File size: 6,948 Bytes
67a2534 | 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78 79 80 81 82 83 84 85 86 87 88 89 90 91 92 93 94 95 96 97 98 99 100 101 102 103 104 105 106 107 108 109 110 111 112 113 114 115 116 117 118 119 120 121 122 123 124 125 126 127 128 129 130 131 132 133 134 135 136 137 138 139 140 141 142 143 144 145 146 147 148 149 150 151 152 153 154 155 156 157 158 159 160 161 162 163 | #!/bin/bash
set -e
# =============================================================================
# Task 21: Phage Genome Characterization
#
# DAG (depth 5, fan-out from assembly):
#
# L0: reads
# L1: fastp (trim)
# L2: shovill (assemble)
# ├────────────────────────────────────────┐
# L3: pharokka (phage annotation) checkv (quality)
# │ │
# L4: functional summary completeness report
# │ │
# ├── abricate VFDB (virulence) │
# └────────────────────────────────────────┘
# L5: MERGE
# =============================================================================
THREADS=$(( $(nproc) > 8 ? 8 : $(nproc) ))
SCRIPT_DIR="$(cd "$(dirname "${BASH_SOURCE[0]}")" && pwd)"
DATA="${SCRIPT_DIR}/data"
REF="${SCRIPT_DIR}/reference"
OUT="${SCRIPT_DIR}/outputs"
RES="${SCRIPT_DIR}/results"
PHAROKKA_DB="/pscratch/sd/l/lingzhi/pharokka_db"
CHECKV_DB="/pscratch/sd/l/lingzhi/checkv_db/checkv-db-v1.5"
log_step() {
echo "=================================================================="
echo "STEP: $1"
echo "$(date)"
echo "=================================================================="
}
mkdir -p "${OUT}"/{trimmed,assembly,pharokka,checkv,vfdb} "${RES}"
# ===========================================================================
# L1: Trimming
# ===========================================================================
log_step "L1: fastp"
if [ ! -f "${OUT}/trimmed/R1.fastq.gz" ]; then
fastp --in1 "${DATA}/reads_R1.fastq.gz" --in2 "${DATA}/reads_R2.fastq.gz" \
--out1 "${OUT}/trimmed/R1.fastq.gz" --out2 "${OUT}/trimmed/R2.fastq.gz" \
--detect_adapter_for_pe --thread ${THREADS} \
--json "${OUT}/trimmed/fastp.json"
else echo "Skipping (exists)"; fi
# ===========================================================================
# L2: Assembly
# ===========================================================================
log_step "L2: MEGAHIT assembly"
if [ ! -f "${OUT}/assembly/final.contigs.fa" ]; then
megahit -1 "${OUT}/trimmed/R1.fastq.gz" -2 "${OUT}/trimmed/R2.fastq.gz" \
-o "${OUT}/assembly" -t ${THREADS} --min-contig-len 500
else echo "Skipping (exists)"; fi
CONTIGS="${OUT}/assembly/final.contigs.fa"
# ===========================================================================
# L3-A: Pharokka phage-specific annotation
# ===========================================================================
log_step "L3-A: pharokka annotation"
if [ ! -f "${OUT}/pharokka/pharokka_final.gff" ]; then
pharokka.py -i "${CONTIGS}" -o "${OUT}/pharokka" \
-d "${PHAROKKA_DB}" -t ${THREADS} --force 2>&1 || {
echo "WARNING: pharokka failed, falling back to prokka --kingdom Viruses"
prokka "${CONTIGS}" --outdir "${OUT}/pharokka" --prefix pharokka_final \
--cpus ${THREADS} --kingdom Viruses --force 2>&1 || true
}
else echo "Skipping (exists)"; fi
# ===========================================================================
# L3-B: CheckV quality assessment
# ===========================================================================
log_step "L3-B: checkv quality"
if [ ! -f "${OUT}/checkv/quality_summary.tsv" ]; then
if [ -d "${CHECKV_DB}" ]; then
checkv end_to_end "${CONTIGS}" "${OUT}/checkv" -d "${CHECKV_DB}" -t ${THREADS} 2>&1 || true
else
echo "WARNING: CheckV database not found"
mkdir -p "${OUT}/checkv"
touch "${OUT}/checkv/quality_summary.tsv"
fi
else echo "Skipping (exists)"; fi
# ===========================================================================
# L3-C: Virulence factor detection
# ===========================================================================
log_step "L3-C: abricate VFDB"
if [ ! -f "${OUT}/vfdb/vfdb_results.tsv" ]; then
abricate "${CONTIGS}" --db vfdb --minid 80 --mincov 60 > "${OUT}/vfdb/vfdb_results.tsv" 2>/dev/null || true
else echo "Skipping (exists)"; fi
# ===========================================================================
# L4: Parse results
# ===========================================================================
log_step "L4: Parse annotation results"
# Pharokka stats
PHAROKKA_GFF="${OUT}/pharokka/pharokka.gff"
if [ -f "${PHAROKKA_GFF}" ]; then
CDS=$(grep -c "CDS" "${PHAROKKA_GFF}" 2>/dev/null || echo "0")
TRNA=$(grep -c "tRNA" "${OUT}/pharokka/pharokka_aragorn.gff" 2>/dev/null || true)
TRNA=${TRNA:-0}
# Count functional categories from pharokka GFF product annotations
LYSIS=$(grep -ic "lysis\|holin\|endolysin\|spanin\|lysozyme" "${PHAROKKA_GFF}" 2>/dev/null || echo "0")
LYSOGENY=$(grep -ic "integrase\|repressor\|lysogeny\|excisionase" "${PHAROKKA_GFF}" 2>/dev/null || echo "0")
REPLICATION=$(grep -ic "helicase\|primase\|polymerase\|replication\|SSB" "${PHAROKKA_GFF}" 2>/dev/null || echo "0")
STRUCTURAL=$(grep -ic "capsid\|tail\|baseplate\|head\|portal\|sheath\|fiber" "${PHAROKKA_GFF}" 2>/dev/null || echo "0")
# Get top MASH hit for taxonomy
TAXONOMY=$(tail -1 "${OUT}/pharokka/pharokka_top_hits_mash_inphared.tsv" 2>/dev/null | cut -f6 || echo "Unknown")
else
CDS=0; TRNA=0; LYSIS=0; LYSOGENY=0; REPLICATION=0; STRUCTURAL=0; TAXONOMY="Unknown"
fi
# CheckV stats
if [ -s "${OUT}/checkv/quality_summary.tsv" ]; then
QUALITY=$(tail -1 "${OUT}/checkv/quality_summary.tsv" | cut -f8 || echo "N/A")
COMPLETENESS=$(tail -1 "${OUT}/checkv/quality_summary.tsv" | cut -f10 || echo "N/A")
CONTAMINATION=$(tail -1 "${OUT}/checkv/quality_summary.tsv" | cut -f12 || echo "N/A")
else
QUALITY="N/A"; COMPLETENESS="N/A"; CONTAMINATION="N/A"
fi
# Assembly stats
NUM_CONTIGS=$(grep -c ">" "${CONTIGS}" || echo "0")
TOTAL_LEN=$(awk '/^>/{if(l)print l; l=0; next}{l+=length}END{print l}' "${CONTIGS}" || echo "0")
GC=$(awk '/^>/{next}{s+=length; gc+=gsub(/[GCgc]/,"&")}END{printf "%.2f",gc/s*100}' "${CONTIGS}" || echo "0")
# Virulence
VF_COUNT=$(tail -n +2 "${OUT}/vfdb/vfdb_results.tsv" 2>/dev/null | wc -l | tr -d ' ' || echo "0")
# ===========================================================================
# L5: MERGE
# ===========================================================================
log_step "L5-MERGE"
cat > "${RES}/phage_report.csv" << CSVEOF
metric,value
genome_length,${TOTAL_LEN}
num_contigs,${NUM_CONTIGS}
gc_content,${GC}
closest_hit,${TAXONOMY}
cds_count,${CDS}
trna_count,${TRNA}
lysis_genes,${LYSIS}
lysogeny_genes,${LYSOGENY}
replication_genes,${REPLICATION}
structural_genes,${STRUCTURAL}
checkv_quality,${QUALITY}
checkv_completeness,${COMPLETENESS}
virulence_factors,${VF_COUNT}
CSVEOF
echo ""
echo "=== Pipeline complete ==="
cat "${RES}/phage_report.csv"
echo ""
ls -lh "${RES}/"
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