Production of nonulosonic acids in the extracellular polymeric substances of “Candidatus Accumulibacter phosphatis”

Abstract

Nonulosonic acids (NulOs) are a family of acidic carbohydrates with a nine-carbon backbone, which include different related structures, such as sialic acids. They have mainly been studied for their relevance in animal cells and pathogenic bacteria. Recently, sialic acids have been discovered as an important compound in the extracellular matrix of virtually all microbial life and in “Candidatus Accumulibacter phosphatis”, a well-studied polyphosphate-accumulating organism, in particular. Here, bioaggregates highly enriched with these bacteria (approx. 95% based on proteomic data) were used to study the production of NulOs in an enrichment of this microorganism. Fluorescence lectin-binding analysis, enzymatic quantification, and mass spectrometry were used to analyze the different NulOs present, showing a wide distribution and variety of these carbohydrates, such as sialic acids and bacterial NulOs, in the bioaggregates. Phylogenetic analysis confirmed the potential of “Ca. Accumulibacter” to produce different types of NulOs. Proteomic analysis showed the ability of “Ca. Accumulibacter” to reutilize and reincorporate these carbohydrates. This investigation points out the importance of diverse NulOs in non-pathogenic bacteria, which are normally overlooked. Sialic acids and other NulOs should be further investigated for their role in the ecology of “Ca. Accumulibacter” in particular, and biofilms in general. Key Points •“Ca. Accumulibacter” has the potential to produce a range of nonulosonic acids. •Mass spectrometry and lectin binding can reveal the presence and location of nonulosonic acids. •The role of nonulosonic acid in non-pathogenic bacteria needs to be studied in detail.

Production of extracellular polymeric substances in granular sludge under selection forAccumulibacterandCompetibacter

“…This chemical functional group should be further investigated in granular sludge. The presence of sialic acid residues in granular sludge and other wastewater biomasses has been previously reported and described (Weissbrodt et al, 2013a;de Graaff et al, 2019;Kleikamp et al, 2020;Tomás-Martínez et al, 2021). These nonulosonic acids linked to a carbohydrate chain can exert a protective role against biodegradation.…”

“…Instead of addressing the broad diversity of EPS metabolic signatures across the biomass, one rational approach can target a set of specific EPS compounds. This was recently started with the analysis of nonulosonic acids in granular sludge by mass spectrometry and metaproteomics (Kleikamp et al, 2020; Kleikamp et al, 2021; Tomás-Martínez et al, 2021). Combination with 13 C substrate labelling experiments help examine their metabolic turnover (Tomás-Martínez et al, 2023b).…”

“…Accumulibacter” has been hypothesized to display metabolic traits for EPS biosynthesis, although often described as a slow-growing chemoorganoheterotroph. First sets of metabolic and enzymatic investigations of EPS formation in PAO enrichments have been recently reported (Tomás-Martínez et al, 2021; 2022; 2023a; 2023b). “ Ca.…”

CRISPR-Cas phage defense systems and prophages inCandidatusAccumulibacter

“…Enhanced biological phosphorous removal (EBPR) is a widely applied process in municipal wastewater treatment plants (WWTPs) (Chen et al, 2022; Mino et al, 1998); it requires polyphosphate accumulating organisms (PAOs) capable of releasing P under anaerobic, and taking up more P than normally required for metabolism and growth, under aerobic conditions (He and McMahon, 2011; Lu et al, 2021; Zhang et al, 2022). Candidatus Accumulibacter represents a key group of PAOs in lab- and full-scale EBPR systems (Martín et al, 2006; Petriglieri et al, 2022; Qiu et al, 2020; Singleton et al, 2022; Srinivasan et al, 2021; Tomás-Martínez et al, 2021). Ensuring the viability of Ca.…”

Enrichment and application of bacterial sialic acids containing polymers from the extracellular polymeric substances of “Candidatus Accumulibacter”

“…Further research with a highly enriched culture of “ Ca . Accumulibacter” revealed its potential to produce different type of NulOs as part of its EPS, primarily the bacterial sialic acids Pse and/or Leg, which have the same molecular weight (Tomás-Martínez et al, 2021). Although the role of these carbohydrates in non-pathogenic environmental bacteria is still unknown, these findings point towards a new potential sustainable source of bacterial sialic acids.…”