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tasks/bacterial-assembly/run_script.sh

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+#!/bin/bash
+set -e
+
+# =============================================================================
+# Bacterial Genome Assembly + Multi-Tool Annotation (Diamond DAG)
+#
+# DAG structure:
+#   FASTQ → fastp → shovill (assemble)
+#                        ↓
+#           ┌────────┬───┼────┬─────────┐
+#         Prokka  ABRicate  mlst  BUSCO  QUAST
+#         (genes) (AMR)   (type) (QC)   (stats)
+#           └────────┴───┼────┴─────────┘
+#                        ↓
+#                  Merge → CSV report
+#
+# Data: MRSA (S. aureus) Illumina paired-end (Hikichi et al. 2019)
+# =============================================================================
+
+THREADS=$(( $(nproc) > 8 ? 8 : $(nproc) ))
+SCRIPT_DIR="$(cd "$(dirname "${BASH_SOURCE[0]}")" && pwd)"
+DATA_DIR="${SCRIPT_DIR}/data"
+OUT_DIR="${SCRIPT_DIR}/outputs"
+RESULTS_DIR="${SCRIPT_DIR}/results"
+
+log_step() {
+    echo "=================================================================="
+    echo "STEP: $1"
+    echo "=================================================================="
+    echo "Started at: $(date)"
+}
+
+mkdir -p "${OUT_DIR}/trimmed" "${OUT_DIR}/assembly" "${OUT_DIR}/prokka" \
+         "${OUT_DIR}/abricate" "${OUT_DIR}/mlst" "${OUT_DIR}/busco" "${OUT_DIR}/quast"
+mkdir -p "${RESULTS_DIR}"
+
+# ==========================================================================
+# STEP 1: Quality trimming with fastp
+# ==========================================================================
+log_step "Quality trimming with fastp"
+
+if [ ! -f "${OUT_DIR}/trimmed/R1.fastq" ]; then
+    fastp \
+        --in1 "${DATA_DIR}/reads_R1.fastq" \
+        --in2 "${DATA_DIR}/reads_R2.fastq" \
+        --out1 "${OUT_DIR}/trimmed/R1.fastq" \
+        --out2 "${OUT_DIR}/trimmed/R2.fastq" \
+        --detect_adapter_for_pe \
+        --cut_front --cut_tail \
+        --cut_mean_quality 20 \
+        --length_required 30 \
+        --thread ${THREADS} \
+        --json "${OUT_DIR}/trimmed/fastp.json" \
+        --html "${OUT_DIR}/trimmed/fastp.html"
+    echo "Trimming complete."
+else
+    echo "Trimmed reads exist, skipping."
+fi
+
+# ==========================================================================
+# STEP 2: Genome assembly with shovill (SPAdes wrapper)
+# ==========================================================================
+log_step "Genome assembly with shovill"
+
+if [ ! -f "${OUT_DIR}/assembly/contigs.fa" ]; then
+    shovill \
+        --R1 "${OUT_DIR}/trimmed/R1.fastq" \
+        --R2 "${OUT_DIR}/trimmed/R2.fastq" \
+        --outdir "${OUT_DIR}/assembly" \
+        --gsize 2914567 \
+        --cpus ${THREADS} \
+        --ram 8 \
+        --force
+    echo "Assembly complete."
+else
+    echo "Assembly exists, skipping."
+fi
+
+CONTIGS="${OUT_DIR}/assembly/contigs.fa"
+
+# === DIAMOND BRANCHES START HERE ===
+# The following 5 tools run independently on the same assembly
+
+# ==========================================================================
+# BRANCH 1: Gene annotation with Prokka
+# ==========================================================================
+log_step "BRANCH 1: Gene annotation with Prokka"
+
+if [ ! -f "${OUT_DIR}/prokka/MRSA.gff" ]; then
+    prokka \
+        "${CONTIGS}" \
+        --outdir "${OUT_DIR}/prokka" \
+        --prefix MRSA \
+        --cpus ${THREADS} \
+        --kingdom Bacteria \
+        --genus Staphylococcus \
+        --species aureus \
+        --force
+    echo "Prokka complete."
+else
+    echo "Prokka output exists, skipping."
+fi
+
+# ==========================================================================
+# BRANCH 2: AMR gene detection with ABRicate
+# ==========================================================================
+log_step "BRANCH 2: AMR gene detection with ABRicate"
+
+if [ ! -f "${OUT_DIR}/abricate/card_results.tsv" ]; then
+    # CARD database for AMR
+    abricate "${CONTIGS}" --db card --minid 80 --mincov 60 \
+        > "${OUT_DIR}/abricate/card_results.tsv"
+    # VFDB for virulence factors
+    abricate "${CONTIGS}" --db vfdb --minid 80 --mincov 60 \
+        > "${OUT_DIR}/abricate/vfdb_results.tsv"
+    echo "ABRicate complete."
+else
+    echo "ABRicate output exists, skipping."
+fi
+
+# ==========================================================================
+# BRANCH 3: MLST typing
+# ==========================================================================
+log_step "BRANCH 3: MLST typing"
+
+if [ ! -f "${OUT_DIR}/mlst/mlst_results.tsv" ]; then
+    mlst "${CONTIGS}" > "${OUT_DIR}/mlst/mlst_results.tsv"
+    echo "MLST complete."
+    cat "${OUT_DIR}/mlst/mlst_results.tsv"
+else
+    echo "MLST output exists, skipping."
+fi
+
+# ==========================================================================
+# BRANCH 4: Assembly completeness with BUSCO
+# ==========================================================================
+log_step "BRANCH 4: Assembly completeness with BUSCO"
+
+if [ ! -f "${OUT_DIR}/busco/short_summary.specific.bacteria_odb10.busco.txt" ]; then
+    busco \
+        -i "${CONTIGS}" \
+        -o busco \
+        --out_path "${OUT_DIR}" \
+        -l bacteria_odb10 \
+        -m genome \
+        -c ${THREADS} \
+        --force
+    echo "BUSCO complete."
+else
+    echo "BUSCO output exists, skipping."
+fi
+
+# ==========================================================================
+# BRANCH 5: Assembly statistics with QUAST
+# ==========================================================================
+log_step "BRANCH 5: Assembly statistics with QUAST"
+
+if [ ! -f "${OUT_DIR}/quast/report.tsv" ]; then
+    quast "${CONTIGS}" -o "${OUT_DIR}/quast" -t ${THREADS}
+    echo "QUAST complete."
+else
+    echo "QUAST output exists, skipping."
+fi
+
+# === DIAMOND MERGE ===
+
+# ==========================================================================
+# STEP 8: Merge all branch results into comprehensive CSV
+# ==========================================================================
+log_step "MERGE: Generating comprehensive results CSV"
+
+# --- Extract QUAST metrics ---
+TOTAL_LENGTH=$(grep "^Total length" "${OUT_DIR}/quast/report.tsv" | head -1 | cut -f2)
+NUM_CONTIGS=$(grep "^# contigs " "${OUT_DIR}/quast/report.tsv" | head -1 | cut -f2)
+N50=$(grep "^N50" "${OUT_DIR}/quast/report.tsv" | cut -f2)
+GC_CONTENT=$(grep "^GC" "${OUT_DIR}/quast/report.tsv" | cut -f2)
+LARGEST_CONTIG=$(grep "^Largest contig" "${OUT_DIR}/quast/report.tsv" | cut -f2)
+
+# --- Extract BUSCO summary ---
+BUSCO_SUMMARY=$(grep "C:" "${OUT_DIR}/busco/short_summary.specific.bacteria_odb10.busco.txt" 2>/dev/null \
+    | head -1 | sed 's/^[[:space:]]*//;s/[[:space:]]*$//' \
+    || echo "N/A")
+
+# --- Extract MLST ---
+MLST_SCHEME=$(cut -f2 "${OUT_DIR}/mlst/mlst_results.tsv")
+MLST_ST=$(cut -f3 "${OUT_DIR}/mlst/mlst_results.tsv")
+
+# --- Extract Prokka gene counts ---
+PROKKA_CDS=$(grep "^CDS" "${OUT_DIR}/prokka/MRSA.txt" 2>/dev/null | awk '{print $2}' || echo "0")
+PROKKA_TRNA=$(grep "^tRNA" "${OUT_DIR}/prokka/MRSA.txt" 2>/dev/null | awk '{print $2}' || echo "0")
+PROKKA_RRNA=$(grep "^rRNA" "${OUT_DIR}/prokka/MRSA.txt" 2>/dev/null | awk '{print $2}' || echo "0")
+
+# --- Write assembly_report.csv (main output) ---
+echo "metric,value" > "${RESULTS_DIR}/assembly_report.csv"
+echo "total_length,${TOTAL_LENGTH}" >> "${RESULTS_DIR}/assembly_report.csv"
+echo "num_contigs,${NUM_CONTIGS}" >> "${RESULTS_DIR}/assembly_report.csv"
+echo "n50,${N50}" >> "${RESULTS_DIR}/assembly_report.csv"
+echo "gc_content,${GC_CONTENT}" >> "${RESULTS_DIR}/assembly_report.csv"
+echo "largest_contig,${LARGEST_CONTIG}" >> "${RESULTS_DIR}/assembly_report.csv"
+echo "busco_summary,${BUSCO_SUMMARY}" >> "${RESULTS_DIR}/assembly_report.csv"
+echo "mlst_scheme,${MLST_SCHEME}" >> "${RESULTS_DIR}/assembly_report.csv"
+echo "mlst_st,${MLST_ST}" >> "${RESULTS_DIR}/assembly_report.csv"
+echo "prokka_cds,${PROKKA_CDS}" >> "${RESULTS_DIR}/assembly_report.csv"
+echo "prokka_trna,${PROKKA_TRNA}" >> "${RESULTS_DIR}/assembly_report.csv"
+echo "prokka_rrna,${PROKKA_RRNA}" >> "${RESULTS_DIR}/assembly_report.csv"
+
+# --- Write AMR results CSV ---
+echo "gene,accession,database,identity,coverage,contig" > "${RESULTS_DIR}/amr_genes.csv"
+if [ -s "${OUT_DIR}/abricate/card_results.tsv" ]; then
+    tail -n +2 "${OUT_DIR}/abricate/card_results.tsv" | \
+        awk -F'\t' 'BEGIN{OFS=","} {print $6,$12,"CARD",$10,$11,$2}' \
+        >> "${RESULTS_DIR}/amr_genes.csv"
+fi
+
+# --- Write virulence results CSV ---
+echo "gene,accession,database,identity,coverage,contig" > "${RESULTS_DIR}/virulence_genes.csv"
+if [ -s "${OUT_DIR}/abricate/vfdb_results.tsv" ]; then
+    tail -n +2 "${OUT_DIR}/abricate/vfdb_results.tsv" | \
+        awk -F'\t' 'BEGIN{OFS=","} {print $6,$12,"VFDB",$10,$11,$2}' \
+        >> "${RESULTS_DIR}/virulence_genes.csv"
+fi
+
+echo ""
+echo "=== Pipeline complete ==="
+echo "=== assembly_report.csv ==="
+cat "${RESULTS_DIR}/assembly_report.csv"
+echo ""
+echo "=== AMR genes found ==="
+wc -l "${RESULTS_DIR}/amr_genes.csv"
+head -5 "${RESULTS_DIR}/amr_genes.csv"
+echo ""
+echo "=== Virulence genes found ==="
+wc -l "${RESULTS_DIR}/virulence_genes.csv"
+head -5 "${RESULTS_DIR}/virulence_genes.csv"
+echo ""
+ls -lh "${RESULTS_DIR}/"