Metabolic Potential of Candidatus Saccharimonadia Including Rare Lineages in Activated Sludge

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.