Ale module #931

PetcuBogdan · 2025-11-16T09:02:25Z

PR checklist

This comment contains a description of changes (with reason).
If you've added a new tool - have you followed the pipeline conventions in the contribution docs
Ensure the test suite passes (nextflow run . -profile test,docker --outdir <OUTDIR>).
Check for unexpected warnings in debug mode (nextflow run . -profile debug,test,docker --outdir <OUTDIR>).
Usage Documentation in docs/usage.md is updated.
Output Documentation in docs/output.md is updated.
CHANGELOG.md is updated.
README.md is updated (including new tool citations and authors/contributors).

Description

This PR adds ALE (Assembly Likelihood Estimator) for assembly quality control in nf-core/mag.
ALE is a probabilistic framework that evaluates assembly quality by computing the likelihood of sequencing reads given an assembly. It provides per-contig quality scores useful for identifying misassemblies, comparing assemblies, and validating quality before binning.

Changes made

Workflow:

Added ALE analysis for short-read assemblies (SPAdes, MEGAHIT)
Runs when binning or ancient DNA analysis is enabled
Reuses existing BAM files from binning preparation

References

GitHub: https://github.com/sc932/ALE
Issue link Integrate the nf-core module for ALE (Assembly Likelihood Evaluation) #925

Release v5.1.0

Release: v5.2.0 Puce Pangolin

jfy133 · 2025-11-16T10:52:43Z

@nf-core-bot fix linting

PetcuBogdan · 2025-11-16T16:26:48Z

Please let me know what I can improve. Thank you!

d4straub

Ci tests fail with various errors. https://github.com/nf-core/mag/actions/runs/19404507607/job/55538896302?pr=931 fails with

    > ERROR ~ Error executing process > 'NFCORE_MAG:MAG:ALE (minigut)'
    > 
    > Caused by:
    >   Process `NFCORE_MAG:MAG:ALE (minigut)` terminated with an error exit status (134)
    > 
    > 
    > Command executed:
    > 
    >   ALE \
    >        \
    >       SPAdesHybrid-minigut-minigut.bam \
    >       SPAdesHybrid-minigut.scaffolds.fa \
    >       minigut_ALEoutput.txt
    >   
    >   cat <<-END_VERSIONS > versions.yml
    >   "NFCORE_MAG:MAG:ALE":
    >       ale: 20180904
    >   END_VERSIONS
    > 
    > Command exit status:
    >   134
    > 
    > Command output:
    >   BAM file: SPAdesHybrid-minigut-minigut.bam
    >   Assembly fasta file: SPAdesHybrid-minigut.scaffolds.fa
    >   ALE Output file: minigut_ALEoutput.txt
    >   Reading in assembly...
    >   Reading in the map and computing statistics...
    >   Insert length and std not given, will be calculated from input map.
    >   Found FR sample avg insert length to be 383.864169 from 28344 mapped reads
    >   Found FR sample insert length std to be 69.336488
    >   Found NOT_PROPER_FR sample avg insert length to be 892.122675 from 66297 mapped reads
    >   Found NOT_PROPER_FR sample insert length std to be 221.969163
    >   There were 99620 total reads, 99620 paired (97898 properly mated), 763 proper singles, 959 improper reads (818 chimeric). (83 reads were unmapped)
    >   Saved library parameters to minigut_ALEoutput.txt.param
    >   Computing read placements and depths
    > 
    > Command error:
    >   WARNING: The following read and its mate do not agree on the contigs and/or positions of their mappings:read1: NC_006347.1_4981 81: 0 0 106315 105875	read2: NC_006347.1_4981 161: 0 0 105578 106537	l: 1.000000 li: 1.000000, s1: 106315, s2: 105875, e1: 106441, e2: -1, c1: 0, c2: 0, NC_006347.1_4981, NOT_PROPER_FR, 0, b1: 34e7c540, b2: 0
    >   ALE: ALElike.c:1892: validateAlignmentMates: Assertion `thisAlignment->start2 == thisReadMate->core.pos' failed.
    >   BAM file: SPAdesHybrid-minigut-minigut.bam
    >   Assembly fasta file: SPAdesHybrid-minigut.scaffolds.fa
    >   ALE Output file: minigut_ALEoutput.txt
    >   Reading in assembly...
    >   Reading in the map and computing statistics...
    >   Insert length and std not given, will be calculated from input map.
    >   Found FR sample avg insert length to be 383.864169 from 28344 mapped reads
    >   Found FR sample insert length std to be 69.336488
    >   Found NOT_PROPER_FR sample avg insert length to be 892.122675 from 66297 mapped reads
    >   Found NOT_PROPER_FR sample insert length std to be 221.969163
    >   There were 99620 total reads, 99620 paired (97898 properly mated), 763 proper singles, 959 improper reads (818 chimeric). (83 reads were unmapped)
    >   Saved library parameters to minigut_ALEoutput.txt.param
    >   Computing read placements and depths
    >   .command.sh: line 6:    34 Aborted                 (core dumped) ALE SPAdesHybrid-minigut-minigut.bam SPAdesHybrid-minigut.scaffolds.fa minigut_ALEoutput.txt
    > 
    > Work dir:
    >   /home/runner/_work/mag/mag/~/tests/b1878932db1a90503becf8394b4ddfd4/work/f4/88d2098735b2dd029b42b1a840ced7
    > 
    > Container:
    >   quay.io/biocontainers/ale:20180904--py27ha92aebf_0
    > 
    > Tip: you can replicate the issue by changing to the process work dir and entering the command `bash .command.run`
    > 
    >  -- Check '/home/runner/_work/mag/mag/~/tests/b1878932db1a90503becf8394b4ddfd4/meta/nextflow.log' file for details
    > ERROR ~ Could not find which method load() to invoke from this list:
    >   public java.lang.Object org.yaml.snakeyaml.Yaml#load(java.io.InputStream)
    >   public java.lang.Object org.yaml.snakeyaml.Yaml#load(java.io.Reader)
    >   public java.lang.Object org.yaml.snakeyaml.Yaml#load(java.lang.String)
    >   public java.lang.Object org.yaml.snakeyaml.Yaml#load(java.io.File)
    >   public java.lang.Object org.yaml.snakeyaml.Yaml#load(java.nio.file.Path)
    > 
    >  -- Check script '/home/runner/_work/mag/mag/subworkflows/nf-core/utils_nfcore_pipeline/main.nf' at line: 82 or see '/home/runner/_work/mag/mag/~/tests/b1878932db1a90503becf8394b4ddfd4/meta/nextflow.log' file for more details
    > ERROR ~ Pipeline failed. Please refer to troubleshooting docs: https://nf-co.re/docs/usage/troubleshooting
    > 
    >  -- Check '/home/runner/_work/mag/mag/~/tests/b1878932db1a90503becf8394b4ddfd4/meta/nextflow.log' file for details
    > -[nf-core/mag] Pipeline completed with errors-
    > WARN: Killing running tasks (1)
    FAILED (481.488s)

Additionally, test https://github.com/nf-core/mag/actions/runs/19404507607/job/55538896283?pr=931 indicates that ALE is run but output files are not published to the results folder:

    2     {                                 2     {                            
    3         "ADJUST_MAXBIN2_EXT": {       3         "ADJUST_MAXBIN2_EXT": {  
    4             "coreutils": 9.5          4             "coreutils": 9.5     
                                        +   5         },                       
                                        +   6         "ALE": {                 
                                        +   7             "ale": 20180904      
    5         },                            8         },                       
    6         "BIN_SUMMARY": {              9         "BIN_SUMMARY": {         
    7             "pandas": "1.4.3",       10             "pandas": "1.4.3",

docs/usage.md

d4straub · 2025-11-17T07:13:01Z

workflows/mag.nf

+
+            1. Enable binning: --skip_binning false
+            2. Enable ancient DNA: --ancient_dna true
+            3. Disable ALE: --skip_ale true


But --skip_ale true will not run ALE?
Maybe rather

Suggested change

3. Disable ALE: --skip_ale true

To avoid that warning disable ALE: --skip_ale

also using --skip_ale is eqivalent to --skip_ale true.

d4straub · 2025-11-17T07:14:36Z

workflows/mag.nf

+    if(!params.skip_ale) {
+        if ( !params.skip_binning || params.ancient_dna) {
+            ch_shortread_assemblies_for_ale = ch_assemblies.filter { meta, assembly ->
+                meta.assembler?.toUpperCase() in ['SPADES', 'SPADESHYBRID', 'MEGAHIT']


I am not a particular fan of defining assembler names here, because in case another short read assembler is added in the future, one has to remember to add that to the list here. Probably there is a better solution?

nextflow_schema.json

Co-authored-by: Daniel Straub <42973691+d4straub@users.noreply.github.com>

prototaxites

Hi @PetcuBogdan, a few thoughts from me!

prototaxites · 2025-11-19T10:25:52Z

conf/modules.config


+    withName: 'NFCORE_MAG:MAG:ALE' {
+        publishDir = [
+            path: { "${params.outdir}/Assembly/${meta.assembler?.toUpperCase() ?: 'UNKNOWN'}/QC/${meta.id}/ALE" },


Suggested change

path: { "${params.outdir}/Assembly/${meta.assembler?.toUpperCase() ?: 'UNKNOWN'}/QC/${meta.id}/ALE" },

path: { "${params.outdir}/Assembly/${meta.assembler}/QC/${meta.id}/ALE" },

If there is ever a case where meta.assembler isn't set, that's a bug. And it will be put in a directory called null in that case, so this is unnecessary.

prototaxites · 2025-11-19T10:26:24Z

conf/modules.config

+            pattern: "*.{ale,txt,log}",
+            saveAs: { filename -> filename.equals('versions.yml') ? null : filename }
+        ]
+        ext.prefix = { "${meta.id}" }


This should be something like this?

Suggested change

ext.prefix = { "${meta.id}" }

ext.prefix = { "${meta.id}-${meta.assembler}" }

prototaxites · 2025-11-19T10:26:58Z

docs/output.md

+
+[ALE (Assembly Likelihood Estimator)](https://github.com/sc932/ALE) is a probabilistic framework that evaluates assembly quality by computing the likelihood of the sequencing reads given an assembly. ALE provides per-contig quality scores and identifies potentially problematic regions in assemblies by analyzing read mapping patterns and insert size distributions. It is particularly useful for comparing assemblies and identifying misassemblies or low-confidence regions.
+
+ALE is run on short-read assemblies (SPAdes, SPAdes hybrid, and MEGAHIT) when binning or ancient DNA analysis is enabled.


If this is just an assembly quality tool, it should run even when binning is off, no?

prototaxites · 2025-11-19T10:28:57Z

workflows/mag.nf

+            log.warn """
+            [nf-core/mag] ALE (Assembly Likelihood Estimator) Warnings
+
+            ALE is enabled (--skip_ale false) but cannot run because:
+            - Binning is disabled (--skip_binning true)
+            - Ancient DNA mode is not enabled (--ancient_dna false)
+
+            To run ALE, choose one of the following options:
+
+            1. Enable binning: --skip_binning false
+            2. Enable ancient DNA: --ancient_dna true
+            3. Disable ALE: --skip_ale true


IMO we don't need to have a long explainer about /why/ ALE doesn't run? It should either not run, silently, or print out a warning on a single line.

prototaxites · 2025-11-19T10:29:39Z

workflows/mag.nf

+    */
+
+    if(!params.skip_ale) {
+        if ( !params.skip_binning || params.ancient_dna) {


We should just pass assemblies into this and not worry about whether binning is enabled, no?

And since this is an assembly QC tool, should this logically be next to the QUAST section in this file?

Hi thanks for the feedback!

Since ALE requires BAM files for the likelihood calculation, it needs access to the mapped reads produced in BINNING_PREPARATION.
Should I move BINNING_PREPARATION earlier in the workflow, or do you have another suggestion?

Oh, of course 🤦‍♂️ In that case, ignore this comment entirely!

PetcuBogdan and others added 6 commits October 23, 2025 10:13

Add ale module and call ale from mag.nf

0eeb3df

Merge pull request nf-core#893 from nf-core/dev

b0bc5ca

Release v5.1.0

add ale result output path

785e7f4

Merge pull request nf-core#914 from nf-core/dev

99859fc

Release: v5.2.0 Puce Pangolin

minor improvements

3779476

Merge branch 'nf-core:main' into ale_module

db0ea09

PetcuBogdan requested review from d4straub, dialvarezs, jfy133, muabnezor and prototaxites as code owners November 16, 2025 09:02

update changelog.md and usage.md

611182a

[automated] Fix code linting

d0a489d

d4straub reviewed Nov 17, 2025

View reviewed changes

PetcuBogdan and others added 2 commits November 18, 2025 20:39

fix duplicate from usage.md

fd550b7

Co-authored-by: Daniel Straub <42973691+d4straub@users.noreply.github.com>

fix misspelled from nextflow_schema

ce4298c

Co-authored-by: Daniel Straub <42973691+d4straub@users.noreply.github.com>

prototaxites reviewed Nov 19, 2025

View reviewed changes

	3. Disable ALE: --skip_ale true

	To avoid that warning disable ALE: --skip_ale

	path: { "${params.outdir}/Assembly/${meta.assembler?.toUpperCase() ?: 'UNKNOWN'}/QC/${meta.id}/ALE" },
	path: { "${params.outdir}/Assembly/${meta.assembler}/QC/${meta.id}/ALE" },

	ext.prefix = { "${meta.id}" }
	ext.prefix = { "${meta.id}-${meta.assembler}" }


		[ALE (Assembly Likelihood Estimator)](https://github.com/sc932/ALE) is a probabilistic framework that evaluates assembly quality by computing the likelihood of the sequencing reads given an assembly. ALE provides per-contig quality scores and identifies potentially problematic regions in assemblies by analyzing read mapping patterns and insert size distributions. It is particularly useful for comparing assemblies and identifying misassemblies or low-confidence regions.

		ALE is run on short-read assemblies (SPAdes, SPAdes hybrid, and MEGAHIT) when binning or ancient DNA analysis is enabled.

Ale module #931

Are you sure you want to change the base?

Ale module #931

Uh oh!

Conversation

PetcuBogdan commented Nov 16, 2025 • edited Loading Uh oh! There was an error while loading. Please reload this page.

Uh oh!

PR checklist

Uh oh!

jfy133 commented Nov 16, 2025

Uh oh!

PetcuBogdan commented Nov 16, 2025

Uh oh!

d4straub left a comment

Choose a reason for hiding this comment

Uh oh!

Uh oh!

Choose a reason for hiding this comment

Uh oh!

Choose a reason for hiding this comment

Uh oh!

Uh oh!

prototaxites left a comment

Choose a reason for hiding this comment

Uh oh!

Choose a reason for hiding this comment

Uh oh!

Choose a reason for hiding this comment

Uh oh!

Choose a reason for hiding this comment

Uh oh!

Choose a reason for hiding this comment

Uh oh!

Choose a reason for hiding this comment

Uh oh!

Choose a reason for hiding this comment

Uh oh!

Choose a reason for hiding this comment

Uh oh!

Choose a reason for hiding this comment

Uh oh!

Reviewers

Assignees

Labels

Projects

Milestone

Development

Uh oh!

6 participants

PetcuBogdan commented Nov 16, 2025 •

edited

Loading