Stochastic processes shape microbial community assembly in biofilters: Hidden role of rare taxa.

Stochastic and deterministic processes are the major themes governing microbial community assembly; however, their roles in bioreactors are poorly understood. Herein, the mechanisms underlying microbial assembly and the effect of rare taxa were studied in biofilters. Phylogenetic tree analysis revealed differences in microbial communities at various stages. Null model analysis showed that stochastic processes shaped the community assembly, and deterministic processes emerged only in the inoculated activated sludge after domestication. This finding indicates the dominant role of stochastic factors (biofilm formation, accumulation, and aging). The Sloan neutral model corroborated the advantages of stochastic processes and mainly attributed these advantages to rare taxa. Cooccurrence networks revealed the importance of rare taxa, which accounted for more than 85% of the keystones. Overall, these results provide good foundations for understanding community assembly, especially the role of rare taxa, and offer theoretical support for future community design and reactor regulation.

Decoding the influence of low temperature on biofilm development: The hidden roles of c-di-GMP.

Biofilms are widely used and play important roles in biological processes. Low temperature of wastewater inhibits the development of biofilms derived from wastewater activated sludge. However, the specific mechanism of temperature on biofilm development is still unclear. This study explored the mechanism of temperature on biofilm development and found a feasible method to enhance biofilm development at low temperature. The amount of biofilm development decreased by approximately 66 % and 55 % at 4 °C and 15 °C, respectively, as compared to 28 °C. The cyclic dimeric guanosine monophosphate (c-di-GMP) concentration also decreased at low temperature and was positively correlated with extracellular polymeric substance (EPS) content, formation, and adhesion strength. Microbial community results showed that low temperature inhibited the normal survival of most microorganisms, but promoted the growth of some psychrophile bacteria like Sporosarcina, Caldilineaceae, Gemmataceae, Anaerolineaceae and Acidobacteriota. Further analysis of functional genes demonstrated that the abundance of functional genes related to the synthesis of c-di-GMP (K18968, K18967 and K13590) decreased at low temperature. Subsequently, the addition of exogenous spermidine increased the level of intracellular c-di-GMP and alleviated the inhibition effect of low temperature on biofilm development. Therefore, the possible mechanism of low temperature on biofilm development could be the inhibition of the microorganism activity and reduction of the communication level between cells, which is the closely related to the EPS content, formation, and adhesion strength. The enhancement of c-di-GMP level through the exogenous addition of spermidine provides an alternative strategy to enhance biofilm development at low temperatures. The results of this study enhance the understanding of the influence of temperature on biofilm development and provide possible strategies for enhancing biofilm development at low temperatures.

Microbial metabolic flexibility guarantees function resilience in response to starvation disturbance.

Starvation disturbance due to nutrient limitation is a common problem in bioreactors. However, an understanding of how microbial systems respond to starvation remains in its infancy. Here the metabolic response mechanism of a biofilm community to starvation was investigated using a well-controlled gaseous toluene treatment biofilter through interruption of its operation. It was found that metabolic characteristics showed significant differences before and after starvation. The dominant carbon source utilization type shifted from amino acids and carboxylic acids to esters and carbohydrates after starvation, which is more conducive to improving energy production. Metagenomic sequencing analysis supported that the changes in the dominant metabolic substrate, enhanced metabolic stability, and flexibility in the mode of energy metabolism could be the main ways to guarantee functional resilience in ecosystems after starvation. The results highlight the microbial metabolic response to starvation, which would be beneficial to the understanding of functional resilience and bioreactor stability.

Engineering a Pseudomonas putida as living quorum quencher for biofilm formation inhibition, benzenes degradation, and environmental risk evaluation.

Bacterial communication interruption based on quorum quenching (QQ) has been proven its potential in biofilm formation inhibition and biofouling control. However, it would be more satisfying if QQ could be combined with the efficient degradation of contaminants in environmental engineering. In this study, we engineered a biofilm of Pseudomonas putida through introducing a QQ synthetic gene, which achieved both biofilm formation inhibition and efficient degradation of benzene series in wastewater. The aiiO gene introduced into the P. putida by heat shock method was highly expressed to produce QQ enzyme to degrade AHL-based signal molecules. The addition of this engineered P. putida reduced the AHLs concentration, quorum sensing gene expression, and connections of the microbial community network in activated sludge and therefore inhibited the biofilm formation. Meanwhile, the sodium benzoate degradation assay indicated an enhanced benzene series removal ability of the engineering bacteria on activated sludge. Besides, we also demonstrated a controllable environmental risk of this engineered bacteria through monitoring its abundance and horizontal gene transfer test. Overall, the results of this study suggest an alternative strategy to solve multiple environmental problems through genetic engineering means and provide support for the application of engineered bacteria in environmental biotechnology.

Nagimycins A and B, Antibacterial Ansamycin-Related Macrolactams from Streptomyces sp. NA07423.

Chemical investigation of Streptomyces sp. NA07423 led to the discovery of two unreported macrolactams, nagimycins A (1) and B (2). Their structures were elucidated by NMR, HRESIMS, X-ray crystallography, and comparison of experimental and theoretical ECD spectra. The nagimycins have a unique butenolide moiety rarely found in ansamycin antibiotics. Genome analysis revealed the putative biosynthetic gene cluster for nagimycins, and a likely biosynthetic pathway was proposed. Notably, compounds 1 and 2 exhibited potent antibacterial activity against two pathogenic Xanthomonas bacteria.

Microbial community evolution and functional trade-offs of biofilm in odor treatment biofilters.

Biofilters inoculated with activated sludge are widely used for odor control in WWTP. In this process, biofilm community evolution plays an important role in the function of reactor and is closely related to reactor performance. However, the trade-offs in biofilm community and bioreactor function during the operation are still unclear. Herein, an artificially constructed biofilter for odorous gas treatment was operated for 105 days to study the trade-offs in the biofilm community and function. Biofilm colonization was found to drive community evolution during the start-up phase (phase 1, days 0-25). Although the removal efficiency of the biofilter was unsatisfactory at this phase, the microbial genera related to quorum sensing and extracellular polymeric substance secretion led to the rapid accumulation of the biofilm (2.3 kg biomass/m3 filter bed /day). During the stable operation phase (phase 2, days 26-80), genera related to target-pollutant degradation showed increases in relative abundance, which accompanied a high removal efficiency and a stable accumulation of biofilm (1.1 kg biomass/m3 filter bed/day). At the clogging phase (phase 3, days 81-105), a sharp decline in the biofilm accumulation rate (0.5 kg biomass/m3 filter bed /day) and fluctuating removal efficiency were observed. The quorum quenching-related genera and quenching genes of signal molecules increased, and competition for resources among species drove the evolution of the community in this phase. The results of this study highlight the trade-offs in biofilm community and functions during the operation of bioreactors, which could help improve bioreactor performance from a biofilm community perspective.

Degradation of trichloroethylene by double dielectric barrier discharge (DDBD) plasma technology: Performance, product analysis and acute biotoxicity assessment.

Trichloroethylene is carcinogenic and poorly degraded by microorganisms in the environment. Advanced Oxidation Technology is considered to be an effective treatment technology for TCE degradation. In this study, a double dielectric barrier discharge (DDBD) reactor was established to decompose TCE. The influence of different condition parameters on DDBD treatment of TCE was investigated to determine the appropriate working conditions. The chemical composition and biotoxicity of TCE degradation products were also investigated. Results showed that when SIE was 300 J L-1, the removal efficiency could reach more than 90%. The energy yield could reach 72.99 g kWh-1 at low SIE and gradually decreased with the increase of SIE. The k of the Non-thermal plasma (NTP) treatment of TCE was about 0.01 L J-1. DDBD degradation products were mainly polychlorinated organic compounds and produced more than 373 mg m-3 ozone. Moreover, a plausible TCE degradation mechanism in the DDBD reactors was proposed. Lastly, the ecological safety and biotoxicity were evaluated, indicating that the generation of chlorinated organic products was the main cause of elevated acute biotoxicity.

A method establishment and application for biofilm quorum quenching activity assay.

The existence of quorum sensing (QS) and quorum quenching (QQ) plays important roles in biofilm formation. However, direct detection of QS ability is difficult due to the low concentrations of signal molecules inside the biofilm. Therefore, QQ activity is typically used to indicate the attribution of QS/QQ to the biofilm. Nevertheless, current detection methods of QQ activity based on biosensors present undesirable operability and accuracy. In this study, the 96-well plate assay based on a specific biosensor, Agrobacterium tumefaciens A136, and a colorimetric substance, X-gal was established. The reliable fitting results were obtained by standardizing the composition of the A136 X-gal assay solution and optimizing the operating conditions. This method improved the accuracy of QQ activity detection and reduced time and cost consumption. Finally, the 96-well plate assay was successfully applied to detect the QQ activities of biofilm samples and explore possible environmental influencing factors. In general, this study provided a new strategy for understanding the QQ effect in biofilm systems.

Microbial community regulation and performance enhancement in gas biofilters by interrupting bacterial communication.

BACKGROUND: Controlling excess biomass accumulation and clogging is important for maintaining the performance of gas biofilters and reducing energy consumption. Interruption of bacterial communication (quorum quenching) can modulate gene expression and alter biofilm properties. However, whether the problem of excess biomass accumulation in gas biofilters can be addressed by interrupting bacterial communication remains unknown. RESULTS: In this study, parallel laboratory-scale gas biofilters were operated with Rhodococcus sp. BH4 (QQBF) and without Rhodococcus sp. BH4 (BF) to explore the effects of quorum quenching (QQ) bacteria on biomass accumulation and clogging. QQBF showed lower biomass accumulation (109 kg/m3) and superior operational stability (85-96%) than BF (170 kg/m3; 63-92%) at the end of the operation. Compared to BF, the QQBF biofilm had lower adhesion strength and decreased extracellular polymeric substance production, leading to easier detachment of biomass from filler surface into the leachate. Meanwhile, the relative abundance of quorum sensing (QS)-related species was found to decrease from 67 (BF) to 56% (QQBF). The QS function genes were also found a lower relative abundance in QQBF, compared with BF. Moreover, although both biofilters presented aromatic compounds removal performance, the keystone species in QQBF played an important role in maintaining biofilm stability, while the keystone species in BF exhibited great potential for biofilm formation. Finally, the possible influencing mechanism of Rhodococcus sp. BH4 on biofilm adhesion was demonstrated. Overall, the results of this study achieved excess biomass control while maintaining stable biofiltration performance (without interrupting operation) and greatly promoted the use of QQ technology in bioreactors. Video Abstract.

Effect of the Early Application of Evolocumab on Blood Lipid Profile and Cardiovascular Prognosis in Patients with Extremely High-Risk Acute Coronary Syndrome.

Proprotein convertase subtilisin/kexin 9 (PCSK9) inhibitors significantly reduce low-density lipoprotein cholesterol (LDL-C) and improve the prognosis of patients with acute coronary syndrome (ACS). However, the feasibility and safety of early application of PCSK9 inhibitors on the basis of statins combined with ezetimibe to strengthen lipid lowering in extremely high-risk coronary heart disease populations are still unknown.This study was a prospective, randomized controlled study. A total of 136 patients with extremely high-risk ACS with LDL-C ≥ 3.0 mmol/L after percutaneous coronary intervention (PCI) treatment were randomly assigned 1:1 to the control group (atorvastatin 40 mg/day and ezetimibe 10 mg/day) or the evolocumab group (evolocumab 140 mg every 2 weeks combined with atorvastatin 40 mg/day and ezetimibe 10 mg/day). We compared the blood lipid profiles, major adverse cardiovascular events (MACEs), and adverse reactions. MACEs included cardiogenic death, nonfatal myocardial infarction, nonfatal stroke, and readmission due to angina. Adverse reactions included allergies, myalgia, poor blood glucose control, and liver damage.Within 1 month, the average level of LDL-C in the evolocumab group decreased from 3.54 to 0.57 mmol/L and that in the control group decreased from 3.52 to 1.26 mmol/L. The LDL-C compliance (< 1.0 mmol/L) rate was significantly increased in the evolocumab group compared with the control group (82.35% versus 22.06%, P < 0.01). The average level of lipoprotein (a) (Lp (a) ) in the control group increased by 9.94 ± 51.93% from baseline after treatment, but evolocumab reduced the Lp (a) level (-38.84 ± 32.40%). Additionally, evolocumab further reduced the levels of apolipoprotein B/A1 (-70.56 ± 22.38% versus -51.29 ± 18.14%), cholesterol (-54.76 ± 18.10% versus -41.16 ± 18.14%), and apolipoprotein B (-66.47 ± 26.89% versus -46.78 ± 24.12%) compared with those in the control group, all P < 0.01. The blood lipid levels of both control and evolocumab groups stabilized after 1 month. During the 3-month follow-up, the incidence of MACEs after PCI was lower in the evolocumab group than in the control group (8.82% versus 24.59%, P = 0.015), and evolocumab combined with statins and ezetimibe did not increase the occurrence of adverse reactions (13.24% versus 11.48%, P = 0.762).In patients with extremely high-risk ACS with high levels of LDL-C, adding evolocumab to their treatment regimen as early as possible may enhance lipid lowering, increase the patient's LDL-C compliance rate in the short term, and improve cardiovascular prognosis but will not increase adverse reactions.

Characterization of the complete plastid genome of Scutellaria microviolacea (Lamiaceae), a species endemic to Yunnan Province of China.

The complete plastid genome of Scutellaria microviolacea C. Y. Wu was firstly reported. The full length of the plastid genome was 152,092 bp and comprised of a large single-copy (LSC) region of 84,090 bp, a small single-copy (SSC) region of 17,534 bp, and two inverted repeat regions (IRs) of 25,234 bp. A total of 131 genes were encoded, including 87 protein-coding genes, 36 tRNA genes, and 8 rRNA genes. The overall GC content of the S. microviolacea plastid genome was 38.3%. Further phylogenetic analysis based on 18 accessions inferred that the genus Scutellaria can be divided into two clades, and S. microviolacea is evolutionarily close to Scutellaria tsinyunensis. Our study provided essential genetic resources for further studies on the evolution and genetic diversity of the genus Scutellaria and its related taxa.

Profile and dynamics of infectious diseases: a population-based observational study using multi-source big data.

BACKGROUND: The current surveillance system only focuses on notifiable infectious diseases in China. The arrival of the big-data era provides us a chance to elaborate on the full spectrum of infectious diseases. METHODS: In this population-based observational study, we used multiple health-related data extracted from the Shandong Multi-Center Healthcare Big Data Platform from January 2013 to June 2017 to estimate the incidence density and describe the epidemiological characteristics and dynamics of various infectious diseases in a population of 3,987,573 individuals in Shandong province, China. RESULTS: In total, 106,289 cases of 130 infectious diseases were diagnosed among the population, with an incidence density (ID) of 694.86 per 100,000 person-years. Besides 73,801 cases of 35 notifiable infectious diseases, 32,488 cases of 95 non-notifiable infectious diseases were identified. The overall ID continuously increased from 364.81 per 100,000 person-years in 2013 to 1071.80 per 100,000 person-years in 2017 (χ2 test for trend, P < 0.0001). Urban areas had a significantly higher ID than rural areas, with a relative risk of 1.25 (95% CI 1.23-1.27). Adolescents aged 10-19 years had the highest ID of varicella, women aged 20-39 years had significantly higher IDs of syphilis and trichomoniasis, and people aged ≥ 60 years had significantly higher IDs of zoster and viral conjunctivitis (all P < 0.05). CONCLUSIONS: Infectious diseases remain a substantial public health problem, and non-notifiable diseases should not be neglected. Multi-source-based big data are beneficial to better understand the profile and dynamics of infectious diseases.

Reduction of biofilm adhesion strength by adjusting the characteristics of biofilms through enzymatic quorum quenching.

Biofilm adhesion to the surface of a carrier plays an essential role during biofilm formation. Quorum quenching (QQ) has been shown to have great potential for delaying biofouling. However, little is known about whether QQ reduces the adhesion strength of biofilms during the formation process to inhibit biomass accumulation. In this study, we explored the effect of enzymatic QQ on the adhesion strength during biofilm development. In addition, a quantitative method was used to measure the adhesion strength of biofilms based on the shear force of water flow. Experimental results showed that QQ enzyme could reduce the adhesion strength of biofilms by at least 37% compared with the control. Furthermore, the biofilm accumulation rates were 0.05673 and 0.08762 h-1 with and without the QQ enzyme, illustrating a negative effect of QQ enzyme on biofilm accumulation. Specifically, QQ was confirmed to reduce extracellular polymeric substances, decrease the relative hydrophobicity, change the zeta potential by degrading signal molecules, and weaken the adhesion strength of biofilms. The successful reduction of the adhesion strength of the biofilm through QQ could provide a new strategy for the management and regulation of biofilm adhesion in the bioreactor.

Mesoporous polydopamine-coated hydroxyapatite nano-composites for ROS-triggered nitric oxide-enhanced photothermal therapy of osteosarcoma.

In this paper, MPDA@hydroxyapatite nanocomposites (MPHA NCs) were prepared and applied to develop a novel reactive oxygen species (ROS)-triggered nitric oxide (NO)-enhanced photothermal therapy nanocomposite system composed of indocyanine green (ICG)/L-arginine-MPDA@HAp (AI-MPHA NCs) for displaying both NO gas therapy and photothermal osteosarcoma treatment. The nanosystem exhibited a mesoporous and core-shell structure and high ICG loading efficiency (about 90%). Under near infrared (NIR) irradiation, the AI-MPHA NCs could not only produce heat but also generate reactive oxygen species (ROS), inducing the catalysis of L-Arg to obtain NO. Under NIR irradiation, the AI-MPHA NCs achieved osteosarcoma ablation by a synergistic combination of photothermal therapy and NO-gas therapy. Additionally, the cell viability of MG-63 cells decreased to 23.6% (co-incubated with AI-MPHA NCs) under irradiation with a power density at 1.0 W cm-2 for 10 min. The study proposed a novel nano-platform for NO-enhanced photothermal therapy of osteosarcoma.

Antioxidant and antiaging effect of traditional Thai rejuvenation medicines in Caenorhabditis elegans.

OBJECTIVE: This study explored the rejuvenation mechanisms of Thai polyherbal medicines using different approaches, including in vitro methods, as well as a well-defined nematode model, Caenorhabditis elegans. METHODS: THP-R-SR012 decoction was selected from 23 polyherbal medicines, based on metal-chelating and chain-breaking antioxidant capacities. The influences of this extract on the survival and some stress biomarkers of C. elegans under paraquat-induced oxidative stress were evaluated. Furthermore, lifespan analysis and levels of lipofuscin accumulation were examined in senescent nematodes. The phytochemical profile of THP-R-SR012 was analyzed. RESULTS: Supplementation with THP-R-SR012 decoction significantly increased the mean lifespan and reduced the oxidative damage to C. elegans under oxidative stress conditions. Further, THP-R-SR012 supplementation slightly influenced the lifespan and the level of lipofuscin accumulation during adulthood. Antioxidant-related phytochemical constituents of THP-R-SR012 decoction were rutin, naringenin, 3,4-dihydroxybenzoic acid, gallic acid, glycyrrhizic acid, demethoxycurcumin and 18α-glycyrrhetinic acid. CONCLUSION: The antioxidant potential of THP-R-SR012 was due to its scavenging properties, its enhancement of antioxidant-related enzyme activities, and the presence of the antioxidant-related compound. These results support the traditional use of THP-R-SR012 decoction as a tonic for nourishing and strengthening the whole body.

The complete plastid genome of Aletris megalantha (Nartheciaceae), an endemic species from Yunnan Province of China.

Aletris megalantha F. T. Wang & Tang is an herbal plant species endemic to Yunnan Province of China. Its complete plastid genome sequence was 154,704 bp in length, with a large single-copy (LSC) region of 83,265 bp, a small single-copy (SSC) region of 18,127 bp, and a pair of inverted repeat regions (IRs) of 26,656 bp. The whole plastid genome encoded 132 genes, including 85 protein-coding genes, 38 tRNA genes, and eight rRNA genes. The overall GC content of A. megalantha plastid genome was 37.4%. Maximum likelihood phylogenetic analysis based on 14 taxa indicated that A. megalantha is evolutionarily close to A. spicata.

Emissions, measurement, and control of odor in livestock farms: A review.

Odor emissions from intensive livestock farms have attracted increased attention due to their adverse impacts on the environment and human health. Nevertheless, a systematic summary regarding the characteristics, sampling detection, and control technology for odor emissions from livestock farms is currently lacking. This paper compares the development of odor standards in different countries and summarizes the odor emission characteristics of livestock farms. Ammonia, the most common odor substance, can reach as high as 4100 ppm in the compost area. Sampling methods for point and area source odor emissions are introduced in this paper, and odor analysis methods are compared. Olfactometers, odorometers, and the triangle odor bag method are usually used to measure odor concentration. Odor control technologies are divided into three categories: physical (activated carbon adsorption, masking, and dilution diffusion), chemical (plant extract spraying, wet scrubbing, combustion, non-thermal plasma, and photocatalytic oxidation), and biological (biofiltration, biotrickling, and bioscrubbing). Each technology is elucidated, and the performance in the removal of different pollutants is summarized. The application scopes, costs, operational stability, and secondary pollution of the technologies are compared. The generation of secondary pollution and long-term operation stability are issues that should be considered in future technological development. Lastly, a case analysis for engineering application is conducted.

[Effects of seasonal temperature variation on nitrogen removal from a tidal flow constructed wetland system with CANON process]. / 季节性温度变化对CANON型潮汐流人工湿地脱氮的影响.

We investigated the effects of seasonal temperature variation on the treatment perfor-mance and underlying mechanisms of nitrogen transformation in a tidal flow constructed wetland (TFCW) with the complete autotrophic nitrogen removal over nitrite (CANON) process. Different temperatures resulted in periodical variations in nitrogen transformation pathways and removal performance of the TFCW with CANON process, which was mainly due to the changes of dominant bacterial communities for nitrogen removal in the system. When temperature was higher than 20.0 ℃, nitrogen transformation and associated microbial characteristics in the TFCW were significantly affected, and the CANON process remained to be the principal pathway for nitrogen removal. The abundance and activity of anammox bacteria experienced different degrees of reduction when temperature dropped below 20.0 ℃. At the temperature of 9.3-20.0 ℃, the proliferation and increased activities of nitrite oxidizing bacteria (NOB) made the nitrification/denitrification process instead of the CANON process became the primary total nitrogen (TN) removal route in the TFCW, and the TN removal efficiency of the system declined to 34.8%±13.0%. Under the temperature range of 2.2-9.0 ℃, anammox bacteria, which was inhibited at the low temperatures, presented competitive advantage in comparison with NOB and denitrifiers, resulting that nitrogen removal in the TFCW relied on the CANON process again. Correspondingly, nitrogen removal rate of the system was 54.8%±4.8%. This study was conductive to the optimization of the TFCW with CANON process, as well as its engineering application.

A short review of bioaerosol emissions from gas bioreactors: Health threats, influencing factors and control technologies.

Bioaerosols have widely been a concern due to their potential harm to human health caused by the carrying and spreading of harmful microorganisms. Biofiltration has been generally used as a green and effective technology for processing VOCs. However, bioaerosols can be emitted into the atmosphere as secondary pollutants from the biofiltration process. This review presents an overview of bioaerosol emissions from gas bioreactors. The mechanism of bioaerosols production and the effect of biofiltration on bioaerosol emissions were analyzed. The results showed that the bioaerosol emission concentrations were generally exceeded 104 CFU m-3, which would damage to human health. Biomass, inlet gas velocity, moisture content, temperature, and some other factors have significant influences on bioaerosol emissions. Moreover, as a result of the analysis done herein, different inactivation technologies and microbial immobilization of bioaerosols were proposed and evaluated as a potential solution for reducing bioaerosols emissions. The purpose of this paper is to make more people realize the importance of controlling the emissions of bioaerosols in the biofiltration process and to make the treatment of VOCs by biotechnology more environmentally friendly. Additionally, the present work intends to increase people's awareness in regards to the control of bioaerosols, including microbial fragment present in bioaerosols.

Performance enhancement of a biofilter with pH buffering and filter bed supporting material in removal of chlorobenzene.

Acidic substances, which produced during chlorinated volatile organic compounds, will corrode the commonly used packing materials, and then affect the removal performance of biofiltration. In this study, three biofilters with different filter bed structure were established to treat gaseous chlorobenzene. CaCO3 and 3D matrix material was added in filter bed as pH buffering material and filter bed supporting material, respectively. A comprehensive investigation of removal performance, biomass accumulation, microbial community, filter bed height, voidage, pressure drops, and specific surface area of the three biofilters was compared. The biofilter with CaCO3 and 3D matrix material addition presented stable removal performance and microbial community, and greater biomass density (209.9 kg biomass/m3 filter bed) and growth rate (0.033 d-1) were obtained by using logistic equation. After 200 days operation, the height, voidage, pressure drop, specific surface area of the filter bed consisted of perlite was 27.4 cm, 0.39, 32.8 Pa/m, 974,89 m2/m3, while those of the filter bed with CaCO3 addition was 28.2 cm, 0.43, 21.3 Pa/m, and 1021.03 m2/m3, and those of the filter bed with CaCO3 and 3D matrix material addition was 28.7 cm, 0.55, 17.4 Pa/m, and 1041.60 m2/m3. All the results verified the biofilter with CaCO3 and 3D matrix material addition is capable of sustaining the long-term performance of biofilters. CaCO3 could limit the changes of removal efficiency, microbial community and filter bed structure by buffering the pH variation. And 3D matrix material could maintain the filter bed structure by supporting the filter bed, regardless of the buffering effect.