Genomics and prevalence of bacterial and archaeal isolates from biogas-producing microbiomes

Abstract

Background: To elucidate biogas microbial communities and processes, the application of high-throughput DNA analysis approaches is becoming increasingly important. Unfortunately, generated data can only partialy be interpreted rudimentary since databases lack reference sequences.

Results: Novel cellulolytic, hydrolytic, and acidogenic/acetogenic Bacteria as well as methanogenic Archaea originating from different anaerobic digestion communities were analyzed on the genomic level to assess their role in biomass decomposition and biogas production. Some of the analyzed bacterial strains were recently described as new species and even genera, namely Herbinix hemicellulosilytica T3/55^T, Herbinix luporum SD1D^T, Clostridium bornimense M2/40^T, Proteiniphilum saccharofermentans M3/6^T, Fermentimonas caenicola ING2-E5B^T, and Petrimonas mucosa ING2-E5A^T. High-throughput genome sequencing of 22 anaerobic digestion isolates enabled functional genome interpretation, metabolic reconstruction, and prediction of microbial traits regarding their abilities to utilize complex bio-polymers and to perform specific fermentation pathways. To determine the prevalence of the isolates included in this study in different biogas systems, corresponding metagenome fragment mappings were done. Methanoculleus bourgensis was found to be abundant in three mesophilic biogas plants studied and slightly less abundant in a thermophilic biogas plant, whereas Defluviitoga tunisiensis was only prominent in the thermophilic system. Moreover, several of the analyzed species were clearly detectable in the mesophilic biogas plants, but appeared to be only moderately abundant. Among the species for which genome sequence information was publicly available prior to this study, only the species Amphibacillus xylanus, Clostridium clariflavum, and Lactobacillus acidophilus are of importance for the biogas microbiomes analyzed, but did not reach the level of abundance as determined for M. bourgensis and D. tunisiensis.

Conclusions: Isolation of key anaerobic digestion microorganisms and their functional interpretation was achieved by application of elaborated cultivation techniques and subsequent genome analyses. New isolates and their genome information extend the repository covering anaerobic digestion community members.

Keywords: Anaerobic digestion; Biomethanation; Defluviitoga tunisiensis; Fragment recruitment; Genome sequencing; Methanoculleus bourgensis.

Fig. 1

Phylogenetic diversity of archaeal and bacterial strains analyzed in this study in relation to the corresponding type species. The program ARB [35] was applied to construct the phylogenetic tree based on the full-length 16S rRNA gene sequences obtained from the strain’s genome sequences and in the case of closely related type species from the SILVA database [34]. The scale bar represents 1% sequence divergence

Fig. 2

Diversity of genes encoding carbohydrate-active enzymes (CAZymes) predicted to be involved in hydrolysis and/or rearrangement of glycosidic bonds for each bacterial isolate studied. The screening for the presence of CAZymes was accomplished applying the HMM-based (Hidden-Markov-Model-based) carbohydrate-active enzyme annotation database dbCAN [40]. The numbers of bacterial genes belonging to a corresponding glycosyl hydrolase (GH) family are given in the fields

Fig. 3

Overview of the four phases of the conversion of biomass into biogas and allocation of the analyzed microbial strains to the different conversion steps. Functional roles of the organisms were determined considering relevant KEGG pathways, namely the propionic acid, ethanol, formic acid, butyric acid, and lactic acid fermentation

Fig. 4

Prevalence of bacterial and archaeal strains within different biogas-producing microbial communities as determined by the fragment recruitment approach. Metagenome sequences derived from the microbial communities of three mesophilic (BGP1-3) and one thermophilic biogas plants (BGP4) described previously [41] were mapped on the genome sequences of the 22 strains analyzed in this study, the four MAGs described previously [41], and 46 publicly available genomes obtained from the RefSeq database [44]. Results for the 25 most abundant organisms are shown in the upper part of the figure. The prevalence of the remaining eight isolates of this study, representing non-abundant organisms, is shown in the lower part of the figure. The x-axis represents the number of GPMs (genomes per million; analogous to TPM = transcripts Per Million), and the y-axis shows the analyzed organisms. Isolates investigated within this study are shown in red, genome bins obtained from a previous study [41] in blue, and genomes obtained from the RefSeq database are visualized in black