Improvement of settling characteristics of activated sludge by addition of a floc-forming bacterium

A floc-forming bacterium, strain no. 5, which made very large and easily precipitated flocs, was added as seed sludge in a bench-scale aeration tank. About one week allowed for production of activated sludge which could treat high-BOD wastewater at high-BOD loading while maintaining very good settling characteristics. The strain no. 5 was present at 3% of the activated sludge. Moreover, the settling characteristics of the activated sludge were much improved when the strain no. 5 culture was added to the bench-scale aeration or a large-scale deep aeration tank.     

Formation of cellulose fibrils by gram-negative bacteria and their role in bacterial flocculation

Summary Exocellular fibrils, consisting of true cellulose, were found to be produced by many bacteria. These bacteria have been selected out of a large number of strains isolated from activated sludge on the basis of their flocculent growth habit in liquid medium.The amount of cellulose, present in the bacterial flocs, varied from 1.0 to 4.0%. In addition to strains isolated from activated sludge, like Pseudomonas, Achromobacter, Alcaligenes and Aerobacter, also strains of the genera Rhizobium, Agrobacterium and Azotobacter were found to give flocculent growth due to the formation of cellulose fibrils.Bacterial flocculation in pure cultures of the strains examined was mainly caused by the production of exocellular fibrils. Apparently, the formation of cellulose fibrils seems to be a common property of Gram-negative, floc-forming bacteria, and may not be restricted to Acetobacter xylinum.     

Effect of sludge flocs on the aeration performance of suction-impellers

The authors have investigated a special aspect of the transfer of oxygen: the aeration performance of small-scale suction-impellers in a suspension of activated sludge floes and a performance comparison with clear water. The principle of the impellers is, that in order to disperse air bubbles into the aforementioned liquids, all that we need do is a simple stirring. Due to the impellers the authors could not show a reduction of the oxygen transfer coefficient in the presence of sludge flocs. Former comparative studies of oxygen transfer, using a fixed orifice, did show a reduction in the presence of sludge flocs. Therefore, it seems probable that the effect of activated sludge flocs on the transfer coefficient is linked with the manner in which air is dispersed, i.e., depends on whether an orifice is fixed or moved. The experimental results allow presumption that by developing a high intensity of shear near the location where air bubbles are produced, we are able to minimize the effect of sludge flocs on the process of bubble formation.     

Two parameters account for the flocculated growth of microbes in biodegradation assays

Microbes in activated sludge tanks mostly occur in flocs rather than in cell suspensions. Flocculation results in a limited supply of substrate to the bacteria inside the flocs, which reduces the biodegradation rate of organic compounds by several orders of magnitude. This article presents a simple two-parameter extension of growth models for cell suspensions to account for the ensuing reduction of the degradation rate. The additional parameters represent floc size at division and diffusion length. The biomass of small flocs initially increases exponentially at a rate equal to that of cell suspensions. After this first phase, the growth rate gradually decreases and finally the radius becomes a linear function of time. At this time flocs are large and have a kernel of dead biomass. This kernel arises when the substrate concentration decreases below the threshold level at which cells are just able to pay their maintenance costs. We deduce an explicit approximative expression for the interdivision time of flocs, and thereby for the growth of flocculated microbial biomass at constant substrate concentrations. The model reveals that the effect of stirring on degradation rates occurs through a reduction of the floc size at division. The results can be applied in realistic biodegradation quantifications in activated sludge tanks as long as substrate concentrations change slowly.     

Comparison between adsorption of poliovirus and rotavirus by aluminum hydroxide and activated sludge flocs.

Adsorption of poliovirus and rotavirus by aluminum hydroxide and activated sludge flocs was studied. Both aluminum hydroxide and activated sludge flocs adsorbed greater amounts of poliovirus than rotavirus. Aluminum hydroxide flocs reduced the titer of poliovirus in tap water by 3 log10, but they only reduced the titer of a simian rotovirus (SA-11) in tap water by 1 log10 or less and did not noticeably reduce the number of human rotavirus particles present in a dilute stool suspension. Activated sludge flocs reduced the titer of added poliovirus by 0.7 to 1.8 log10 and reduced the titer of SA-11 by 0.5 log10 or less. These studies indicate that a basic difference in the adsorptive behavior of enteroviruses and rotaviruses exists and that water and wastewater treatment processes that are highly effective in removal of enteroviruses may not be as effective in removing other viral groups such as rotaviruses.     

Comparison between adsorption of poliovirus and rotavirus by aluminum hydroxide and activated sludge flocs.

Adsorption of poliovirus and rotavirus by aluminum hydroxide and activated sludge flocs was studied. Both aluminum hydroxide and activated sludge flocs adsorbed greater amounts of poliovirus than rotavirus. Aluminum hydroxide flocs reduced the titer of poliovirus in tap water by 3 log10, but they only reduced the titer of a simian rotovirus (SA-11) in tap water by 1 log10 or less and did not noticeably reduce the number of human rotavirus particles present in a dilute stool suspension. Activated sludge flocs reduced the titer of added poliovirus by 0.7 to 1.8 log10 and reduced the titer of SA-11 by 0.5 log10 or less. These studies indicate that a basic difference in the adsorptive behavior of enteroviruses and rotaviruses exists and that water and wastewater treatment processes that are highly effective in removal of enteroviruses may not be as effective in removing other viral groups such as rotaviruses.     

The quorum-sensing effect of aerobic granules on bacterial adhesion, biofilm formation, and sludge granulation

Quorum sensing (QS) through signal chemical molecules is known to be essential to bacterial adhesion and biofilm formation. In this study, the QS ability of aerobic granules—a special form of biofilms used for biological wastewater treatment—was investigated and compared with that of conventional activated sludge flocs. A novel sectional membrane bioreactor was used together with a flow-cell to evaluate the possible influence of signal chemicals produced by the source sludge on the growth mode of bacterial cells. The results demonstrate the apparent production of QS chemicals from granules and its impact on initial cell attachment and granule formation. When granules were used as the signal-producing biomass, the attached-growth mode was dominant for the free cells, and the biofilm formation rate in the flow-cell was about ten times faster than in cases which used activated sludge as the signal source biomass. In addition, the intracellular extract from mature granules significantly accelerated the sludge granulation process. It is argued that the production and expression of QS signal chemicals from granules and granule precursors might have induced the gene expression of bacteria in suspension for attached growth rather than suspended growth, leading to granule formation and its stable structure.     

Effects of synthetic polymer on the filamentous bacteria in activated sludge

Filamentous bulking is one of the solid-liquid separation problems always seen in activated sludge process. The addition of synthetic polymer is always one of the popular ways for the treatment plant operator to immediately solve the poor sludge settling problem. Therefore, it may be interesting to understand the effects of synthetic polymer on the filamentous bacteria in activated sludge. In this study, synthetic polymer was applied to a lab-scale wastewater treatment system with the filamentous bulking problem. The population structure of filamentous bacteria and sludge characteristics were investigated under different conditions. When synthetic polymer was added into the system, it was found that poor sludge settleability caused by filamentous bulking was temporarily solved and filamentous branches growing outside the flocs were damaged or inhibited. However, filamentous growth was still observed inside the flocs. After the addition of polymer was halted, filamentous branches extended out of the flocs immediately. Very serious filamentous bulking occurred and sludge settleability became much worse than that occurring before the addition of polymer. And, it took several weeks for the system to return to normal operation.     

Characterization of non-flocculent cells isolated from a culture of flocculent Saccharomyces cerevisiae NCYC 1001

During cultivation of a flocculent yeast, Saccharomyces cerevisiae 1001, two cell fractions, flocs and free cells, appeared in the medium. Free cells contained cells with a normal ability to flocculate, less flocculent cells and not-flocculent cells. When the non-flocculent cells and not-flocculent cells. When the non-flocculent cell fraction from the postexponential phase of growth was collected and used as an inoculum, the culture showed synchronous growth. The floc forming ability of the yeast cells from this culture increased gradually with the number of divisions.     

Involvement of Ca2+ in the flocculation of Kluyvera cryocrescens KA-103

Kluyvera cryocrescens KA-103 showed a dispersed growth in Ca²⁺-free Polypepton medium, but formed flocs on addition of a sufficient concentration of Ca²⁺ to the bacterial cell suspension. Therefore, calcium adsorption properties and flocculation conditions were investigated using bacterial cells cultured in the Ca²⁺-free Polypepton medium. The bacterium required 1.5 mM Ca²⁺ or more for good flocculation (F>90%), but a cooperative effect of Na⁺ and Ca²⁺ on good flocculation was observed at lower concentrations of Ca²⁺. The Langmuir adsorption isotherm was used to describe the adsorption of Ca²⁺ by the bacterial cells.     

Deflocculation of Activated Sludge by the Dissimilatory Fe(III)-Reducing Bacterium Shewanella alga BrY

The influence of microbial Fe(III) reduction on the deflocculation of autoclaved activated sludge was investigated. Fe(III) flocculated activated sludge better than Fe(II). Decreasing concentrations of Fe(III) caused an increase in sludge bulk water turbidity, while bulk water turbidity remained relatively constant over a range of Fe(II) concentrations. Cells of the dissimilatory metal-reducing bacterium Shewanella alga BrY coupled the oxidation of H(inf2) to the reduction of Fe(III) bound in sludge flocs. Cell adhesion to the Fe(III)-sludge flocs was a prerequisite for Fe(III) reduction. The reduction of Fe(III) in sludge flocs by strain BrY caused an increase in bulk water turbidity, suggesting that the sludge was deflocculated. The results of this study support previous research suggesting that microbial Fe(III) respiration may have an impact on the floc structure and colloidal chemistry of activated sludge.     

Deoxyribonuclease-susceptible Floc Forming Pseudomonas sp.

A bacterium belonging to Pseudomonas which was isolated from activated sludge formed flocs in glycerol-containing medium. The flocs were deflocculated by deoxyribonuclease treatment in the presence of magnesium ions. Flocs were also deflocculated by 2 m NaCl, heating at temperatures higher than 50°C, and at pH below 1 or above 11. The observations suggest that deoxyribonucleic acid is directly involved in the association of cells and that ionic bonds are responsible for the flocculation of cells.     

Production and release of surfactant byCorynebacterium lepus in hydrocarbon and glucose media

Summary Corynebacterium lepus produced a considerable amount of extracellular surfactant during growth in a mineral salts medium containing hexadecane as the sole carbon source. The study revealed that the bacterium also produced a large amount of surfactant when grown on glucose, but in this case the surface active agent was cell bound. The surfactant was released from the cells when they were treated with hexadecane after growth. Tetradecane also showed a good capability for release of the surfactant. Decane and octane were less effective than hexadecane and tetradecane.     

Removal of Escherichia coli in wastewater by activated sludge.

Removal of bacteria from wastewater treated with activated sludge was studied by the use of a streptomycin-resistant Escherichia coli strain. The removal appeared to be a biphasic process. A rapid sorption of bacteria to the sludge flocs took place in the first hour after seeding mixed liquor with E. coli. Thereafter, slower elimination of E. coli was observed. The latter process was due to predation on E. coli by ciliated protozoa. This was shown by: (i) appearance of fluorescent food vacuoles of ciliates when fluorescent E. coli cells were added to mixed liquor; (ii) inhibition of predation either in the presence of cycloheximide or under anaerobic conditions; and (iii) absence of predation in bulking and washed sludge.     

Enhancement of L+-lactic acid production using mycelial flocs of Rhizopus oryzae

L(+)-Lactic acid production was enhanced in the culture of Rhizopus oryzae using mycelial flocs formed by addition of 3 g/L mineral support and 5 ppm polyethylene oxide. By addition of the mineral support, an electrostatic repulsion between mycelia increased by 3.5-fold compared to that of mycelia, which allowed a dispersed growth of R. oryzae in the early growth phase. In conventional culture the morphology of R. oryzae is that of a pellet-like cake, however, when support and polyethylene oxide are added to the culture, the morphology of R. oryzae takes on a cotton-like appearance. The formation of these cotton-like mycelial flocs was induced by the addition of 5 ppm polyethylene oxide into a 14 h culture containing the mineral support before the formation of the conventional pellet morphology. The cotton-like flocs were also formed in cultures grown in a fermentor. This morphology allowed effective mass transfer inside the flocs and effective fluidity of culture broth in the reactor. L(+)-Lactic acid concentration produced by mycelial flocs in fermentor, with the support and polyethylene oxide, was 103.6 g/L with the yield of 0.86 using 120 g/L of glucose as the substrate for this cultures without both, the concentration was 65.2 g/L. It demonstrates that cotton-like mycelial flocs are the optimal morphology in the culture of R. oryzae. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 461-470, 1997.     

Removal of Escherichia coli in wastewater by activated sludge.

Removal of bacteria from wastewater treated with activated sludge was studied by the use of a streptomycin-resistant Escherichia coli strain. The removal appeared to be a biphasic process. A rapid sorption of bacteria to the sludge flocs took place in the first hour after seeding mixed liquor with E. coli. Thereafter, slower elimination of E. coli was observed. The latter process was due to predation on E. coli by ciliated protozoa. This was shown by: (i) appearance of fluorescent food vacuoles of ciliates when fluorescent E. coli cells were added to mixed liquor; (ii) inhibition of predation either in the presence of cycloheximide or under anaerobic conditions; and (iii) absence of predation in bulking and washed sludge.     

Distribution of extracellular polysaccharides and flotation of anaerobic sludge

 Extracellular polysaccharides (EPS) were quantified in dense granules and loose flocs by chemical analysis of the uronic acid content. Their distribution within the aggregates was determined by microscopic staining. Granules contained a higher amount of EPS (1–1.6 mg/g volatile suspended solids, VSS) than flocs (0.3 mg/g VSS). In granules approximately 50% of the total amount of EPS was present in a 40-μm-thick zone on the surface. The remainder was dispersed in the rest of the aggregate. In flocs the highest concentration was present in the centre and the EPS layer on the surface was not found. Tests showed that flocculent sludge was very sensitive to flotation, while the studied granules did not float. The lower susceptibility to flotation of granules as compared to flocs was attributed to the presence of the hydrophilic EPS coating that prevents attachment of gas bubbles. Received: 21 November 1995/Received last revision: 15 April 1996/Accepted: 22 April 1996     

Addition of Al and Fe salts during treatment of paper mill effluents to improve activated sludge settlement characteristics

Metal salts, ferrous sulphate and aluminium chloride, were added to laboratory-scale activated sludge plant treating paper mill effluents to investigate the effect on settlement characteristics. Before treatment the sludge was filamentous, had stirred sludge volume index (SSVI) values in excess of 300 and was moderately hydrophobic. The use of FeSO4.7H2O took three weeks to reduce the SSVI to 90. Microscopic examination showed that Fe had converted the filamentous flocs into a compact structure. When the iron dosing was stopped, the sludge returned to its bulking state within four weeks. In a subsequent trial, the addition of AlCl3 initially resulted in an improvement of the settlement index but then caused deterioration of the sludge properties. It is possible that aluminium was overdosed and caused charge reversal, increasing the SSVI.     

Enhanced phosphate uptake and polyphosphate accumulation in Burkholderia cepacia grown under low pH conditions

Of bacterial cells in a sample of activated sludge, 34% contained detectable intracellular polyphosphate inclusions following Neisser staining, when grown on glucose/mineral salts medium at pH 5.5; at pH 7.5 only 7% of cells visibly accumulated polyphosphate. In a sludge isolate of Burkholderia cepacia chosen for further study, maximal removal of phosphate and accumulation of polyphosphate occurred at pH 5.5; levels were up to 220% and 330% higher, respectively, than in cells grown at pH 7.5. During the early stationary phase of growth at pH 5.5 a maximum level of intracellular polyphosphate that comprised 13.6% of cellular dry weight was reached. Polyphosphate kinase activity was detected in actively growing cells only when cultured at pH 5.5. The phenomenon of acid-stimulated phosphate uptake and polyphosphate accumulation in this environmental bacterial population parallels observations previously made by us in the yeast Candida humicola and may thus represent a widespread microbial response to low external pH values.     

Isolation and characterization of solvent-tolerant Pseudomonas putida strain T-57, and its application to biotransformation of toluene to cresol in a two-phase (organic-aqueous) system

Pseudomonas putida T-57 was isolated from an activated sludge sample after enrichment on mineral salts basal medium with toluene as a sole source of carbon. P. putida T-57 utilizes n-butanol, toluene, styrene, m-xylene, ethylbenzene, n-hexane, and propylbenzene as growth substrates. The strain was able to grow on toluene when liquid toluene was added to mineral salts basal medium at 10-90% (v/v), and was tolerant to organic solvents whose log P(ow) (1-octanol/water partition coefficient) was higher than 2.5. Enzymatic and genetic analysis revealed that P. putida T-57 used the toluene dioxygenase pathway to catabolize toluene. A cis-toluene dihydrodiol dehydrogenase gene (todD) mutant of T-57 was constructed using a gene replacement technique. The todD mutant accumulated o-cresol (maximum 1.7 g/L in the aqueous phase) when cultivated in minimal salts basal medium supplemented with 3% (v/v) toluene and 7% (v/v) 1-octanol. Thus, T-57 is thought to be a good candidate host strain for bioconversion of hydrophobic substrates in two-phase (organic-aqueous) systems.