Additional file 1: Figure S1. of The rumen microbiome as a reservoir of antimicrobial resistance and pathogenicity genes is directly affected by diet in beef cattle

Relative abundance (%) of 20 groups of functional genes representing 204 selected genes (number of animals, n = 50 samples). The sum of the relative abundance (%) of genes grouping within the same function is shown in this figure. Figure S2A. Total abundance of 204 selected genes based on diet treatments (n = 50). *P value < 0.05. Figure S2B. Shannon index diversity of 204 selected genes based on diet treatments (n = 50). *P value < 0.05, °P value < 0.1. Figure S3. Canonical Variate analysis (CVA) on the structure of 204 genes selected based on breed, age, weight, Proteobacteria ratio, FCR and methane grouping (n = 50). Figure S4. Factors explaining the significant differences observed for Proteobacteria ratio (n = 50). Figure S5. Microbial community composition at the phylum level (n = 50). Table S1. Characteristics of the cattle used in the experiment. Table S2. Groups of AMR genes significantly correlated with abundance of the Proteobacteria phylum and Proteobacteria ratio. Table S3. The relative abundance of AMR genes. Table S4, Proteobacteria populations strongly correlated with the Proteobacteria ratio. Table S5. Functional genes significantly correlated with Proteobacteria ratio (PLS). Table S6. Cluster distribution of functional genes significantly different between diets. (DOCX 60 kb

Multiple rarefaction collectors curves.

<p>Observed number of OTUs for different sequencing and analysis methods: A. SA <i>rrn</i>. B. IM <i>rrn</i>. C. IM <i>mcrA</i>. D. IA <i>rrn.</i></p

Phylogenetic analysis of SA <i>rrn</i> OTUs (RINH01–RINH21) and IA <i>rrn</i> OTUs (T01–T10).

<p>Placement of representative sequences of the present study in clades indicated with additional reference sequences obtained from GenBank. A sequence related to the Crenarchaeota phylum (Acc. No. AF418935) was used as an outgroup. Full multiple alignment using ClustalW and a consensus tree was constructed using the Neighbor-Joining method with the Jukes-Cantor substitution model. The trees were bootstrap resampled 1000 times with branch values ≥50% shown. Scale shows 0.05 nucleotide substitutions per nucleotide position.</p

Relative distribution of methanogenic archaeal clades for different sequencing and analysis methods: A. SA <i>rrn</i>. B. IM <i>rrn</i>. C. IM <i>mcrA</i>. D. IA <i>rrn.</i>

<p>Relative distribution of methanogenic archaeal clades for different sequencing and analysis methods: A. SA <i>rrn</i>. B. IM <i>rrn</i>. C. IM <i>mcrA</i>. D. IA <i>rrn.</i></p

Comparison of methanogenic archaeal diversity and depth of coverage for Sheep A and Sheep B.

<p>Shannon index (H′), Chao1 estimated number of species and Good’s statistic (C).</p><p>Comparison of methanogenic archaeal diversity and depth of coverage for Sheep A and Sheep B.</p

Archaea sequence data

Primers used for PCR amplification of bacteria and archaea 16S rRNA genes, ciliate protozoa 18S rRNA genes and anaerobic fungi ITS1 genes were designed in silico using ecoPrimers, the OBITools software suite (http://www.grenoble.prabi.fr/trac/OBITools) and a database created from sequences stored in GenBank. PCR amplicons were combined in equal volumes and purified (QIAquick PCR purification kit, Qiagen, Germany). After library preparation using a standard protocol with only five PCR cycles, amplicons were sequenced using the MiSeq technology from Illumina (Fasteris, SA, Geneva, Switzerland), which produced 250-base paired-end reads for all markers, other than for archaea (100-base paired-end reads). Alignment of paired-end reads, sample assignment and removal of sequences with ambiguous nucleotides and sequences of lengths outside the empirical sequence length distribution were performed with the OBITools software suite. Primer sequences and metadata for each sample are provided in the file 'Sequence metadata.xlsx', that is also part of this Dryad data package

Fungi sequence data

Primers used for PCR amplification of bacteria and archaea 16S rRNA genes, ciliate protozoa 18S rRNA genes and anaerobic fungi ITS1 genes were designed in silico using ecoPrimers, the OBITools software suite (http://www.grenoble.prabi.fr/trac/OBITools) and a database created from sequences stored in GenBank. PCR amplicons were combined in equal volumes and purified (QIAquick PCR purification kit, Qiagen, Germany). After library preparation using a standard protocol with only five PCR cycles, amplicons were sequenced using the MiSeq technology from Illumina (Fasteris, SA, Geneva, Switzerland), which produced 250-base paired-end reads for all markers, other than for archaea (100-base paired-end reads). Alignment of paired-end reads, sample assignment and removal of sequences with ambiguous nucleotides and sequences of lengths outside the empirical sequence length distribution were performed with the OBITools software suite. Primer sequences and metadata for each sample are provided in the file 'Sequence metadata.xlsx', that is also part of this Dryad data package

Protozoa sequence data

Primers used for PCR amplification of bacteria and archaea 16S rRNA genes, ciliate protozoa 18S rRNA genes and anaerobic fungi ITS1 genes were designed in silico using ecoPrimers, the OBITools software suite (http://www.grenoble.prabi.fr/trac/OBITools) and a database created from sequences stored in GenBank. PCR amplicons were combined in equal volumes and purified (QIAquick PCR purification kit, Qiagen, Germany). After library preparation using a standard protocol with only five PCR cycles, amplicons were sequenced using the MiSeq technology from Illumina (Fasteris, SA, Geneva, Switzerland), which produced 250-base paired-end reads for all markers, other than for archaea (100-base paired-end reads). Alignment of paired-end reads, sample assignment and removal of sequences with ambiguous nucleotides and sequences of lengths outside the empirical sequence length distribution were performed with the OBITools software suite. Primer sequences and metadata for each sample are provided in the file 'Sequence metadata.xlsx', that is also part of this Dryad data package

Sequence metadata

This file contains information on animal ID's, primer sequences, sampling periods and diets

Bacteria sequence data

Primers used for PCR amplification of bacteria and archaea 16S rRNA genes, ciliate protozoa 18S rRNA genes and anaerobic fungi ITS1 genes were designed in silico using ecoPrimers, the OBITools software suite (http://www.grenoble.prabi.fr/trac/OBITools) and a database created from sequences stored in GenBank. PCR amplicons were combined in equal volumes and purified (QIAquick PCR purification kit, Qiagen, Germany). After library preparation using a standard protocol with only five PCR cycles, amplicons were sequenced using the MiSeq technology from Illumina (Fasteris, SA, Geneva, Switzerland), which produced 250-base paired-end reads for all markers, other than for archaea (100-base paired-end reads). Alignment of paired-end reads, sample assignment and removal of sequences with ambiguous nucleotides and sequences of lengths outside the empirical sequence length distribution were performed with the OBITools software suite. Primer sequences and metadata for each sample are provided in the file 'Sequence metadata.xlsx', that is also part of this Dryad data package