Bioinformatic analyses using the MG-RAST pipeline.

Unassembled DNA and cDNA sequences were loaded onto the Metagenomics Rapid Annotation (MG-RAST) server (v4.0) (http://metagenomics.anl.gov) to enable phylogenetic and metabolic reconstructions and to perform a protein similarity analysis, including both functional annotation and function classification (23, 24). Quality control and assembly information are available in Fig. S4 and MG-RAST.

Taxonomic profiles were calculated according to the best hit classification at an E value cutoff of 10⁻⁵ with a minimum alignment length of 50 bp on the basis of all annotation source databases available in MG-RAST (including RefSeq, IMG, TrEMBL, SEED, KEEG, and GenBank). The distributions of the taxonomic domains, namely, the phyla, orders, families, genera, species, and strains, for the annotations were analyzed in detail. For taxonomic profiles, the percentages shown in the study refer to classifications at a specific taxonomic level.

Functional profiling was conducted by performing gene annotation with the Subsystems, Clusters of Orthologous Group (COG), and KO databases using hierarchical classification at an E value cutoff of 10⁻⁵ and a minimum alignment length of 17 amino acids (47, 48). Detailed statistical information regarding hits against different databases is summarized in Table S2.