Comparison of microbiome community structure and dynamics during anaerobic digestion of different renewable solid wastes

Abstract

This study analysed the effect of the different lignocellulose composition of two crop substrates on the structure and dynamics of bacterial communities during anaerobic digestion (AD) processes for biogas production. To this end, cereal grains and grape pomace biomasses were analysed in parallel in an experimental AD bench-scale system to define and compare their metagenomic profiles for different experimental time intervals. The bacterial community structure and dynamics during the AD process were detected and characterised using high-resolution whole metagenomic shotgun analyses. Statistical evaluation identified 15 strains as specific to two substrates. Some strains, like Clostridium isatidis, Methanothermobacter wolfeii, and Methanobacter sp. MB1 in cereal grains, and Acetomicrobium hydrogeniformans and Acetomicrobium thermoterrenum in grape pomace, were never before detected in biogas reactors. The presence of bacteria such as Acetomicrobium sp. and Petrimonas mucosa, which degrade lipids and protein-rich substrates, along with Methanosarcina sp. and Peptococcaceae bacterium 1109, which tolerate high hydrogen pressures and ammonia concentrations, suggests a complex syntrophic community in lignin-cellulose-enriched substrates. This finding could help develop new strategies for the production of a tailor-made microbial consortium to be inoculated from the beginning of the digestion process of specific lignocellulosic biomass.

Keywords: Anaerobic digestion; Biogas production; Lignin biodegradation; Lignocellulosic biomasses; Metagenomics.

Graphical abstract

Fig. 1

Biogas yield from cereal grains and grape pomace AD, expressed as Nm³/tonne FM. Each data point is the average (± standard deviation) of measurements from six reactors.

Fig. 2

Diversity inside different matrices during time: 2A) alpha-diversity index, 2B) J Pielou's Evenness index, 2C) Richness index (d).

Fig. 3

Percentage of unclassified reads retrieved in taxonomical analyses using BLASTN (yellow), MetaPhlAn3 (blue) and MetaPhlAn4 (pink).

Fig. 4

Dynamics of microbial community during biogas production process represented by: 4A) Neighbour Joining tree from BLASTN output; 4B) plot of principal coordinates analysis (PCoA) of microbial composition from MetaPhlAn3 output, the analysis was based on Bray-Curtis matrix distance (species).

Fig. 5

Inoculum composition: 5A) phyla, 5B) species obtained with MetaPhlAn3.

Fig. 6

Taxonomic profiles, in terms of phyla, detected from time 0 to day 30 in the two different substrates (cereal grain and grape pomace) and identified with MetaPhlAn3.

Fig. 7

- Taxonomic profile obtained with MetaPhlAn3, in terms of species, detected from time 0 to day 30 from both cereal grains (cg0–30) and grape pomace (gp0–30), plus inoculum.

Fig. 8

8A) Cladogram plotted from LEfSe analysis, comparing all CG samples with all GP samples. Taxonomic levels are represented by rings with phyla in the outermost ring and species in the innermost ring. Each circle represents a member within that level, the dimension of each node is proportional to relative abundance of the represented taxon. Taxa with enriched level in samples from CG are coloured in red, those enriched in samples from GP are coloured in green; unenriched taxa are ochre coloured; 8B) Relative abundance of strains resulted significantly different in LEfSe analysis between substrates, represented as an heatmap. Abundancy values were showed with a shade colour scale from white (0 %) to red (100 %).

Fig. 9

Functional groups of microorganisms observed in hydrolysis, acidogenesis, acetogenesis and methanogenesis processes. Microorganisms that increased in abundance in CG are marked in red, and those that increased in abundance in GP are marked in green.

Fig. 10

Dynamic of functional profiles detected from time 0 to day 30 from both cereal grains (cg0–30), grape pomace (gp0–30) and inoculum. The heatmap shows only the MetaCyc pathways resulted significantly (p < 0.05) enriched across the digestion period in each matrix.