Microbiological characterization and specific methanogenic activity of anaerobe sludges used in urban solid waste treatment

This study presents the microbiological characterization of the anaerobic sludge used in a two-stage anaerobic reactor for the treatment of organic fraction of urban solid waste (OFUSW). This treatment is one alternative for reducing solid waste in landfills at the same time producing a biogas (CH(4) and CO(2)) and an effluent that can be used as biofertilizer. The system was inoculated with sludge from a wastewater treatment plant (WWTP) (Río Frío Plant in Bucaramanga-Colombia) and a methanogenic anaerobic digester for the treatment of pig manure (Mesa de los Santos in Santander). Bacterial populations were evaluated by counting groups related to oxygen sensitivity, while metabolic groups were determined by most probable number (MPN) technique. Specific methanogenic activity (SMA) for acetate, formate, methanol and ethanol substrates was also determined. In the acidogenic reactor (R1), volatile fatty acids (VFA) reached values of 25,000 mg L(-1) and a concentration of CO(2) of 90%. In this reactor, the fermentative population was predominant (10(5)-10(6)MPN mL(-1)). The acetogenic population was (10(5)MPN mL(-1)) and the sulphate-reducing population was (10(4)-10(5)MPN mL(-1)). In the methanogenic reactor (R2), levels of CH(4) (70%) were higher than CO(2) (25%), whereas the VFA values were lower than 4000 mg L(-1). Substrate competition between sulphate-reducing (10(4)-10(5)MPN mL(-1)) and methanogenic bacteria (10(5)MPN mL(-1)) was not detected. From the SMA results obtained, acetoclastic (2.39 g COD-CH(4)g(-1)VSS(-1)day(-1)) and hydrogenophilic (0.94 g COD-CH(4)g(-1)VSS(-1)day(-1)) transformations as possible metabolic pathways used by methanogenic bacteria is suggested from the SMA results obtained. Methanotrix sp., Methanosarcina sp., Methanoccocus sp. and Methanobacterium sp. were identified.     

Effect of substrate feeding frequencies on the methane production and microbial communities of laboratory-scale anaerobic digestion reactors

Even though full-scale digesters have been designed based on laboratory-scale tests, the substrate feeding modes of laboratory-scale tests might be different from those of full-scale digesters. The effect of substrate feeding frequencies on the performance and microbial community of laboratory-scale anaerobic digestion reactors was investigated. Feeding frequencies of twice a day, once a day, and every two days were tested in three 2-L reactors with an organic loading rate of 0.5 g-glucose/L/day under mesophilic condition. According to the results of this study, all the reactors showed similar methane production rates and SCOD removal efficiencies after sufficient time of acclimation, but frequently feeding promoted more stable digestion. Although there was no significant difference in microbial diversities from pyrosequencing analyses, the changes of archaeal community composition were observed. The decrease in feeding frequency appeared to cause shifts from acetoclastic methanogens affiliated with Methanosaeta to H₂-utilizing methanogens. The increase of Methanosaeta at a frequently feeding might contribute to the stability of reactor operation. Since this study uses glucose as the substrate, there is still possibility of different results for more complex substrates, such as sludge, food waste, etc.     

Anaerobic co-digestion of tannery waste water and tannery solid waste using two-stage anaerobic sequencing batch reactor: focus on performances of methanogenic step

In this study, anaerobic co-digestion of the tannery waste water (TWW) and tannery solid waste (TSW) with four TWW to TSW mixing ratios (100:0, 75:25, 50:50 and 25:75) was carried out using semi-continuous two-phase anaerobic sequencing batch reactor system under mesophilic temperature (38?±?2 °C). During the experimental study, effluents resulted from previously optimized acidogenic reactors were used to feed subsequent methanogenic reactors and then operated at hydraulic retention time (HRT) of 20, 15 and 10 days and equivalent organic loading rate. The findings revealed that methanogenic reactor of 50:50 (TWW:TSW) treating the effluent from previously optimized acidogenic step exhibits best process performances in terms of daily biogas (415 ml/day), methane production (251 ml/day), methane content (60.5%) and COD removal efficiency (75%) when operated at HRT of 20 days. Process stability of methanogenic step also evaluated and the obtained results showed suitable pH (6.8), no VFA accumulation, i.e., VFA/Alkalinity (0.305), alkalinity (3210 mgCaCO₃/l) and ammonia (246 mg/l with in optimum operating range). In general, improved process stability as well as performance was achieved during anaerobic co-digestion of TWW with TSW compared to mono-digestion of TWW.     

Evolution of butyric acid and the methanogenic microbial population in a thermophilic dry anaerobic reactor

Knowledge of the relationship between the evolution of butyric acid, the main precursor of methane, and the methanogenic microbial population, quantified by fluorescent in situ hybridization (FISH), is important for understating and controlling the anaerobic digestion of solid waste. In this study, a statistical analysis has been made of this relationship in a laboratory-scale continuously stirred tank reactor (CSTR) without recycling of biomass. The CSTR has been operated at four organic loading rates (OLR) between 4.42 and 7.50 kg VS m^?3 day^?1, using the Organic Fraction of Municipal Solid Waste (OFMSW) as the substrate. During the start-up stage, the consumption of butyric acid is related to the population of H₂-utilising methanogens (R² = 0.774), whereas in the stabilization stage, it is related to that of acetoclastic methanogens (R² = 0.716). Therefore, it can be concluded that the methanogenic population dynamics are closely correlated with the concentration of volatile fatty acids, and in particular, with that of butyric acid.     

Long-term anaerobic digestion of food waste stabilized by trace elements

The purpose of this study was to examine if long-term anaerobic digestion of food waste in a semi-continuous single-stage reactor could be stabilized by supplementing trace elements. Contrary to the failure of anaerobic digestion of food waste alone, stable anaerobic digestion of food waste was achieved for 368 days by supplementing trace elements. Under the conditions of OLR (organic loading rates) of 2.19-6.64 g VS (volatile solid)/L day and 20-30 days of HRT (hydraulic retention time), a high methane yield (352-450 mL CH(4)/g VS(added)) was obtained, and no significant accumulation of volatile fatty acids was observed. The subsequent investigation on effects of individual trace elements (Co, Fe, Mo and Ni) showed that iron was essential for maintaining stable methane production. These results proved that the food waste used in this study was deficient in trace elements.     

Anaerobic digestion of municipal solid waste: Thermophilic vs. mesophilic performance at high solids

High solids anaerobic digestion of the mechanically sorted organic fraction of municipal solid waste under mesophilic and thermophilic conditions is reported. The semi-dry thermophilic process has a gas production rate two to three times the mesophilic process and nearly complete biodegradation. A 3 m³ stirred digester, feeding organic waste at 16–23% solids, was operated at hydraulic (volumetric) retention times decreasing from 15-8 days, and at organic loading rates increasing from 6 to 14 kg volatile solids m⁻³ day⁻¹. An economic evaluation favours the thermophilic over the mesophilic process.     

Treatment of a high-strength sulphate-rich alkaline leachate using an anaerobic filter

The research looks at the feasibility of treating an alkaline sulphate-rich leachate arising from the co-disposal of municipal solid waste with cement kiln dust by means of an anaerobic filter (AF). This type of leachate with a high sulphate concentration is commonly prohibited for discharge to sewer and requires an on-site treatment solution. The AF used had a working volume of 4l and contained reticulated polyurethane foam as the biomass support material. The filters were operated over a 152 day experimental period during which the COD loading onto the filter was increased from 0.76 to 7.63kgCODm(-3)d(-1). In the early stages of operation at low loading, soluble sulphides accumulated that inhibited methanogenic activity. This was restored by dosing FeCl(3) to the reactor. The continued dosing allowed efficient COD removal of between 75% and 90% until the nominal retention time in the reactor was 3 days, at which point reactor performance declined significantly. The main mechanism for COD removal was by sulphate-reducing bacteria, which also resulted in up to 88% sulphate removal from the leachate. The average methane generation rate was 0.10lg(-1) COD removed. The results indicate the potential for using this approach as a pre-treatment that could significantly reduce the COD load to a second stage treatment process, but problems associated with the implementation of the technology at a larger scale have been identified.     

Two-phase anaerobic digestion within a solid waste/wastewater integrated management system

A two-phase, wet anaerobic digestion process was tested at laboratory scale using mechanically pre-treated municipal solid waste (MSW) as the substrate. The proposed process scheme differs from others due to the integration of the MSW and wastewater treatment cycles, which makes it possible to avoid the recirculation of process effluent. The results obtained show that the supplying of facultative biomass, drawn from the wastewater aeration tank, to the solid waste acidogenic reactor allows an improvement of the performance of the first phase of the process which is positively reflected on the second one. The proposed process performed successfully, adopting mesophilic conditions and a relatively short hydraulic retention time in the methanogenic reactor, as well as high values of organic loading rate. Significant VS removal efficiency and biogas production were achieved. Moreover, the methanogenic reactor quickly reached optimal conditions for a stable methanogenic phase. Studies conducted elsewhere also confirm the feasibility of integrating the treatment of the organic fraction of MSW with that of wastewater.     

Effect of long chain fatty acids removal as a pretreatment on the anaerobic digestion of food waste

This study investigated the effect of long chain fatty acids (LCFAs) removal as a pretreatment prior to anaerobic digestion on the production of methane from food waste. The results showed that the anaerobic digestion of food waste containing 1.6 g COD/L of LCFAs was not inhibited (4 days lag-time, 78.3 % methane recovery in 35 days) compared to that of lipid free food waste (3 days lag time, 72.5 % methane recovery in 35 days); however, some unsaturated LCFAs, which are toxic to microorganism, were accumulated in the batch anaerobic digestion reactor. Meanwhile, in a methanogenic activity study, the activity of methanogens was observed to be linearly inhibited by the presence of more than 1 g COD/L of LCFAs. The possibility of the accumulation of unsaturated LCFAs in the reactor should be considered when operating a large-scale continuous system.     

Report: anaerobic digestion of a sulphate-rich high-strength landfill leachate: the effect of differential dosing with FeCl3.

The effect of different dose concentrations of FeCl3 on the performance of a mesophilic anaerobic digester treating a highly alkaline, high-strength and sulphate-rich landfill leachate was tested. The results indicated that sulphate reduction was the predominant reaction and methanogenic processes were entirely inhibited in the reactor without FeCl3 addition. Adding FeCl3 into the reactor restored some methanogenic activity and also improved the rates of sulphate reduction. A combination of sulphate reduction and methanogenic activity resulted in up to 75% chemical oxygen demand (COD) removal and 85% sulphate removal. Sulphate reduction remained the principle mechanism by which COD removal took place with a methane yield of only between 0.14 and 0.18 m3 CH4 kg(-1) COD removed being achieved. The process was, however, stable and offered advantages for the further treatment or conveyance of the anaerobically treated leachate.     

Kinetics of microbial landfill methane oxidation in biofilters

A methane oxidizing biofilter system fitted to the passive venting system of a harbor sludge landfill in Germany was characterized with respect to the the methanotrophic population, methane oxidizing capacity, and reaction kinetics. Methanotrophic cell counts stabilized on a high level with 1.3 x 10(8) to 7.1 x 10(9) cells g dw(-1) about one year after first biofilter operation, and a maximum of 1.2 x 10(11) cells g dw(-1). Potential methane oxidizing activity varied between 5.3 and 10.7 microg h(-1) g dw(-1). Cell numbers correlated well with methane oxidation activities. Extrapolation of potential activities gave methane removal rates between 35 and 109 g CH4 h(-1) m(-3), calculated for 30 degrees C. Optimum temperature was 38 degrees C for freshly sampled biofilter material and 22 degrees C for a methanotrophic enrichment culture grown at 10 degrees C incubation temperature. Substrate kinetics revealed the presence of a low-affinity methane oxidizing community with a high Vmax of 1.78 micromol CH4 h(-1) g ww(-1) and a high K(M) of 15.1 microM. K(MO2) for methane oxidation was 58 microM. No substantial methane oxidizing activity was detected below 1.7-2.6 vol.-% O2 in the gaseous phase. Methane deprivation led to a decrease in methane oxidation activity within 5-9 weeks but could still be detected after 25 weeks of substrate deprivation and was fully restored within 3 weeks of continuous methane supply. Very high salt loads are leached from the novel biofilter material, expanded clay, yielding electric conductivity values of up to 15 mS cm(-1) in the leachate. Values > 6 mS cm(-1) were shown to depress methane consumption. Water retention characteristics of the material proved to be favourable for methane oxidizing systems with a gas permeable volume of 78% of bulk volume at field capacity water content. Correspondingly, no influence of water content on methane oxidation activity could be detected at water contents between 2.5 and 20 vol.-%.     

Investigation into the effect of high concentrations of volatile fatty acids in anaerobic digestion on methanogenic communities

A study was conducted to determine whether differences in the levels of volatile fatty acids (VFAs) in anaerobic digester plants could result in variations in the indigenous methanogenic communities. Two digesters (one operated under mesophilic conditions, the other under thermophilic conditions) were monitored, and sampled at points where VFA levels were high, as well as when VFA levels were low. Physical and chemical parameters were measured, and the methanogenic diversity was screened using the phylogenetic microarray ANAEROCHIP. In addition, real-time PCR was used to quantify the presence of the different methanogenic genera in the sludge samples. Array results indicated that the archaeal communities in the different reactors were stable, and that changes in the VFA levels of the anaerobic digesters did not greatly alter the dominating methanogenic organisms. In contrast, the two digesters were found to harbour different dominating methanogenic communities, which appeared to remain stable over time. Real-time PCR results were inline with those of microarray analysis indicating only minimal changes in methanogen numbers during periods of high VFAs, however, revealed a greater diversity in methanogens than found with the array.     

A dynamic simulation of methane fermentation process receiving heterogeneous food wastes and modelling acidic failure

Microbial response on volatile fatty acids (VFAs) is a key for methane fermentation processes since accumulation of VFAs often causes an acidic failure, especially treating such organics as food wastes composed of mostly readily biodegradable materials. To evaluate the impact of VFA accumulation, a lab-scale continuous experiment was performed for 110 days with sequential feeding of heterogeneous food wastes. When the volumetric loading rate was increased from 6 to 8 kg-COD/m³/day, a sudden decrease of methane production was observed with an accumulation of acetate and propionate in the fermenter. After discontinuation of feeding for 10 days, the digestate in the fermenter was centrifuged and washed with tap water to reduce the VFAs to be acceptable concentration below 1000 mg-COD/L. Nevertheless, no recovery of methane production was observed and VFA concentrations consistently increased. To model the event, a modification of ADM1 was made assuming the methanogens in the fermenter were irreversibly inactivated under very high VFA. Also considering the different nature of the fed food wastes over 11 samples, decomposition kinetics of individual food wastes were manipulated. The modified ADM1 with methanogenic activity decay reasonably reproduced the responses for soluble material concentrations and methane gas production rate over the experimental period.     

两相厌氧工艺处理高浓度丙烯酸生产废水

利用两相厌氧工艺处理高浓度丙烯酸生产废水。实验结果表明：在较高进水COD和容积负荷的条件下,系统具有良好、稳定的处理效果;在负荷提高及稳定运行阶段,将生活污水与丙烯酸生产废水的体积比调整为5∶1,容积负荷最大提高至12.3 kg/（m3·d）,两相厌氧反应器可长期稳定运行,总COD去除率基本维持在90%以上,出水COD小于323 mg/L;当进水甲醛质量浓度为800~1 733 mg/L时,总甲醛去除率基本稳定在95.6%~99.3%;在负荷提高及稳定运行阶段,水解酸化相反应器和产甲烷相反应器的出水pH分别为6.2~7.6和7.6~8.1,出水总碱度分别为1 220~1 820 mg/L和1 800~2 620 mg/L。     

ENUMERATION OF ANAEROBIC REFUSE-DECOMPOSING MICRO-ORGANISMS ON REFUSE CONSTITUENTS

Hydrolytic, acetogenic and methanogenic bacteria are required for the conversion of refuse to methane in landfills. In order to identify sources of these trophic groups in refuse, the total anaerobic population and the sub-populations of cellulolytic, hemicellulolytic, butyrate catabolizing acetogenic, and acetate- and H₂-CO₂-utilizing methanogenic bacteria as present on grass, leaves, branches, food waste, whole refuse and two landfill cover soils were enumerated by the most probable number (MPN) technique. Total anaerobes ranged from 10³cells per dry gram in cover soil to 10⁹in grass, food waste and fresh refuse. Hemicellulolytics ranged from 160 cells per dry gram in cover soil to 10⁹in grass. The highest cellulolytic population was measured on branches (316 cells per dry gram), while the maximum acetogenic population was measured on leaves (2.5×10⁴). The highest methanogen populations were measured on leaves (6.3×10³) and one of two fresh refuse samples (10⁵). Yard waste was the major carrier of the trophic groups required for refuse decomposition, while the cover soils tested did not represent major inputs of the requisite bacteria to landfills.     

Anaerobic biodegradation of cellulosic material: Batch experiments and modelling based on isotopic data and focusing on aceticlastic and non-aceticlastic methanogenesis

Utilizing stable carbon isotope data to account for aceticlastic and non-aceticlastic pathways of methane generation, a model was created to describe laboratory batch anaerobic decomposition of cellulosic materials (office paper and cardboard). The total organic and inorganic carbon concentrations, methane production volume, and methane and CO₂ partial pressure values were used for the model calibration and validation. According to the fluorescent in situ hybridization observations, three groups of methanogens including strictly hydrogenotrophic methanogens, strictly aceticlastic methanogens (Methanosaeta sp.) and Methanosarcina sp., consuming both acetate and H₂/H₂CO₃ as well as acetate-oxidizing syntrophs, were considered. It was shown that temporary inhibition of aceticlastic methanogens by non-ionized volatile fatty acids or acidic pH was responsible for two-step methane production from office paper at 35 °C where during the first and second steps methane was generated mostly from H₂/H₂CO₃ and acetate, respectively. Water saturated and unsaturated cases were tested. According to the model, at the intermediate moisture (150%), much lower methane production occurred because of full-time inhibition of aceticlastic methanogens. At the lowest moisture, methane production was very low because most likely hydrolysis was seriously inhibited. Simulations showed that during cardboard and office paper biodegradation at 55 °C, non-aceticlastic syntrophic oxidation by acetate-oxidizing syntrophs and hydrogenotrophic methanogens were the dominant methanogenic pathways.     

Archaeal community composition affects the function of anaerobic co-digesters in response to organic overload

Microbial community diversity in two thermophilic laboratory-scale and three full-scale anaerobic co-digesters was analysed by genetic profiling based on PCR-amplified partial 16S rRNA genes. In parallel operated laboratory reactors a stepwise increase of the organic loading rate (OLR) resulted in a decrease of methane production and an accumulation of volatile fatty acids (VFAs). However, almost threefold different OLRs were necessary to inhibit the gas production in the reactors. During stable reactor performance, no significant differences in the bacterial community structures were detected, except for in the archaeal communities. Sequencing of archaeal PCR products revealed a dominance of the acetoclastic methanogen Methanosarcina thermophila, while hydrogenotrophic methanogens were of minor importance and differed additionally in their abundance between reactors. As a consequence of the perturbation, changes in bacterial and archaeal populations were observed. After organic overload, hydrogenotrophic methanogens (Methanospirillum hungatei and Methanoculleus receptaculi) became more dominant, especially in the reactor attributed by a higher OLR capacity. In addition, aggregates composed of mineral and organic layers formed during organic overload and indicated tight spatial relationships between minerals and microbial processes that may support de-acidification processes in over-acidified sludge.Comparative analyses of mesophilic stationary phase full-scale reactors additionally indicated a correlation between the diversity of methanogens and the VFA concentration combined with the methane yield. This study demonstrates that the coexistence of two types of methanogens, i.e. hydrogenotrophic and acetoclastic methanogens is necessary to respond successfully to perturbation and leads to stable process performance.     

The effect of advanced oxidation processes on leachate biodegradation in recycling lysimeters.

Landfill processes were simulated in laboratory-scale bioreactors--lysimeters. The changes in leachate characteristics as well as the influence of advanced oxidation processes (AOPs) on the processes taking place in the sanitary landfill were investigated. Lysimeters were filled with material simulating municipal waste in the city of Lodz, Poland. Compost in the amount of 30% w/w and the methanogens inoculum were added in order to enhance development of a methanogenic phase. The leachate produced in lysimeters was recirculated. In order to investigate the influence of AOPs implementation on processes taking place in landfills two runs in lysimeters were performed, each lasting about 250 days. The leachate composition and biogas composition and production changes showed trends that confirmed that the bench-scale lysimeters appeared suitable to simulate processes taking place in the landfill. The application of AOPs to the leachate recirculated into the lysimeters did not bring about unequivocally positive effects. The ozonation of the leachate, implemented at the beginning of the methanogenic phase, caused slight acceleration (about 2 weeks) of the biodegradation, whereas employment of H2O2/UV led to the inhibition of anaerobic processes.     

Effect of the organic loading rate on the performance of anaerobic acidogenic fermentation of two-phase olive mill solid residue

A study of the effect of the organic loading rate (OLR) on the anaerobic acidogenic fermentation of two-phase olive mill solid residue (OMSR) derived from fruits with a low ripening index was carried out in a laboratory-scale completely stirred tank reactor at mesophilic temperature (35 degrees C). Eight experimental runs were carried out at OLRs of 3.2, 5.6, 7.4, 9.6, 11.0, 12.9, 14.0 and 15.1g T-COD/ld, which were equivalent to hydraulic retention times of 50.0, 28.8, 21.8, 16.9, 14.7, 12.4, 11.5 and 10.7d, respectively. The experimental results obtained demonstrated that the optimum value of OLR for the acidogenic fermentation process was 12.9 g T-COD/ld, for which a maximum production of acetic acid was achieved. It was found that inhibition of the process occurred at OLRs higher than 12.9 g T-COD/ld. This was characterized by a significant decrease in the acetic acid concentration in the effluent and an increase in the concentration of other volatile acids that may affect the methanogenic step. The process inhibition was also characterized by the plateau in the curves of the effluent substrate concentration versus the OLR applied. It was found that a first-order kinetics satisfactorily described the influence of non-acetic acid soluble organic matter concentration (S-COD( *)) on the rate of soluble organic matter conversion to acetic acid (R(S-COD)( *)), and the influence of acetic acid concentration (AcH) on the rate of acetic acid production (R(AcH)), while a potential equation type adequately described the influence of acetic acid concentration on the volumetric hydrogen production (R(ACH) ). The kinetic constant for soluble organic matter removal was 0.145 d(-1), while the constant for acetic acid formation was found to be 0.075 d(-1).     

Comparative performance between temperature-phased and conventional mesophilic two-phased processes in terms of anaerobically produced bioenergy from food waste.

Comparative evaluation of bioenergy production from food waste was carried out with both a temperature-phased and a conventional mesophilic two-phased process at different organic loading rates (OLRs). No methane was detected in the temperature-phased thermophilic-acidogenic fermenter at all the OLRs tested. However, a significant amount of methane content was detected in the conventional two-phased mesophilic-acidogenic fermenter, with increments depending on the organic loading rate [from 17% at 3 g VS L(-1) day(-1) to 25% at 8 g VS L(-1) day(-1) (VS, volatile solid)]. Acetate and butyrate were the main volatile fatty acids (VFAs) in the temperature-phased thermophilic-acidogenic fermenter; conversely propionate was a major VFA in the conventional two-phased mesophilic-acidogenic fermenter. Through the chemical oxygen demand (COD) balance of both temperature-phased and conventional mesophilic two-phased processes, the fraction of the feed-COD converted to the hydrogen-COD in the thermophilic-acidogenic fermenter within the former process was estimated from 7.9 to 9.3%, with a peak at ORL of 6 g VS L(-1) day(-1), whereas it was quantified from 0.3 to 0.9% in the mesophilic-acidogenic fermenter within the latter one. Moreover, the fraction of the feed-COD converted to the methane-COD in the mesophilic-acidogenic fermenter within the conventional two-phased process ranged from 5.4 to 7.9%. On the other hand, conversion of the feed-COD to the methane-COD in the mesophilic-methanogenic fermenter of both temperature-phased and conventional mesophilic two-phased processes ranged from 66.2 to 72.3% and from 63.5 to 70.5%, respectively, with decrements related to the increase of organic loading rate.