Differences of the protozoan biomass and grazing during spring and summer in the Indian sector of the Southern Ocean

The dynamics of protozoa were investigated during two cruises in the Indian sector of the Southern Ocean: the early spring ANTARES 3 cruise (28 September to 8 November 1995) and the late summer ANTARES 2 cruise (6 February to 8 March 1994). Biomass and feeding activity of protozoa were measured as well as the biomass of their potential prey – bacteria and phototrophic flagellates – along the 62°E meridian. The sampling grid extended from the Polar Frontal region to the Coastal and Continental Shelf Zone in late summer and to the ice edge in spring, crossing the Antarctic Divergence. Protozoan biomass, although low in absolute terms, contributed 30% and 20% to the total microbial biomass (bacteria, phytoplankton and protozoa) in early spring and late summer, respectively. Nanoprotozoa dominated the total protozoan biomass. The geographical and seasonal distribution of protozoan biomass was correlated with that of phototrophic flagellates. However, bacterial and phototrophic flagellate biomass were inversely correlated. Phototrophic flagellates dominated in the Sea Ice Zone whereas bacteria were predominant at the end of summer in the Polar Frontal region and Coastal and Continental Shelf Zone. Furthermore, bacteria were the most important component of the microbial community (57% of the total microbial biomass) in late summer. Phototrophic flagellates were ingested by both nano-and microprotozoa. In contrast, bacteria were only ingested by nanoprotozoa. Protozoa controlled up to 90% of the daily bacterial production over the period examined. The spring daily protozoan ingestion controlled more than 100% of daily phototrophic flagellate production. This control was less strong at the end of summer when protozoan grazing controlled 42% of the daily phototrophic flagellate production. Accepted: 30 October 1999     

Anaerobic Methane Oxidation: Occurrence and Ecology

Anoxic sediments and digested sewage sludge anaerobically oxidized methane to carbon dioxide while producing methane. This strictly anaerobic process showed a temperature optimum between 25 and 37°C, indicating an active microbial participation in this reaction. Methane oxidation in these anaerobic habitats was inhibited by oxygen. The rate of the oxidation followed the rate of methane production. The observed anoxic methane oxidation in Lake Mendota and digested sewage sludge was more sensitive to 2-bromoethanesulfonic acid than the simultaneous methane formation. Sulfate diminished methane formation as well as methane oxidation. However, in the presence of iron and sulfate the ratio of methane oxidized to methane formed increased markedly. Manganese dioxide and higher partial pressures of methane also stimulated the oxidation. The rate of methane oxidation in untreated samples was approximately 2% of the CH₄ production rate in Lake Mendota sediments and 8% of that in digested sludge. This percentage could be increased up to 90% in sludge in the presence of 10 mM ferrous sulfate and at a partial pressure of methane of 20 atm (2,027 kPa).     

Phototrophic purple and green bacteria in a sewage treatment plant

In all purification stages of a biological sewage treatment plant, phototrophic bacteria were detected by the method of viable cell counts. The predominant species identified belonged to the genus Rhodopseudomonas of purple nonsulfur bacteria. The number of phototrophic bacteria was highest in wastewater containing sludge. In activated sludge, an average of 10(5) viable cells/ml was found; the number depended upon concentration of sludge rather than on seasonal changes in light conditions in the course of a year. Bacteriochlorophyll a was extracted from activated sludge. Relative to the viable counts of phototrophic bacteria, the content of bacteriochlorophyll a was 5- to 10-fold higher than that of three representative pure cultures. By incubation of activated and digester sludge under different environmental conditions, it was shown that phototrophic bacteria can complete with other bacteria only under anaerobic conditions in the light.     

Phototrophic purple and green bacteria in a sewage treatment plant.

In all purification stages of a biological sewage treatment plant, phototrophic bacteria were detected by the method of viable cell counts. The predominant species identified belonged to the genus Rhodopseudomonas of purple nonsulfur bacteria. The number of phototrophic bacteria was highest in wastewater containing sludge. In activated sludge, an average of 10(5) viable cells/ml was found; the number depended upon concentration of sludge rather than on seasonal changes in light conditions in the course of a year. Bacteriochlorophyll a was extracted from activated sludge. Relative to the viable counts of phototrophic bacteria, the content of bacteriochlorophyll a was 5- to 10-fold higher than that of three representative pure cultures. By incubation of activated and digester sludge under different environmental conditions, it was shown that phototrophic bacteria can complete with other bacteria only under anaerobic conditions in the light.     

Synthesis of bacteriochlorophyll a by the purple nonsulfur bacterium Rhodobacter capsulatus

The ability of purple nonsulfur bacteria Rhodobacter capsulatus B10 to synthesize bacteriochlorophyll under phototrophic and dark conditions was studied. The modes for cultivation in the dark with oxygen limitation in a continuous culture at D = 0.1 h(-1) were selected. The yield of biomass reached 20 g/l; the bacteriochlorophyll a output of the process amounted to 16.6 mg/l h(-1).     

Conversion of municipal solid waste to carboxylic acids using a mixed culture of mesophilic microorganisms

Waste biomass was anaerobically converted to carboxylate salts by using a mixed culture of acid-forming microorganisms. Municipal solid waste (MSW) was the energy source (carbohydrates) and sewage sludge (SS) was the nutrient source (minerals, metals, and vitamins). Four fermentors were arranged in series and solids and liquids were transferred countercurrently in opposite directions, which allows both high conversions and high product concentrations. Fresh biomass was added to Fermentor 1 (highest carboxylic acid concentration) and fresh media was added to Fermentor 4 (most digested biomass). All fermentations were performed at 40 degrees C. Calcium carbonate was added to the fermentors to neutralize the acids to their corresponding carboxylate salts. Iodoform was used to inhibit methane production and urea was added as a nitrogen source. Product concentrations were up to 25 g/L, with productivities up to 1.4 g total acid/(L liquid d). Mass balances with closure between 93% and 105% were obtained for all systems. Continuum particle distribution modeling (CPDM) was applied to correlate batch fermentation data to countercurrent fermentation data and predict product concentration over a wide range of solids loading rates and residence times. CPDM for lime-treated MSW/SS fermentation system predicted the experimental total acid concentration and conversion within 4% and 16% respectively.     

Membrane biofouling by extracellular polymeric substances or soluble microbial products from membrane bioreactor sludge

This study extracted the soluble microbial products and loosely bound and tightly bound extracellular polymeric substances (EPS) from suspended sludge from a membrane bioreactor, original and aerobically/anaerobically digested, and compared their fouling potentials on a microfiltration membrane. The resistance of cake layer accounts for 95–98% of the total filtration resistances when filtering the whole sludges, with anaerobically digested sludge presenting the highest resistance among the three tested sludges. The tightly bound EPS has the highest potential to foul the membrane; however, the loosely bound EPS contribute most of the filtration resistances of the whole sludges. The foulants corresponding to the irreversible fouling have chemical fingerprints similar to those from loosely bound EPS, which have a greater predilection to proteins and humic substances than to polysaccharides.     

Transformation of indole by methanogenic and sulfate-reducing microorganisms isolated from digested sludge

In the present study, mineralization of an aromaticN-heterocyclic molecule, indole, by microorganisms present in anaerobically digested sewage sludge was examined. The first step in indole mineralization was the formation of a hydroxylated intermediate, oxindole. The rate of transformation of indole to oxindole and its subsequent disappearance was dependent on the concentration of inoculum and indole and the incubation temperature. Methanogenesis appeared to be the dominant process in the mineralization of indole in 10% digested sludge even in the presence of high concentrations of sulfate. Enrichment of the digested sludge with sulfate as an electron acceptor allowed the isolation of a metabolically stable mixed culture of anaerobic bacteria which transformed indole to oxindole and acetate, and ultimately to methane and carbon dioxide. This mixed culture exhibited a predominance of sulfate-reducers over methanogens with more than 75% of the substrate mineralized to carbon dioxide. The investigation demonstrates that indole can be transformed by both methanogenic and sulfate-reducing microbial populations.     

Thermochemical liquidization and anaerobic treatment of dewatered sewage sludge

Dewatered sewage sludge was thermochemically liquidized at 175°C and the liquidized sludge was separated by centrifugation to 57.7% (w/w) supernatant [moisture, 92.3%; volatile solid (VS), 7.0%] and 42.3% precipitate (moisture, 71.6%; VS, 18.9%). The supernatant was successfully anaerobically digested. Biogas yield from the supernatant at organic loading concentrations of 1.9–2.2 g VS/l during 9 days' incubation was 440 ml/g-added VS and digestion ratio was 66% (w/w). Biogas yield in the case of dewatered sewage sludge was 257% ml/g-added VS and digestion ratio was 45%. Digestion of the supernatant resulted in high biogas productivity and a high digestion ratio compared with that of the dewatered sewage sludge. Moreover, it was suggested that the precipitate can be incinerated without the need for any supplemental fuel.     

Performance characteristics of a two-stage dark fermentative system producing hydrogen and methane continuously

The performance of a mesophilic two-stage system generating hydrogen and methane continuously from sucrose (10-30 g/L) was investigated. A hydrogen-generating CSTR followed by an upflow anaerobic filter were both inoculated with anaerobically digested sewage sludge, and ORP, pH, gas output, %H(2), %CH(4) and %CO(2) monitored. pH was controlled with NaOH, KOH or Ca(OH)(2). Using NaOH as alkali with 10 g/L sucrose, yields of 1.62 +/- 0.2 mol H(2)/mol hexose added and 323 mL CH(4)/gCOD added to the hydrogen and methane reactors respectively were achieved. The overall chemical oxygen demand (COD) reduction was 92.6% with 0.90 +/- 0.1 g/L sodium and 316 +/- 40 mg/L residual acetate in the methane reactor. Operation at 20 g/L sucrose and NaOH as alkali led to impaired volatile fatty acid (VFA) degradation in the methane reactor with 2.23 +/- 0.2 g/L sodium, 1,885 mg/L residual acetate, a hydrogen yield of 1.47 +/- 0.1 mol/mol hexose added, a methane yield of 294 mL/gCOD added and an overall COD reduction of 83%. Using Ca(OH)(2) as alkali with 20 g/L sucrose gave a hydrogen yield of 1.29 +/- 0.3 mol/mol hexose added, a methane yield of 337 mL/gCOD added and improved the overall COD reduction to 91% with residual acetate concentrations of 522 +/- 87 mg/L. Operation at 30 g/L sucrose with Ca(OH)(2) gave poorer overall COD reduction (68%), a hydrogen yield of 1.47 +/- 0.2 mol/mol hexose added, a methane yield of 138 mL/gCOD added and residual acetate 7,343 +/- 715 mg/L. It was shown that sodium toxicity and overloading are important issues for successful anaerobic digestion of effluent from biohydrogen reactors in high rate systems.     

Peatland microhabitat heterogeneity drives phototrophic microbe distribution and photosynthetic activity

Phototrophic microbes are widespread in soils, but their contribution to soil carbon (C) uptake remains underexplored in most terrestrial systems, including C-accreting systems such as peatlands. Here, by means of metabarcoding and ecophysiological measurements, we examined how microbial photosynthesis and its biotic (e.g., phototrophic community structure, biomass) and abiotic drivers (e.g., Sphagnum moisture, light intensity) vary across peatland microhabitats. Using a natural gradient of microhabitat conditions from pool to forest, we show that the structure of phototrophic microbial communities shifted from a dominance of eukaryotes in pools to prokaryotes in forests. We identified five groups of co-occurring phototrophic operational taxonomic units with specific environmental preferences across the gradient. Along with such structural changes, we found that microbial C uptake was the highest in the driest and shadiest microhabitats. This study renews and improves current views on phototrophic microbes in peatlands, as the contribution of microbial photosynthesis to peatland C uptake has essentially been studied in wet microhabitats.     

Characterization of new denitrifying Rhodobacter strains isolated from photosynthetic sludge for wastewater treatment

New denitrifying strains of phototrophic bacteria isolated from photosynthetic sludge reactors for wastewater treatment were characterized. All of the new isolates were mesophilic, nonhalophilic, facultative photoheterotrophs that were able to grow by anaerobic photosynthesis, aerobic respiration, or nitrate respiration. They had ovoid cells that were motile by single polar flagella, formed vesicular photosynthetic membranes together with bacteriochlorophyll a and carotenoids of the spheroidene series, required biotin, thiamine, and biotin as growth factors, and utilized a wide variety of organic compounds as electron donor and carbon sources. In these respects, the isolates most closely resembled Rhodobacter sphaeroides. However, they differed from this species in utilizing malonate and dulcitol but not tartrate as carbon sources and in their inability to grow anaerobically in darkness with trimethylamine N-oxide or dimethylsulfoxide as a terminal oxidant. Partial sequencing of 16S rRNA genes provided evidence for genetic differences between the new isolates and R. sphaeroides or other members of the genus Rhodobacter. Activities of nitrate reductase, nitrite reductase, and nitrous oxide reductase were detected in intact cells of one of the new isolates. All these enzyme activities were induced by cultivation with nitrate.     

Settleability and kinetics of a nitrifying sludge in a sequencing batch reactor

A physiological study of a nitrifying sludge was carried out in a sequencing batch reactor (SBR). Pseudo steady-state nitrification conditions were obtained with an ammonium removal efficiency of 99% +/- 1% and 98% +/- 2% conversion of NH4+-N to NO3 - -N. The rate of biomass production was negligible (1.3 +/- 0.1 mg microbial protein-N.L(-1).d(-1)). The sludge presented good settling properties with sludge volume index values lower than 20 mL.g(-1) and an exopolymeric protein/carbohydrate ratio of 0.53 +/- 0.34. Kinetic results indicated that the nitrifying behavior of the sludge changed with the number of cycles. After 22 cycles, a decrease in the specific rate of NO3- -N production coupled with an increase in the NO2- -N accumulation were observed. These results showed that the activity of the nitrite oxidizing bacteria decreased at a longer operation time. Ammonia oxidizing bacteria were found to exhibit the best stability. After 4 months of operation, the specific rates of NH4+-N consumption and NO3- -N production were 1.72 NH4+-N per microbial protein-N per hour (g.g(-1).h(-1)) and 0.54 NO3- -N per microbial protein-N per hour (g.g(-1).h(-1)), respectively.     

Synthesis of bacteriochlorophyll <Emphasis Type="Italic">a</Emphasis> by the purple nonsulfur bacterium <Emphasis Type="Italic">Rhodobacter capsulatus</Emphasis>

The ability of purple nonsulfur bacteria Rhodobacter capsulatus B10 to synthesize bacteriochlorophyll under phototrophic and dark conditions was studied. The modes for cultivation in the dark with oxygen limitation in a continuous culture at D = 0.1 h^?1 were selected. The yield of biomass reached 20 g/l; the bacteriochlorophyll a output of the process amounted to 16.6 mg/l h^?1.     

Light environment and synthesis of bacteriochlorophyll by populations of Chromatium okenii under natural environmental conditions

Abstract: In the meromictic alpine Lake Cadagno a dense layer of phototrophic bacteria, mainly Chromatium okenii and Amoebobacter purpureus , develop annually at the chemocline at about 10 to 11 m depth. Radiometric spectral profiles of the incident sunlight demonstrate different attenuation coefficients in the mixolimnion and in the chemocline not only for the visible light effective at each depth (photosynthetically available radiation), but also for selected photosynthetically active wavelengths used by oxygenic and anoxygenic phototrophs. Phototrophic bacteria sampled from the upper part of the layer at the maximum of cell concentration were incubated in transparent bottles at the sampling depth and at a lower depth where the light intensity is only a few percent of the one at the sampling depth. Within 4 h the specific bacteriochlorophyll concentration (Bchl protein⁻¹) increased up to 50% depending on the difference in light intensity between the sampling and the incubation depth. The specific bacteriochlorophyll concentration in the upper part of the layer remained constant (53.0 mg Bchl g⁻¹ protein, S.D. = 4.8) in spite of large changes in cell concentrations in the lake water over the season. These observations illustrate the phenomenon of light-regulated pigment synthesis under natural conditions.     

Nitrogen removal performance of a hybrid anammox reactor

The anaerobic ammonium oxidation (anammox) process has attracted considerable attention in recent years as an alternative to conventional nitrogen removal technologies. In this study, an innovative hybrid reactor combining fluidized and fixed beds for anammox treatment was developed. The fluidized bed was mechanically stirred and the gaseous product could be rapidly released from the anammox sludge to prevent washout of the sludge caused by floatation. The fixed bed comprising a non-woven biomass carrier could efficiently catch sludge to reduce washout. During the operation, nitrogen loading rates to the reactor were increased to 27.3 kg N/m³/d, with total nitrogen removal efficiencies of 75%. The biomass concentration in the fluidized bed reached 26-g VSS/L. Anammox granules were observed in the reactors, with settling velocities and sludge volumetric index of 27.3 ± 6.5 m/h and 23 mL/g, respectively. Quantification of extracellular polymeric substances revealed the anammox granules contained a significant amount of extracellular proteins.     

Enhancing post anaerobic digestion of full-scale anaerobically digested sludge using free nitrous acid treatment

In some wastewater treatment plants (WWTPs), the ever increasing production of sludge with the expanding population overloaded the anaerobic digestion which compromises the sludge reduction efficiency. Post anaerobic digestion of anaerobically digested sludge (ADS) has been applied to enhance sludge reduction, however, to a very limited extent. This study verified the effectiveness of free nitrous acid (FNA i.e. HNO₂) pre-treatment on enhancing full-scale ADS degradation in post anaerobic digestion. The ADS collected from a full-scale WWTP was subject to FNA treatment at concentrations of 0.77, 1.54, 2.31, 3.08, and 3.85 mg N/L for 24 h followed by biochemical methane potential tests. The FNA treatment at all concentrations resulted in an increase (from 1.5–3.1 % compared to the control) in sludge reduction with the highest improvement achieved at 0.77 mg HNO₂-N/L. The FNA treatment at this concentration also resulted in the highest increase in methane production (40 %) compared to the control. The economic analysis indicates that FNA treatment is economically attractive for enhancing post anaerobic digestion of full-scale ADS.     

Nitrogen transformations under different conditions in open ponds by means of microalgae-bacteria consortium treating pig slurry

Four open ponds inoculated with microalgae-bacteria consortium treating different swine slurries (fresh and anaerobically digested) were evaluated in terms of nitrogen transformation under optimal and real conditions of temperature and illumination. Ammonium complete depletion was not achieved. Ponds operated under real conditions presented lower ammonium removal. Elimination capacities were around 26 mg N/L d and were subsequently increased with increasing inlet ammonium loading rate. Different nitrogen transformation was observed depending on substrate source. When anaerobically digested slurry was fed to the ponds, nitrification followed by biomass uptake and denitrification were the main nitrogen transformation taking place depending on inlet ammonium loading rate and operational conditions. Ponds fed with fresh slurry exhibited denitrification as the main nitrogen removal mechanism for the pond operated under real conditions while under optimal conditions stripping, denitrification and biomass uptake contributed similarly. Therefore, this study confirmed that the so-claimed nitrogen recovery by microalgae biomass is frequently overestimated.     

A multiple extraction--centrifugation method for the recovery of viruses from waste water treatment plant effluents and sludges

Absorption of 14C-labelled poliovirus-2 to sedimentable solids of primary sludge samples collected from a secondary treatment facility during a 6-month period averaged 94%; for anaerobically digested sludge, 99%. The extent of virus adsorption was influenced by the amount of solids. Maximal adsorption occurred at or above 0.5% solids with sludge diluted with deionized water and above 1.5% solids when diluted with the respective particle-free sludge supernatants. A Tris-HCl buffer containing NaCl, glycerol, and serum was found to efficiently elute poliovirus-2 from primary sludge solids. By means of re-extraction and concentration by centrifugation (the TEC procedure), the average recoveries of poliovirus-2 were 92-94% based upon either infectivity or radioactivity analyses. Similarly, recoveries were 90-92% for poliovirus-2 in digested sludge. Maximum elution was dependent upon all four TEC buffer components and the restriction of solids to less than or equal to 1.0%. The procedure was found to be more efficient than glycine-NaOH and Freon procedures or elution with beef embryo extract. As adapted for effluents the procedure increased the yield and improved the consistency of virus recovery. The arithmetic mean titers and obtained during a monitoring study for primary and digested sludge were 4.2 X 10(5) and 5.1 X 10(3) plaque-forming units (pfu)/L; for primary, secondary, and final effluents 2.3 X 10(5), 4.7 X 10(3), and 4.7 X 10(2) pfu/L, respectively.     

Nitrogen dynamics under Lolium perenne after a single application of three different sewage sludge types from the same treatment stream

Three sludge types from the same treatment stream (undigested liquid, anaerobically digested liquid and dewatered, anaerobically digested cake) were used in a field based tub study. Amendments (4, 8, and 16 Mg dry solid (ds)ha(-1)) were incorporated into the upper 15 cm of a sandy loam soil prior to sowing with rye-grass (Lolium perenne L.). Nitrogen transformations in the soil were determined for the 80 d period following incorporation. Nitrogen uptake and crop yield were measured in the cut sward 35 and 70 d after sowing. The study showed that application of sewage sludge at rates as low as 4 Mgha(-1) can have a nutritional benefit to rye-grass over the two harvests. Differences in N transformation, and hence crop nutritional benefit, between sludge types were evident throughout the experiment. In particular, the dewatering process changed the mineral N characteristics of the anaerobically digested sludge, which, when not dewatered, outperformed the other sludges in terms of yield and mineralisation rate at both harvests. The dewatered sludge produced the lowest yield of rye-grass. The undigested liquid sludge had the lowest foliar N and soil NO(3)-N concentrations, possibly immobilised as the large oxidisable C component of this sludge was metabolised by the microbial biomass. Correlation data support the concept of preferential uptake of NH(4)-N over NO(3)-N in Lolium perenne. Results are discussed in the context of managing sludge type and application for a plant nutrient source and NO(3)-N release.