Publications

4592

ABSTRACT Candidatus Saccharimonadia is a class‐level lineage of ultrasmall bacteria within the phylum Minisyncoccota (formerly Candidate Phyla Radiation or Ca . Patescibacteria), commonly found in activated sludge processes treating municipal wastewater. In this study, we aimed to elucidate the metabolic potential of Ca . Saccharimonadia by using shotgun metagenomic sequencing combined with a filtration‐based size‐fractionation approach for activated sludge from five wastewater treatment plants. A total of 65 high‐quality metagenomic bins were recovered, belonging to four orders and 19 families of Ca . Saccharimonadia, including previously unreported lineages in activated sludge. These bins had small genomes (approximately 0.46–1.73 Mbp) with limited metabolic capabilities, indicating dependency on other microorganisms. Notably, the order Ca . Saccharimonadales retained a type IV secretion system and effector gene cluster for parasitic interactions with the hosts, suggesting that Ca . Saccharimonadales bacteria may exhibit a parasitic lifestyle. Co‐occurrence network analysis showed that members of the order Ca . Saccharimonadales were significantly correlated with multiple lineages, including Actinobacteriota , for which a parasitic relationship has been previously demonstrated. Our results shed light on the potential ecophysiology of the diverse members of Ca . Saccharimonadia, providing a comprehensive understanding of Ca . Saccharimonadia in activated sludge.

ABSTRACT Biogas, a mix of CO 2 , CH 4 and small proportions of other gases, is a biofuel obtained by anaerobic digestion (AD). Biogas production is often considered a black box process, as the role and dynamics of some of the microorganisms involved remain undisclosed. Previous metataxonomic studies in the frame of the MICRO4BIOGAS project ( www.micro4biogas.eu ) revealed that MBA03, an uncharacterised and uncultured bacterial taxon belonging to phylum Bacillota , was very prevalent and abundant in industrial full‐scale AD plants. Despite the efforts, this taxon has not yet been cultivated, which makes the analysis of its taxonomy, ecology and metabolism even more challenging. In the present work, 30 samples derived from anaerobic digesters were sequenced, allowing the reconstruction of 108 metagenome‐assembled genomes (MAGs) potentially belonging to MBA03. According to phylogenetic analyses and genomic similarity indices, MBA03 was classified as a new bacterial order, proposed as ‘ Candidatus Darwinibacteriales’ ord. nov., which includes ‘ Candidatus Darwinibacter acetoxidans’ gen. nov., sp. nov. of ‘ Candidatus Darwinibacteriaceae’ fam. nov., along with ‘ Candidatus Wallacebacter cryptica’ gen. nov., sp. nov. of the ‘ Candidatus Wallacebacteriaceae’ fam. nov. Ecotaxonomic studies determined that AD processes are the main ecological niche of ‘ Candidatus Darwinibacteriales’. Moreover, metabolic predictions identified Darwinibacteraceae members as putative syntrophic acetate‐oxidising bacteria (SAOB), as they encode for the reversed Wood–Ljungdahl (W–L) pathway coupled to the glycine cleavage system. This suggests that Darwinibacteraceae members could work in collaboration with hydrogenotrophic methanogenic archaea to produce methane in industrial biogas plants. Overall, our findings present ‘ Candidatus Darwinibacteriales’ as a potential key player in anaerobic digestion and pave the way towards the complete characterisation of this newly described bacterial taxon, which has not yet been cultured.