## Commands used to analyse bird microbiome ## Meta-transcriptomics reveals a diverse antibiotic resistance gene pool in avian microbiomes #$r1 and $r2 refers to forwrad and reverse reads ### Quality control with kneaddata and trommomatic: kneaddata -i "$r1" -i "$r2" -o "$o" -db db/knead_gallus/gallus_db -t 20 -p 10 --trimmomatic-options="ILLUMINACLIP:adapters.fa:2:30:10" --trimmomatic-options="LEADING:30" --trimmomatic-options="TRAILING:30" --trimmomatic-options="MINLEN:50" ### rRNA removal with sortmerna: merge-paired-reads.sh "$r1" "$r2" "$o" sortmerna --reads $o --ref $all_dbs --aligned $o.rRNA --other $o.mRNA --paired_in --fastx --log --num_alignments 1 -a 20 -v ### Identify resistance genes with KMA: kma -i $r12 -o $o -t_db $db -NW -1t1 ### Functional profiling with humann2: humann2 --input $r12 --output $o --protein-database db/humann2_UniProt/uniref_90_diam/ --nucleotide-database db/humann2_chocophlan_viruless/ --bypass-prescreen --threads 20 humann2_renorm_table --input $file --output $file.cpm.tsv --units cpm humann2_renorm_table --input $file --output $file.cpm.tsv --units cpm ### Count host reads with Bowtie2 and HTSeq bowtie2 -x $bird_index -1 $r1 -2 $r2 --very-sensitive -p 20 --no-unal -S $o samtools view -bS $f -@ 10 > $f.bam htseq-count $f -a 0