. + The graph nodes can be colored by a marker. The marker can be a (list of) marker(s) or "pixel_type". + Example usage: plot_2d_graph(pxl, component=["PXLCMP0000000"], marker=["HLA-ABC", "HLA-RA"]). + + :param pxl_data: The pixel dataset to plot. + :param component: The component(s) to plot. Defaults to None. + :param marker: The marker attribute to use for coloring the nodes. Defaults to "pixel_type". + :param layout_algorithm: The layout algorithm to use. Defaults to None (checking for pre-computed). + :param show_edges: Whether to show the edges in the graph. Defaults to False. + :param log_scale: Whether to use a logarithmic scale for the marker attribute. Defaults to True. + :param node_size: The size of the nodes. Defaults to 10.0. + :param edge_width: The width of the edges. Defaults to 1.0. + :param show_b_nodes: Whether to show the B-nodes. Defaults to False. + :param cmap: The colormap to use for coloring the nodes. Defaults to "cool". + :param alpha: The alpha value for the nodes. Defaults to 0.7. + :param cache_layout: Whether to cache the layout coordinates. Defaults to False. + :param random_seed: The random seed to use for the layout algorithm. Defaults to None. + + :return: The figure and axes objects of the plot. + :rtype: Tuple[plt.Figure, plt.Axes] + + :raises: AssertionError if the marker is not found in the component graph. + :raises: AssertionError if no nodes are found with the specified marker. + :raises: AssertionError if "pixel_type" is in the markers together with other markers. + + """ + if isinstance(component, str): + component = [component] + + if isinstance(marker, str): + marker_list = [marker] + else: + marker_list = marker + + fig, ax = plt.subplots(nrows=len(marker_list), ncols=len(component)) + + include_colorbar = False + vmax = 0 # maximum value for colorbar + for i_c, comp in enumerate(component): + coordinates, edgelist, component_graph = _get_coordinates( + pxl_data=pxl_data, + component=comp, + layout_algorithm=layout_algorithm, + cache_layout=cache_layout, + show_b_nodes=show_b_nodes, + random_seed=random_seed, + ) + + for i_m, mark in enumerate(marker_list): + crnt_ax = _get_current_axis(ax, i_m, i_c, marker_list, component) + color_val, vmax = _get_color_values(mark, coordinates, log_scale, vmax) + if mark != "pixel_type": + include_colorbar = True + + im = nx.draw_networkx( + component_graph.raw, + nodelist=coordinates.index, + pos=coordinates.loc[:, ["x", "y"]].T.to_dict("list"), + ax=crnt_ax, + node_size=node_size, + node_color=color_val, + cmap=cmap, + width=edge_width, + with_labels=False, + edgelist=edgelist if show_edges else [], + label="_nolegend_", + alpha=alpha, + ) + + if mark == "pixel_type": + # Re-plot one point from each pixel type to add a legend + _plot_for_legend(coordinates, crnt_ax, show_b_nodes, cmap, node_size) + legend_ax = crnt_ax + else: + legend_ax = None + + crnt_ax.grid(False) + crnt_ax.spines[:].set_visible(False) + crnt_ax.set_xticks([]) + crnt_ax.set_yticks([]) + + _decorate_plot( + im=im, + include_colorbar=include_colorbar, + vmax=vmax, + cmap=cmap, + ax=ax, + fig=fig, + legend_ax=legend_ax, + ) + + return fig, ax + + +def plot_3d_from_coordinates( + coordinates: pd.DataFrame, + node_size: float = 3.0, + opacity: float = 0.4, + cmap: str = "Inferno", + suppress_fig: bool = False, +) -> go.Figure: + """Plot a 3D graph from the given coordinates. + + :param coordinates: The coordinates to plot. + :param node_size: The size of the nodes. Defaults to 3.0. + :param opacity: The opacity of the nodes. Defaults to 0.4. + :param cmap: The colormap to use for coloring the nodes. Defaults to "Inferno". + :param suppress_fig: Whether to suppress (i.e. not plot) the figure. Defaults to False. + :return: The plotted 3D graph. + :rtype: go.Figure + + """ + fig = go.Figure( + data=[ + go.Scatter3d( + x=coordinates["x"], + y=coordinates["y"], + z=coordinates["z"], + mode="markers", + marker=dict( + size=node_size, + color=coordinates["color"], + colorscale=cmap, + opacity=opacity, + colorbar=dict(thickness=20, title="color"), + ), + ) + ] + ) + + if not suppress_fig: + fig.show() + return fig + + +def plot_3d_graph( + pxl_data: PixelDataset, + component: str, + marker: Union[list, None] = None, + layout_algorithm: SupportedLayoutAlgorithm | None = None, + log_scale: bool = True, + normalize: bool = False, + node_size: float = 3.0, + opacity: float = 0.4, + show_b_nodes: bool = False, + cmap: str = "Inferno", + cache_layout: bool = False, + suppress_fig: bool = False, +) -> go.Figure: + """Plot a 3D graph of the specified component in the given PixelDataset. + + :param pxl_data: The PixelDataset containing the data. + :param component: The component to plot. + :param marker: The marker to use for coloring the nodes. Defaults to None. + :param layout_algorithm: The layout algorithm to use for positioning the nodes. Defaults to "fruchterman_reingold_3d". + :param log_scale: Whether to apply logarithmic scaling to the marker values. Defaults to True. + :param normalize: Whether to normalize the coordinates. Defaults to False. + :param node_size: The size of the nodes. Defaults to 3.0. + :param opacity: The opacity of the nodes. Defaults to 0.4. + :param show_b_nodes: Whether to show nodes of type B. Defaults to False. + :param cmap: The colormap to use for coloring the nodes. Defaults to "Inferno". + :param cache_layout: Whether to cache the layout coordinates. Defaults to False. + :param suppress_fig: Whether to suppress (i.e. not plot) the figure. Defaults to False. + :return: The plotted 3D graph. + :rtype: go.Figure + + """ + coordinates, _, _ = _get_coordinates( + pxl_data=pxl_data, + component=component, + layout_algorithm=layout_algorithm, + cache_layout=cache_layout, + show_b_nodes=show_b_nodes, + ) + + if marker is not None: + if log_scale: + coordinates["color"] = np.log1p(coordinates[marker]) + else: + coordinates["color"] = coordinates[marker] + else: + coordinates["color"] = Color.SKYBLUE3 + + if normalize: + coordinates[["x", "y", "z"]] = coordinates[["x_norm", "y_norm", "z_norm"]] + + fig = plot_3d_from_coordinates( + coordinates=coordinates, + node_size=node_size, + opacity=opacity, + cmap=cmap, + suppress_fig=True, + ) + + fig.update_layout(title=component) + + if not suppress_fig: + fig.show() + + return fig + + +@experimental +def plot_3d_heatmap( + component_graph: Graph, + marker: str, + distance_cutoff: float, + layout_algorithm: Literal[ + "fruchterman_reingold_3d", "kamada_kawai_3d" + ] = "fruchterman_reingold_3d", + cache_layout: bool = False, +) -> Tuple[plt.Figure, plt.Axes]: + """Plot a 3D heatmap for the marker in the provided component. + + :param component_graph: A component graph to plot for. + :param marker: marker to plot this for. + :param distance_cutoff: a distance cutoff to use for determining size of + area to consider as close in the density calculation. + :param layout_algorithm: Layout algorithm to use. Options are: + "fruchterman_reingold_3d" and "kamada_kawai_3d" + :param cache_layout: set this to `True` to cache the layout + or faster computations on subsequent calls. This comes at the + cost of additional memory usage. + :return: A matplotlib 3D heatmap figure, and it's associated Axes instance + :rtype: Tuple[plt.Figure, plt.Axes] + :raises: AssertionError if the provided `layout_algorithm` is not valid, + or there are no with markers for the provided `marker` + """ + coordinates = component_graph.layout_coordinates( + layout_algorithm=layout_algorithm, cache=cache_layout + ) + coordinates = coordinates[coordinates[marker] > 0] + coordinates = np.array(coordinates[["x_norm", "y_norm", "z_norm"]]) + if len(coordinates) < 1: + raise AssertionError(f"No nodes found with {marker}.") + + horizontal_resolution = 120 + vertical_resolution = 80 + X, Y, Z = _unit_sphere_surface( + horizontal_resolution=horizontal_resolution, + vertical_resolution=vertical_resolution, + ) + + densities = _calculate_densities( + coordinates=coordinates, + distance_cutoff=distance_cutoff, + unit_sphere_surface=(X, Y, Z), + ) + + fig = plt.figure(figsize=(6, 6)) + ax = fig.add_subplot(1, 1, 1, projection="3d") + color_scale = cm.inferno( # type: ignore + densities.reshape(horizontal_resolution, -1) + ) + ax.plot_surface( # type: ignore + X, + Y, + Z, + cstride=1, + rstride=1, + facecolors=color_scale, + shade=False, + ) + ax.set_axis_off() + ax.set_box_aspect([1.0, 1.0, 1.0]) # type: ignore + return fig, ax diff --git a/src/pixelator/plot/spatial_analysis_plots.py b/src/pixelator/plot/spatial_analysis_plots.py new file mode 100644 index 00000000..16296fb8 --- /dev/null +++ b/src/pixelator/plot/spatial_analysis_plots.py @@ -0,0 +1,325 @@ +"""Functions for creating spatial analysis plots for MPX data. + +Copyright © 2024 Pixelgen Technologies AB. +""" + +from typing import Optional, Tuple, Union + +import matplotlib.pyplot as plt +import numpy as np +import pandas as pd +import seaborn as sns + +from pixelator.analysis.colocalization import get_differential_colocalization + + +def _pivot_colocalization_data( + colocalization_data: pd.DataFrame, + value_col: str = "pearson", + markers: Union[list, None] = None, +): + colocalization_data_pivot = pd.pivot_table( + colocalization_data, + index="marker_1", + columns="marker_2", + values=value_col, + fill_value=0, + ) + + if markers is not None: + colocalization_data_pivot = colocalization_data_pivot.loc[markers, markers] + + for m in colocalization_data_pivot.index: + colocalization_data_pivot.loc[m, m] = 0 # remove autocorrelations + + colocalization_data_pivot = colocalization_data_pivot.fillna(0) + + return colocalization_data_pivot + + +def _make_colocalization_symmetric( + colocalization_data: pd.DataFrame, + value_col: str = "pearson", +): + colocalization_data = pd.DataFrame( + np.concatenate( + [ + colocalization_data[["marker_1", "marker_2", value_col]].to_numpy(), + colocalization_data[["marker_2", "marker_1", value_col]].to_numpy(), + ], + ), + columns=["marker_1", "marker_2", value_col], + ) + colocalization_data = ( + colocalization_data.groupby(["marker_1", "marker_2"])[value_col] + .apply(lambda x: np.mean(x)) + .reset_index() + ) + + return colocalization_data + + +def plot_colocalization_heatmap( + colocalization_data: pd.DataFrame, + markers: Union[list, None] = None, + cmap: str = "vlag", + use_z_scores: bool = False, +) -> Tuple[plt.Figure, plt.Axes]: + """Plot a colocalization heatmap based on the provided colocalization data. + + The colocalization_data DataFrame should contain the columns "marker_1", "marker_2", "pearson", "pearson_z". + Example usage: plot_colocalization_heatmap(pxl.colocalization). + + :param colocalization_data: The colocalization data to plot. The colocalization data frame that can be found in a pixel variable "pxl" through pxl.colocalization. The data frame should contain the columns "marker_1", "marker_2", "pearson", "pearson_z", and "component". + :param markers: The markers to include in the heatmap. Defaults to None. + :param cmap: The colormap to use for the heatmap. Defaults to "vlag". + :param use_z_scores: Whether to use z-scores. Defaults to False. + + :return: The figure and axes objects of the plot. + :rtype: Tuple[plt.Figure, plt.Axes] + + """ + if use_z_scores: + value_col = "pearson_z" + else: + value_col = "pearson" + + colocalization_data = _make_colocalization_symmetric(colocalization_data, value_col) + + colocalization_data_pivot = _pivot_colocalization_data( + colocalization_data, value_col, markers=markers + ) + sns.clustermap( + colocalization_data_pivot, + yticklabels=True, + xticklabels=True, + linewidths=0.1, + method="complete", + cmap=cmap, + ) + + return plt.gcf(), plt.gca() + + +def _get_top_marker_pairs( + colocalization_data: pd.DataFrame, + n_top_marker_pairs: int, + value_col: str = "pearson", +) -> list: + colocalization_data["abs_val"] = colocalization_data[value_col].abs() + top_marker_pairs = colocalization_data.nlargest(n_top_marker_pairs, "abs_val") + top_markers = list( + set(top_marker_pairs["marker_1"]).union(set(top_marker_pairs["marker_2"])) + ) + + return top_markers + + +def plot_colocalization_diff_heatmap( + colocalization_data: pd.DataFrame, + target: str, + reference: str, + contrast_column: str = "sample", + markers: Union[list, None] = None, + n_top_marker_pairs: Union[int, None] = None, + cmap: str = "vlag", + use_z_score: bool = True, +) -> Tuple[plt.Figure, plt.Axes]: + """Plot the differential colocalization between reference and target components. + + Example usage: plot_colocalization_diff_heatmap(pxl.colocalization, target:"stimulated", reference:"control", contrast_column="sample"). + + :param colocalization_data: The colocalization data frame that can be found in a pixel variable "pxl" through pxl.colocalization. The data frame should contain the columns "marker_1", "marker_2", "pearson", "pearson_z", and the contrast_column. + :param target: The label for target components in the contrast_column. + :param reference: The label for reference components in the contrast_column. + :param contrast_column: The column to use for the contrast. Defaults to "sample". + :param markers: The markers to include in the heatmap. Defaults to None. At most only one of n_top_marker_pairs or markers should be provided. + :param n_top_marker_pairs: The number of top marker pairs to include in the heatmap. Defaults to None. At most only one of n_top_marker_pairs or markers should be provided. + :param cmap: The colormap to use for the heatmap. Defaults to "vlag". + :param use_z_score: Whether to use the z-score. Defaults to True. + + :return: The figure and axes objects of the plot. + :rtype: Tuple[plt.Figure, plt.Axes] + """ + assert ( + markers is None or n_top_marker_pairs is None + ), "Only one of markers or n_top_marker_pairs can be provided." + + if use_z_score: + value_col = "pearson_z" + else: + value_col = "pearson" + + if markers is not None: + filter_mask = (colocalization_data["marker_1"].isin(markers)) & ( + colocalization_data["marker_2"].isin(markers) + ) + colocalization_data = colocalization_data[filter_mask] + + differential_colocalization = get_differential_colocalization( + colocalization_data, + target=target, + reference=reference, + contrast_column=contrast_column, + use_z_score=use_z_score, + ) + + differential_colocalization = differential_colocalization.fillna(0).reset_index() + + if n_top_marker_pairs is not None: + top_markers = _get_top_marker_pairs( + differential_colocalization, n_top_marker_pairs, "median_difference" + ) + else: + top_markers = None + + # Making the differential colocalization symmetric + differential_colocalization = _make_colocalization_symmetric( + differential_colocalization, "median_difference" + ) + + pivoted_differential_colocalization = _pivot_colocalization_data( + differential_colocalization, + "median_difference", + markers=top_markers, + ) + + max_value = np.max(np.abs(pivoted_differential_colocalization.to_numpy().flatten())) + sns.clustermap( + pivoted_differential_colocalization, + yticklabels=True, + xticklabels=True, + method="complete", + linewidths=0.1, + vmin=-max_value, + vmax=max_value, + cmap=cmap, + ) + + return plt.gcf(), plt.gca() + + +def _add_top_marker_labels( + differential_colocalization, + ax, + n_top_pairs: int = 5, + min_log_p: float = 5.0, +): + differential_colocalization = differential_colocalization.sort_values( + "median_difference" + ) + differential_colocalization = differential_colocalization.loc[ + -np.log10(differential_colocalization["p_adj"]) > min_log_p, : + ] + + ## Labels for marker pair withs highest negative differential colocalization scores + for _, row in differential_colocalization.head(n_top_pairs).iterrows(): + x, y = row[["median_difference", "p_adj"]] + y = -np.log10(y) + if x > 0: + continue + ax.text(x, y, row["markers"], horizontalalignment="left", fontsize="xx-small") + + ## Labels for marker pair with highest positive differential colocalization scores + for _, row in differential_colocalization.tail(n_top_pairs).iterrows(): + x, y = row[["median_difference", "p_adj"]] + y = -np.log10(y) + if x < 0: + continue + ax.text(x, y, row["markers"], horizontalalignment="left", fontsize="xx-small") + + +def _add_target_mean_colocalizations( + differential_colocalization, + target_coloc, + value_col, +): + differential_colocalization = differential_colocalization.fillna(0).reset_index() + + target_values = ( + target_coloc.groupby(["marker_1", "marker_2"])[value_col].mean().reset_index() + ) + differential_colocalization = pd.merge( + target_values, + differential_colocalization, + on=["marker_1", "marker_2"], + how="inner", + ) + + return differential_colocalization + + +def plot_colocalization_diff_volcano( + colocalization_data: pd.DataFrame, + target: str, + reference: str, + contrast_column: str = "sample", + cmap: str = "vlag", + use_z_score: bool = True, + n_top_pairs: int = 5, + min_log_p: float = 5.0, + ax: Optional[plt.Axes] = None, +) -> Tuple[plt.Figure, plt.Axes]: + """Generate the volcano plot of differential colocalization between reference and target components. + + Example usage: `plot_colocalization_diff_volcano(pxl.colocalization, target:"stimulated", reference:"control", contrast_column="sample")`. + + :param colocalization_data: The colocalization data frame that can be found in a pixel variable + "pxl" through pxl.colocalization. The data frame should contain the + columns "marker_1", "marker_2", "pearson", "pearson_z", and the contrast_column. + :param target: The label for target components in the contrast_column. + :param reference: The label for reference components in the contrast_column. + :param contrast_column: The column to use for the contrast. Defaults to "sample". + :param cmap: The colormap to use for the heatmap. Defaults to "vlag". + :param use_z_score: Whether to use the z-score. Defaults to True. + :param n_top_pairs: Number of high value marker-pairs to label from positive and negative sides. + :param min_log_p: marker-pairs only receive a label if -log10 of their p-value is higher than + this parameter. + + :return: The figure and axes objects of the plot. + :rtype: Tuple[plt.Figure, plt.Axes] + """ + if use_z_score: + value_col = "pearson_z" + else: + value_col = "pearson" + + differential_colocalization = get_differential_colocalization( + colocalization_data, + target=target, + reference=reference, + contrast_column=contrast_column, + use_z_score=use_z_score, + ) + + differential_colocalization = _add_target_mean_colocalizations( + differential_colocalization, + target_coloc=colocalization_data.loc[ + colocalization_data[contrast_column] == target, : + ], + value_col=value_col, + ) + + if ax is None: + fig, ax = plt.subplots() + + p = ax.scatter( + x=differential_colocalization["median_difference"], + y=-np.log10(differential_colocalization["p_adj"]), + c=differential_colocalization[value_col], + s=20, + marker="o", + cmap=cmap, + ) + + ax.set(xlabel="Median difference", ylabel=r"$-\log_{10}$(adj. p-value)") + fig = plt.gcf() + fig.colorbar(p, label="Mean target colocalization score", cmap=cmap) + _add_top_marker_labels( + differential_colocalization, + n_top_pairs=n_top_pairs, + min_log_p=min_log_p, + ax=ax, + ) + + return fig, ax diff --git a/src/pixelator/report/common/reporting.py b/src/pixelator/report/common/reporting.py index 9b2ee3a7..43c48a4e 100644 --- a/src/pixelator/report/common/reporting.py +++ b/src/pixelator/report/common/reporting.py @@ -55,6 +55,9 @@ def __init__(self, workdir: Path | PixelatorWorkdir): else: self.workdir = PixelatorWorkdir(workdir) + # all pixelator commands in defined order + self._command_list: list[str] | None = None + def samples(self) -> set[str]: """Return a list of all samples encountered from command metadata files.""" return self.workdir.samples() @@ -353,7 +356,6 @@ def cli_invocation_info(self, sample: str, cache: bool = True) -> CLIInvocationI :raises WorkdirOutputNotFound: If no commandline metadata is found for the sample """ # Function scope import to avoid circular dependencies - from pixelator.cli import main_cli as click_context try: metadata_files = self.workdir.metadata_files(sample, cache=cache) @@ -361,12 +363,42 @@ def cli_invocation_info(self, sample: str, cache: bool = True) -> CLIInvocationI e.message = f'No command line metadata found for sample: "{sample}"' raise - data_flat: list[CommandInfo] = [] - order = list(click_context.commands["single-cell"].commands.keys()) # type: ignore - + data_flat = [] for f in metadata_files: command_info_flat = CommandInfo.from_json(f) data_flat.append(command_info_flat) - data_flat.sort(key=lambda x: order.index(x.command.split(" ")[-1])) + if self._command_list is None: + self._command_list = _ordered_pixelator_commands() + + # mypy cannot detect that self._command_list cannot be None + # anymore at this point so we cast it explicitly. + data_flat.sort( + key=lambda x: typing.cast("list[str]", self._command_list).index(x.command) + ) return CLIInvocationInfo(data_flat, sample_id=sample) + + +def _ordered_pixelator_commands() -> list[str]: + """Return a list of pixelator CLI commands in depth-first order.""" + # local imports to avoid circular dependency issues + import click + + from pixelator.cli import main_cli as click_context + + def build_command_list(obj: typing.Any, prefix=None): + """Recursively go through groups and commands to create a list of commands.""" + prefix = prefix or [] + + if isinstance(obj, click.Group): + res = [] + new_prefix = prefix + [obj.name] + + for subcommand in obj.commands.values(): + res.extend(build_command_list(subcommand, prefix=new_prefix)) + + return res + + return (" ".join(prefix + [obj.name]),) + + return build_command_list(click_context) diff --git a/src/pixelator/report/models/annotate.py b/src/pixelator/report/models/annotate.py index c7d9f3ba..4b49c5e5 100644 --- a/src/pixelator/report/models/annotate.py +++ b/src/pixelator/report/models/annotate.py @@ -117,7 +117,7 @@ def pixel_count(self) -> int: # noqa: D102 # ------------------------------------------------------------------------------- # aggregate_count: int | None = pydantic.Field( - description="The number of components identified as aggregates and removed from results." + description="The number of components identified as aggregates." ) molecules_in_aggregates_count: int | None = pydantic.Field( @@ -128,6 +128,23 @@ def pixel_count(self) -> int: # noqa: D102 description="The total number of reads for unique molecules in aggregates." ) + @pydantic.computed_field( + return_type=float, + description=textwrap.dedent( + """The fraction of components identified as aggregates or + None if aggregate recovery was disabled. + """ + ), + ) + def fraction_aggregate_components(self) -> float | None: + """Return the fraction of the total cell count that is marked as aggregates. + + Returns None if aggregate recovery was disabled during analysis. + """ + if self.aggregate_count is not None and self.cell_count > 0: + return self.aggregate_count / self.cell_count + return None + @pydantic.computed_field( return_type=float, description=textwrap.dedent( diff --git a/src/pixelator/report/qcreport/builder.py b/src/pixelator/report/qcreport/builder.py index c6b1cee0..e55101bb 100644 --- a/src/pixelator/report/qcreport/builder.py +++ b/src/pixelator/report/qcreport/builder.py @@ -46,7 +46,7 @@ class QCReportBuilder: """ _JSON_OPTIONS: ClassVar[Dict[str, Any]] = {"indent": None, "separators": (",", ":")} - VERSIONS_CONSTRAINTS: ClassVar[List[str]] = ["<0.14.0", ">=0.10.0"] + VERSIONS_CONSTRAINTS: ClassVar[List[str]] = ["<0.15.0", ">=0.10.0"] def __init__(self, template: Union[str, Path] = DEFAULT_QC_REPORT_TEMPLATE): """Construct a QC report builder given a html template. diff --git a/src/pixelator/report/qcreport/main.py b/src/pixelator/report/qcreport/main.py index d0dafab7..456a5220 100644 --- a/src/pixelator/report/qcreport/main.py +++ b/src/pixelator/report/qcreport/main.py @@ -93,6 +93,7 @@ def create_dynamic_report( # Map pixelator metrics to QC report metrics metrics = Metrics( number_of_cells=annotate_metrics.cell_count, + fraction_outlier_cells=annotate_metrics.fraction_aggregate_components, average_reads_usable_per_cell=int( annotate_metrics.read_count_per_cell_stats.mean ), diff --git a/src/pixelator/report/qcreport/template.html b/src/pixelator/report/qcreport/template.html index 68d1b459..eba575e4 100644 --- a/src/pixelator/report/qcreport/template.html +++ b/src/pixelator/report/qcreport/template.html @@ -1,4 +1,4 @@ -
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