Genome-centric metatranscriptomes and ecological roles of the active microbial populations during cellulosic biomass anaerobic digestion

Abstract

Background: Although anaerobic digestion for biogas production is used worldwide in treatment processes to recover energy from carbon-rich waste such as cellulosic biomass, the activities and interactions among the microbial populations that perform anaerobic digestion deserve further investigations, especially at the population genome level. To understand the cellulosic biomass-degrading potentials in two full-scale digesters, this study examined five methanogenic enrichment cultures derived from the digesters that anaerobically digested cellulose or xylan for more than 2 years under 35 or 55 °C conditions.

Results: Metagenomics and metatranscriptomics were used to capture the active microbial populations in each enrichment culture and reconstruct their meta-metabolic network and ecological roles. 107 population genomes were reconstructed from the five enrichment cultures using a differential coverage binning approach, of which only a subset was highly transcribed in the metatranscriptomes. Phylogenetic and functional convergence of communities by enrichment condition and phase of fermentation was observed for the highly transcribed populations in the metatranscriptomes. In the 35 °C cultures grown on cellulose, Clostridium cellulolyticum-related and Ruminococcus-related bacteria were identified as major hydrolyzers and primary fermenters in the early growth phase, while Clostridium leptum-related bacteria were major secondary fermenters and potential fatty acid scavengers in the late growth phase. While the meta-metabolism and trophic roles of the cultures were similar, the bacterial populations performing each function were distinct between the enrichment conditions.

Conclusions: Overall, a population genome-centric view of the meta-metabolism and functional roles of key active players in anaerobic digestion of cellulosic biomass was obtained. This study represents a major step forward towards understanding the microbial functions and interactions at population genome level during the microbial conversion of lignocellulosic biomass to methane. The knowledge of this study can facilitate development of potential biomarkers and rational design of the microbiome in anaerobic digesters.

Keywords: Anaerobic digestion; Ecological roles; Lignocellulosic biomass; Metagenomics; Metatranscriptomics.

Fig. 1

Schematic representation of the development of the enrichment cultures and sample collection for metagenomic and metatranscriptomic analyses

Fig. 2

a Transcriptional level of the reconstructed PGs from each enrichment culture during batch experiments. The relative abundance was calculated based on the total sum of TPM values of all the ORFs of a PG divided by the sum of TPM values of all the ORFs of a culture at a particular time point. b Relative abundance of PGs in the metagenome of each enrichment culture along the enrichment process. The relative abundance was calculated based on the coverage of contigs of each PG within a metagenome. Only the highly transcribed or highly abundant PGs were labeled in each plot and the taxonomic classification of the PGs was based on their phylogenetic placement by PhyloPhlAn (see Fig. 3a)

Fig. 3

a Phylogenetic tree showing the placement of selected highly transcribed PGs. While 3737 reference genomes were used for alignment, only a subset of the aligned references was displayed. Blue diamonds indicate collapsed monophyletic clades with the number of reference genomes indicated in the brackets, black squares represent PGs from cultures amended with cellulose, and black triangles with xylan. Monophyletic clades of interest have been highlighted and bootstrap values (based on 100 iterations) are shown on internal nodes. The tree was midpoint-rooted. The scale bar indicates the evolutionary distance (substitution/site). PGs were color-coded by their enrichment culture, indigo for GZ-C-35, purple for SWH-C-35, red for GZ-X-35, green for SWH-X-35, and orange for SWH-C-55. b A heatmap showing the transcriptional level of the major energy conservation complexes and hydrogenases based on the TPM values of the corresponding genes. Time points in the order of day 5, mid-exponential growth phase, day 10 and day 15 were arranged as columns in the heatmap for each gene

Fig. 4

Heatmap showing the transcriptional profiles of major functions in AD process for the highly transcribed bacterial and methanogenic PGs. The PGs were organized by enrichment culture and color-coded as in Fig. 3. A few key functions are boxed and highlighted

Fig. 5

Schematic representation of the functional and ecological roles of the highly transcribed PGs in the GZ-C-35 enrichment culture. PGs were colored following the color code in Fig. 2. The schematics for the other cultures are shown in Additional file 8