Gene abundance of fecal metagenomes for B-GUS and GDH activities

To investigate the distribution of b-gus and gdh in fecal metagenomes, we reanalyzed metagenomes of healthy donors (n = 103), and of CRC state I-IV patients (n = 53) of previously generated datasets from a French cohort [25]. The dereplicated gene catalogue generated in [25] was aligned against the bacterial RefSeq database (Release 85, downloaded at 04.01.2018) [51] using DIAMOND (v0.9.13, BlastX in sensitive mode) [52]. Alignments with a bitScore lower than 99% of the best alignment were removed. Furthermore, alignments with a query and reference coverage < 80% or > 130% were filtered, and those with less than 50% positives were removed. Genes aligning to b-gus and gdh were extracted, leading to a reduced gene catalogue containing 420 candidate genes. The corresponding B-GUS were assigned to EC 3.2.1.31 while GDH were assigned to EC 4.2.1.28 and 4.2.1.30. Taxonomic annotation of candidates was derived using the RefSeq sequence information. For each metagenome candidate gene abundance was extracted from the gene length normalized gene abundance matrix [25]. Abundance was multiplied with 1000 and further normalized by median marker gene abundance [25, 53] to report gene copies per thousand cells (GC).