No abstract

Phylogenetically diverse anammox bacteria have been detected in most of anoxic natural and engineered ecosystems and thus regarded as key players in the global nitrogen cycle. However, ecological niche differentiation of anammox bacteria remains unresolved despite its ecological and practical importance. In this study, the microbial competitions for a common substrate (nitrite) among three anammox species (i.e. "Candidatus Brocadia sinica", "Candidatus Jettenia caeni" and "Candidatus Kuenenia stuttgartiensis") were systematically investigated in nitrite-limited gel-immobilized column reactors (GICR) and membrane bioreactors (MBRs) under different nitrogen loading rates (NLRs). 16 S rRNA gene-based population dynamics revealed that "Ca. J. caeni" could proliferate only at low NLRs, whereas "Ca. B. sinica" outcompeted other two species at higher NLRs in both types of reactors. Furthermore, FISH analysis revealed that "Ca. J. caeni" was mainly present as spherical microclusters at the inner part (low NO environment), whereas "Ca. B. sinica" was present throughout the gel beads and granules. This spatial distribution supports the outcomes of the competition experiments. However, the successful competition of "Ca. J. caeni" at low NLR could not be explained with the Monod model probably due to inaccuracy of kinetic parameters such as half saturation constant (K) for nitrite and a difference in the maintenance rate (m). In addition, the growth of "Ca. K. stuttgartiensis" could not be observed in any experimental conditions, suggesting possible unknown factor(s) is missing. Taken together, NLR was one of factors determining ecological niche differentiation of "Ca. B. sinica" and "Ca. J. caeni".

We successfully enriched a novel anaerobic ammonium-oxidizing (anammox) bacterium affiliated with the genus 'Candidatus Brocadia' with high purity (>90%) in a membrane bioreactor (MBR). The enriched bacterium was distantly related to the hitherto characterized 'Ca. Brocadia fulgida' and 'Ca. Brocadia sinica' with 96% and 93% of 16S ribosomal RNA gene sequence identity, respectively. The bacterium exhibited the common structural features of anammox bacteria and produced hydrazine in the presence of hydroxylamine under anoxic conditions. The temperature range of anammox activity was 20-45°C with a maximum activity at 37°C. The maximum specific growth rate (μ) was 0.0082h at 37°C, corresponding to a doubling time of 3.5 days. The half-saturation constant (K) for nitrite was 5±2.5μM. The anammox activity was inhibited by nitrite (IC=11.6mM) but not by formate and acetate. The major respiratory quinone was identified to be menaquinone-7 (MK-7). The enriched anammox bacterium shared nearly half of genes with 'Ca. Brocadia sinica' and 'Ca. Brocadia fulgida'. The enriched bacterium showed all known physiological characteristics of anammox bacteria and can be distinguished from the close relatives by its 16S rRNA gene sequence. Therefore, we proposed the name 'Ca. Brocadia sapporoensis' sp. nov.

Anaerobic ammonium-oxidizing (anammox) bacteria are well known for their aggregation ability. However, very little is known about cell surface physicochemical properties of anammox bacteria and thus their aggregation abilities have not been quantitatively evaluated yet. Here, we investigated the aggregation abilities of three different anammox bacterial species: "Candidatus Brocadia sinica", "Ca. Jettenia caeni" and "Ca. Brocadia sapporoensis". Planktonic free-living enrichment cultures of these three anammox species were harvested from the membrane bioreactors (MBRs). The physicochemical properties (e.g., contact angle, zeta potential, and surface thermodynamics) were analyzed for these anammox bacterial species and used in the extended DLVO theory to understand the force-distance relationship. In addition, their extracellular polymeric substances (EPSs) were characterized by X-ray photoelectron spectroscopy and nuclear magnetic resonance. The results revealed that the "Ca. B. sinica" cells have the most hydrophobic surface and less hydrophilic functional groups in EPS than other anammox strains, suggesting better aggregation capability. Furthermore, aggregate formation and anammox bacterial populations were monitored when planktonic free-living cells were cultured in up-flow column reactors under the same conditions. Rapid development of microbial aggregates was observed with the anammox bacterial population shifts to a dominance of "Ca. B. sinica" in all three reactors. The dominance of "Ca. B. sinica" could be explained by its better aggregation ability and the superior growth kinetic properties (higher growth rate and affinity to nitrite). The superior aggregation ability of "Ca. B. sinica" indicates significant advantages (efficient and rapid start-up of anammox reactors due to better biomass retention as granules and consequently stable performance) in wastewater treatment application.

The anaerobic ammonium-oxidizing (anammox) bacterium “Candidatus Brocadia sp. 40” demonstrated the fastest growth rate compared to others in this taxon. Here, we report the 2.93-Mb draft genome sequence of this bacterium, which has 2,565 gene-coding regions, 41 tRNAs, and a single rrn operon.