biodiversitycellatlas · bonitavw · Feb 6, 2026 · Feb 5, 2026 · Feb 5, 2026 · Feb 6, 2026
diff --git a/bin/calculate_rrna_mtdna.sh b/bin/calculate_rrna_mtdna.sh
@@ -45,6 +45,24 @@ echo -e "Number of ribosomal RNA reads in uniquely mapped reads,$rrna" >> $outfi
 perc_rrna=$(awk -v r="$rrna" -v u="$uniquely_mapped" 'BEGIN {printf "%.4f", (r/u)}')
 echo -e "Percentage of rRNA reads (of uniquely mapped reads),$perc_rrna" >> $outfile
 
+# Count rRNA in multimapped primary alignments
+featureCounts -t "${grep_rrna}" -a ${ref_gtf} -o feat_counts_rRNA_mmpa.txt multimapped_primealign.bam
+rrna_mmpa=$(cat feat_counts_rRNA_mmpa.txt.summary | grep -E "Assigned" | awk '{print $2}')
+echo -e "rRNA counts in Multimapped reads (primary alignment),$rrna_mmpa" >> $outfile
+
+# Percentage of rRNA in multimapped primary alignments
+perc_rrna_mmpa=$(awk -v r="$rrna_mmpa" -v tot="$total_mmpa" 'BEGIN {printf "%.4f", (r/tot)}')
+echo -e "Percentage of rRNA in multimapped reads (primary alignment),$perc_rrna_mmpa" >> $outfile
+
+# Count rRNA in multimapped all alignments
+featureCounts -t "${grep_rrna}" -a ${ref_gtf} -o feat_counts_rRNA_mmaa.txt multimapped_allalign.bam
+rrna_mmaa=$(cat feat_counts_rRNA_mmaa.txt.summary | grep -E "Assigned" | awk '{print $2}')
+echo -e "rRNA counts in Multimapped reads (all alignments),$rrna_mmaa" >> $outfile
+
+# Percentage of rRNA in multimapped all alignments
+perc_rrna_mmaa=$(awk -v r="$rrna_mmaa" -v tot="$total_mmaa" 'BEGIN {printf "%.4f", (r/tot)}')
+echo -e "Percentage of rRNA in multimapped reads (all alignments),$perc_rrna_mmaa" >> $outfile
+
 
 # ------------------------------------------------------------------
 # Calculate mtDNA metrics

diff --git a/modules/local/custom/demultiplex/parsebio_demultiplex/main.nf b/modules/local/custom/demultiplex/parsebio_demultiplex/main.nf
@@ -10,12 +10,11 @@ process PARSEBIO_CUSTOM_DEMUX {
     tuple val(meta), path(fastq_cDNA), path(fastq_BC_UMI), path(fastq_indices), path(input_file)
 
     output:
-    tuple val(meta), path("*group*_R1*"), path("*group*_R2*"), path(input_file), emit: splitted_files
+    tuple val(meta), path("*group*_R1*"), path("*group*_R2*"), path(fastq_indices), path(input_file), emit: splitted_files
 
     script:
     """
     echo "\n\n==================  Parse Biosciences: Custom Demultiplexing  =================="
-    echo "Conda environment: \$CONDA_DEFAULT_ENV"
     echo "Processing sample: ${meta}"
     echo "Fastq files: ${fastq_cDNA}, ${fastq_BC_UMI}"
     echo "Group id: ${meta.id}, wells (rt): ${meta.rt}"

diff --git a/modules/local/tools/cellbender/main.nf b/modules/local/tools/cellbender/main.nf
@@ -3,10 +3,7 @@ process CELLBENDER {
     tag "${meta.id}"
     label 'process_high_memory'
 
-    container "${ task.ext.use_gpu ? 'us.gcr.io/broad-dsde-methods/cellbender:0.3.2' :
-        workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://community-cr-prod.seqera.io/docker/registry/v2/blobs/sha256/eb/ebcf140f995f79fcad5c17783622e000550ff6f171771f9fc4233484ee6f63cf/data':
-        'community.wave.seqera.io/library/cellbender_webcolors:156d413fdfc16cdb' }"
+    container "oras://community.wave.seqera.io/library/cellbender:0.3.2--4f86af6695399b4f"
 
     input:
     tuple val(meta), path(mapping_files)

diff --git a/modules/local/tools/cutadapt/main.nf b/modules/local/tools/cutadapt/main.nf
@@ -4,9 +4,7 @@ process RM_VARBASES {
 
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://community-cr-prod.seqera.io/docker/registry/v2/blobs/sha256/17/1758869538eb8e658077cc14cd7a4e76fd9b6d73d3a68f85a70bf292e39e27c5/data' :
-        'community.wave.seqera.io/library/cutadapt:5.0--991bbd2e184b7014' }"
+    container "oras://community.wave.seqera.io/library/cutadapt:5.2--5505472a18e9cce0"
 
     input:
     tuple val(meta), path(fastq_cDNA), path(fastq_BC_UMI), path(fastq_indices), path(input_file)

diff --git a/modules/local/tools/fastp/main.nf b/modules/local/tools/fastp/main.nf
@@ -5,9 +5,7 @@ process FASTP {
     debug true
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://community-cr-prod.seqera.io/docker/registry/v2/blobs/sha256/88/889a182b8066804f4799f3808a5813ad601381a8a0e3baa4ab8d73e739b97001/data' :
-        'community.wave.seqera.io/library/fastp:0.24.0--62c97b06e8447690' }"
+    container "oras://community.wave.seqera.io/library/fastp:0.24.0--0397de619771c7ae"
 
     input:
     tuple val(meta), path(fastq_cDNA), path(fastq_BC_UMI), path(fastq_indices), path(input_file)

diff --git a/modules/local/tools/fastqc/main.nf b/modules/local/tools/fastqc/main.nf
@@ -4,9 +4,7 @@ process FASTQC {
     label 'process_low'
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://depot.galaxyproject.org/singularity/fastqc:0.12.1--hdfd78af_0' :
-        'quay.io/biocontainers/fastqc:0.12.1--hdfd78af_0' }"
+    container "oras://community.wave.seqera.io/library/fastqc:0.12.1--104d26ddd9519960"
 
     input:
     tuple val(meta), path(fastq_cDNA), path(fastq_BC_UMI), path(fastq_indices), path(input_file)

diff --git a/modules/local/tools/kraken/kraken_classify/main.nf b/modules/local/tools/kraken/kraken_classify/main.nf
@@ -8,9 +8,7 @@ process KRAKEN {
     tuple val(meta), path(mapping_files)
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://community-cr-prod.seqera.io/docker/registry/v2/blobs/sha256/29/29ed8f68315625eca61a3de9fcb7b8739fe8da23f5779eda3792b9d276aa3b8f/data' :
-        'community.wave.seqera.io/library/kraken2_coreutils_pigz:45764814c4bb5bf3' }"
+    container "oras://community.wave.seqera.io/library/kraken2_samtools_coreutils_pigz:8961943c277652a6"
 
     output:
     path("*")

diff --git a/modules/local/tools/krona/main.nf b/modules/local/tools/krona/main.nf
@@ -4,9 +4,7 @@ process KRONA {
 
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://depot.galaxyproject.org/singularity/krona:2.8--pl5262hdfd78af_2' :
-        'quay.io/biocontainers/krona:2.8--pl5262hdfd78af_2' }"
+    container "oras://community.wave.seqera.io/library/krona:2.8--8fc5aef4acd456a6"
 
     input:
     val(trigger)

diff --git a/modules/local/tools/multiqc/main.nf b/modules/local/tools/multiqc/main.nf
@@ -4,9 +4,7 @@ process MULTIQC {
 
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://depot.galaxyproject.org/singularity/multiqc:1.30--pyhdfd78af_1' :
-        'quay.io/biocontainers/multiqc:1.30--pyhdfd78af_1' }"
+    container "oras://community.wave.seqera.io/library/multiqc:1.30--d3e586af7b974fba"
 
     input:
     val(trigger)

diff --git a/modules/local/tools/salmon_alevin/alevin-fry/main.nf b/modules/local/tools/salmon_alevin/alevin-fry/main.nf
@@ -5,9 +5,7 @@ process ALEVIN_FRY {
 
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://depot.galaxyproject.org/singularity/simpleaf:0.19.4--ha6fb395_0':
-        'quay.io/biocontainers/simpleaf:0.19.4--ha6fb395_0' }"
+    container "oras://community.wave.seqera.io/library/alevin-fry_piscem_salmon_simpleaf_pruned:c71cfb476b003414"
 
     input:
     tuple val(meta), path(fastq_cDNA), path(fastq_BC_UMI), path(fastq_indices), path(input_file)

diff --git a/modules/local/tools/salmon_alevin/salmon_index/main.nf b/modules/local/tools/salmon_alevin/salmon_index/main.nf
@@ -4,9 +4,7 @@ process SALMON_INDEX {
 
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://depot.galaxyproject.org/singularity/simpleaf:0.19.4--ha6fb395_0':
-        'quay.io/biocontainers/simpleaf:0.19.4--ha6fb395_0' }"
+    container "oras://community.wave.seqera.io/library/salmon:1.10.3--726401738a281398"
 
     input:
     tuple val(meta), path(fastq_cDNA), path(fastq_BC_UMI), path(fastq_indices), path(input_file)

diff --git a/modules/local/tools/samtools/samtools_index/main.nf b/modules/local/tools/samtools/samtools_index/main.nf
@@ -4,9 +4,7 @@ process SAMTOOLS_INDEX {
     label 'process_single'
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://depot.galaxyproject.org/singularity/samtools:1.22.1--h96c455f_0' :
-        'quay.io/biocontainers/samtools:1.22.1--h96c455f_0' }"
+    container "oras://community.wave.seqera.io/library/samtools:1.22.1--9a10f06c24cdf05f"
 
     input:
     tuple val(meta), path(mapping_files)

diff --git a/modules/local/tools/samtools/samtools_view/main.nf b/modules/local/tools/samtools/samtools_view/main.nf
@@ -4,9 +4,7 @@ process SAMTOOLS_VIEW {
     label 'process_single'
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://depot.galaxyproject.org/singularity/samtools:1.22.1--h96c455f_0' :
-        'quay.io/biocontainers/samtools:1.22.1--h96c455f_0' }"
+    container "oras://community.wave.seqera.io/library/samtools:1.22.1--9a10f06c24cdf05f"
 
     input:
     tuple val(meta), path(mapping_files)

diff --git a/modules/local/tools/scirocket/scirocket_demux/main.nf b/modules/local/tools/scirocket/scirocket_demux/main.nf
@@ -4,8 +4,8 @@ process SCIROCKET_DEMUX {
     label 'process_medium'
     debug true
 
-
     conda "${moduleDir}/environment.yml"
+    container "oras://community.wave.seqera.io/library/fastp_pysam_sambamba_star_pruned:ac8b1ecd993405dd"
 
     input:
     tuple val(meta), path(fastq_cDNA), path(fastq_BC_UMI), path(fastq_indices), path(input_file)
@@ -29,7 +29,6 @@ process SCIROCKET_DEMUX {
 
     """
     echo "\n\n==================  DEMULTIPLEXING FASTQ FILES  =================="
-    echo "Conda environment: \$CONDA_DEFAULT_ENV"
     echo "Sample ID: ${meta.id}"
     echo "FASTQ cDNA: ${fastq_cDNA}"
     echo "FASTQ BC & UMI: ${fastq_BC_UMI}"

diff --git a/modules/local/tools/star/starsolo_genome_generate/main.nf b/modules/local/tools/star/starsolo_genome_generate/main.nf
@@ -7,9 +7,7 @@ process STARSOLO_INDEX {
     path ref_gtf
 
     conda "${moduleDir}/environment.yml"
-    container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://community-cr-prod.seqera.io/docker/registry/v2/blobs/sha256/26/268b4c9c6cbf8fa6606c9b7fd4fafce18bf2c931d1a809a0ce51b105ec06c89d/data' :
-        'community.wave.seqera.io/library/htslib_samtools_star_gawk:ae438e9a604351a4' }"
+    container "oras://community.wave.seqera.io/library/htslib_samtools_star_gawk:f196f82abbbc8871"
 
     output:
     path("*")