Integration of anaerobic digestion and electrodialysis for methane yield promotion and in-situ ammonium recovery.

Ammonia inhibition is a common issue encountered in anaerobic digestion (AD) when treating nitrogen-rich substrates. This study proposed a novel approach, the electrodialysis-integrated AD (ADED) system, for in-situ recovery of ammonium (NH4+) while simultaneously enhancing AD performance. The ADED reactor was operated at two different NH4+-N concentrations (5,000 mg/L and 10,000 mg/L) to evaluate its performance against a conventional AD reactor. The results indicate that the ADED technology effectively reduced the NH4+-N concentration to below 2,000 mg/L, achieving this with a competitive energy consumption. Moreover, the ADED reactor demonstrated a 1.43-fold improvement in methane production when the influent NH4+-N was 5,000 mg/L, and it effectively prevented complete inhibition of methane production at the influent NH4+-N of 10,000 mg/L. The life cycle impact assessment reveals that ADED technology offers a more environmentally friendly alternative by recovering valuable fertilizer from the AD system.

A novel entropy-driven dual-output mode integrated with DNAzyme for enhanced microRNA detection.

Accurate microRNA (miRNA) detection is pivotal in the diagnosis and monitoring of cancer. Entropy-driven catalysis (EDC) has attracted widespread attention as an enzyme-free, isothermal technique for miRNA detection owing to its inherent simplicity and reliability. However, conventional EDC is a single-output mode, limiting the efficiency of signal amplification. In this study, a novel EDC dual-output mode was employed in conjunction with DNAzyme, resulting in the development of an EDC dual-end DNAzyme (EDC-DED) approach for highly sensitive miRNA detection. In this system, miRNA-21 initiated the EDC reaction, producing a large amount of catalytically active dual-end Mg2+-dependent DNAzyme. The DNAzyme further cleaved the reporter cyclically, generating a notably amplified fluorescence signal. The proposed method achieved a low detection limit of 2 pM. Compared with the traditional EDC single-end DNAzyme (EDC-SED) strategy, the present method exhibited superior amplification efficiency, enhancing detection sensitivity by approximately 46.5-fold. Furthermore, this platform demonstrated ideal specificity, satisfactory reproducibility and acceptable detection capabilities in clinical serum samples. Therefore, the straightforward and convenient strategy is a potential tool for miRNA analysis, which may provide a new perspective for biological analysis and clinical application.

Building a stable and robust anti-interference DNA dissipation system by eliminating the accumulation of systemic specified errors.

BACKGROUND: The emergence of DNA nanotechnology has enabled the systematic design of diverse bionic dissipative behaviors under the precise control of nucleic acid nanodevices. Nevertheless, when compared to the dissipation observed in robust living systems, it is highly desirable to enhance the anti-interference for artificial DNA dissipation to withstand perturbations and facilitate repairs within the complex biological environments. RESULTS: In this study, we introduce strategically designed "trash cans" to facilitate kinetic control over interferences, transforming the stochastic binding of individual components within a homogeneous solution into a competitive binding process. This approach effectively eliminates incorrect binding and the accumulation of systemic interferences while ensuring a consistent pattern of energy fluctuation from response to silence. Remarkably, even in the presence of numerous interferences differing by only one base, we successfully achieve complete system reset through multiple cycles, effectively restoring the energy level to a minimum. SIGNIFICANCE: The system was able to operate stably without any adverse effect under conditions of irregular interference, high-abundance interference, and even multiplex interferences including DNA and RNA crosstalk. This work not only provides an effective paradigm for constructing robust DNA dissipation systems but also greatly broadens the potential of DNA dissipation for applications in high-precision molecular recognition and complex biological reaction networks.

An improved technology for monitoring groundwater flow velocity and direction in fractured rock system based on colloidal particles motion.

The colloidal borescope, using colloidal particle motion, is used to monitor the flow velocities and directions of groundwater. It integrates advanced techniques such as microscopy, high-speed photography, and big data computing and enjoys high sensitivity at the micron level. However, In the same well, the groundwater flow velocity monitored by colloidal hole mirror is varies greatly from that obtained by conventional hydrogeological monitoring, such as pumping test. In order to solve this problem, the stability catcher and stratified packer are designed to control the interference of the vertical flow in drilling, and to monitor the flow velocity and direction of groundwater velocity at the target aquifer and target fracture. Five wells with different aquifers and different groundwater types were selected for monitoring in south-central China. The instantaneous velocity and direction are converted into east-west component and north-south component, the average velocity and direction is calculated according to the time of 10 min, and the particle trajectory diagram is established. Based on these results, it proposed a concept of cumulative flow velocity. Using curve-fitting equations, the limits of cumulative flow velocities as the monitoring time tends to infinity were then calculated as the actual flow velocities of the groundwater. The permeability coefficient of aquifer is calculated by using the fissure ratio of aquifer, hydraulic slope and flow velocity, and compared with the permeability coefficient obtained by pumping test. The results are as follows: (1) The variation coefficient of the instantaneous flow velocity measured at the same depth in the same well at different times is greater than that of the time average flow velocity and greater than that of the cumulative flow velocity. The variation coefficient of the actual velocity is the smallest, indicating that the risk of using the actual flow velocity is lower. (2) The variation coefficient of the flow rate monitored at different depths in the same well is mainly controlled by the properties of the aquifer. The more uniform water storage space in the aquifer, the smaller the variation coefficient. (3) The comparison between the permeability coefficient obtained by monitoring and the permeability coefficient obtained by pumping test shows that the flow of structural fissure water controlled by planar fissure is more surface flow, and the results are consistent. When the groundwater flow is controlled by pores and solution gaps, the flow channel is complicated, which is easy to produce turbulent flow, and the result consistency is poor. (4) According to different research accuracy requirements, different monitoring and calculation methods can be selected for different aquifers and groundwater types. Researches show that, the permeability coefficient calculated for the actual flow velocity in well DR01 is the same as that calculated for the pumping test. The aquifer characteristics reflected by the coefficient of variation of the actual flow velocity in the same aquifer are more realistic. The pumping test method obtains the comprehensive parameters of a certain aquifer, and this method can be used to monitor a certain fissure. In this paper, the new technology developed for monitoring, and the new algorithm established for data processing, can accurately obtain the flow velocity and direction of groundwater, using capsule hole mirror monitoring method. The key parameters of hydrogeology can be obtained by using one well, which can reduce the time and cost input and improve the work efficiency.

New insights into syntrophic ethanol oxidation: Effects of operational modes and solids retention times.

Anaerobic ethanol oxidation relies on syntrophic interactions among functional microorganisms to become thermodynamically feasible. Different operational modes (sequencing batch reactors, SBRs, and continuous flow reactors, CFRs) and solids retention times (SRT, 25 days and 10 days) were employed in four ethanol-fed reactors, named as SBR25d, SBR10d, CFR25d, and CFR10d, respectively. System performance, syntrophic relationships, microbial communities, and metabolic pathways were examined. During the long-term operation, 2002.7 ± 56.0 mg COD/L acetate was accumulated in CFR10d due to the washout of acetotrophic methanogens. Microorganisms with high half-saturation constants were enriched in reactors of 25-day SRT. Moreover, ethanol oxidizing bacteria and acetotrophic methanogens with high half-saturation constants could be acclimated in SBRs. In SBRs, Syner-01 and Methanothrix dominated, and the low SRT of 10 days increased the relative abundance of Geobacter to 38.0%. In CFRs, the low SRT of 10 days resulted in an increase of Desulfovibrio among syntrophic bacteria, and CFR10d could be employed in enriching hydrogenotrophic methanogens like Methanobrevibacter.

Microplastic pollution in the groundwater under a bedrock island in the South China sea.

Groundwater is the only freshwater resource on islands. Research on microplastic pollution in groundwater on islands is scarce. This study is the first to explore microplastic pollution in the groundwater under a bedrock island (Dawanshan Island) located in the South China Sea. The influence of hydrogeological factors on the distribution, source, and ageing features of microplastics in the groundwater were investigated. Despite the small scale of industrial and agricultural activities on the island, the amount of microplastics in the groundwater ranged from 34 to 64 particles/L, with over 80% of the microplastics being polyester fibres with diameters smaller than 2 mm, which is comparable to those in coastal cities. These microplastics were originated from inland plastic usage, rather than from the surrounding sea, which was confirmed by the lack of seawater intrusion on the island. Owing to the low permeability of granite, microplastics were mainly distributed in the water of the loose layer of porous sediment, and their quantity decreased with depth. In addition, the abundance of microplastics in pore groundwater increased with an increase in the velocity of groundwater flow. The severity of microplastic pollution in the groundwater increased with an increase and decrease in the content of total dissolved solids and dissolved oxygen, respectively. The microplastics originated from plastic waste disposed of on the island, rather than from seawater intrusion. Also, through groundwater infiltration into exposed soil at recharge areas, artificial wells at residential areas, and water exchange with surface water at valley areas. Microplastics buried in the groundwater aged faster along the migration path of the groundwater. These microplastics threaten the safety of people and plants on the island through exposure resulting from the extraction of groundwater for irrigation, while they endanger marine life through submarine groundwater discharge.

Multifunctional dumbbell probes-based logic circuits: microRNAs logic detection and tumor cells identification.

BACKGROUND: The powerful logic processing capability of DNA logic circuits over multiple input signals perfectly meets the demands of multi-biomarker-based clinical diagnostics. As important biomarkers for cancer diagnosis and treatment, the orthogonal differential expression of microRNAs (miRNAs) in different diseases and different cancer cells makes the precise logical detection of multiple miRNAs particularly critical. RESULTS: Therefore, we constructed two fundamental "AND" and "OR" logic gates and one "AND-OR" logic gate on the basis of our proposed multifunctional dumbbell probes. These logic gates allowed for the logical profiling of multiple cancer-associated miRNAs. In addition, by making simple adjustments to the functional modules of multifunctional dumbbell probes, the three logic gates we proposed could be easily transformed without the use of sophisticated probe design. Remarkably, these logic gates, in particular the "AND-OR" logic gate, were able to compute several miRNAs simultaneously, demonstrating excellent cell identification capabilities. SIGNIFICANCE: Overall, this work provided a new idea for accurately distinguishing multiple cell types and showed great application prospects.

Unmodificated stepless regulation of CRISPR/Cas12a multi-performance.

As CRISPR technology is promoted to more fine-divided molecular biology applications, its inherent performance finds it increasingly difficult to cope with diverse needs in these different fields, and how to more accurately control the performance has become a key issue to develop CRISPR technology to a new stage. Herein, we propose a CRISPR/Cas12a regulation strategy based on the powerful programmability of nucleic acid nanotechnology. Unlike previous difficult and rigid regulation of core components Cas nuclease and crRNA, only a simple switch of different external RNA accessories is required to change the reaction kinetics or thermodynamics, thereby finely and almost steplessly regulating multi-performance of CRISPR/Cas12a including activity, speed, specificity, compatibility, programmability and sensitivity. In particular, the significantly improved specificity is expected to mark advance the accuracy of molecular detection and the safety of gene editing. In addition, this strategy was applied to regulate the delayed activation of Cas12a, overcoming the compatibility problem of the one-pot assay without any physical separation or external stimulation, and demonstrating great potential for fine-grained control of CRISPR. This simple but powerful CRISPR regulation strategy without any component modification has pioneering flexibility and versatility, and will unlock the potential for deeper applications of CRISPR technology in many finely divided fields.

Universal probe-based SNP genotyping with visual readout: a robust and versatile method.

Detection of single nucleotide polymorphisms (SNPs) is critical for personalized clinical diagnosis, treatment, and medication. Current clinical detection methods suffer from primer dimerization and require the redesigning of reaction systems for different targets, resulting in a time-consuming and laborious process. Here, we present a robust and versatile method for SNP typing by using tailed primers and universal small molecule probes in combination with a visualized lateral flow assay (LFA). This approach enables not only rapid typing of different targets, but also eliminates the interference of primer dimers and enhances the accuracy and reliability of the results. Our proposed universal assay has been successfully applied to the typing of four SNP loci of clinical samples to verify the accuracy and universality, and the results are consistent with those obtained by Sanger sequencing. Therefore, our study establishes a new universal "typing formula" using nucleic acid tags and small molecule probes that provides a powerful genotyping platform for genetic analysis and molecular diagnostics.

Detection of rare CTCs by electrochemical biosensor built on quaternary PdPtCuRu nanospheres with mesoporous architectures.

Circulating tumor cells (CTCs) are promising liquid biopsy biomarkers for early cancer detection and anti-cancer therapy evaluation. The ultra-low abundance of CTCs in blood samples requires highly sensitive and accurate detection ways. In this study, we propose the design of a dual-recognition electrochemical biosensor to improve both the specificity and signal response. PdPtCuRu mesoporous nanospheres (PdPtCuRu MNSs) with excellent three dimensions (3D) nanopore structures were synthesized by one-pot method and connected to mucin 1 (MUC1) aptamer to serve as signal amplification probe. Besides, superconductive carbon black, Ketjen Black (KB), and gold nanoparticles (AuNPs) modified organometallic frame (CeMOF-Au) were combined to work as signal transducer. The characteristic branching structure of KB provides abundant contact points to load CeMOF-Au to heighten the interface electron transfer rate. In addition, AuNPs were reduced on the surface of CeMOF, which could effectively bind the capture antibody and further enhance the conductivity. Under the optimized condition, the limit of detection (LOD) of the as-constructed biosensor was less than 10 cells mL-1 for model A549 cells, and showed good specificity and accuracy in spiked serum samples. We envision the as-proposed electrochemical biosensor would alternate as a useful tool for the clinical detection of CTCs for cancer diagnosis.

Distinguishing responses of acetoclastic and hydrogenotrophic methanogens to ammonia stress in mesophilic mixed cultures.

A shift from the acetoclastic to the hydrogenotrophic pathway in methanogenesis under ammonia inhibition is a common observation in anaerobic digestion. However, there are still considerable knowledge gaps concerning the differential ammonia tolerance of acetoclastic and hydrogenotrophic methanogens (AMs and HMs), their responses to different ammonia species (NH4+, NH3), and their recoverability after ammonia inhibition. With the successful enrichment of mesophilic AMs and HMs cultures, this study aimed at addressing the above knowledge gaps through batch inhibition/recovery tests and kinetic modeling under varying total ammonia (TAN, 0.2-10 g N/L) and pH (7.0-8.5) conditions. The results showed that the tolerance level of HMs to free ammonia (FAN, IC50=1345 mg N/L) and NH4+ (IC50=6050 mg N/L) was nearly 11 times and 3 times those of AMs (NH3, IC50=123 mg N/L; NH4+, IC50=2133 mg N/L), respectively. Consistent with general belief, the AMs were more impacted by FAN. However, the HMs were more adversely affected by NH4+ when the pH was ≤8.0. A low TAN (1.0-4.0 g N/L) could cause irreversible inhibition of the AMs due to significant cell death, whereas the activity of HMs could be fully or even over recovered from severe ammonia stress (FAN≤ 0.9 g N/L or TAN≤10 g N/L; pH ≤8.0). The different tolerance responses of AMs and HMs might be associated with the cell morphology, multiple energy-converting systems, and Gibbs free energy from substrate-level phosphorylation.

Role of iron(II) sulfide in autotrophic denitrification under tetracycline stress: Substrate and detoxification effect.

Autotrophic denitrification using inorganic compounds as electron donors has gained increasing attention in the field of wastewater treatment due to its numerous advantages, such as no need for exogenous organic carbon, low energy input, and low sludge production. Tetracycline (TC), a refractory contaminant, is often found coexisting with nutrients (NO3- and PO43-) in wastewater, which can negatively affect the biological nutrient removal process because of its biological toxicity. However, the performance of autotrophic denitrification under TC stress has rarely been reported. In this study, the effects of TC on autotrophic denitrification with thiosulfate (Na2S2O3) and iron (II) sulfide (FeS) as the electron donors were investigated. With Na2S2O3 as the electron donor, TC slowed down the nitrate removal rate, which decreased from 1.32 to 0.18 d-1, when TC concentration increased from 0 mg/L to 50 mg/L. When TC concentration was higher than 2 mg/L, nitrite reduction was seriously inhibited, leading to nitrite accumulation. With FeS as the electron donor, nitrate removal was much more efficient under TC-stressed conditions, and no distinct nitrite accumulation was observed when the initial TC concentration was as high as 10 mg/L, indicating the effective detoxification of FeS. The detoxification effects in the FeS autotrophic denitrification system mainly resulted from the rapid adsorption of TC by FeS and effective degradation of TC, as proven by a relatively higher living biomass area. This study offers new insights into the response of sulfur-based autotrophic denitrifiers to TC stress and demonstrates that the FeS-based autotrophic denitrification process is a promising technology for the treatment of wastewater containing emerging contaminants and nutrients.

OGT as potential novel target: Structure, function and inhibitors.

O-linked N-acetylglucosamine transferase (OGT), a key protein in O-linked N-acetylglucosamine (O-GlcNAc) post-translational modification, catalyzes O-GlcNAcylation by linking GlcNAc from glycosyl donors to protein Ser/Thr residues to regulate multiple biological processes. OGT has become a popular research topic in the field of glycobiology as a potential target. This report consists of three parts, namely: the basic information of OGT, the relationship between OGT and diseases, and the research progress of OGT inhibitors. This report aims to improve the understanding of OGT-related fields from the perspective of its structure. OGT inhibitors could aid the effective and efficient investigation of the O-GlcNAcylation mechanism, and provide new ideas and methods for the treatment of related diseases.

Stimulatory effects of biochar addition on dry anaerobic co-digestion of pig manure and food waste under mesophilic conditions.

The stimulatory effect of biochar addition on dry anaerobic digestion (AD) has been rarely investigated. In this study, the effects of commonly used biochars (bamboo, rice husk, and pecan shell) on dry co-AD were investigated using mesophilic batch digesters fed with pig manure and food waste as substrates. The results show that the specific methane yield was mildly elevated with the addition of biochars by 7.9%, 9.4%, and 12.0% for bamboo, rice husk, and pecan shell-derived biochar additions, respectively. Biochar did facilitate the degradation of poorly biodegradable organics. In comparison, there was no significant effect on the peak methane production rate by the supplementation of the selected biochars. Among the three mechanisms of enhancing methanogenesis by biochar (buffering, providing supporting surface, and enhancing electron transfer), the first two mechanisms did not function significantly in dry co-AD, while the third mechanism (i.e., enhancing electron transfer) might play an important part in dry AD process. It is recommended that the utilization of biochar for the enhancement of biomethanation in dry AD should be more focused on mono digestion in future studies.

Improved reduction of antibiotic resistance genes and mobile genetic elements from biowastes in dry anaerobic co-digestion.

This study investigated the performance of anaerobic co-digestion (AcoD) of pig manure and food waste on the reduction of antibiotic resistomes under wet and dry AcoD conditions. High-throughput quantitative PCR technology was utilized for a comprehensive assessment of the performances of the two processes. The results show that dry AcoD with a total solids (TS) content of 20% effectively reduced total antibiotic resistance genes (ARGs) by 1.24 log copies/g wet sample, while only 0.54 log copies/g wet sample was reduced in wet AcoD with a TS content of 5%. Dry AcoD was more efficient in reduction of aminoglycosides, multidrug and sulfonamide resistance genes compared with the reduction of other classes of ARGs. Dry AcoD caused a significant reduction of ARGs with resistance mechanisms of efflux pump and antibiotic deactivation. In contrast, there was no obvious difference in reductions of ARGs with different resistance mechanisms in wet AcoD. Network analysis showed that ARGs were significantly correlated with mobile genetic elements (MGEs) (Spearman's r > 0.8, P < 0.05), as well as microbial communities. Enrichment of ARGs and MGEs was found at the early period of AcoD processes, indicating some ARGs and MGEs increased during the hydrolysis and acidogenesis stages. But after a long retention time, their abundances were effectively reduced by dry AcoD in the subsequent stages.

Filamentous microalgae as an advantageous co-substrate for enhanced methane production and digestate dewaterability in anaerobic co-digestion of pig manure.

This study aimed at exploring filamentous microalgae (Tribonema sp.) as an advantageous co-substrate for anaerobic digestion (AD) of pig manure. Its impacts on the AD performance were assessed in terms of methane yield, energy conversion efficiency, digestion kinetics, and digestate dewaterability. The microalgae substantially improved methane yield, AD kinetics, and digestate dewaterability of the AD process. The enhancement in methane yield ranged from 2 to 27.4%, with the maximum enhancement (corresponding to an energy conversion efficiency of 81%) occurring at a mixing ratio of 1:1 (VS basis). The AD kinetics was improved as indicated by the increased hydrolysis rate constants and diminished lag time. The specific resistance to filtration (SRF) of the digestate decreased significantly with the increasing proportion of the microalgae in the co-substrates, which would facilitate digestate processing and valorisation. Subsequently, the high biomass productivity of the microalgae (441 mg/L/d) in liquid digestate would enable sustainable bioenergy production through nutrient recycling.

Impact of total solids content on anaerobic co-digestion of pig manure and food waste: Insights into shifting of the methanogenic pathway.

Dry anaerobic digestion (AD) has advantages over wet AD in treating high-solid organic wastes like livestock and food wastes, but an elevated total solids (TS) content would affect the AD performances. In this study, methane production of digesters co-digesting pig manure (PM) and food waste (FW) at different TS contents (R1, TS 5%; R2, TS 10%; R3, TS 15%; and R4, TS 20%) was assessed. The results showed the specific methane yield had no significant difference with the increase of TS contents from 5% to 15% (278.8-291.7 NmL/g VSadded), while it was reduced at a 20% TS content (259.8 NmL/g VSadded). Two peaks of total volatile fatty acids and daily methane production were observed in the high-solid digesters (R2-R4), while only one peak occurred in wet AD (R1). A new kinetics model was developed to describe the two-peak methane production behavior at high TS contents. The analysis on the microbial community structure clearly showed the different evolutions of methanogenic pathways in low and high solids content systems. In dry AD (R4), there was a general shifting from the acetoclastic pathway, to mixotrophic pathway and hydrogenotrophic pathway, with the dominance of mixotrophic and hydrogenotrophic methanogens.

Design, recombinant expression, and antibacterial activity of a novel hybrid magainin-thanatin antimicrobial peptide.

Antimicrobial peptides are small molecule polypeptides with biological activity, which can avoid the drug resistance. Magainin and thanatin are antimicrobial peptides with a broad spectrum of inhibitory microbes, and the core sequence of magainin is linked to a core sequence of thanatin. Here, the hybrid magainin-thanatin (MT) antimicrobial peptide was designed through bioinformatics analysis. The recombinant MT antimicrobial peptide was successfully expressed and purified in Escherichia coli BL21 (DE3). The molecular weight of the hybrid MT antimicrobial peptide was about 3.35 kDa. Moreover, the target protein indeed has an inhibitory effect on Staphylococcus aureus, E. coli DH5α, and Bacillus subtilis, with the minimum inhibitory concentrations 16.5, 20, and 9 µM, respectively. The rational designed hybrid MT antimicrobial peptide will hopefully provide large-scale fermentable antimicrobial peptides in the industrial production in the future.

Impacts of environmental factors on microbial diversity, distribution patterns and syntrophic correlation in anaerobic processes.

Anaerobic processes are widely used for treating high-strength organic wastewater. Understanding the ecological patterns of the microorganisms involved and the effect of environmental factors on microbial community are important to manage the performance of anaerobic processes. Microbial communities of 12 anaerobic sludge samples acclimated under different environmental conditions were investigated. Genera detected from these anaerobic sludge samples generally presented three distribution patterns: frequently detected with high abundance, frequently detected with low abundance and occasionally detected with permanently low abundance. The type of feed stock was one of the most important process parameters affecting the shape of microbial community (e.g., Syntrophus, Methylomonas and Methylobacillus). Dye wastewater (Bacteroides) and the supplement of conductive materials (genus T78) were also found to shape the microbial community. Some syntrophic bacteria and methanogens were rare in many anaerobic samples. However, correlation analysis suggested that rare genera are potential syntrophic partners and are responsible for syntrophic methanogenesis.

Color and nitrogen removal from synthetic dye wastewater in an integrated mesophilic hydrolysis/acidification and multiple anoxic/aerobic process.

Dye wastewater is one kind of refractory pollutant and it is commonly treated by the integrated anaerobic and aerobic process. A new integrated hydrolysis/acidification and multiple anoxic/aerobic (AO) process was proposed for the removal of color and nitrogen from azo dye wastewater. System performance, the degradation pathway of azo dye and nitrogen metabolic pathway were investigated with quadrupole-time-of-flight and metagenomic analyses. The proposed process removed color and nitrogen efficiently, with the removal percentages of 89.4% and 54.0%, respectively. A colorful intermediate C16H11N3O7S2 during the degradation of azo dye was detected. Controlling a low dissolved oxygen concentration in the multiple AO process could enhance nitrogen removal. The detected bacteria possessing azoreductase for the azo dye degradation included Desulfovibrio aminophilus, Thermoanaerobacter, Lactococcus raffinolactis, Ruminiclostridium and Rhodopirellula. The nitrifying genes of amo and hao were mainly detected in Nitrosomonas, while the denitrifying genes were detected in Thauera, Candidatus Accumulibacter and Rhodothermus marinus.