Abundance, Diversity, and Dynamics of Viruses on Microorganisms in Activated Sludge Processes

We examined the abundance of viruses on microorganisms in activated sludge and the dynamics of their community structure. Direct counting with epifluorescence microscopy and pulsed-field gel electrophoresis (PFGE) were applied to 20 samples from 14 full-scale wastewater treatment plants (wwtps) treating municipal, industrial, or animal wastewater. Furthermore, to observe the dynamics of viral community structure over time, a laboratory-scale sequencing batch reactor was operated for 58 days. The concentrations of virus particles in the wwtps, as quantified by epifluorescence microscopy, ranged from 4.2 × 10⁷ to 3.0 × 10⁹ mL⁻¹. PFGE, improved by the introduction of a higher concentration of Tris–EDTA buffer in the DNA extraction step, was successfully used to profile DNA viruses in the activated sludge. Most of the samples from different wwtps commonly had bands in the 40–70 kb range. In the monitoring of viral DNA size distribution in the laboratory-scale reactor, some bands were observed stably throughout the experimental period, some emerged during the operation, and others disappeared. Rapid emergence and disappearance of two intense bands within 6 days was observed. Our data suggest that viruses—especially those associated with microorganisms—are abundant and show dynamic behavior in activated sludge.     

A study on using fireclay as a biomass carrier in an activated sludge system

By adding a biomass carrier to an activated sludge system, the biomass concentration will increase, and subsequently the organic removal efficiency will be enhanced. In this study, the possibility of using excess sludge from ceramic and tile manufacturing plants as a biomass carrier was investigated. The aim of this study was to determine the effect of using fireclay as a biomass carrier on biomass concentration, organic removal and nitrification efficiency in an activated sludge system. Experiments were conducted by using a bench scale activated sludge system operating in batch and continuous modes. Artificial simulated wastewater was made by using recirculated water in a ceramic manufactutring plant. In the continuous mode, hydraulic detention time in the aeration reactor was 8 and 22 h. In the batch mode, aeration time was 8 and 16 h. Fireclay doses were 500, 1,400 and 2,250 mg l⁻¹, and were added to the reactors in each experiment separately. The reactor with added fireclay was called a Hybrid Biological Reactor (HBR). A reactor without added fireclay was used as a control. Efficiency parameters such as COD, MLVSS and nitrate were measured in the control and HBR reactors according to standard methods. The average concentration of biomass in the HBR reactor was greater than in the control reactor. The total biomass concentration in the HBR reactor (2.25 g l⁻¹ fireclay) in the continuous mode was 3,000 mg l⁻¹ and in the batch mode was 2,400 mg l⁻¹. The attached biomass concentration in the HBR reactor (2.25 g l⁻¹ fireclay) in the continuous mode was 1,500 mg l⁻¹ and in the batch mode was 980 mg l⁻¹. Efficiency for COD removal in the HBR and control reactor was 95 and 55%, respectively. In the HBR reactor, nitrification was enhanced, and the concentration of nitrate was increased by 80%. By increasing the fireclay dose, total and attached biomass was increased. By adding fireclay as a biomass carrier, the efficiency of an activated sludge system to treat wastewater from ceramic manufacturing plants was increased.     

Degrading high-strength phenol using aerobic granular sludge

Aerobic granules were adopted to degrade high-strength phenol wastewater in batch experiments. The acclimated granules effectively degraded phenol at a concentration of up to 5,000 mg l⁻¹ without severe inhibitory effects. The biodegradation of phenol by activated sludge was inhibited at phenol concentrations >3,000 mg l⁻¹. The granules were composed of cells embedded in a compact extracellular matrix. After acid or alkaline pretreatment, the granules continued to degrade phenol at an acceptable rate. The polymerase chain reaction-denaturing gradient gel electrophoresis technique was employed to monitor the microbial communities of the activated sludge and the aerobic granules following their being used to treat high concentrations of phenol in batch tests.     

Reclamation of an activated-sludge microbial consortium by selective biostimulation

Our previous study showed that an activated-sludge process broke down at the phenol-loading rate of 1.5 g l⁻¹ day⁻¹, when non-flocculating bacteria (called R6T and R10) overgrew the sludge, resulting in a sludge washout. In this study, we attempted to circumvent this breakdown problem by reclaiming the consortium structure. Activated sludge was fed phenol, and the phenol-loading rate was increased stepwise from 0.5 g l⁻¹ day⁻¹ to 1.0 g l⁻¹ day⁻¹ and then to 1.5 g l⁻¹ day⁻¹. Either galactose or glucose (at 0.5 g l⁻¹ day⁻¹) was also supplied to the activated sludge from the phenol-loading rate of 1.0 g l⁻¹ day⁻¹. Pure culture experiments have suggested galactose to be a preferential substrate for a floc-forming bacterium (R6F) that predominantly degrades phenol under low phenol-loading conditions. Supplying galactose allowed sustainment of the R6F population and suppression of the overgrowth of R6T and R10 at the phenol-loading rate of 1.5 g l⁻¹ day⁻¹. This measure allowed the activated-sludge process to treat phenol at a phenol-loading rate up to 1.5 g l⁻¹ day⁻¹, although it broke down at 2.0 g l⁻¹ day⁻¹. In contrast, supplying glucose reduced the R6F population and allowed the activated-sludge process to break down at the phenol-loading rate of 1.0 g l⁻¹ day⁻¹. This study demonstrated that reclamation of the activated-sludge consortium by selective biostimulation of the floc-forming population improved the phenol-treating ability of the process. Received: 13 January 2000 / Received revision: 10 March 2000 / Accepted: 7 April 2000     

Degradation of cyanide in agroindustrial or industrial wastewater in an acidification reactor or in a single-step methane reactor by bacteria enriched from soil and peels of cassava

During cassava starch production, large amounts of cyanoglycosides were released and hydrolysed by plant-borne enzymes, leading to cyanide concentrations in the wastewater as high as 200 mg/l. For anaerobic degradation of the cyanide during pre-acidification or single-step methane fermentation, anaerobic cultures were enriched from soil residues of cassava roots and sewage sludge. In a pre-acidification reactor this culture was able to remove up to 4 g potassium cyanide/l of wastewater at a hydraulic retention time (t _HR) of 4 days, equivalent to a maximal cyanide space loading of 400 mg CN⁻ l⁻¹ day⁻¹. The residual cyanide concentration was 0.2–0.5 mg/l. Concentrated cell suspensions of the mixed culture formed ammonia and formate in almost equimolar amounts from cyanide. Little formamide was generated by chemical decay. A concentration of up to 100 mmol ammonia/l had no inhibitory effect on cyanide degradation. The optimal pH for cyanide degradation was 6–7.5, the optimal temperature 25–37 °C. At a pH of 5 or lower, cyanide accumulated in the reactor and pre-acidification failed. The minimal t _HR for continuous cyanide removal was 1.5 days. The enriched mixed culture was also able to degrade cyanide in purely mineralic wastewater from metal deburring, either in a pre-acidification reactor with a two-step process or in a one-step methanogenic reactor. It was necessary to supplement the wastewater with a carbon source (e.g. starch) to keep the population active enough to cope with any possible inhibiting effect of cyanide. Received: 29 April 1998 / Received revision: 8 June 1998 / Accepted: 14 June 1998     

Degradation of 2,4,5-trichlorophenol and 2,3,5,6-tetrachlorophenol by combining pulse electric discharge with bioremediation

Degradation of 2,4,5-trichlorophenol (2,4,5-TCP) and 2,3,5,6-tetrachlorophenol (TeCP) was studied using a two-stage approach that utilized efficient pulse electric discharge (PED) followed by biological degradation with a consortium from acclimated return activated sludge. The chlorinated phenols were treated in the PED reactor as an aerosol at a voltage of 55–60 kV, a frequency of 385 Hz, a current of 50–60, and with a 200-ns pulse. As determined by gas chromatography and mass spectrometry (GC/MS), the first stage converted 500 ppm 2,4,5-TCP to 163 ppm 2,4,5-TCP and dimethyldecene, dichloronaphthalenol, octyl acetate, and silyl esters. The total carbon content of 2,4,5-TCP after PED treatment was determined to be 228 ± 35 ppm. The remaining 2,4,5-TCP and the products formed were then mineralized by the acclimated activated sludge in shake flasks; the initial rate of degradation of 2,4,5-TCP was calculated to be 5 nmol min⁻¹ mg protein⁻¹ at 163 ppm initial concentration (three orders of magnitude higher than the only rate found in the literature). By combining the two techniques, a synergistic effect (2.3-fold increase in the concentration of 2,4,5-TCP degraded and 3.3-fold increase in total carbon degraded) was observed, in that bacteria without any treatment degraded a maximum of 70 ppm 2,4,5-TCP but after PED treatment 163 ppm 2,4,5-TCP was degraded. TeCP was also mineralized by the acclimated activated sludge after treatment with PED. This two-stage approach was also evaluated using a continuous 1-l fluidized-bed reactor. Received: 3 November 1998 / Received revision: 28 February 1999 / Accepted: 14 March 1999     

Treatment of cyanide-containing wastewater from the food industry in a laboratory-scale fixed-bed methanogenic reactor

During the process of producing cassava starch from Manihot esculenta roots, large amounts of cyanoglycosides were released, which rapidly decayed to CN⁻ following enzymatic hydrolysis. Depending on the varying cyanoglycoside content of the cassava varieties, the cyanide concentration in the wastewater was as high as 200 mg/l. To simulate anaerobic stabilization, a wastewater with a chemical oxygen demand (COD) of about 20 g/l was prepared from cassava roots and was fermented in a fixed-bed methanogenic reactor. The start-up phase for a 99% degradation of low concentrations of cyanide (10 mg/l) required about 6 months. After establishment of the biofilm, a cyanide concentration of up to 150 mg CN⁻/l in the fresh wastewater was degraded during anaerobic treatment at a hydraulic retention time of 3 days. All nitrogen from the degraded cyanide was converted to organic nitrogen by the biomass of the effluent. The cyanide-degrading biocoenosis of the anaerobic reactor could tolerate shock concentrations of cyanide up to 240 mg CN⁻/l for a short time. Up to 5 mmol/l NH₄Cl (i.e. 70 mg N/l = 265 mg NH₄Cl/l) in the fresh wastewater did not affect cyanide degradation. The bleaching agent sulphite, however, had a negative effect on COD and cyanide removal. For anaerobic treatment, the maximum COD space loading was 12 g l⁻¹ day⁻¹, equivalent to a hydraulic retention time of 1.8 days. The COD removal efficiency was around 90%. The maximum permanent cyanide space loading was 50 mg CN⁻ l⁻¹ day⁻¹, with tolerable shock loadings up to 75 mg CN⁻ l⁻¹day⁻¹. Under steady-state conditions, the cyanide concentration of the effluent was lower than 0.5 mg/l. Received: 15 August 1997 / Received revision: 10 October 1997 / Accepted: 14 October 1997     

Change in ammonia-oxidizing microorganisms in enriched nitrifying activated sludge

In this study, sludge was taken from a municipal wastewater treatment plant that contained a nearly equal number of archaeal amoA genes (5.70 × 10⁶ ± 3.30 × 10⁵ copies mg sludge⁻¹) to bacterial amoA genes (8.60 × 10⁶ ± 7.64 × 10⁵ copies mg sludge⁻¹) and enriched in three continuous-flow reactors receiving an inorganic medium containing different ammonium concentrations: 2, 10, and 30 mM NH₄⁺–N (28, 140, and 420 mg N l⁻¹). The abundance and communities of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in enriched nitrifying activated sludge (NAS) were monitored at days 60 and 360 of the operation. Early on, between day 0 and day 60 of reactor operation, comparative abundance of AOA amoA genes to AOB amoA genes varied among the reactors depending on the ammonium levels found in the reactors. As compared to the seed sludge, the number of AOA amoA genes was unchanged in the reactor with lower ammonium level (0.06 ± 0.04 mgN l⁻¹), while in the reactors with higher ammonium levels (0.51 ± 0.33 and 0.25 ± 0.10 mgN l⁻¹), the numbers of AOA amoA genes were deteriorated. By day 360, AOA disappeared from the ammonia-oxidizing consortiums in all reactors. The majority of the AOA sequences from all NASs at each sampling period fell into a single AOA cluster, however, suggesting that the ammonium did not affect the AOA communities under this operational condition. This result is contradictory to the case of AOB, where the communities varied significantly among the NASs. AOB with a high affinity for ammonia were present in the reactors with lower ammonium levels, whereas AOB with a low affinity to ammonia existed in the reactors with higher ammonium levels.     

Towards a reduction in excess sludge production in activated sludge processes: biomass physicochemical treatment and biodegradation

To decrease activated sludge production, microbial cell lysis can be amplified to enhance cryptic growth (biomass growth on lysates). Cell breakage techniques (thermal, alkaline, acid) were studied to generate Alcaligenes eutrophus and sludge lysates and to evaluate their biodegradability. Gentle treatment conditions produced the best results. Complete cell deactivation was obtained for temperatures higher than 55 °C. The release kinetics were similar for temperatures varying from 60 °C to 100 °C. A 20-min incubation was suitable for reaching 80% of the maximum releasable carbon. In thermal-chemical hydrolysis, NaOH was the most efficient for inducing cell lysis. Carbon release was a two-step process. First an immediate release occurred, which was of the same order of magnitude for A. eutrophus and sludge [100–200 mg dissolved organic C (DOC) g total suspended solids (TSS)⁻¹], followed by a post-treatment release. The second step was virtually equivalent to the first for sludge, and weaker for A. eutrophus (<50 mg DOC g TSS⁻¹). The biodegradability of the soluble fraction, both the immediate and the post-treatment carbon release, was investigated. The optimal degradation yield, obtained with sludge cells, reached 55% after 48 h of incubation and 80% after 350 h. The most consistent lysis and biodegradation results occurred at pH 10 and 60 °C after a 20-min incubation. Received: 30 October 1998 / Received revision: 16 February 1999 / Accepted: 20 February 1999     

Effect of feed composition and upflow velocity on aggregate characteristics in anaerobic upflow reactors

Two upflow anaerobic hybrid reactors treated lactose and a mixture of ethanol, propionate and butyrate, respectively, at a volumetric loading rate of 3.7 kg chemical oxygen demand (COD) m⁻³day⁻¹, a hydraulic retention time of 5 days and a liquid upflow velocity of 0.01 m/h. Under steady-state conditions, the lactose-fed sludge had much higher (20%–100%) specific methanogenic conversion rates than the volatile-fatty acid␣(VFA)/ethanol-fed sludge for all substrates tested, including VFA. In both reactors, a flocculant sludge developed, although a much higher content of extracellular polysaccharide was measured in the lactose-fed sludge [1900 μg compared to 305 μg uronic acid/g volatile suspended solids (VSS)]. When the liquid upflow velocity of a third, VFA/ethanol-fed reactor was increased to 0.5 m/h, granulation of the sludge occurred, accompanied by a large increase (200%–500%) in the specific methanogenic conversion rates for the syntrophic and methanogenic substrates studied. Granulation reduced the susceptibility of the sludge to flotation. Glucose was degraded at a high rate (100 mg glucose gVSS⁻¹h⁻¹) by the sludge from the third reactor, despite not having been exposed to a sugar-containing influent for 563␣days. Received: 7 June 1996 / Received revision: 23 September 1996 / Accepted: 29 September 1996     

Color removal of real textile wastewater by sequential anaerobic and aerobic reactors

Textile wastewater from the Pusan Dyeing Industrial Complex (PDIC) was treated utilizing a two-stage continuous system, composed of an upflow anaerobic sludge blanket reactor and an activated sludge reactor. The effects of color and organic loading rates were studied by varying the hydraulic retention time and influent glucose concentration. The maximum color load to satisfy the legal discharge limit of color intensity in Korea (400 ADMI, unit of the American Dye Manufacturers Institute) was estimated to be 2,700 ADMI·L⁻¹ day⁻¹. This study indicates that the two-stage anaerobic/aerobic reaction system is potentially useful in the treatment of textile wastewater.     

DNA Extraction from Activated Sludges

To optimize the cell lysis step for DNA extraction from activated sludge samples, two floc dispersion methods (sonication versus stirring with a cation exchange resin), and three cell lysis treatments (lysozyme + SDS, sonication in a water bath, and thermal shock) were tested. For dispersion, stirring with cation exchange resin was more efficient than sonication. The cell lysis procedures were applied in two sequences, and DNA was quantified after each cell lysis treatment. Lysozyme + SDS was the most effective step in the cell lysis procedures. The cell lysis treatment sequences giving the highest DNA yields were not the same for all the sludges. The differences in sludge microbial compositions and floc structures required specifically adapted cell lysis protocols. The proposed protocols were highly efficient for DNA extraction, yielding about 50 mg DNA g⁻¹ volatile suspended solids, and allowed PCR amplification of 16S rDNA. Received: 26 September 1998 / Accepted: 13 February 1999     

Real-time PCR assay for the simultaneous quantification of nitrifying and denitrifying bacteria in activated sludge

In order to improve wastewater treatment processes, a need exists for tools that rapidly give detailed insight into the community structure of activated sludge, supplementary to chemical and physical data. In this study, the advantages of microarrays and quantitative polymerase chin reaction (PCR) methods were combined into a real-time PCR assay that allows the simultaneous quantification of phylogenetic and functional genes involved in nitrification and denitrification processes. Simultaneous quantification was possible along a 5-log dynamic range and with high linear correlation (R ² > 0.98). The specificity of the assay was confirmed by cloning and sequencing analyses of PCR amplicons obtained from activated sludge. The real-time assay was validated on mixed liquid samples of different treatment plants, which varied in nitrogen removal rate. The abundance of ammonia oxidizers was in the order of magnitude of 10⁶ down to 10⁴ ml⁻¹, whereas nitrite oxidizers were less abundant (10³–10¹ order of magnitude). The results were in correspondence with the nitrite oxidation rate in the sludge types. As for the nirS, nirK, and nosZ gene copy numbers, their abundance was generally in the order of magnitude of 10⁸–10⁵. When sludge samples were subjected to lab-scale perturbations, a decrease in nitrification rate was reflected within 18 h in the copy numbers of nitrifier genes (decrease with 1 to 5 log units), whereas denitrification genes remained rather unaffected. These results demonstrate that the method is a fast and accurate tool for the analysis of the (de)nitrifying community structure and size in both natural and engineered environmental samples. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Microbial decolorization of azo dyes by Proteus mirabilis

A bacterium identified as Proteus mirabilis was isolated from acclimated sludge from a dyeing wastewater treatment plant. This strain rapidly decolorized a deep red azo dye solution (RED RBN). Features of the decolorizing process related to biodegradation and biosorption were also studied. Although P. mirabilis displayed good growth in shake culture, color removal was best in anoxic static cultures. For color removal, the optimal pH and temperature were 6.5–7.5 and 30–35°C, respectively. The organism exhibited a remarkable color removal capability, even at a high concentration of azo dye. More than 95% of azo dye was reduced within 20 h at a dye concentration of 1.0 g L⁻¹. Decolorization appears to proceed primarily by enzymatic reduction associated with a minor portion, 13–17%, of biosorption to inactivated microbial cells. Received 06 January 1999/ Accepted in revised form 22 April 1999     

Biosorption and biodegradation of pentachlorophenol (PCP) in an upflow anaerobic sludge blanket (UASB) reactor

In order to understand the fate of PCP in upflow anaerobic sludge blanket reactor (UASB) more completely, the sorption and biodegradation of pentachlorophenol (PCP) by anaerobic sludge granules were investigated. The anaerobic granular sludge degrading PCP was formed in UASB reactor, which was seeded with anaerobic sludge acclimated by chlorophenols. At the hydraulic retention time (HRT) of 20–22 h, and PCP loading rate of 200–220 mg l⁻¹ d⁻¹, UASB reactor exhibited good performance in treating wastewater which containing 170–180 mg l⁻¹ PCP and the PCP removal rate of 99.5% was achieved. Sequential appearance of tetra-, tri-, di-, and mono-chlorophenol was observed in the reactor effluent after 20 mg l⁻¹ PCP introduction. Sorption and desorption of PCP on the anaerobic sludge granules were all fitted to the Freundlich isotherm equation. Sorption of PCP was partly irreversible. The Freundlich equation could describe the behavior of PCP amount sorbed by granular sludge in anaerobic reactor reasonably well. The results demonstrated that the main mechanism leading to removal of PCP on anaerobic granular sludge was biodegradation, not sorption or volatization.     

Microbial biodegradation of organic wastes containing surfactants in a continuous-flow reactor

In a continuous flow bioreactor seeded with microbes from municipal activated sludge, complete organic carbon oxidation of simulated graywater (wastewater produced in human residences, excluding toilet wastes) was achieved at dilution rates up to 0.36 h⁻¹ in the presence of 64.1 μ M linear alkylbenzenesulfonate (LAS) L⁻¹. At LAS concentrations of 187 μ M, the system functioned only at dilution rates up to 0.23 h⁻¹, and the biomass yield was two-fold lower. There were physiological changes in the microbial communities under different operating conditions, as measured by specific contents of ATP and extracellular hydrolases as well as the respiratory potential of the biomass. LAS inhibited the activity of LAS-degrading microbes at >150  μ M LAS, and the activity of other microbes at >75 μ M LAS. Chemical analysis of graywater indicated that samples consisted primarily of biological polymers (proteins and polysaccharides) and lower concentrations of surfactants. Biological remediation of graywater is possible, although treatment efficiency is influenced by the operating conditions and wastestream composition. Received 08 July 1996/ Accepted in revised form 14 November 1996     

Nitrifying granules cultivation in a sequencing batch reactor at a low organics-to-total nitrogen ratio in wastewater

It is possible to cultivate aerobic granular sludge at a low organic loading rate and organics-to-total nitrogen (COD/N) ratio in wastewater in the reactor with typical geometry (height/diameter = 2.1, superficial air velocity = 6 mm/s). The noted nitrification efficiency was very high (99%). At the highest applied ammonia load (0.3 ± 0.002 mg NH₄⁺–N g total suspended solids (TSS)⁻¹ day⁻¹, COD/N = 1), the dominating oxidized form of nitrogen was nitrite. Despite a constant aeration in the reactor, denitrification occurred in the structure of granules. Applied molecular techniques allowed the changes in the ammonia-oxidizing bacteria (AOB) community in granular sludge to be tracked. The major factor influencing AOB number and species composition was ammonia load. At the ammonia load of 0.3 ± 0.002 mg NH₄⁺–N g TSS⁻¹ day⁻¹, a highly diverse AOB community covering bacteria belonging to both the Nitrosospira and Nitrosomonas genera accounted for ca. 40% of the total bacteria in the biomass.     

Increase of xylitol yield by feeding xylose and glucose in Candida tropicalis

Candida tropicalis, a strain isolated from the sludge of a factory manufacturing xylose, produced a high xylitol concentration of 131 g/l from 150 g/l xylose at 45 h in a flask. Above 150 g/l xylose, however, volumetric xylitol production rates decreased because of a lag period in cell growth. In fed-batch culture, the volumetric production rate and xylitol yield from xylose varied substantially with the controlled xylose concentration and were maximum at a controlled xylose concentration of 60 g/l. To increase the xylitol yield from xylose, feeding experiments using different ratios of xylose and glucose were carried out in a fermentor. The maximum xylitol yield from 300 g/l xylose was 91% at a glucose/xylose feeding ratio of 15%, while the maximum volumetric production rate of xylitol was 3.98 g l⁻¹ h⁻¹ at a glucose/xylose feeding ratio of 20%. Xylitol production was found to decrease markedly as its concentration rose above 250 g/l. In order to accumulate xylitol to 250 g/l, 270 g/l xylose was added in total, at a glucose/xylose feeding ratio of 15%. Under these conditions, a final xylitol production of 251 g/l, which corresponded to a yield of 93%, was obtained from 270 g/l xylose in 55 h. Received: 20 April 1998 / Received revision: 29 May 1998 / Accepted: 19 June 1998     

Bioaugmentation in activated sludge: current features and future perspectives

Bioaugmentation of activated sludge systems with specialised bacterial strains could be a powerful tool to improve several aspects in wastewater treatment processes, such as improved flocculation and degradation of recalcitrant compounds. This review focuses on the addition of strains to activated sludge to enhance the biodegradation of recalcitrant compounds, either through the activity of the inoculated strain or after transfer of degradative plasmids to activated sludge bacteria. Different factors that improve the aggregation of the sludge flocs and their influence on biodegradation are described. This review further deals with the role of bacterial plasmids in natural genetic exchange between inoculated and indigenous sludge bacteria, and in the construction of new genetically modified organisms. The few successful cases of bioaugmentation described in this review, together with future research, must lead to a better understanding of sludge bioaugmentation. Received: 5 January 1998 / Received revision: 20 April 1998 / Accepted: 20 April 1998     

Decolorization of triphenylmethane, azo, and anthraquinone dyes by a newly isolated Aeromonas hydrophila strain

A broad-spectrum dye-decolorizing bacterium, strain DN322, was isolated from activated sludge of a textile printing wastewater treatment plant. The strain was characterized and identified as a member of Aeromonas hydrophila based on Gram staining, morphology characters, biochemical tests, and nearly complete sequence analysis of 16S rRNA gene and the gyrase subunit beta gene (gyrB). Strain DN322 decolorized a variety of synthetic dyes, including triphenylmethane, azo, and anthraquinone dyes. For color removal, the most suitable pH and temperature were pH 5.0–10.0 and 25–37°C, respectively. Triphenylmethane dye, e.g., Crystal Violet, Basic Fuchsin, Brilliant Green, and Malachite Green (50 mg l⁻¹) were decolorized more than 90% within 10 h under aerobic culture condition and Crystal Violet could be used as sole carbon source and energy source for cell growth. The color removal of triphenylmethane dyes was due to a soluble cytosolic enzyme, and the enzyme was an NADH/NADPH-dependent oxygenase; For azo and anthraquinone dyes, e.g., Acid Amaranth, Great Red GR, Reactive Red KE-3B, and Reactive Brilliant Blue K-GR (50 mg l⁻¹) could be decolorized more than 85% within 36 h under anoxic condition. This strain may be useful for bioremediation applications.