Anammox Community Research Articles

Important role of Chloroflexi: Improved the cooperation of anammox community under PFOS stress

Perfluorooctane sulfonic acid (PFOS) has received extensive attention in recent years due to its persistence and biotoxicity. As an advanced biological nitrogen removal process, anaerobic ammonium oxidation (anammox) is considered to be the most promising process to replace traditional nitrogen removal technology in the future. However, it is still unclear how PFOS affects microbial community and network structure in anammox system. The study found that PFOS had little influence on the nitrogen removal efficiency. Through microbial community and network analysis, the results showed that the typical anammox bacteria (Candidatus_Kuenenia, Candidatus_Jettenia, and Candidatus_Brocadia) enriched with the increasing concentrations of PFOS. Overall network complexity and proportion of positive links was significantly increased under 10 mg/L PFOS. Especially the subnetwork of Chloroflexi owned 100 % positive links under 10 mg/L PFOS, which accounted for 53.62 % of the corresponding overall network. The genus OLB13 and norank_f__Anaerolineaceae from Chloroflexi could cooperate with anammox bacteria to resist the impact of PFOS and further maintained the stability of anammox system. This study provided new insights into the effects of perfluoroalkyl compounds on anammox system.

Deciphering the anammox microbial community succession with humic acid exposure to optimize large anammox granules for robust nitrogen removal

The robustness of the anaerobic ammonia oxidation (anammox) process in treating wastewater with high concentrations of humic acids (HAs), including landfill leachate and sludge anaerobic digestion liquid, has been paid great attention. This study revealed that the anammox sludge granule size of 1.0–2.0 mm could be robust under the HA exposure with high concentrations. The total nitrogen removal efficiency (NRE) was 96.2% at the HA concentration of 20–100 mg/L, while the NRE was 88.5% at the HA concentration of 500 mg/L, with reduced by 7.7%. The increased extracellular polymeric substances (EPS) content which was stimulated by the HA exposure favored the formation of large granules (1.0–2.0 mm) by enveloping medium and micro granules (0.2–1.0 mm). The abundance of anammox bacteria Candidatus Brocadia was found to be higher (14.2%) in large anammox granules sized 1.0–2.0 mm, suggesting a potentially high anammox activity. However, the abundance of denitrifiers Denitratisoma increased by 4.3% in ultra-large anammox granules sized >2.0 mm, which could be attributed to the high EPS content for heterotrophic denitrifiers metabolism as organic matter. The feedback mechanism of the anammox community for maintaining the ecological function under the HA exposure resulted in a closely related microbial community, with positive and negative correlations in the ecological network increased by 64.3%. This study revealed that the HA exposure of the anammox system resulted in the anammox granules of 1.0–2.0 mm size being the dominant granules with robust nitrogen removal, providing significant guidance for the optimization of anammox granules for an efficient treatment of HA-containing wastewater in anammox applications.

Metagenomic evidence of a novel anammox community in a cold aquifer with high nitrogen pollution

The process of anaerobic ammonium oxidation by nitrite (anammox) is a globally essential part of N cycle. To date, 8 Candidatus genera and more than 22 species of anammox bacteria have been discovered in various anthropogenic and natural habitats, including nitrogen-polluted aquifers. In this work, anammox bacteria were detected for the first time in the groundwater ecosystem with high anthropogenic nitrogen pollution (up to 1760 mg NO3−-N/L and 280 mg NH4+-N/L) and low year-round temperature (7–8 °C) in the zone of a uranium sludge repository. Further metagenomic analysis resulted in retrieval of metagenome-assembled genomes of 4 distinct anammox bacteria: a new genus named Ca. Frigussubterria, new species in Ca. Kuenenia, and two strains of a new species in Ca. Scalindua. Analysis of the genomes revealed essential genes involved in anammox metabolism. Both strains of Ca. Scalindua chemeplantae had a high copy number of genes encoding the cold shock proteins CspA/B, which can also function as an antifreeze protein (CspB). Ca. Kuenenia glazoviensis and Ca. Frigussubterria udmurtiae were abundant in less N-polluted site, while Ca. Scalindua chemeplantae inhabited both sites. Genes for urea utilization, reduction of insoluble Fe2O3 or MnO2, assimilatory sulfate reduction, reactive oxygen detoxification, nitrate reduction to ammonium, and putatively arsenate respiration were found. These findings enrich knowledge of the functional and phylogenetic diversity of anammox bacteria and improve understanding of the nitrogen cycle in polluted aquifers.

Mechanisms of anammox granular sludge reactor effluent as biostimulant: Shaping microenvironment for anammox metabolism

Effluent from anammox granular sludge (AnGS) bioreactor contains microbes and microbial products. This study explored mechanisms of utilizing AnGS-effluent as biostimulant for anammox process enhancement. Compared with no AnGS-effluent supplemented control reactor, 5.0 and 1.3 times higher ammonium nitrogen and total inorganic nitrogen removal rates, respectively were obtained with continuous AnGS-effluent supplementation after 98 days’ operation. Anammox bacteria from Candidatus Brocadia accounted for 0.1 % (DNA level) and 1.3 %–1.5 % (RNA level) in control reactor, and 2.9 % (DNA level) and 54.5 %–55.4 % (RNA level) in the AnGS-effluent-fed reactor. Influent microbial immigration evaluation showed that bacterial immigration via AnGS-effluent supplementation was not the main contributor to active anammox community development. Amino acids biosynthesis, B-vitamins and coenzymes metabolism related pathways were facilitated by AnGS-effluent supplementation. AnGS-effluent supplementation aided anammox metabolic activity by shaping microenvironment and microbial interactions. This study provides insights into enhancing anammox bacterial metabolism with AnGS-effluent microbial products as biostimulant.

Biosynthetic potential of uncultured anammox community bacteria revealed through multi-omics analysis

Microbial secondary metabolites (SMs) and their derivatives have been widely used in medicine, agriculture, and energy. Growing needs for renewable energy and the challenges posed by antibiotic resistance, cancer, and pesticides emphasize the crucial hunt for new SMs. Anaerobic ammonium-oxidation (anammox) systems harbor many uncultured or underexplored bacteria, representing potential resources for discovering novel SMs. Leveraging HiFi long-read metagenomic sequencing, 1,040 biosynthetic gene clusters (BGCs) were unearthed from the anammox microbiome with 58% being complete and showcasing rich diversity. Most of them showed distant relations to known BGCs, implying novelty. Members of the underexplored lineages (Chloroflexota and Planctomycetota) and Proteobacteria contained lots of BGCs, showcasing substantial biosynthetic potential. Metaproteomic results indicated that Planctomycetota members harbored the most active BGCs, particularly those involved in producing potential biofuel—ladderane. Overall, these findings underscore that anammox microbiomes could serve as valuable resources for mining novel BGCs and discovering new SMs for practical application.

New insights into functional divergence and adaptive evolution of uncultured bacteria in anammox community by complete genome-centric analysis

Anaerobic ammonium-oxidation (anammox) bacteria play a crucial role in global nitrogen cycling and wastewater nitrogen removal, but they share symbiotic relationships with various other microorganisms. Functional divergence and adaptive evolution of uncultured bacteria in anammox community remain underexplored. Although shotgun metagenomics based on short reads has been widely used in anammox research, metagenome-assembled genomes (MAGs) are often discontinuous and highly contaminated, which limits in-depth analyses of anammox communities. Here, for the first time, we performed Pacific Biosciences high-fidelity (HiFi) long-read sequencing on the anammox granule sludge sample from a lab-scale bioreactor, and obtained 30 accurate and complete metagenome-assembled genomes (cMAGs). These cMAGs were obtained by selecting high-quality circular contigs from initial assemblies of long reads generated by HiFi sequencing, eliminating the need for Illumina short reads, binning, and reassembly. One new anammox species affiliated with Candidatus Jettenia and three species affiliated with novel families were found in this anammox community. cMAG-centric analysis revealed functional divergence in general and nitrogen metabolism among the anammox community members, and they might adopt a cross-feeding strategy in organic matter, cofactors, and vitamins. Furthermore, we identified 63 mobile genetic elements (MGEs) and 50 putative horizontal gene transfer (HGT) events within these cMAGs. The results suggest that HGT events and MGEs related to phage and integration or excision, particularly transposons containing tnpA in anammox bacteria, might play important roles in the adaptive evolution of this anammox community. The cMAGs generated in the present study could be used to establish of a comprehensive database for anammox bacteria and associated microorganisms. These findings highlight the advantages of HiFi sequencing for the studies of complex mixed cultures and advance the understanding of anammox communities.

Rhizosphere-Associated Anammox Bacterial Diversity and Abundance of Nitrogen Cycle-Related Functional Genes of Emergent Macrophytes in Eutrophic Wetlands.

Rhizospheric microbial community of emergent macrophytes plays an important role in nitrogen removal, especially in the eutrophic wetlands. The objective of this study was to identify the differences in anammox bacterial community composition among different emergent macrophytes and investigate revealed the the main factors affecting on the composition, diversity, and abundance of anammox bacterial community. Results showed that the composition, diversity, and abundance of the anammox community were significantly different between the vegetated sediments of three emergent macrophytes and unvegetated sediment. The composition of the anammox bacterial community was different in the vegetated sediments of different emergent macrophytes. Also, the abundance of nitrogen cycle-related functional genes in the vegetated sediments was found to be higher than that in the unvegetated sediment. Canonical correspondence analysis (CCA) and structural equation models analysis (SEM) showed that salinity and pH were the main environmental factors influencing the composition and diversity of the anammox bacterial community and NO2--N indirectly affected anammox bacterial community diversity by affecting TOC. nirK-type denitrifying bacteria abundance had significant effects on the bacterial community composition, diversity, and abundance of anammox bacteria. The community composition of anammox bacteria varies with emergent macrophyte species. The rhizosphere of emergent macrophytes provides a favorable environment and promotes the growth of nitrogen cycling-related microorganisms that likely accelerate nitrogen removal in eutrophic wetlands.

Influence of C/N ratio and ammonia on nitrogen removal and N2O emissions from one-stage partial denitrification coupled with anammox

The development of low carbon treatment processes is an important issue worldwide. Partial denitrification coupled with anammox (PD/A) is a novel strategy to remove nitrogen and reduce N2O emissions. The influence of C/N ratio and NH4+ concentration on nitrogen removal and N2O emissions was investigated in batch reactors filled with PD/A coupled sludge. A C/N ratio of 2.1 was effective for nitrogen removal and N2O reduction; higher ammonia concentration might make anammox more active and indirectly reduce N2O emissions. Long-term operation further confirmed that a C/N ratio of 2.1 resulted in a minimum effluent N2O concentration (mean value of 0.94 μmol L−1); as the influent NH4+ concentration decreased to 50 mg L−1 (NH4+-N/NO3−-N: 1), the nitrogen removal rate increased to 82.41%. Microbial analysis showed that anammox bacteria (Candidatus Jettenia and Ca. Brocadia) were enriched in the PD/A system and Ca. Brocadia gradually dominated the anammox community, with the relative abundance increasing from 1.69% to 18.44% between days 97 and 141. Finally, functional gene analysis indicated that the abundance of nirS/K and hao involved in partial denitrification and anammox, respectively, increased during long-term operation of the reactor; this change benefitted nitrogen metabolism in anammox, which could indirectly reduce N2O emissions.

Effect of Sulfide on the Processes of Transformation of Nitrogen Compounds and the Microbial Community Composition in the Anammox Bioreactor

Anammox is one of the most important processes in the global nitrogen cycle and the basis for an efficient technology of nitrogen removal from wastewater. The effect of the presence of sulfide in wastewater on the transformation of nitrogen compounds by the anammox community has been insufficiently studied. The present work dealt with the effect of sulfide on nitrogen removal efficiency and the dynamics of nitrogen species in a laboratory sequencing batch bioreactor modeling the functioning of the anammox community carrying out ammonium oxidation via nitritation and anammox and nitrite oxidation. The 16S rRNA gene profiling of the community of the anammox-activated sludge attached to the stationary carrier revealed members of the key physiological groups: ammonium oxidizers of the genus Nitrosomonas, nitrite oxidizers of the genus Nitrosospira, and anammox bacteria of the genera Candidatus Brocadia and Ca. Jettenia, as well as members of other bacterial genera. Nitrate removal was not sensitive to sulfide at concentrations up to 50 mg S/L and decreased by 17% at 100 mg/L. The threshold of sulfide sensitivity for group I nitrifiers was ~50 mg/L, while anammox bacteria were resistant to sulfide concentrations of up to 100 mg S/L in the incoming water. Group II nitrifiers were probably the most sulfide-sensitive components of the community. A drastic increase in the abundance of members of the family Hydrogenophilaceae at elevated sulfide concentrations, together with the precipitation of elemental sulfur, may indicate sulfide oxidation either by molecular oxygen or via nitrate reduction; this finding requires further investigation. This is the first report on the different effects of sulfide on the growth rate of members of the nitrifying genus Nitrosomonas, increasing/decreasing or not affecting it for different phylotypes at elevated sulfide concentrations.

Affinity difference determines the assembly and interaction mode of anammox community reconstructed by siderophores

Anammox community is the core of anammox process. The constancy of the anammox community determines the stability of the anammox process and the ability of withstand environmental impact. Community stability is influenced by the assembly and interaction mode of the community. This study aimed to explore the assembly, interaction mode, and stability of anammox community influenced by two siderophores (enterobactin and putrebactin) specific for Ca. Brocadia and Ca. Kuenenia as produced in our previous research. Siderophores improved the stability of the anammox community, among which vulnerability dropped by 30.02 % and 72.53 % respectively. Enterobactin and putrebactin altered the succession speed and assembly pattern of communities, with a respective increase of 9.77 % and 80.87 % in the deterministic process of anammox community assembly, respectively. Enterobactin and putrebactin reduced the dependence of Ca. Brocadia and Ca. Kuenenia on companion bacteria by 60 items and 27 items respectively. The affinity of different siderophore-Fe with bacterial membrane receptors caused variations in community reconstruction, with Ca. Brocadia and Ca. Kuenenia exhibiting the highest affinity with enterobactin-Fe (−11.4 kcal/mol) and putrebactin-Fe (−9.0 kcal/mol), respectively. This study demonstrated how siderophores can enhance the stability of anammox process by regulating assembly and interaction mode of anammox community, while also revealing the underlying molecular mechanisms.

The long-term effect of glutaraldehyde on the bacterial community in anaerobic ammonium oxidation reactor

A 160-day incubation was performed with two anammox reactors (GA and CK) to investigate the effect of glutaraldehyde. The results indicated that anammox bacteria were very sensitive when glutaraldehyde in GA reactor increased to 40 mg/L, the nitrogen removal efficiency sharply decreased to 11%, only one-quarter of CK. Glutaraldehyde changed spatial distribution of exopolysaccharides, caused anammox bacteria (Brocadia CK_gra75) to disassociate from granules (24.70% of the reads in CK but only 14.09% in GA granules). Metagenome analysis indicated glutaraldehyde led to the denitrifier community succession from strains without nir (nitrite reductase) and nor (nitric oxide reductases) genes to those with them, and the rapid growth of denitrifiers with NodT (an outer membrane factor)-related efflux pumps replacing those with another TolC -related ones. Meanwhile, Brocadia CK_gra75 lacks the NodT proteins. This study provides important insight into community adaptation and potential resistance mechanism in an active anammox community after exposure to disinfectant.

Fast start-up of anammox process: Effects of extracellular polymeric substances addition on performance, granule properties, and bacterial community structure

The slow startup is the major obstacle to the application of anaerobic ammonium oxidation (anammox) process in mainstream wastewater treatment. Extracellular polymeric substances (EPS) are one potential resource for stable anammox reactor operation. Response surface analysis was used to optimize the specific anammox activity (SAA) with the addition of EPS; SAA was maximum at a temperature of 35 °C and the EPS concentration of 4 mg/L. By comparing the nitrogen removal of anammox reactors with no EPS (R0), immobilized EPS (EPS-alginate beads) (R1), and liquid EPS (R2), we found that EPS-alginate beads significantly speed up the startup of anammox process and enable the start time to be shortened from 31 to 19 days. As a result of the higher MLVSS content, higher zeta potential, and lower SVI30, anammox granules of R1 exhibited a stronger capacity to aggregate. Moreover, EPS extracted from R1 had higher flocculation efficiencies than EPS derived from R0 and R2. Phylogenetic analysis of 16S rRNA genes revealed that the main anammox species in R1 is Kuenenia taxon. To clarify the relative significance of stochastic vs deterministic processes in the anammox community, neutral model and network analysis are employed. In R1, community assembly became more deterministic and stable than in other cultures. Our results show that EPS might inhibit heterotrophic denitrification and thereby promote anammox activity. This study suggested a quick start-up strategy for the anammox process based on resource recovery, which is helpful for environmentally sustainable and energy-efficient wastewater treatment.

Nitrogen-loss and associated microbial communities in sediments from the Yangtze Estuary and adjacent sea

IntroductionDenitrification and anammox play the crucial role for N removal processes in coastal ecosystems, but the ecological distribution of denitrifying and anammox microorganisms and their N removal rates in the Yangtze Estuary and its adjacent sea are required in-depth analysis.MethodsHere, we utilized high-throughput sequencing, qPCR, and 15N isotope to reveal the community structure and function of denitrifying and anammox microorganisms in the surface sediments from Yangtze Estuary and adjacent sea.ResultsThe results suggested that the gene abundances of nirS and nirK for denitrifiers were higher than AMX 16S rRNA for anammox bacteria. The genera composition of nirS- and nirK-encoding denitrifiers communities showed different distribution patterns. Furthermore, Candidatus Anammoximicrobium dominated the anammox community, implying the anammox oxidation capacity of the other genera should be noted in marine sediments.DiscussionCompared to anammox, denitrification was the dominant contributor of nitrogen removal process and contributed 73.5% on average. The sediment Chla was the key factor to regulate denitrification and anammox rates, indicating the fresh organic matter was more labile and easier to be utilized by NOx− removal processes.

Carrier type affects anammox community assembly, species interactions and nitrogen conversion

The impacts of carrier type on anammox community assembly, species interactions and nitrogen conversion were studied in this work. It was found that in addition to shared species with higher abundance, different carrier types recruited rare species by imposing selection pressure. Results from co-occurrence networks revealed that carrier type strongly influenced interactions between keystone species inhabiting within anammox biofilm through potentially inducing niche differences. Overall, elastic cubic sponges would lead to closer cooperation between different populations, whereas plastic hollow cylinders would trigger fiercer competition. Meanwhile, the results based on metagenomics sequencing showed carrier type significantly affected nitrogen conversion related genes abundances, and higher reads number was detected on the elastic cubic sponges. The information obtained in this work could provide some valuable information for the selection and optimization of carrier type in the anammox process.

Effect of Mineral Carriers on Biofilm Formation and Nitrogen Removal Activity by an Indigenous Anammox Community from Cold Groundwater Ecosystem Alone and Bioaugmented with Biomass from a "Warm" Anammox Reactor.

Simple SummaryDuring more than 50 years of exploitation of the sludge repositories near Chepetsky Mechanical Plant (Glazov, Udmurtia, Russia) containing solid wastes of uranium and processed polymetallic concentrate, the soluble compounds entered the upper aquifer due to infiltration. Nowadays, this has resulted in a high level of pollution of the groundwater with reduced and oxidized nitrogen compounds. In this work, quartz, kaolin, and bentonite clays from various deposits were shown to induce biofilm formation and enhance nitrogen removal by an indigenous microbial community capable of anaerobic ammonium oxidation with nitrite (anammox) at low temperatures. The addition of a “warm” anammox community was also effective in further improving nitrogen removal and expanding the list of mineral carriers most suitable for creating a permeable reactive barrier. It has been suggested that the anammox activity is determined by the presence of essential trace elements in the carrier, the morphology of its surface, and most importantly, competition from rapidly growing microbial groups. Future work was discussed to adapt the “warm” anammox community to cold and provide the anammox community with nitrite through a partial denitrification route within the scope of sustainable anammox-based bioremediation of a nitrogen-polluted cold aquifer. In this unique habitat, novel species of anammox bacteria that are adapted to cold and heavy nitrogen pollution can be discovered.The complex pollution of aquifers by reduced and oxidized nitrogen compounds is currently considered one of the urgent environmental problems that require non-standard solutions. This work was a laboratory-scale trial to show the feasibility of using various mineral carriers to create a permeable in situ barrier in cold (10 °C) aquifers with extremely high nitrogen pollution and inhabited by the Candidatus Scalindua-dominated indigenous anammox community. It has been established that for the removal of ammonium and nitrite in situ due to the predominant contribution of the anammox process, quartz, kaolin clays of the Kantatsky and Kamalinsky deposits, bentonite clay of the Berezovsky deposit, and zeolite of the Kholinsky deposit can be used as components of the permeable barrier. Biofouling of natural loams from a contaminated aquifer can also occur under favorable conditions. It has been suggested that the anammox activity is determined by a number of factors, including the presence of the essential trace elements in the carrier and the surface morphology. However, one of the most important factors is competition with other microbial groups that can develop on the surface of the carrier at a faster rate. For this reason, carriers with a high specific surface area and containing the necessary microelements were overgrown with the most rapidly growing microorganisms. Bioaugmentation with a “warm” anammox community from a laboratory reactor dominated by Ca. Kuenenia improved nitrogen removal rates and biofilm formation on most of the mineral carriers, including bentonite clay of the Dinozavrovoye deposit, as well as loamy rock and zeolite-containing tripoli, in addition to carriers that perform best with the indigenous anammox community. The feasibility of coupled partial denitrification–anammox and the adaptation of a “warm” anammox community to low temperatures and hazardous components contained in polluted groundwater prior to bioaugmentation should be the scope of future research to enhance the anammox process in cold, nitrate-rich aquifers.

Genome-centric metagenomics insights into functional divergence and horizontal gene transfer of denitrifying bacteria in anammox consortia

Denitrifying bacteria with high abundances in anammox communities play crucial roles in achieving stable anammox-based systems. Despite the relative constant composition of denitrifying bacteria, their functional diversity remains to be explored in anammox communities. Herein, a total of 77 high-quality metagenome-assembled genomes (MAGs) of denitrifying bacteria were recovered from the anammox community in a full-scale swine wastewater treatment plant. Among these microbes, a total of 26 MAGs were affiliated with the seven dominant denitrifying genera that have total abundances higher than 1%. A meta-analysis of these species suggested that external organics reduced the abundances of genus Ignavibacterium and species MAG.305 of UTPRO2 in anammox communities. Comparative genome analysis revealed functional divergence across different denitrifying bacteria, largely owing to their distinct capabilities for carbohydrate (including endogenous and exogenous) utilization and vitamin (e.g., pantothenate and thiamine) biosynthesis. Serval microbes in this system contained fewer genes encoding biotin, pantothenate and methionine biosynthesis compared with their related species from other habitats. In addition, the genes encoding energy production and conversion (73 genes) and inorganic ion transport (53 genes) putatively transferred from other species to denitrifying bacteria, while these denitrifying bacteria (especially genera UTPRO2 and SCN-69–89) likely donated the genes encoding nutrients (e.g., inorganic ion and amino acid) transporter (64 genes) for other members to utilize new metabolites. Collectively, these findings highlighted the functional divergence of these denitrifying bacteria and speculated that the genetic interactions within anammox communities through horizontal gene transfer may be one of the reasons for their functional divergence.

Insight into the microbial and genetic response of anammox biomass to broad range concentrations of different antibiotics: Linking performance and mechanism

Antibiotics have become emerging pollutants occurring in wastewater, influencing the activity of microorganisms responsible for wastewater treatment. Moreover, the potential application of the anammox process in the treatment of antibiotic-containing wastewater has paid much attention. A common antibiotic, OTC (oxytetracycline), CIP (ciprofloxacin), and CLA (clarithromycin) are recognized to be monitored in wastewater due to their relatively high concentration and hazardous impact on the environment. However, their effect on the anammox process remains unknown. Therefore, this paper presents the study concerning the long-term effects of a successive concentration of three antibiotics (OTC, CIP, CLA) on the anammox process, with special emphasis on treatment efficiency, resistance mechanism, bacteria cell morphology, and activated sludge community structure. It is worth noting that the influence of a successive concentration of CIP and CLA has been studied for the first time. Results revealed that anammox community could adapt to CIP, OTC, and CLA at low concentrations (<1 mg L-1), while the high concentration of antibiotics (100 mg L-1) reduced the nitrogen removal rate (NRR) by 27 % (OTC), 30 % (CIP), and 56 % (CLA). Community structure analysis showed that the abundance of Planctomycetes increased with the increase of CIP and CLA concentration while decreasing under OTC stress. On contrary, other nitrogen-cycle bacteria (e.g., Nitrospira) contributed to the nitrogen removal, especially during antibiotic suppression. The abundance of corresponding ARGs (OTC resistance gens: tetX, tetC, tetW, CIP resistance gens: qnrB4, qnrS, CLA resistance gens: mphA) generally increased under antibiotic suppression. In addition, co-occurrence analysis showed that anammox bacteria might participate in the transfer of macrolide resistance genes. The findings of this study are essential in understanding the mechanisms of three antibiotics commonly occurring in wastewater during the anammox process. Moreover, the results have implications for using the anammox process for antibiotic-containing wastewater treatment and provide the operational guidance for stable conducting of the anammox process under antibiotics suppression.

Micron-scale biogeography reveals conservative intra anammox bacteria spatial co-associations

Micron-scale resolution can help to reliably identify true taxon-taxon interactions in complex microbial communities. Despite widespread recognition of the critical role of metabolic interactions in anaerobic ammonium oxidation (anammox) system performance, no studies have examined microbial interactions at the micron-scale in anammox consortia. To fill this gap, we extensively sampled (totally 242 samples) the consortia of a lab-scale anammox reactor at different length scales, including bulk-scale (∼cm), macro-scale (300–500 µm) and micron-scale (70–100 µm). We firstly observed evident micron-scale heterogeneity in anammox consortia, with the relative abundance of anammox bacteria fluctuated greatly across individual clusters (2.0%–79.3%), indicating that the biotic interactions play a significant role in the assembly of anammox communities under well-controlled and well-mixed condition. Importantly, by mapping the spatial associations in anammox consortia at micron-scale, we demonstrated that the conserved co-associations for anammox bacteria were restricted to three different Brocadia species over time, and their co-associations with heterotrophs were random, implying that there was no statistically significant symbiotic interaction between anammox bacteria and other heterotrophic populations. Further metagenomic binning revealed that the quorum sensing with secondary messenger c-di-GMP potentially holding on the conservative metabolic cooperation among Brocadia species. These results shed new light on the social behavior of the anammox community. Overall, delineating of biological structures at micron-scale opens a new way of monitoring the microbial spatial structure and interactions, paving the way for improved community engineering of biotreatment systems.

Response and resilience of anammox consortia to nutrient starvation

BackgroundIt is of critical importance to understand how anammox consortia respond to disturbance events and fluctuations in the wastewater treatment reactors. Although the responses of anammox consortia to operational parameters (e.g., temperature, dissolved oxygen, nutrient concentrations) have frequently been reported in previous studies, less is known about their responses and resilience when they suffer from nutrient interruption.ResultsHere, we investigated the anammox community states and transcriptional patterns before and after a short-term nutrient starvation (3 days) to determine how anammox consortia respond to and recover from such stress. The results demonstrated that the remarkable changes in transcriptional patterns, rather than the community compositions were associated with the nutritional stress. The divergent expression of genes involved in anammox reactions, especially the hydrazine synthase complex (HZS), and nutrient transportation might function as part of a starvation response mechanism in anammox bacteria. In addition, effective energy conservation and substrate supply strategies (ATP accumulation, upregulated amino acid biosynthesis, and enhanced protein degradation) and synergistic interactions between anammox bacteria and heterotrophs might benefit their survival during starvation and the ensuing recovery of the anammox process. Compared with abundant heterotrophs in the anammox system, the overall transcription pattern of the core autotrophic producers (i.e., anammox bacteria) was highly resilient and quickly returned to its pre-starvation state, further contributing to the prompt recovery when the feeding was resumed.ConclusionsThese findings provide important insights into nutritional stress-induced changes in transcriptional activities in the anammox consortia and would be beneficial for the understanding of the capacity of anammox consortia in response to stress and process stability in the engineered ecosystems.7cebb1f-VYhKkDnFTSZAj9Video

Specific Denitrifying and Dissimilatory Nitrate Reduction to Ammonium Bacteria Assisted the Recovery of Anammox Community From Nitrite Inhibition.

The anaerobic ammonium oxidation (anammox) by autotrophic anaerobic ammonia-oxidizing bacteria (AnAOB) is a biological process used to remove reactive nitrogen from wastewater. It has been repeatedly reported that elevated nitrite concentrations can severely inhibit the growth of AnAOB, which renders the anammox process challenging for industrial-scale applications. Both denitrifying (DN) and dissimilatory nitrate reduction to ammonium (DNRA) bacteria can potentially consume excess nitrite in an anammox system to prevent its inhibitory effect on AnAOB. However, metabolic interactions among DN, DNRA, and AnAOB bacteria under elevated nitrite conditions remain to be elucidated at metabolic resolutions. In this study, a laboratory-scale anammox bioreactor was used to conduct an investigation of the microbial shift and functional interactions of AnAOB, DN, and DNRA bacteria during a long-term nitrite inhibition to eventual self-recovery episode. The relative abundance of AnAOB first decreased due to high nitrite concentration, which lowered the system’s nitrogen removal efficiency, but then recovered automatically without any external interference. Based on the relative abundance variations of genomes in the inhibition, adaptation, and recovery periods, we found that DN and DNRA bacteria could be divided into three niche groups: type I (types Ia and Ib) that includes mainly DN bacteria and type II and type III that include primarily DNRA bacteria. Type Ia and type II bacteria outcompeted other bacteria in the inhibition and adaptation periods, respectively. They were recognized as potential nitrite scavengers at high nitrite concentrations, contributing to stabilizing the nitrite concentration and the eventual recovery of the anammox system. These findings shed light on the potential engineering solutions to maintain a robust and efficient industrial-scale anammox process.