Release 5.0.0

.nf-core.yml

@@ -24,4 +24,4 @@ template:
     - fastqc
     - multiqc
     - igenomes
-  version: 4.1.2
+  version: 5.0.0

CHANGELOG.md

@@ -3,6 +3,39 @@
 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).
 
+## [[5.0.0](https://github.com/nf-core/pixelator/releases/tag/5.0.0)] - 2026-07-13
+
+### Changed
+
+- **(breaking change)** Rename `pna_sample_calling_confidence_threshold` to `pna_sample_calling_enrichment_threshold` to match pixelator 0.29.0. By @elhb [#227](https://github.com/nf-core/pixelator/pull/227)
+- Change default layout algorithm from `wpmds_3d` to `coarsened_pmds_3d`. By @elhb [#227](https://github.com/nf-core/pixelator/pull/227)
+- Change `pna_graph_refinement_stage_leiden_resolution` and `pna_graph_refinement_stage_max_edges_to_remove` default values
+  to 0.5, and 20 respectively based on latest evaluations. By @ptajvar [#226](https://github.com/nf-core/pixelator/pull/226)
+- Set `pna_sample_calling_save_undetermined` by default and deliver the undetermined PXL file at the `pixelator/` output root, next to the final (layout) PXL files. By @Aratz [#229](https://github.com/nf-core/pixelator/pull/229)
+
+### Enhancements & fixes
+
+- Update pixelator container to 0.29.0
+- Update pixelatorES to 0.11.2
+
+### Software dependencies
+
+| Dependency    | Old version | New version |
+| ------------- | ----------- | ----------- |
+| `pixelator`   | 0.28.0      | 0.29.0      |
+| `pixelatorES` | 0.10.4      | 0.11.2      |
+
+### Parameters
+
+| Old parameter                             | New parameter                             |
+| ----------------------------------------- | ----------------------------------------- |
+| `pna_sample_calling_confidence_threshold` | `pna_sample_calling_enrichment_threshold` |
+
+> [!NOTE]
+> Parameter has been **updated** if both old and new parameter information is present.
+> Parameter has been **added** if just the new parameter information is present.
+> Parameter has been **removed** if new parameter information isn't present.
+
 ## [[4.1.2](https://github.com/nf-core/pixelator/releases/tag/4.1.2)] - 2026-06-04
 
 ### Enhancements & fixes

conf/modules.pna.config

@@ -160,15 +160,24 @@ process {
             [
                 params.pna_sample_calling_remove_incompatible ? "--remove-incompatible" : '',
                 params.pna_sample_calling_save_undetermined ? "--save-undetermined" : '',
-                params.pna_sample_calling_confidence_threshold != null ? "--confidence-threshold ${params.pna_sample_calling_confidence_threshold}" : '',
+                params.pna_sample_calling_enrichment_threshold != null ? "--enrichment-threshold ${params.pna_sample_calling_enrichment_threshold}" : '',
             ].join(' ').trim()
         }
         publishDir = [
             [
+                // Deliver the undetermined PXL file next to the final (layout) PXL files
+                path: { "${params.outdir}/pixelator" },
+                mode: params.publish_dir_mode,
+                pattern: 'sample_calling/*undetermined*.pxl',
+                saveAs: { filename -> params.pna_sample_calling_save_undetermined || params.save_all ? new File(filename).name : null },
+            ],
+            [
+                // The undetermined PXL file is delivered at the pixelator root above, so
+                // exclude it here to avoid publishing it twice.
                 path: { "${params.outdir}/pixelator" },
                 mode: params.publish_dir_mode,
                 pattern: 'sample_calling/*.pxl',
-                saveAs: { params.save_pna_sample_calling_pixelfile || params.save_all ? it : null },
+                saveAs: { (params.save_pna_sample_calling_pixelfile || params.save_all) && !it.contains('undetermined') ? it : null },
             ],
             [
                 path: { "${params.outdir}/pixelator/logs/${meta.id}" },

conf/test_proxiome_v1.config

@@ -17,9 +17,15 @@ params {
 
     // Input data
     input                                  = params.pipelines_testdata_base_path + '/samplesheet/pna/samplesheet_proxiome_v1.csv'
-    input_basedir                          = params.pipelines_testdata_base_path + '/testdata/pna'
+    input_basedir                          = params.pipelines_testdata_base_path + '/new-test-data/pna'
 
-    pna_graph_component_size_min_threshold = 100
+    // Synthetic data was generated to pass with the test with lower than default resolution
+    pna_graph_refinement_stage_leiden_resolution = 0.01
+    pna_graph_component_size_min_threshold       = 100
+
+    // Synthetic nf-core test datasets include components too small for
+    // coarsened_pmds_3d. Keep wpmds_3d in test profiles.
+    pna_layout_layout_algorithm            = "wpmds_3d"
 
     skip_layout                            = false
     technology                             = "proxiome-v1"

conf/test_proxiome_v2.config

@@ -16,10 +16,16 @@ params {
     config_profile_description             = 'Minimal test dataset to check pipeline function'
 
     // Input data
-    input                                  = params.pipelines_testdata_base_path + '/samplesheet/pna/samplesheet_proxiome_v2.csv'
-    input_basedir                          = params.pipelines_testdata_base_path + '/testdata/pna'
+    input                                  = params.pipelines_testdata_base_path + '/new-test-data/samplesheet/pna/samplesheet_proxiome_v2.csv'
+    input_basedir                          = params.pipelines_testdata_base_path + '/new-test-data/pna'
 
-    pna_graph_component_size_min_threshold = 100
+    // Synthetic data was generated to pass with the test with lower than default resolution
+    pna_graph_refinement_stage_leiden_resolution = 0.01
+    pna_graph_component_size_min_threshold       = 100
+
+    // Synthetic nf-core test datasets include components too small for
+    // coarsened_pmds_3d. Keep wpmds_3d in  test profiles.
+    pna_layout_layout_algorithm            = "wpmds_3d"
 
     skip_layout                            = false
     technology                             = "proxiome-v2"

docs/output.md

@@ -151,19 +151,26 @@ This step uses the `pixelator single-cell-pna sample-calling` command. It takes
 the graph PXL file and the sample sheet as input and assigns each component to
 the most likely sample in pooled experiments.
 
-Components with assignment confidence below the selected threshold are marked
-as undetermined and can be saved to a separate output file. The resulting
-per-sample dehashed PXL files are then used as input for the analysis step.
+Components with sample enrichment below `pna_sample_calling_enrichment_threshold`
+(default 10.0) are marked as undetermined and can be saved to a separate output
+file. The resulting per-sample dehashed PXL files are then used as input for
+the analysis step.
+
+The undetermined components do not continue through the rest of the pipeline, so
+their PXL file is delivered directly at the `pixelator/` root, next to the final
+(layout) PXL files. This is only done when `pna_sample_calling_save_undetermined = true`
+(the default). The intermediate per-sample dehashed PXL files continue downstream
+and are only copied to the `sample_calling` folder when `save_pna_sample_calling_pixelfile = true`.
 
 <details markdown="1">
 <summary>Output files</summary>
 
 - `pixelator`
+  - `<pool-id>_undetermined.dehashed.pxl`: The pixel file containing data that could not be associated with any sample. This is only output when `pna_sample_calling_save_undetermined = true`.
   - `sample_calling`
-    - `<sample-id>.dehashed.pxl`: The pixel file containing the data associated with each samples.
+    - `<sample-id>.dehashed.pxl`: The pixel file containing the data associated with each samples. Only saved when `save_pna_sample_calling_pixelfile = true`.
     - `<sample-id>.meta.json`: Command invocation metadata.
     - `<sample-id>.report.json`: QC metrics for the sample calling step.
-    - `<pool-id>_undetermined.dehashed.pxl`: The pixel file containing data that could not be associated with any sample. This is only output when `pna_sample_calling_save_undetermined = true`.
     - `<pool-id>_undetermined.meta.json`: Command invocation metadata.
     - `<pool-id>_undetermined.report.json`: QC metrics for the sample calling step.
 
@@ -257,6 +264,8 @@ generated by the pipeline (with the most "complete" information) and an interact
 The PXL dataset files can be from either the `graph`, `analysis` or `layout` step.
 
 With default parameters, the `<sample-id>.layout.pxl` will be copied to the output directory.
+The `<pool-id>_undetermined.dehashed.pxl` file is also delivered here by default, since these
+components are not processed any further by the pipeline.
 
 Various flags are available to store intermediate files and are described in the input parameter documentation. Alternatively, you can keep all intermediate files using `--save_all`.
 
@@ -275,3 +284,4 @@ Below is an example output structure for a pipeline run using the default settin
   - `pbmcs_unstimulated.qc-report.html`
   - `uropod_control.layout.pxl`
   - `uropod_control.qc-report.html`
+  - `<pool-id>_undetermined.dehashed.pxl`

modules/local/experiment_summary/main.nf

@@ -4,8 +4,8 @@ process EXPERIMENT_SUMMARY {
     label "error_retry"
 
     container "${params.experiment_summary_container?: workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container
-        ? 'quay.io/pixelgen-technologies/pixelatores:0.10.4'
-        : 'quay.io/pixelgen-technologies/pixelatores:0.10.4'}"
+        ? 'quay.io/pixelgen-technologies/pixelatores:0.11.2'
+        : 'quay.io/pixelgen-technologies/pixelatores:0.11.2'}"
 
     input:
     path samplesheet_path
@@ -22,17 +22,24 @@ process EXPERIMENT_SUMMARY {
     assert results_data instanceof List : "Expected results_data to be a List, got ${results_data?.getClass()?.simpleName ?: 'null'}"
     assert results_data.size() == result_stages.size(): "Mismatch between result files (${results_data.size()}) and stage labels (${result_stages.size()})"
 
-    def stageCopies = [result_stages, results_data].transpose().collect { stage, file ->
-        """
-        mkdir -p "results/${stage}"
-        ln -s "../../${file}" "results/${stage}/"
-        """
-    }.join('\n')
+    def stageArray = result_stages.collect { "\"${it}\"" }.join(' ')
     """
     # Copy the full quarto dir from the read-only image into the workdir
     cp -r /workspace/inst/quarto/ ./quarto/
     mkdir -p results
-    ${stageCopies}
+
+    # Stage each result file into results/<stage>/. Files are staged into
+    # results_raw/1, results_raw/2, ... in the same order as the stage names.
+    stages=(${stageArray})
+    for i in "\${!stages[@]}"; do
+        idx=\$((i + 1))
+        dest="results/\${stages[\$i]}"
+        mkdir -p "\$dest"
+        for f in "results_raw/\${idx}"/*; do
+            ln -s "../../\$f" "\$dest/"
+        done
+    done
+
     quarto render ./quarto/pixelatorES.qmd \\
         -P sample_sheet="\$PWD/${samplesheet_path}" \\
         -P data_folder="\$PWD/results/" \\