Assisted Selection of Biomarkers by Linear Discriminant Analysis Effect Size (LEfSe) in Microbiome Data

There is growing attention toward closed biological genomes in the environment and in health. To explore and reveal the intergroup differences among ...

North China Electric Power University

The discovery of complete ammonia oxidizing bacteria (CAOB) has fundamentally overturned the traditional recognition of nitrification. However, little was known about the transcriptional activity and diversity of the newly recognized ammonia oxidizing prokaryote in engineered ecosystems. To fill this gap, transcriptional investigations of CAOB amoA genes were carried out comparatively with the canonical ammonia oxidizing bacteria (AOB) and archaea (AOA) in eight full-scale wastewater treatment plants (WWTPs). Remarkably, qPCR results revealed the transcriptional levels of CAOB amoA gene were unexpectedly high in most of samples with the highest 24-fold that of AOB amoA, suggesting CAOB were actively participating in ammonia oxidation while they were previously overlooked. This result also well explained the confusing high abundances of genus Nitrospira which were frequently detected in WWTPs. Furthermore, phylogenetic analysis based on high throughput sequencing indicated the CAOB amoA gene sequences formed three well-supported clusters and Nitrospira nitrosa cluster accounted for 97% of all the retrieved sequences, which was supposed to be the dominant taxon of CAOB in the ammonia-intensive environment due to niche partitioning. This study highlighted the significance of including the newly discovered ammonia oxidizing bacterial member when assessing the nitrification process and ecological function in the future.

Transcriptional activity and diversity of comammox bacteria as a previously overlooked ammonia oxidizing prokaryote in full-scale wastewater treatment plants.

With the extensive application of graphene oxide (GO), its leakage and release into wastewater treatment plants become inevitable. However, the toxicity of graphene oxide (GO) on nitrification process and the underlying mechanisms still remain unclear. In this study, the toxic effects of GO at concentration of 10 and 100 mg/L in 4 h and 10 days were evaluated with sealed reactors operated in sequencing batch mode. In the initial 4 h, both GO concentrations showed no negative effect on nitrogen conversion. However, the exposure to 100 mg/L GO significantly weakened the NH+ 4-N and NO- 2-N conversion capabilities and intensified the nitrous oxide (NO) generation after 10 days. Extracellular polymeric substance (EPS) analysis suggested that 100 mg/L GO decreased the protein content of the nitrifying activated sludge. Moreover, reactive oxygen species (ROS) level was promoted by 100 mg/L GO owing to the impaired endogenous antioxidant enzymes including superoxide dismutase (SOD) and catalase (CAT), which caused oxidative stress to bacteria. Finally, quantitative PCR results confirmed that nitrite-oxidizing bacteria (NOB) and complete ammonia oxidizing bacteria (CAOB) were more sensitive to GO, which was the primary cause for the significant promotion of NO generation in the high GO concentration. This study offered new insights in the toxicity of GO on nitrification and NO generation in the terms of dose and exposure time.

The effects of graphene oxide on nitrification and NO emission: Dose and exposure time dependent.

Complete ammonia oxidizing bacteria (CAOB) have been recognized as a new member of ammonia-oxidizing microorganisms (AOMs) due to its single-step nitrification capability. However, the abundance and diversity of CAOB in environmental ecosystems were still far from known owing to the lack of specific molecular marker. Herein, a universal primer set specifically targeting both clades of CAOB amoA gene with high specificity and coverage was successfully designed. Intriguingly, real-time quantitative PCR tests revealed that CAOB were ubiquitous and unexpectedly abundant in agricultural soils, river sediments, intertidal zones, drinking water and wastewater treatment systems. Phylogenetic analysis indicated that clade A existed in all the five types of ecosystems, while clade B were only detected in soil and sediment samples. Four sub-clusters were further classified within clade A, in which N. nitrosa cluster dominated CAOB amoA in activated sludge samples while the new recognized soil cluster was the primary constitute in soils. Moreover, the niche specialization between different CAOB species and the environmental conditions were supposed to be the primary driven force to shape the diversity and community of CAOB. This study provided a strong evidence in support of the ubiquities and high abundances of CAOB in various environmental ecosystems and highlighted the significance of including CAOB as the new member of AOMs to re-evaluate the biogeochemical nitrogen cycle.

Abundance and community composition of comammox bacteria in different ecosystems by a universal primer set.

Nitrous oxide (NO) emission from wastewater treatment plants (WWTPs) has become a focus of attention due to its significant greenhouse effect. In this study, the role of sludge retention time (SRT) in mitigation of NO emission from a pilot-scale oxidation ditch was systematically investigated. The activated sludge system that operated at SRT of 25 days demonstrated significantly lower NO emission factor, higher resistance to ammonia overload and aeration failure shock than those obtained at SRT of 15 days no matter which hydraulic retention time (HRT) was adopted. Batch experiments revealed that nitrifier denitrification (ND) was the primary mechanism of NO generation. However, more microbes affiliated with Nitrospira genera were harbored in the system at SRT 25 d, which could effectively avoid nitrite accumulation, a key factor promoting NO generation by ND. PICRUSt results further suggested the system at SRT 25 d possessed higher genetic potential for NO reduction reflected by the more abundant nitrous-oxide reductase.

Role of sludge retention time in mitigation of nitrous oxide emission from a pilot-scale oxidation ditch.

Nitrous oxide (NO) generated from wastewater treatment plants (WWTPs) has drawn attention due to its high emission load and significant greenhouse effect. In the present study, NO emissions from a pilot-scale Carrousel oxidation ditch under various chemical oxygen demand (COD) to nitrogen ratio (COD/N) and aeration rates were systematically investigated. The highest NO emission factor was 0.142 ± 0.013%, at COD/N of 5 and aeration rate of 1.8 m h, which was much lower than the majority of previous studies. The results could be attributed to the high internal recycle ratio of the oxidation ditch process which lightened the burden of influent load to the system. The profiles of NO emissions and dissolved NO concentration along the channels showed a distinct spatial variation that NO emissions primarily occurred in the aeration zones due to the air stripping effect. However, both the aeration and anoxic zones contributed to NO generation due to autotrophic nitrification (AN), which was considered to be the main NO generation process. In addition, two simulated shock-load conditions, ammonia overload shock and aeration failure shock, were carried out to explore the response of the biological nitrogen removal (BNR) system. The results indicated that both shock-loads lead to excessive NO emissions, especially at higher aeration rates, which could be explained by the improved NO generation by AN process during the shock-load period. This study offered new insights into the role of operational parameters to NO emission and the alternative approach for NO mitigation during both the steady-state operation and shock-load conditions in the oxidation ditch process.

Nitrous oxide (NO) emissions from a pilot-scale oxidation ditch under different COD/N ratios, aeration rates and two shock-load conditions.

Ammonia-oxidizing bacteria (AOB) and archaea (AOA) are two microbial groups mediating nitrification, yet little is presently known about their abundances and community structures at the transcriptional level in wastewater treatment systems (WWTSs). This is a significant issue, as the numerical abundance of AOA or AOB at the gene level may not necessarily represent their functional role in ammonia oxidation. Using amoA genes as molecular markers, this study investigated the transcriptional abundance and community structure of active AOA and AOB in 14 WWTSs. Quantitative PCR results indicated that the transcriptional abundances of AOB amoA (averaged: 1.6 × 10 copies g dry sludge) were higher than those of AOA (averaged: 3.4 × 10 copies g dry sludge) in all WWTSs despite several higher abundances of AOA amoA at the gene level. Moreover, phylogenetic analysis demonstrated that Nitrosomonas europaea and unknown clusters accounted for 37.66% and 49.96% of the total AOB amoA transcripts, respectively, suggesting their dominant role in driving ammonia oxidation. Meanwhile, AOA amoA transcripts were only successfully retrieved from 3 samples, and the Nitrosospaera sister cluster dominated, accounting for 83.46%. Finally, the substrate utilization kinetics of different AOA and AOB species might play a fundamental role in shaping their niche differentiation, community composition, and functional activity. This study provides a basis for evaluating the relative contributions of ammonia-oxidizing microorganisms (AOMs) to nitrogen conversions in WWTSs.

Shishi He

Key Laboratory of Regional Energy Systems Optimization,
Ministry of Education,
College of Environmental Science and Engineering,
Key Laboratory of Regional Energy Systems Optimization, Ministry of Education, College of Environmental Science and Engineering

North China Electric Power University

Assisted Selection of Biomarkers by Linear Discriminant Analysis Effect Size (LEfSe) in Microbiome Data