Heterotrophic Nitrification-aerobic Denitrification Bacteria Research Articles

Ammonia removal through combined methane oxidation and nitrification-denitrification and the interactions among functional microorganisms

It would be highly beneficial to use the methane produced by anaerobic digestion, which is low cost and accessible, as the carbon source in the removal of nitrogenous contaminants in wastewater. However, there is a knowledge gap regarding coupling systems that entail methane oxidation, nitrification, and denitrification, which restricts their industrial application. In this study, we acclimated a mixed culture to deal with simultaneous nitrification-denitrification coupled to methane oxidation in a laboratory-scale hollow-fiber membrane biofilm reactor, which achieved a steady ammonia removal rate of 38.09 mg N/(L•d). Furthermore, a series of batch experiments were conducted to test methane oxidation coupled to nitrate denitrification (AME-D3), nitrite denitrification (AME-D2), and simultaneous nitrification and denitrification (ME-SND). The molar ratio between methane consumed and nitrate reduced (C/N) equals 10 and 5 mol CH4C mol−1 NO3N in AME-D3 and AME-D2, averagely and respectively. Without methane injection, the removal of nitrates and nitrites was very low, indicating that the coupling of nitrate/nitrite denitrification and methane oxidation was beneficial. The average ammonia removal rates in the 20% O2 and 25% O2 groups were 20.06 and 22.03 mg N/(L•d) in the ME-SND system, respectively. Without methane, the ammonia oxidation rate declined, and large amounts of nitrite accumulated. As traditional ammonia and nitrite oxidation approaches are autotrophic, we proposed the possibility of heterotrophic nitrification-aerobic denitrification (HN-AD). To study the coupling systems, the microbial communities and functional bacteria were analyzed. The results indicated that the system contained a guild of methanotrophs (mainly Methylobacter) and HN-AD bacteria (mainly Chrysobacterium and Comamonas).

Performance and microbial ecology of a novel moving bed biofilm reactor process inoculated with heterotrophic nitrification-aerobic denitrification bacteria for high ammonia nitrogen wastewater treatment

To overcome long start-up time, poor ammonia tolerance and removal performance of traditional moving bed biofilm reactor (MBBR) inoculated with activated sludge for high-ammonia wastewater treatment, a novel MBBR based on heterotrophic nitrification-aerobic denitrification (HN-AD) was proposed. Start-up of MBBR was firstly performed via inoculated with HN-AD bacteria. Start-up time was shortened from 39 d to 15 d, NH4+ tolerance was enhanced from 200 mg/L to 1000 mg/L, and TN removal was increased from 30.4% to 80.7%. The carrier types and NH4+ concentration had significant effects on nitrogen removal and microbial ecology. When the NH4+ concentration was increased to 900 mg/L in MBBR using polyvinyl alcohol gel as carrier, the TN removal, the abundance of HN-AD bacteria Acinetobacter, Pseudomonas and Paracoccus, which played a key role in TN removal and ammonia tolerance, and the abundance of genes related to nitrogen removal were much higher than those of MBBR using kaldness.

Identification, interactions, nitrogen removal pathways and performances of culturable heterotrophic nitrification-aerobic denitrification bacteria from mariculture water by using cell culture and metagenomics

The rapid expansion of aquaculture industry brings about significant environmental concerns, especially nitrogen pollution. Compared to nitrogen bioconversion implemented by the conventional autotrophic nitrifiers and anaerobic denitrifiers, bacteria capable of heterotrophic nitrification-aerobic denitrification (HNAD) in mariculture environments have yet to be well understood. In this study, twenty-five species of new halophilic HNAD bacteria were isolated and identified from mariculture water. By these strains co-cultured in the synthetic mariculture water (ammonia: 5 mg/L, C/N: 5, salinity: 30‰), microbial dynamic analysis showed that ammonia were mainly removed by dominant genera of Marinomonas, Marinobacterium, Halomonas, and Cobetia which simultaneously had positive correlations to total nitrogen removal. Metagenomic annotations revealed that inorganic-N was converted into gaseous-N and organic-N by these HNAD bacteria through nitrogen metabolism pathways of assimilation, partial nitrification, nitroalkane oxidation, nitrate/nitrite dissimilation reduction, and denitrification. Among them, due to the interspecific coexistence and cooperation, Marinomonas communis &Halomonas titanicae, Marinomonas communis &Cobetia marina, Marinomonas aquimarina &Halomonas titanicae, and Marinomonas aquimarina &Cobetia marina exhibited significantly better inorganic-N removal efficiency and stability. The four novel bacterial consortia could transform approximately 60% of initial ammonia into intracellular organic-N (18–20%) and gaseous-N (36–38%), which were significantly higher than those of their single strains. These findings will contribute to understanding and developing the culturable HNAD bacteria as promising candidates for nitrogen pollution control and water bioremediation in mariculture or other saline environments.

Intensified nitrogen removal by heterotrophic nitrification aerobic denitrification bacteria in two pilot-scale tidal flow constructed wetlands: Influence of influent C/N ratios and tidal strategies

This study investigated the influence of C/N ratios and tidal strategies on nitrogen removal and bacterial communities in two pilot-scale tidal flow constructed wetlands (TFCWs) with simultaneous nitrification–denitrification process. Heterotrophic nitrification aerobic denitrification (HNAD) was the main nitrogen transformation pathway in both TFCWs. High C/N ratios and effluent circulation at low temperature promoted HNAD in TFCWs with high nitrogen removal efficiencies (72.6%–95.5% for NH4+-N and 70.9%~91.8% for TN). Effluent circulation had more influence on bacterial community structure and diversity than C/N ratios. Among 16 detected genera related to nitrogen removal, HNAD bacteria (HNADB) were abundant. Especially, some dominant HNADB (e.g. Aeromonas, Hydrogenophage and Gemmobacter) were core genera, showing positive interactions with other genera related to nitrogen removal. Tidal strategies had more contribution to the shifts in these genera than C/N ratios. This study highlights the importance of HNADB in pilot-scale TFCWs and their responses to C/N ratios and tidal strategies.

Advances in heterotrophic nitrification-aerobic denitrifying bacteria for nitrogen removal under extreme conditions

Heterotrophic nitrification-aerobic denitrification (HN-AD) is an enrichment and breakthrough theory of traditional autotrophic nitrification heterotrophic denitrification. Heterotrophic nitrification-aerobic denitrifiers with the feature of wide distribution, strong adaptability and unique metabolic mechanism have many special advantages, including fast-growing, rapid biodegradability and long lasting activity, which can rapidly remove ammonia nitrogen, nitrate nitrogen (NO₃⁻-N) and nitrite nitrogen (NO₂⁻-N) under aerobic conditions simultaneously. Therefore, HN-AD bacteria show the important potential for denitrification under extreme conditions with high-salt, low-temperature or high-ammonia nitrogen environment, and HN-AD bacteria attract extensive attention in the field of biological denitrification of wastewater. In this review, we first introduce the previously reported HN-AD bacterial species which have denitrification performance in the extreme environments and state their typical metabolic mechanism. Then, we systematically analyze the nitrogen removal characteristics and potential under extreme conditions. We also briefly describe the progress in the application of HN-AD bacterial. Finally, we outlook the application prospects and research directions of HN-AD denitrification technology.

New insight into the nitrogen metabolism of simultaneous heterotrophic nitrification-aerobic denitrification bacterium in mRNA expression.

Here, the draft genome of simultaneous nitrification-denitrification strain (SND) Klebsiella sp. KSND revealed possible existence of genes involved in N-assimilation and -dissimilation pathways. The change levels of genes under defined N-sources were analyzed by Quantitative Real-Time PCR. It suggested that NH4+-assimilation via NADP-glutamate dehydrogenase pathway would occur preferentially. NirBD genes were tightly regulated in a lower level, so that nitrite was rapidly consumed for detoxication by denitrification. Three types of nitrate reductase homologues are surprisingly present in KSND, whereas the dominant nitrate reduction for assimilation and denitrification processes mediates by NapA-type nitrate reductase. Nitric oxide reductase homologues FlRd and FlRd-red provide an adequate capacity for NO detoxification. The recombinant hydroxylamine reductase showed high activity in hydroxylamine to generate ammonium, which might contribute to detoxification mechanism in nitrogen cycling. Overall, this study firstly provides valuable insights into the genes expression and enzyme action, which helps understanding the mechanism of SND processes.

Highly efficient nitrogen removal of a coldness-resistant and low nutrient needed bacterium, Janthinobacterium sp. M-11

A novel heterotrophic nitrification-aerobic denitrification bacterium, identified as Janthinobacterium sp. M-11, was isolated from the Songhua River. When the initial ammonium concentration was 5 mg·L−1, 98% of ammonium was removed under cold condition (2 °C) with the C/N ratio of 5 at initial pH 7 and aerobic condition, which demonstrated the significant ammonium removal capacity of M-11 with low nutrient consumption at cold temperature. Denitrification processes under aerobic and anaerobic conditions were also investigated. 89% of nitrite and 89% of nitrate were removed under aerobic condition. Under anaerobic condition, 93% of nitrite and 98% of nitrate were removed. Interestingly, a high amount of nitrite accumulation was observed in the mid-stage of anaerobic denitrification for nitrate. This special phenomenon was probably because of the existence of narG gene amplified in the strain M-11, which would encode membrane-bound nitrate reductase and accelerate the nitrate conversion rate of M-11 under anaerobic condition.

The potential multiple mechanisms and microbial communities in simultaneous nitrification and denitrification process treating high carbon and nitrogen concentration saline wastewater

A simultaneous nitrification and denitrification (SND) process in sequencing batch biofilm reactor (SBBR) was established to treat high carbon and nitrogen saline wastewater in this study. Acetate, glucose and an organic mixture were applied as organic sources in three SBBRs, achieving average total nitrogen removal efficiency of 97.15%, 63.94% and 94.99% during 120days’ operation, respectively. The underlying nitrogen removal mechanisms were investigated by 16S rRNA sequencing and batch tests. Results showed different carbon sources had great impact on microbial communities, and led to different nitrogen removal mechanism. Autotrophic and heterotrophic nitrification together contributed to the well performance of nitrification process. And denitrification was carried out by a combined anoxic and aerobic denitrificans. Furthermore, the SND process was mainly via nitrite not nitrate. Compared with ammonia-oxidizing bacteria, ammonia-oxidizing archaea with a much higher abundance contributed more to autotrophic nitrification. Pseudomonas_stutzeri and Bacillus_cereus were the predominant detected heterotrophic nitrification-aerobic denitrification bacterium.

Microbial Community Structure and Diversity in Cellar Water by 16S rRNA High-throughput Sequencing

This study was aimed to explore the bacterial diversity of cellar water as well as to study the relationship between the bacterial diversity and environmental factors. The MiSeq high-throughput sequencing was used to analyze and compare the bacterial diversity and community composition of samples from different cellar water samples. Overall 1605 optimized reads were obtained from four samples based on high-throughput sequencing of the V4 region of the 16S rRNA gene. Bacterial species detected in these samples covered 22 phyla,42 classes,71 orders,115 families, 146 genera. Analysis showed that the bacterial diversity was very high in these samples, and there were differences among different samples. The distribution characteristics of the dominant bacteria showed patterns of a large number of rare species and a few common types. Taxonomic assignment analysis indicated that Bacteroidetes,Proteobacteria,Actinobacteria,Verrucomicrobia,OD1 dominated in the Cellar water, and accounted for 87.1% to 94.8% at phylum level. The predominant groups were Actinobacteria,Acidimicrobiia,Cytophagia, Flavobacteriia, Sphingobacteriia,α-Proteobacteria,β-Proteobacteria,γ-Proteobacteria,Opitutae, Verrucomicrobiae,Pedosphaerae and ZB2 at class level. At genus level Rhodobacter,Dechloromonas,Flavobacterium,Acinetobacter,Comamonas,Pseudomonas,Hydrogenophaga,et al were the abundant taxa, which were mainly denitrifying bacteria and heterotrophic nitrification-aerobic denitrification bacteria. The result of RDA suggested that the influences of different environmental factors on different microbes were different. Bacterial community Ⅱ had significant positive correlation with UV254,permanganate index,BOD5,and Bacterial community Ⅲ had significant positive correlation with TN,NO2--N,NO3--N,TP,NH4+-N. This research should deepen the understanding on microbial community in Cellar water, and provide references for the association of bacterial composition and diversity with environmental factors.

Characteristics and metabolic pathway of Alcaligenes sp. TB for simultaneous heterotrophic nitrification-aerobic denitrification.

A novel heterotrophic nitrification-aerobic denitrification bacterium, Alcaligenes sp. TB (GenBank accession no. JQ044686), was isolated from a rotating drum biofilter for NO removal. Its characteristics and metabolic pathway for NO removal were comprehensively investigated. Experimental results showed that the nitrification and denitrification efficiency reached 99.42 and 96.44%, respectively, after 44h under the conditions of pH 7.2, 30°C, and 120rpm. The tests with the addition of Pb2+ and Na2WO4 as the reductase inhibitor revealed that nitrite was the key intermediate to produce the nitrogen gas as the final product in the simultaneous heterotrophic nitrification and denitrification by strain TB. Based on the experimental results, the metabolic pathway of strain TB has been proposed that it carries out shortcut/complete simultaneous nitrification and denitrification with nitrite as an intermediate and nitrogen gas as a final product. The two potential metabolic pathways existing in strain TB can be described as NH4+→NH2OH→NO2-→N2O→N2 and NH4+→NH2OH→NO2-→NO3-→NO2-→N2O→N2. This work indicates that the strain TB may be a good candidate for the denitrification of the sewage.

Ammonium removal by Agrobacterium sp. LAD9 capable of heterotrophic nitrification–aerobic denitrification

Characteristics of ammonium removal by a newly isolated heterotrophic nitrification-aerobic denitrification bacterium Agrobacterium sp. LAD9 were systematically investigated. Succinate and acetate were found to be the most favorable carbon sources for LAD9. Response surface methodology (RSM) analysis demonstrated that maximum removal of ammonium occurred under the conditions with an initial pH of 8.46, C/N ratio of 8.28, temperature of 27.9°C and shaking speed of 150rpm, where temperature and shaking speed produced the largest effect. Further nitrogen balance analysis revealed that 50.1% of nitrogen was removed as gas products and 40.8% was converted to the biomass. Moreover, the occurrence of aerobic denitrification was evidenced by the utilization of nitrite and nitrate as nitrogen sources, and the successful amplifications of membrane bound nitrate reductase and cytochrome cd(1) nitrite reductase genes from strain LAD9. Thus, the nitrogen removal in strain LAD9 was speculated to comply with the mechanism of heterotrophic nitrification coupled with aerobic denitrification (NH(4)(+)-NH(2)OH-NO(2)(-)-N(2)O-N(2)), in which also accompanied with the mutual transformation of nitrite and nitrate. The findings can help in applying appropriate controls over operational parameters in systems involving the use of this kind of strain.

The characteristics of a novel heterotrophic nitrification–aerobic denitrification bacterium, Bacillus methylotrophicus strain L7

Bacillus methylotrophicus strain L7, exhibited efficient heterotrophic nitrification–aerobic denitrification ability, with maximum NH4+-N and NO2--N removal rate of 51.58mg/L/d and 5.81mg/L/d, respectively. Strain L7 showed different gaseous emitting patterns from those strains ever described. When 15NH4Cl, or Na15NO2, or K15NO3 was used, results of GC–MS indicated that N2O was emitted as the intermediate of heterotrophic nitrification or aerobic denitrification, while GC–IRMS results showed that N2 was produced as end product when nitrite was used. Single factor experiments suggested that the optimal conditions for heterotrophic nitrification were sodium succinate as carbon source, C/N 6, pH 7–8, 0g/L NaCl, 37°C and a wide range of NH4+-N from 80 to 1000mg/L. Orthogonal tests showed that the optimal conditions for aerobic denitrification were C/N 20, pH 7–8, 10g/L NaCl and DO 4.82mg/L (shaking speed 50r/min) when nitrite was served as substrate.