In addition to benefits in terms of improved taxonomic description, shotgun metagenomic sequencing enables a more detailed assessment of the functional potential of the microbial community through comparison with databases of metabolic pathways (such as the Kyoto Encyclopedia of Genes and Genomes) [ 21].
Gene functions of dominant OTUs were predicted using PICRUSt [ 49], a tool that predicts the gene function of a microbial community using an existing database of microbial genomes.
Here we adopt the MetaPathways annotation and analysis pipeline and Pathway Tools to construct environmental pathway/genome databases (ePGDBs) that describe microbial community metabolism using MetaCyc, a highly curated database of metabolic pathways and components covering all domains of life.
Matching the sequencing reads from BE-1 and BEY to sequences collected in NT and NR databases, we dissected microbial community structure of the two libraries (excluding the reads with no-hits; Figure 2B), showing that at the domain level there is significant difference between the two libraries in the proportion of reads assigned to bacterial, archaeal, viral, and eukaryotic sequences.
To assess the functional capacity of microbial community, metagenomic sequence reads need alignment to a database of known genes and can be achieved with several well-characterized functional databases including KEGG orthology (KO) [ 16] and COGs [ 17].
These observations showed that analysis of sequence signatures provides an additional tool to evaluate patterns in microbial community data, and this tool is not subject to the biases in existing databases.
Composition of the biogas-producing microbial community was obtained by taxonomic classification of reads base on the M5NR database on the MG-RAST platform.
The taxonomic assignment at phylum level using the M5NR database revealed dominance of the Bacteroidetes phyla in the rumen microbial community of liquid fraction of Green Roughage (GL) as well as the Dry Roughage (DL) diet fed animals (Fig. 1a).
To obtain gene profile characteristic for the anaerobic microbial community, the total sequencing reads were annotated based on BLASTX analysis against the database of Clusters of Orthologous Groups of proteins (COG) [ 72] with an E-value cut-off of 10−5.
Meanwhile, we also analyzed microbial community compositions based on the assembled contigs, with BLASTN (version 2.2.13) against the NT and NR databases with E-value cutoff of 10-5.
