Visualization of active biomass distribution in a BGAC fluidized bed reactor using GFP tagged Pseudomonas putida F1

A favorable microenvironment for biofilm growth on GAC particles was shown using green fluorescent protein (GFP) as a marker for a phenol degrading bacterium, Pseudomonas putida F1. The dispersion of P. putida F1 in a biofilm covering granulated activated carbon (GAC) particles was monitored and compared to a biofilm on non-activated granular carbon particles. Laser scanning confocal microscopy (LSCM) micrographs of the biofilms taken from two fluidized bed reactors operating under identical conditions, showed higher fluorescent green areas in the GAC biofilm, especially close to the GAC surface. Quantitative analysis of the biofilm by COMSTAT, a three-dimensional biofilm structure analysis program, showed higher biomass concentration and higher viability in the GAC covered biofilm vs. the non-activated carbon biofilm. In addition, better effluent quality was measured for the BGAC reactor, which strongly suggests a significantly larger biofilm surface area available to the substrate, as opposed to that of the non-activated carbon carrier reactor.     

Mixing in upflow anaerobic filters and its influence on performance and scale-up

Tracer studies were carried out in laboratory-scale and pilot-scale upflow anaerobic filters to determine the effect of liquid velocity, gas production and media depth on mixing patterns. A computer simulation model was developed to analyse tracer-response curves. In water studies at laboratory scale, gas production was shown to have a significantly greater effect on mixing than liquid upflow velocity. A reduction in the quantity of media also resulted in greater mixing due to the greater void space in which synthetic gas bubbles could cause turbulence. In the presence of sludge during reactor operation, at pilot and laboratory-scale, gas production had a significant influence on mixing. However, liquid velocity played an important role in solids distribution in the filter, in conjunction with media depth. At pilot-scale, at a low solids concentration, a high liquid velocity lifted the sludge “bed”, raising the source of gas production. The absence of gas below the sludge bed resulted in a plug flow regime which the incoming substrate entered. A reduction in the quantity of media increased the degree of mixing for a given liquid velocity and gas surface load. Lower liquid upflow velocities are required at a reduced media depth to prevent excessive biomass loss. Shear rates increase at high liquid and gas velocities, resulting in detachment of solids from the media and biomass washout. A close correlation was established between mixing and process performance which led to the development of a programme for start-up and operation of the filter to maintain optimum biomass/substrate contact. A strategy for scale-up was proposed through the development of correlations obtained from laboratory-scale filter studies which were used to predict pilot-scale mixing characteristics. This research highlighted the important factors influencing mixing patterns and scale-up in anaerobic upflow filters.     

低浓度污水厌氧污泥颗粒化促进技术研究

向低浓度生活污水中投中絮凝剂（壳聚糖）和颗粒活性炭（GAC）的试验结果表明，这两种物质对厌氧污泥的颗粒化均有促进作用，但不如中、高浓度污水中明显。二者相比，GAC的效果更好一些（反应器的运行稳定，耐冲击能力强，出水水质良好），各反应器中形成的颗粒污泥均以产甲烷丝状菌为主。     

Effects of increase modes of shear force on granule disruption in upflow anaerobic reactors

Sludge washout is listed among the top practical problems of the high rate upflow anaerobic reactors. This study investigated quantitatively two sludge washout processes operated under different hydrodynamic shear increase modes with the intervals of 1 and 10 days respectively. The results reveal that the sludge washout accompanying with large-scale granule disruption could lead to performance failure with heavy sludge loss ratio of about 46.1% at sludge loss rate about 0.35 gVSS L⁻¹ d⁻¹ during the process with shear increase interval of 1 day, while the highest sludge loss rate was only 0.12 gVSS L⁻¹ d⁻¹ during the process with 10-day interval. The intensified shear conditions could weaken the granules through inhibiting the extracellular polymers production and bioactivity. As consequences, an outbreak of large-scale granule disruption would raise and then significantly accelerate the sludge washout. Since long interval could provide the granules the opportunity to recover from these negative effects to some extent, the shear increase strategy of long interval over 10 days is favorably recommended to operate full-scale reactors during the start-up and shock load periods. The pioneer use of the micro particle image velocimetry in this study offers the possibility to discover the real hydrodynamic conditions around granules at microscale for the first time and reveals that the shear force exerts directly on the granular surface as a mechanical disruption force and big granules undergo high disruption force. The granule disruption is a result of the competition between the granule and the ambient hydrodynamic shear conditions rather than a process with shear force as a sole dominant factor. These could facilitate the understanding of the complicated interactions between the hydrodynamics and reactor performance and favor then a better control of the full-scale reactors.     

Variation of bulk properties of anaerobic granules with wastewater type

Development of a granular sludge with high strength, high biological activity and a narrow settling distribution is necessary for optimal operation of high-rate upflow anaerobic treatment systems. Several studies have compared granules produced from different wastewaters but these have largely been from laboratory-fed reactors or compared granules from full-scale reactors fed similar wastewater types. Though two authors have commented on the inferiority of granules produced by a protein-based feed, the properties of these granules have not been characterised. In this paper, granules from full-scale reactors treating fruit and vegetable cannery effluent, two brewery effluents and a pig abattoir (slaughterhouse) were compared in terms of basic composition, size distribution, density, settling velocity, shear strength, and EPS content. The results supported previous qualitative observations by other researchers that indicate granule properties depend more on wastewater type rather than reactor design or operating conditions such as pre-acidification level. The cannery-fed granules had excellent shear strength, settling distribution and density. Granules from the two brewery-fed reactors had statistically the same bulk properties, which were still acceptable for upflow applications. The protein-grown granule had poor strength and settling velocity.     

Effect of settling on performance of the upflow anaerobic sludge bed reactors

The effects of settler volume on the start-up and steady-state performance of 41. laboratory upflow sludge bed reactors treating bean blanching waste of 10,000 mg COD l⁻¹ were determined. The rate of start-up, as well as the maximum loading rate, increased with increased settler volume and performance. A loading rate of 30 kg COD m⁻³ day⁻¹ (based on reactor volume alone) and a COD removal of 95% was obtained with a 21. settling flask and a 4 to 1 recirculation rate. Without a settler, the maximum loading rate was 10 kg COD m⁻³ day⁻¹. The sludge was flocculent rather than granular. Sludge profiles and characteristics in the reactors and settlers were determined.     

Oxygen transfer and uptake, nutrient removal, and energy footprint of parallel full-scale IFAS and activated sludge processes

Integrated fixed-film activated sludge (IFAS) processes are becoming more popular for both secondary and sidestream treatment in wastewater facilities. These processes are a combination of biofilm reactors and activated sludge processes, achieved by introducing and retaining biofilm carrier media in activated sludge reactors. A full-scale train of three IFAS reactors equipped with AnoxKaldnes media and coarse-bubble aeration was tested using off-gas analysis. This was operated independently in parallel to an existing full-scale activated sludge process. Both processes achieved the same percent removal of COD and ammonia, despite the double oxygen demand on the IFAS reactors. In order to prevent kinetic limitations associated with DO diffusional gradients through the IFAS biofilm, this systems was operated at an elevated dissolved oxygen concentration, in line with the manufacturer’s recommendation. Also, to avoid media coalescence on the reactor surface and promote biofilm contact with the substrate, high mixing requirements are specified. Therefore, the air flux in the IFAS reactors was much higher than that of the parallel activated sludge reactors. However, the standardized oxygen transfer efficiency in process water was almost same for both processes. In theory, when the oxygen transfer efficiency is the same, the air used per unit load removed should be the same. However, due to the high DO and mixing requirements, the IFAS reactors were characterized by elevated air flux and air use per unit load treated. This directly reflected in the relative energy footprint for aeration, which in this case was much higher for the IFAS system than activated sludge.     

Chemisorption of oxygen onto activated carbon can enhance the stability of biological perchlorate reduction in fixed bed biofilm reactors

Fixed bed biofilm reactors with granular activated carbon (GAC) or glass beads as support media were used to evaluate the influence of short-term (12h) and long-term (23 days) increases of influent dissolved oxygen (DO) concentrations on biological perchlorate removal. The goal was to evaluate the extent by which chemisorption of oxygen to GAC can enhance the stability of biological perchlorate reduction. Baseline influent concentrations were 50 microg/L of perchlorate, 2 mg/L of acetate as C, and 1mg/L of DO. Perchlorate removal in the glass bead reactor seized immediately after increasing influent DO concentrations from 1 to 4 mg/L since glass beads have no sorptive capacity. In the biologically active carbon (BAC) reactor, chemisorption of oxygen to GAC removed a substantial fraction of the influent DO, and perchlorate removal was maintained during short-term increases of influent DO levels up to 8 mg/L. During long-term exposure to influent DO concentrations of 8.5mg/L, effluent perchlorate and DO concentrations increased slowly. Subsequent exposure of the BAC reactor bed to low DO concentrations partially regenerated the capacity for oxygen chemisorption. Microbial analyses indicated similar microbial communities in both reactors, which confirmed that the differences in reactor performance during dynamic loading conditions could be attributed to the sorptive properties of GAC. Using a sorptive biofilm support medium can enhance biological perchlorate removal under dynamic loading conditions.     

Biodegradation of 2-fluorobenzoate and dichloromethane under simultaneous and sequential alternating pollutant feeding

Two up-flow fixed-bed reactors (UFBRs), inoculated with activated sludge and operated for 162 days, were fed 1mmolL(-1)d(-1) with two model halogenated compounds, 2-fluorobenzoate (2-FB) and dichloromethane (DCM). Expanded clay (EC) and granular activated carbon (GAC) were used as biofilm carrier. EC did not have any adsorption capacity for both model compounds tested, whereas GAC could adsorb 1.3mmolg(-1) GAC for 2-FB and 4.5mmolg(-1) GAC for DCM. Both pollutants were degraded in both reactors under simultaneous feeding. However, biodegradation in the EC reactor was more pronounced, and re-inoculation of the GAC reactor was required to initiate 2-FB degradation. Imposing sequential alternating pollutant (SAP) feeding caused starvation periods in the EC reactor, requiring time-consuming recovery of 2-FB biodegradation after resuming its feeding, whereas DCM degradation recovered significantly faster. The SAP feeding did not affect performance in the GAC reactor as biodegradation of both pollutants was continuously observed during SAP feeding, indicating the absence of true starvation.     

Competition between planktonic and fixed microorganisms during the start-up of methanogenic biofilm reactors

The influence of the hydraulic retention time (HRT) on the start-up phase of a methanogenic inverse turbulent bed bioreactor was investigated. Two identical reactors were monitored, the only differing parameter being the HRT: one of the reactors was fed with a diluted wastewater at a constant HRT of 1 day, the organic loading rate (OLR) being increased by decreasing the substrate dilution; the second reactor was fed at a constant influent concentration of 20 g COD L(-1), the OLR being increased by decreasing the HRT from 40 days to 1 day. After 45 days of start-up, both reactors were operated at an OLR of 20 g COD L(-1)d(-1) and a HRT of 1 day. However, strong differences were observed on biofilm growth. In the reactor operated at a constant short HRT, biofilm concentration was 4.5 as high as in the reactor operated at an increasing HRT. This difference was attributed to the competition between planktonic and biofilm microorganisms in the latter reactor, whereas suspended biomass was quickly washed out in the former reactor because of the low HRT.     

Granulation in high-load denitrifying upflow sludge bed (USB) pulsed reactors

In this work, the effect of the application of a pulse system to anoxic upflow sludge bed (USB) denitrifying reactors for enhancing sludge granulation was studied. In all, three 0.8 L reactors (two operated with flow pulsation, P1 with effluent recycling and P2 without recycling, and one without pulsation and effluent recycling, no pulsation (NP)) were fed with a mixture of NaNO3 and glucose and inoculated with methanogenic granular sludge. The organic loading rate (OLR) and the nitrogen loading rate (NLR) were progressively increased and, at the end of the experiment, extremely high values were obtained (67.5 kgCOD/m3d and 11.25 kgN-NO3-/m3 d). Ammonia and nitrite accumulation in reactor NP were important in the maturation stage, decreasing the denitrification efficiency to 90%, while in reactor P1 only low nitrite values were obtained in the last few days of the experiment. In reactor P2, nitrogen removal was 100% most of the time. Several operational problems (flotation and the subsequent wash out of biomass) appeared in the NP reactor when working at high denitrifying loading rates, while in reactors P1 and P2 there were no notable problems, mainly due to the good characteristics of the sludge developed and the efficient degasification produced by the pulsing flow. The sludge formed in the NP reactor presented a flocculent structure and a total disintegration of the initial methanogenic granules occurred, while a small-sized granular biomass with a high specific density was developed in the pulsed reactors due to the shear stress produced.     

Treatment of a colored groundwater by ozone-biofiltration: pilot studies and modeling interpretation

Pilot studies investigated the fates of color, dissolved organic carbon (DOC), and biodegradable organic matter (BOM) by the tandem of ozone plus biofiltration for treating a source water having significant color (50 cu) and DOC (3.2 mg/l). Transferred ozone doses were from 1.0 to 1.8 g O3/g C. Rapid biofilters used sand, anthracite, or granular activated carbon as media with empty-bed contact time (EBCT) up to 9 min. The pilot studies demonstrated that ozonation plus biofiltration removed most color and substantial DOC, and increasing the transferred ozone dose enhanced the removals. For the highest ozone dose, removals were as high as 90% for color and 38% for DOC. While most of the color removal took place during ozonation, most DOC removal occurred in the biofilters, particularly when the ozone dose was high. Compared to sand and anthracite biofilters, the GAC biofilter gave the best performance for color and DOC removal, but some of this enhanced performance was caused by adsorption, since the GAC was virgin at the beginning of the pilot studies. Backwashing events had no noticeable impact of the performance of the biofilters. The Transient-State, Multiple-Species Biofilm Model (TSMSBM) was used to interpret the experimental results. Model simulations show that soluble microbial products, which comprised a significant part of the effluent BOM, offset the removal of original BOM, a factor that kept the removal of DOC relatively constant over the range of EBCTs of 3.5-9 min. Although improved biofilm retention, represented by a small detachment rate, allowed more total biofilm accumulation and greater removal of original BOM, it also caused more release of soluble microbial products and the build up of inert biomass in the biofilm. Backwashing had little impact on biofilter performance, because it did not remove more than 25% of the biofilm under any condition simulated.     

Influence of hydrodynamic conditions on the start-up of methanogenic inverse turbulent bed reactors

The influence of hydrodynamic conditions on the start-up phase of an inverse turbulent bed bioreactor was investigated. Two identical reactors, differing only by the gas velocity ensuring the carrier fluidization and generating the main hydrodynamic strengths (attrition), were monitored. Regarding the first 96 days, the experiments showed better performances for the reactor having the lowest hydrodynamic strengths. These results were correlated to lower attached biomass and higher biofilm specific removal rate for the reactor subjected to strong hydrodynamic strengths. Once the start-up was completed and the reactor stabilized at an organic loading rate of 6g(COD)L(-1)d(-1), the same hydrodynamic strengths were applied (gas velocity was equalized). The results proved that hydrodynamic conditions make it possible to control the biofilm characteristics. Consequently, it should be recommended to apply minimal strengths to enhance the biofilm growth during the early start-up phase, and then to increase them to control biofilm thickness and to optimize the reactor performances.     

Methane yield as a monitoring parameter for the start-up of anaerobic fixed film reactors

This paper describes the variation of the methane yield during the start-up period of an anaerobic fluidized bed reactor. After a lag phase, with acclimatized sludge, the methane yield increased with time during biofilm development up to the theoretical steady yield value, reported to be around 0.351 CH4/g CODdeg. The establishment of the biofilm required a high consumption of organic material through the microbial synthesis (anabolism), thereby reducing the proportion of substrate converted to methane. As a result, this yield could be an indirect metabolic parameter for evaluating a start-up operation. It could provide vital information about bacterial fixation processes and is easy to be applied to any biofilm reactor, such as anaerobic filters, where biomass sampling is impracticable. Monitoring this parameter could also give useful dynamic information about the different steps of colonization and biomass attachment, which could be used to improve the start-up performance.     

Control of start-up and operation of anaerobic biofilm reactors: An overview of 15 years of research

Anaerobic biofilm reactors have to be operated in a way that optimizes on one hand the start-up period by a quick growth of an active biofilm, on the other hand the regular operation by an active control of the biofilm to avoid diffusion limitations and clogging. This article is an overview of the research carried out at INRA-LBE for the last 15 years. The start-up of anaerobic biofilm reactors may be considerably shortened by applying a short inoculation period (i.e. contact between the inoculum and the support media). Then, the increase of the organic loading rate should be operated at a short hydraulic retention time and low hydrodynamic constraints in order to favor biofilm growth. After the start-up period, biofilm growth should be controlled to maintain a high specific activity and prevent clogging. This can be done in particulate biofilm systems by using hydrodynamics to increase or decrease shear forces and attrition but is much more difficult in anaerobic fixed bed reactors.     

Kinetic model of autotrophic denitrification in sulphur packed-bed reactors

Autotrophic denitrification of synthetic wastewater by Thiobacillus denitrificans in upflow sulphur packed-bed reactors was studied in order to establish the process kinetics for prediction of effluent concentration. Elemental sulphur particles of different size served as energy substrate as well as the physical support for the microbial biofilm. Experiments were performed under operating conditions of (i) different flow rates at constant influent nitrate concentration; and (ii) different influent nitrate concentrations at constant flow rate. The experimental results show that autotrophic denitrification rates in upflow sulphur packed-bed reactors can be described by a half-order kinetic model for biofilms. It was found that the half-order kinetic constants of upflow packed-bed reactors are 2.94-3.60, 1.47-2.04, and 1.12-1.29 mg1/2/L1/2 h for sulphur particle sizes of 2.8-5.6, 5.6-11.2, and 11.2-16 mm, respectively. The half-order kinetic constants could be related to the specific surface area of the reactor media by a simple equation. Successful application of the half-order reaction rate model was demonstrated for an actual wastewater (nitrified leachate). A comparison with the literature showed that the half-order reaction rate constants for autotrophic denitrification using elemental sulphur are approximately one order of magnitude lower than those of heterotrophic denitrification. An improved stoichiometric equation for autotrophic denitrification using elemental sulphur as electronic donor is also proposed.     

The roles of calcium in sludge granulation during UASB reactor start-up

Six upflow anaerobic sludge blanket (UASB) reactors were concurrently operated for 146 d to examine the effects of calcium on the sludge granulation process during start-up. Introduction of Ca2+ at concentrations from 150 to 300 mg/l enhanced the biomass accumulation and granulation process. The calcium concentration in the granules was nearly proportional to the calcium concentration in the feed, and calcium carbonate was the main calcium precipitate in the granules. The specific activity of granules decreased with increasing influent calcium concentration. The optimum calcium concentration for the granulation was from 150 to 300 mg/l. The addition of low-concentration calcium to the UASB reactors appeared to enhance the three steps of sludge granulation: adsorption, adhesion and multiplication, but it did not lead to a different proliferation of predominant microorganisms in the granules.     

DEGRADATION OF AROCLOR 1242 IN A SINGLE-STAGE COUPLED ANAEROBIC/AEROBIC BIOREACTOR

Degradation of Aroclor 1242 was studied in granular biofilm reactors with limited aeration. An aerobic biphenyl degrader, Rhodococcus sp. M5, was used to supplement a natural bacterial population present in a “bioaugmented” reactor, while the “non-bioaugmented” reactor only contained natural granular sludge. The bioaugmentation, however appeared to have no effect on the reactor performance. Aroclor measurements showed its disappearance in both reactors with only 16–19% of Aroclor recovered from the reactor biomass and effluent. Simultaneously, a chlorine balance indicated that dechlorination occurred at a specific rate of 1.43 mg PCB (g volatile suspended solids)⁻¹ d⁻¹, which was comparable to the observed rate of Aroclor disappearance. Intermediates detected in both reactors were biphenyl, benzoic acid, and mono-hydroxybiphenyls. This suggests that a near-complete mineralization of Aroclor can be achieved in a single-stage anaerobic/aerobic system due to a combination of reductive and oxidative degradation mechanisms.     

Evaluation of the impact of bioaugmentation and biostimulation by in situ hybridization and microelectrode

Three rotating disk biofilm reactors were operated to evaluate whether bioaugmentation and biostimulation can be used to improve the start-up of microbial nitrification. The first reactor was bioaugmented during start-up period with an enrichment culture of nitrifying bacteria, the second reactor received a synthetic medium containing NH(4)(+) and NO(2)(-) to facilitate concomitant proliferation of ammonia- and nitrite-oxidizing bacteria, and the third reactor was used as a control. To evaluate the effectiveness of bioaugmentation and biostimulation approaches, time-dependent developments of nitrifying bacterial community and in situ nitrifying activity in biofilms were monitored by fluorescence in situ hybridization (FISH) technique and microelectrode measurements of NH(4)(+), NO(2)(-), NO(3)(-), and O(2). In situ hybridization results revealed that addition of the enrichment culture of nitrifying bacteria significantly facilitated development of dense nitrifying bacterial populations in the biofilm shortly after, which led to a rapid start-up and enhancement of in situ nitrification activity. The inoculated bacteria could proliferate and/or survive in the biofilm. In addition, the addition of nitrifying bacteria increased the abundance of nitrifying bacteria in the surface of the biofilm, resulting in the higher nitrification rate. On the other hand, the addition of 2.1mM NO(2)(-) did not stimulate the growth of nitrite-oxidizing bacteria and did inhibit the proliferation of ammonia-oxidizing bacteria instead. Thus, the start-up of NO(2)(-) oxidation was unchanged, and the start-up of NH(4)(+) oxidation was delayed. In all the three biofilm reactors, data sets of time series analyses on population dynamics of nitrifying bacteria determined by FISH, in situ nitrifying activities determined by microelectrode measurements, and the reactor performances revealed an approximate agreement between the appearance of nitrifying bacteria and the initiation of nitrification activity, suggesting that the combination of these techniques was a very powerful monitoring tool to evaluate the effectiveness of bioaugmentation and biostimulation strategies.