Eukaryotic diversity in an anaerobic aquifer polluted with landfill leachate

Eukaryotes may influence pollutant degradation processes in groundwater ecosystems by activities such as predation on bacteria and recycling of nutrients. Culture-independent community profiling and phylogenetic analysis of 18S rRNA gene fragments, as well as culturing, were employed to obtain insight into the sediment-associated eukaryotic community composition in an anaerobic sandy aquifer polluted with landfill leachate (Banisveld, The Netherlands). The microeukaryotic community at a depth of 1 to 5 m below the surface along a transect downgradient (21 to 68 m) from the landfill and at a clean reference location was diverse. Fungal sequences dominated most clone libraries. The fungal diversity was high, and most sequences were sequences of yeasts of the Basidiomycota. Sequences of green algae (Chlorophyta) were detected in parts of the aquifer close (<30 m) to the landfill. The bacterium-predating nanoflagellate Heteromita globosa (Cercozoa) was retrieved in enrichments, and its sequences dominated the clone library derived from the polluted aquifer at a depth of 5 m at a location 21 m downgradient from the landfill. The number of culturable eukaryotes ranged from 10(2) to 10(3) cells/g sediment. Culture-independent quantification revealed slightly higher numbers. Groundwater mesofauna was not detected. We concluded that the food chain in this polluted aquifer is short and consists of prokaryotes and fungi as decomposers of organic matter and protists as primary consumers of the prokaryotes.     

Comparison of long-term performances and final microbial compositions of anaerobic reactors treating landfill leachate

Laboratory scale anaerobic upflow filter, sludge blanket and hybrid bed reactors were operated for 860 days in the treatment of high ammonia landfill leachate. Organic loading was gradually increased from 1.3 to 23.5 kg COD/m3 day in the start-up period and then fluctuated according to the COD concentration of raw leachate. To prevent free ammonia inhibition, influent pH was reduced to 4.5 after Day 181 and consequently COD removal efficiencies above 80% were achieved in all reactors. However, the anaerobic filter and hybrid bed reactor were generally found slightly more efficient and stable than the UASB reactor. In addition to conventional anaerobic reactor control parameters, the complementary techniques of denaturing gradient gel electrophoresis (DGGE), cloning and fluorescent in situ hybridization (FISH) were used to identify and compare the microbial profiles in the reactors at Day 830. Molecular analyses revealed that acetoclastic Methanosaeta species were prevalent in all reactors and configuration did not have an impact on microbial diversity in the long-term.     

Anaerobic digestion of a petrochemical effluent using an anaerobic hybrid digester

Summary An anaerobic hybrid reactor was used in the anaerobic treatment of an acidic petrochemical effluent. An organic loading rate of 20.04 kg COD/(m³d) at a HRT of 17 hours was obtained with a volatile fatty acid removal of 91%, and COD removal of 84%. A final reactor effluent containing 44 mg/l ammonia nitrogen and 12.3 mg/l PO₄-P was produced.     

Temperature-induced changes in treatment efficiency and microbial structure of aerobic granules treating landfill leachate

This paper investigates the effect of temperature on nitrogen and carbon removal by aerobic granules from landfill leachate with a high ammonium concentration and low concentration of biodegradable organics. The study was conducted in three stages; firstly the operating temperature of the batch reactor with aerobic granules was maintained at 29 °C, then at 25 °C, and finally at 20 °C. It was found that a gradual decrease in operational temperature allowed the nitrogen-converting community in the granules to acclimate, ensuring efficient nitrification even at ambient temperature (20 °C). Ammonium was fully removed from leachate regardless of the temperature, but higher operational temperatures resulted in higher ammonium removal rates [up to 44.2 mg/(L h) at 29 °C]. Lowering the operational temperature from 29 to 20 °C decreased nitrite accumulation in the GSBR cycle. The highest efficiency of total nitrogen removal was achieved at 25 °C (36.8 ± 10.9 %). The COD removal efficiency did not exceed 50 %. Granules constituted 77, 80 and 83 % of the biomass at 29, 25 and 20 °C, respectively. Ammonium was oxidized by both aerobic and anaerobic ammonium-oxidizing bacteria. Accumulibacter sp., Thauera sp., cultured Tetrasphaera PAO and Azoarcus–Thauera cluster occurred in granules independent of the temperature. Lower temperatures favored the occurrence of denitrifiers of Zooglea lineage (not Z. resiniphila), bacteria related to Comamonadaceae, Curvibacter sp., Azoarcus cluster, Rhodobacter sp., Roseobacter sp. and Acidovorax spp. At lower temperatures, the increased abundance of denitrifiers compensated for the lowered enzymatic activity of the biomass and ensured that nitrogen removal at 20 °C was similar to that at 25 °C and significantly higher than removal at 29 °C.     

Functional stability of a hybrid anaerobic digester/microbial fuel cell system treating municipal wastewater

A thermophilic anaerobic digester (AD) was combined with a microbial fuel cell (MFC) to evaluate whether either component had increased stability when operated in combination as a hybrid system, perturbed by the addition of acetic acid. The MFC and the anaerobic digester were able to operate effectively together. The MFC was more susceptible to high acetic acid load than the AD. The hybrid system did not have increased resilience compared to the solitary systems in the conditions tested. However, the low pH had a relatively delayed effect on the MFC compared to the AD, allowing the hybrid system to have a more stable energy output. Also, at very low pH, when operating as a hybrid, the AD component was able to recover pH to normal levels when the MFC component failed. These results demonstrate that there are synergies that can be gained from this hybrid system.     

The development of granulation in an upflow floc digester and an upflow anaerobic sludge blanket digester treating cane juice stillage

Summary The use of synthetic polyelectrolytes in the Upflow Floc digester during the treatment of high strength cane juice stillage, resulted in a more rapid accumulation of biomass and promoted granule formation at an earlier stage compared to a control upflow anaerobic sludge blanket digester (UASB). In the Upflow Floc reactor the granules were composed of rod shaped organisms, whereas in the UASB the granules were primarily filamentous. Both types of granules had good settling properties and high activities.     

Short communication: Butyrate-degrading syntrophic culture from an anaerobic digester treating cattle waste

A co-culture of bacteria responsible for the conversion of butyrate to methane and CO₂ was isolated from a cattle-waste treatment plant. The non-methanogenic partner of the co-culture was Syntrophomonas wolfei and the methanogenic partner was Methanobacterium formicicum. Although butyrate degradation occurred at pH<6.0 and below 45°C, methanogenesis was observed at pH>6.5 and above 40°C.     

Stability and efficiency of biofilms for landfill leachate treatment

The objective of this work was to assess the feasibility of a fixed-film biological aerated filtration process for the treatment of the leachate produced at Harnhill landfill site (South Gloucestershire, UK). The laboratory scale plant consisted of four identical biological aerated filters (a triplicate and a “control” column) packed with small brick fragments. Biofilm formed within 24 h of immersion of the support material in the reactor, and had a high resistance to antibiotics and other toxic agents. The plant maintained a stable operation in the 20–45 °C temperature range, showing the best results (35.4 ± 6.6% COD removal and 73.9 ± 5.5% BOD₅ reduction) at 40 °C. The lowest COD and BOD₅ values obtained at the outlet of the columns were 7067 mg/L and 1050 mg/L, respectively.     

Nitrogen removal through different pathways in an aged refuse bioreactor treating mature landfill leachate

In this study, an aged refuse bioreactor was constructed to remove nitrogen in a mature landfill leachate. The nitrogen removal efficiency and the microbial community composition in the bioreactor were investigated. The results showed that the aged refuse bioreactor removed more than 90 % of total nitrogen in the leachate under the nitrogen loading rate (NLR) of 0.74 g/kg (vs) day, and the total nitrogen removal rate decreased to 62.2 % when NLR increased up to 2.03 g/kg (vs) day. Quantitative polymerase chain reaction results showed that the average cell number of ammonia-oxidizing bacteria in the bioreactor was 1.58?×?10⁸ cells/g, which accounted for 0.41 % of total bacteria. The number of anammox bacteria in the reactor was 1.09?×?10⁸ cells/g, which accounted for 0.27 % of total bacteria. Isotopic ¹⁵N tracing experiments showed that nearly 10 % of nitrogen was removed by anammox. High-throughout 454 pyrosequencing revealed that the predominant bacteria in the bioreactor were Proteobacteria, Chloroflexi, Actinobacteria, Bacteroidetes, and Gemmatimonadetes, including various nitrifiers and denitrifiers with diverse heterotrophic and autotrophic metabolic pathways, supporting that nitrogen was removed through different pathways in this aged refuse bioreactor.     

A pilot scale two-stage anaerobic digester treating food waste leachate (FWL): Performance and microbial structure analysis using pyrosequencing

Food waste leachate (FWL) from the food waste recycling facilities in Korea is a serious environmental problem. Much research was done on anaerobic digestion of FWL in a lab-scale; however, there is little information on a large scale anaerobic digestion system (ADS). In this study, a two-phase ADS in a pilot scale was operated using FWL and the ADS performance and microbial structure dynamics using pyrosequencing were investigated. The ADS was operated for 136 days using FWL containing a high concentration of volatile fatty acid (12,435 ± 2203 mg/L), exhibiting volatile acid (VS) removal efficiency of 74–89% and CH₄ yield of 0.39–0.85 Nm³/kg of reduced VS. The microbial structure at 76, 101, and 132 days indicated the methanogen population shift from acetoclastic methanogens (Methanosarcina and Methanosaeta) to hydrogenotrophic methanogens (Methanobacterium and Methanoculleus). The bacterial community also shifted to the taxa syntrophically related with hydrogenotrophic methanogens (Clostridia). The statistical analysis revealed the positive correlation of VS removal efficiency with Methanosarcina, but the negative correlation with Methanobacterium. The results presented here suggest that acetoclastic methanogens and their associated bacteria were more efficient for VS removal in the pilot scale ADS system, providing useful information for FWL treatment in a large scale ADS.     

Characterization of aerobic and facultative anaerobic bacteria from the liquid phase of an anaerobic fixed-bed digester treating a cheese whey substrate

Bacterial counts on the liquid phase of an anaerobic, fixed-bed digester, treating a deproteinated, prefermented cheese whey substrate, were conducted on two different media under aerobic and facultative conditions. Average counts of 16.6×10⁶ and 26.5×10⁶ ml^–1 were obtained on the two media, with the nutritionally poorer of the two media giving the highest average count. Seventy-five isolates from both media, incubated aerobically as well as in anaerobic jars, were obtained. These isolates as well as 13 reference strains were phenotypically characterized. The similarities between cultures were calculated using the similarity coefficient of Sokal and Michener [16]. The organisms were clustered using the unweighted pair group method, and the results presented as a simplified dendrogram. The isolates could be divided into 3 main groups: gram-negative fermentative rods, mainlyEnterobacter, Klebsiella, andCitrobacter, withKlebsiella as the predominant genus; gram-positive bacteria, mainly enterococci; and gram-negative nonfermentive rods of the generaPseudomonas, Alcaligenes, andAcinetobacter. All the enterobacteria and enterococci were able to produce acetic acid, an intermediate in methanogenesis.     

Membrane fouling and micro-pollutant removal of membrane bioreactor treating landfill leachate

Reviews in Environmental Science and Bio/Technology - The characteristics of landfill leachate from solid waste disposal vary, depending on waste composition, waste age, and landfilling technique....     

Behavior of methanogens during start-up of farm-scale anaerobic digester treating swine wastewater

The aim of this study was to monitor the changes in methanogenic community structures in an anaerobic digester (250 m³ working volume) during start-up including prolonged starvation periods. Redundancy analysis was performed to investigate the correlations between environmental variables and microbial community structures. The anaerobic digester was operated for 591 days at alternating operating temperatures. In initial start-up period at stage I (35 °C), growth of various species of mesophilic aceticlastic methanogens (AMs) and hydrogenotrophic methanogens (HMs) was observed. Methanobacteriales species survived better than other methanogens under long-term starvation conditions. In stage II (50 °C), HMs became dominant over AMs as the operating temperature changed from mesophilic to thermophilic due to increase of ammonia inhibition. In stage III (35 °C), only the Methanomicrobiales population significantly increased during 50 days of HRT while Methanobacteriales dominated over 15 days of HRT. The influent pH negatively correlated with all methanogenic populations especially in stage II.     

Microbial community structure of a pilot-scale thermophilic anaerobic digester treating poultry litter

The microbial community structure of a stable pilot-scale thermophilic continuous stirred tank reactor digester stabilized on poultry litter was investigated. This 40-m³ digester produced biogas with 57 % methane, and chemical oxygen demand removal of 54 %. Bacterial and archaeal diversity were examined using both cloning and pyrosequencing that targeted 16S rRNA genes. The bacterial community was dominated by phylum Firmicutes, constituting 93 % of the clones and 76 % of the pyrotags. Of the Firmicutes, class Clostridia (52 % pyrotags) was most abundant followed by class Bacilli (13 % pyrotags). The bacterial libraries identified 94 operational taxonomic units (OTUs) and pyrosequencing identified 577 OTUs at the 97 % minimum similarity level. Fifteen OTUs were dominant (≥2 % abundance), and nine of these were novel unclassified Firmicutes. Several of the dominant OTUs could not be classified more specifically than Clostridiales, but were most similar to plant biomass degraders, including Clostridium thermocellum. Of the rare pyrotag OTUs (<0.5 % abundance), 75 % were Firmicutes. The dominant methanogen was Methanothermobacter which has hydrogenotrophic metabolism, and accounted for >99 % of the archaeal clones. Based on the primary methanogen, as well as digester chemistry (high VA and ammonia levels), we propose that bacterial acetate oxidation is the primary pathway in this digester for the control of acetate levels.     

Influence of the acetate concentration on the recovery time of a perturbed anaerobic digester

Summary Imbalance in a fluidized bed reactor was simulated by injecting pure or mixed acetate and propionate salts into the digester. For acetate levels higher than 13–15 mM, propionate degradation was inhibited. In these cases, the time required by the system to reduce the propionate concentration up to steady-state values was approximately 60–70 % higher than the recovery time found in the cases in which this inhibition was not present.     

Performance and bacterial compositions of aged refuse reactors treating mature landfill leachate

Aged landfill leachates become more refractory over time and difficulty to treat. Recently, aged refuse bioreactors show great promise in treating leachates. In this study, aged refuse bioreactors were constructed to simulate landfill leachate degradation process. The characteristics of leachate were: COD_cr, ∼2200 mg/L; BOD₅, ∼280 mg/L; total nitrogen, ∼2030 mg/L; and ammonia, ∼1900 mg/L. Results showed that bioreactor could remove leachate pollutants effectively at hydraulic loading of 20 L/m³ d. The removal rate reduced when hydraulic loading doubled or temperature lowered. Effluent recirculation could alleviate the temperature effect. Combining aged refuse and slag biofilters could treat leachate more efficiently. Pyrosequencing analysis indicated that bacteria from Pseudomonas, Lysobacter, Bacillus and δ-proteobacter, Flexibacteraceae were more abundant in the samples. The Shannon index decreased at lower temperature, while evenness and equitability increased with recirculation. We suggest that filter medium and temperature may be the main factors for shaping bacterial community structure.     

N-butyrate catabolism in the treatment of a semi-synthetic landfill leachate on anaerobic filter under sequential feeding conditions

Calculations of the kinetics of volatile fatty acids removal in an anaerobic filter showed that methane production from n-butyrate was not consistent with β-oxidation when the reactor was operated under sequential feeding conditions. Under these conditions, n-butyrate might be anaerobically catabolised by decarboxylation. Iso-butyrate was formed at about 25% of the total amount of n-butyrate removed. After 13 days of continuous feeding, the material balance was consistent with the β-oxidation mechanism and the formation of iso-butyrate became negligible. N-butyrate decarboxylation may therefore be a transient catabolic pathway, although the reason why decarboxylation is preferred to β-oxidation under conditions such as sequential feeding is not clear. Results of similar experiments on acetate removal were consistent with theoretical expectations under conditions of both sequential and continous feeding.     

Treatment of leachate from a solid waste landfill site using a two-stage anaerobic filter

Raw leachate was treated using a two-stage upflow anaerobic filter process. Leachate from a solid waste landfill site, which received both municipal and industrial wastes, contained high organic matter (17-21 g/L COD, 13-14 g/L BOD, and 3.5-4.6 g/L volatile acids), and low metal (Zn and Fe) concentrations. Depending on sampling time, leachate composition and characteristics varied considerably. At an organic loading up to 4 g COD/day(2) media area, the BOD and COD removal percentages were 98 and 91%, respectively. The biofilters were also effective for metal removal. However, the filter effluent contained a high concentration of ammonia. System overloading was characterized by the accumulation of large quantities of volatile acids and by a now ratio of alkalinity/volatile acids, resulting in low COD removal and reduced gas production. Once the first filter was upset, the second stage could only partially respond to the volatile acids accumulated in the effluent of first filter.     

Nitrogen mass balance and microbial analysis of constructed wetlands treating municipal landfill leachate

Experiments were conducted to investigate the feasibility of applying constructed wetlands to treat a sanitary landfill leachate containing high nitrogen and bacterial contents. Under a tropical condition (temperature of about 30 degrees C), the constructed wetland units operating at the hydraulic retention time of 8d yielded the best treatment efficiencies with BOD(5), TN and fecal coliforms removal of 91%, 96% and more than 99%, respectively. Cadmium removal in the SFCW bed was 99.7%. Mass balance analysis, based on total nitrogen contents of the plant biomass and dissolved oxygen and oxidation-reduction potential values, suggested that 88% of the input total nitrogen were uptaken by the plant biomass. Fluorescence in situ hybridization results revealed the predominance of bacteria, including heterotrophic and autotrophic, responsible for BOD(5) removal. Nitrifying bacteria was not present in the constructed wetland beds.