Microbial community analysis was carried out on ruminal digesta obtained directly via rumen fistula and buccal fluid, regurgitated digesta (bolus) and faeces of dairy cattle to assess if non-invasive samples could be used as proxies for ruminal digesta. Samples were collected from five cows receiving grass silage based diets containing no additional lipid or four different lipid supplements in a 5 x 5 Latin square design. Extracted DNA was analysed by qPCR and by sequencing 16S and 18S rRNA genes or the fungal ITS1 amplicons. Faeces contained few protozoa, and bacterial, fungal and archaeal communities were substantially different to ruminal digesta. Buccal and bolus samples gave much more similar profiles to ruminal digesta, although fewer archaea were detected in buccal and bolus samples. Bolus samples overall were most similar to ruminal samples. The differences between both buccal and bolus samples and ruminal digesta were consistent across all treatments. It can be concluded that either proxy sample type could be used as a predictor of the rumen microbial community, thereby enabling more convenient large-scale animal sampling for phenotyping and possible use in future animal breeding programs aimed at selecting cattle with a lower environmental footprint.
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.
archaea.tar.gz
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.
bacteria.tar.gz
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.
fungi.tar.gz
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.
protozoa.tar.gz
Sequence metadata
This file contains information on animal ID's, primer sequences, sampling periods and diets.
