This study revealed the presence of nitrifying bacteria in influent municipal wastewaters reaching full-scale biological wastewater treatment plants. Respirometric assays showed that the influent nitrifiers were active following a 5- to 8-hour period of metabolic induction. Diversity analyses by pyrosequencing of functional gene PCR (polymerase chain reaction) amplicon suggested that the nitrifiers in the influent stream likely seeded activated sludge bioreactors since the most abundant operational taxonomic units in the influent and mixed liquor were the same. Based on the estimated seeding intensity of 0.3 g of nitrifiers per day per gram of nitrifiers already present, the absolute minimum solids retention time (SRT) was reduced by approximately 56% at 5 °C as compared to non-seeding conditions. This can have important repercussions on the design and sizing of bioreactors operating in cold climates and calls for a need to fine-tune process modelling by considering the contribution of autotrophic nitrifying biomass from municipal influent streams.

Effect of ecological variables on community assembly of heterotrophic bacteria at eight full‐scale and two pilot‐scale activated sludge wastewater treatment plants (AS‐WWTPs) were explored by pyrosequencing of 16S rRNA gene amplicons. In total, 39 samples covering a range of abiotic factors spread over space and time were analyzed. A core bacterial community of 24 families detected in at least six of the eight AS‐WWTPs was defined. In addition to the core families, plant‐specific families (observed at <50% AS‐WWTPs) were found to be also important in the community structure. Observed beta diversity was partitioned with respect to ecological variables. Specifically, the following variables were considered: influent wastewater characteristics, season (winter vs. summer), process operations (conventional, oxidation ditch, and sequence batch reactor), reactor sizes (pilot‐scale vs. full‐scale reactors), chemical stresses defined by ozonation of return activated sludge, interannual variation, and geographical locations. Among the assessed variables, influent wastewater characteristics and geographical locations contributed more in explaining the differences between AS‐WWTP bacterial communities with a maximum of approximately 26% of the observed variations. Partitioning of beta diversity is necessary to interpret the inherent variability in microbial community assembly and identify the driving forces at play in engineered microbial ecosystem.

Wastewater treatment plants (WWTPs) are host to diverse microbial communities and receive a constant influx of microbes from influent wastewater, however the impact of immigrants on the structure and activities of the activated sludge (AS) microbial community remains unclear. To gain insight on this phenomenon known as perpetual community coalescence, the current study utilised controlled manipulative experiments that decoupled the influent wastewater composition from the microbial populations to reveal the fundamental mechanisms involved in immigration between sewers and AS-WWTP. The immigration dynamics of heterotrophs were analysed by harvesting wastewater biomass solids from 3 different sewer systems and adding to synthetic wastewater. Immigrating influent populations were observed to contribute up to 25 % of the sequencing reads in the AS. By modelling the net growth rate of taxa, it was revealed that immigrants primarily exhibited low or negative net growth rates. By developing a protocol to reproducibly grow AS-WWTP communities in the lab, we have laid down the foundational principals for the testing of operational factors creating community variations with low noise and appropriate replication. Understanding the processes that drive microbial community diversity and assembly is a key question in microbial ecology. In the future, this knowledge can be used to manipulate the structure of microbial communities and improve system performance in WWTPs.

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This work aimed at demonstrating the natural bioaugmentation of biological activated sludge systems with nitrifying biomass from influent wastewater in lab-scale reactors. Three sequencing batch reactors (SBR) were fed with sterile synthetic wastewater. While nitrification was complete at a temperature of 8 °C and a SRT of 20 days, it failed when the temperature was lowered to 5 °C, and the SRT decreased to 7 days. In the test period, the sterile synthetic wastewater fed to the Test Reactor was supplemented by influent solids harvested at a full-scale treatment facility at a total suspended solids concentration of 100 mg/L, which corresponded to approximately 5 mg-COD/L of nitrifying biomass. Upon this addition, nitrification was restores. Subsequent halting the supply of influent solids to the Test Reactor led a rapid failure of nitrification and washout of nitrifiers from the SBR. Reproducibility was demonstrated by switching the feed composition between the Test and Negative control reactors. PCR-based amplicon sequencing analyses targeting the amoA, and nxrB genes of the Nitrospira genus have shown that the influent wastewater governed the structure and composition of the activated sludge nitrifying populations. The most abundant ammonia-oxidizing bacteria (AOB) and Nitrospira-related nitrite-oxidizing bacteria (NOB) in the influent seeds occurred as the most dominant ones in the activated sludge. This pattern was observed even when the influent seeds varied over time. The heterotrophic populations were less affected by the influent seeds with the activated sludge and raw sewage showing distinct microbial populations based on principal coordinate analysis (PCoA). However, the immigrant populations appeared to modulate the structure of the activated sludge heterotrophic communities to some extent. These findings provide concrete evidence of the presence of active nitrifiers in raw wastewater capable of supporting nitrification in an otherwise non-conducive environment. This may have important implications on process design, operation and optimization of wastewater treatment systems.