Acetate treatment in 70°C upflow anaerobic sludge-blanket (UASB) reactors: start-up with thermophilic inocula and the kinetics of the UASB sludges

This study focused on the use of thermophilic anaerobic granulae in the start-up of 70°C acetate-fed upflow anaerobic sludge-blanket (UASB) reactors and the kinetics of granulae grown at 70°C. In the UASB reactors, chemical oxygen demand removal commenced within 48 h of the start-up. The maximum reduction in chemical oxygen demand was 84% with the feed containing yeast and 71% without a yeast supplement. In the bioassays, the yeast-grown sludge converted 98% of the acetate consumed to methane as compared to 92% for the sludge grown without yeast. The highest initial specific methane production rate (µ_{CH 4}) of the UASB sludges grown at 70°C was 0.088 h^–1 at an acetate concentration of 4.6 mM. The higher initial acetate concentration was found to prolong the lag-phase in methane production significantly and to decrease the µ_{CH 4}. The half-saturation constant (K _s), the inhibition constant (K _i), the inhibition response coefficient (n), and the µ_{CH 4} ^max, calculated according to a modified Haldane equation, were 1.5 mM, 2.8 mM, 0.8, and 0.28 h^–1, respectively. The prolonged starvation of the 70°C sludge (15 days) decreased the µ_{CH 4} from about 0.022 h^–1 to 0.011 h^–1 and increased the lag phase in methane production from 6 h to 24 h as compared to non-starved sludge.     

Microbial community dynamics during start-up operation of flowerpot-using fed-batch reactors for composting of household biowaste

Microbial community changes during start-up operation of flowerpot-using fed-batch reactors for composting of household biowaste were studied by quinone profiling, rRNA-targeted fluorescence in situ hybridization (FISH) and cultivation methods. Total and plate counts of bacteria and quinone contents in the reactors increased sharply with time during the start-up period. These increase patterns had two phases; the first increase occurred during 3-4 weeks from the start of waste loading and the second increase was found during the subsequent 4 weeks. The microbial biomass was temporally reduced between the two succession phases. Ubiquinones predominated at the beginning of operation but decreased sharply with time, whereas partially saturated menaquinones became predominant at the fully acclimated stage. These data indicated that the major constituents of microbial populations changed from ubiquinone-containing Proteobacteria to Actinobacteria during the period of operation. Neighbour-joining dendrograms constructed based on the quinone profile data suggested that at least one month is required to establish a stable community structure with the Actinobacteria predominating. The characteristic population shift in the start-up process was also demonstrated by FISH probing and 16S rDNA sequence comparisons of bacterial strains isolated.     

Impact of long-term alternations of discharge and spate on the chironomid community in the lowland Widawka River (Central Poland)

Spatial and temporal distribution, abundance and production of macroinvertebrate communities were estimated over two years in a fifth-order section of the Widawka River. Discharge of this river has been increased artificially by coal mine water inputs. Additionally, during the second year, one of the highest discharges of the current 20-year period was recorded. Chironomidae were co-dominant in macrobenthos, both in a straight reach (WIA) and in a meandering site (WIB). More mosaic habitats resulted in higher densities of midges, reaching 6215 ind.m^–2 in year 1 and 1141 ind.m^–2 in year 2 at WIA, while at WIB 896 densities were ind.m^–2 and 257 ind.m^–2, respectively. Flooding affected the distribution and abundance of the chironomid assemblages. Recolonization by psammophilous Polypedilum began after the various microhabitats were buried with sand. Chironomid production was estimated on a species-specific basis for the dominant taxa. In year 1 (mean annual water temperature 10.0° C) chironomid production was 12.4 g dry wt m^–2 yr^–1 1 at WIA and 1.9 g dry wt m^–2 yr^–1 at WIB. These values sharply decreased in year 2 (mean annual water temperature 9.8° C) reaching 1.9 g dry wt m^–2 yr^–1 at WIA and 0.4 g dry wt m^–2 yr^–1 at WIB, as effects of the high spate.     

Microbial populations and sludge characteristics in thermophilic aerobic sewage sludge digestion

Summary Estimates of bacterial numbers from raw sewage sludge and sludge treated by thermophilic aerobic digestion were compared with simple indicators of sludge quality and concentrations of potential substrates. Significant differences were found between sludge types for all but one of the variables examined (frequency of dividing cells). During a stable period of digestor operation, the average number of viable obligate thermophiles present in digested sludge (1.63 × 10⁶ ml^–1) was approximately 10²-fold greater than in feed sludge (1.10 × 10⁴ ml^–1). Total numbers of bacteria were slightly greater in digested sludge (3.24 × 10¹⁰ ml^–1) than in feed sludge (2.39 × 10 ml^–10), as were viable counts of bacteria at incubation temperatures of 37°C and 55°C. Significant correlation was found between viable counts of bacteria at 37°C and 55°C for digested sludge, and 65°C and 55°C for feed sludge. The numbers of obligate thermophiles present and the total of bacteria present were related to the temperature and pH of the digested sludge and inversely related to the numbers ofEscherichia coli and coliforms present, which were not detected at temperatures greater than 50°C.     

Temperature Effect on Carbachol-Induced Depression of Spontaneous Quantal Transmitter Release in Frog Neuromuscular Junction

The effects of carbachol (CCh) on the frequency (f) of the miniature endplate potentials were tested at temperatures between 5 and 30°C. Higher CCh concentrations, 1 × 10^–5 and 5 × 10^–6 M, reduced the f to 60% and the temperature dependence was negligible. However, an inverse temperature dependence was found when low concentrations 3 × 10^–7 and 6 × 10^–7 M were applied. The depression of f was 40–50% in 5–10°C but only 10–20% of the control in the 25 and 30°C. During application of CCh, the new steady of f was reached at temperatures between 5 and 30°C within 17–20 min (Q₁₀ = 1.07). Much greater temperature dependence of recovery was observed during washing out CCh (Q₁₀ = 1.6). The temperature-independence of the steady state effects of CCh, good agreement with Langmuir adsorption-desorption theory and non-steady kinetics indicate that physical rather than receptor-mediated events are responsible for the depression of f.     

Microbiological evaluation of processed rice consumed in Japan

Samples of processed rice from four different brands showed counts of mesophilic aerobic bacteria from 1 × 10² to 5 × 10³ for milled rice and from 1 × 10⁶ to 6.8 × 10⁶ cells g^–1 for brown rice. Rice seeds contaminated by milling had extensive counts including faecal coliforms. In cooked rice, no microbial growth was noted during 4 days at room temperature (spring season, 15–20 °C) and at least 2 weeks at 4 °C. No contamination was detected in cooked rice packs. A rapid and highly reliable procedure for detection of microorganisms in cooked rice is proposed.     

The influence of variable organic loading on the start-up of an anaerobic fluidised bed reactor

Summary A stepped-loading start-up regime utilising variable organic influent concentrations in the range 1650–11600 mgCOD1^–1 was applied to an anaerobic fluidised bed bioreactor at 37°C. The reactor was sensitive to variable influent COD concentrations, but the stepped-loading aided rapid recovery from transient organic loading shocks. Variable effluent COD levels were produced but a COD removal efficiency of 76% was obtained at a final HRT of 0.5 d and an organic loading rate of 5.3 kg COD m^–3 d^–1.     

Rapid granulation and sludge retention for tetrachloroethylene removal in an upflow anaerobic sludge blanket reactor

A laboratory scale upflow anaerobic sludge blanket (UASB) reactor was operated at 35 °C for over 200 days to investigate the granulation mechanism during tetrachloroethylene (TCE) biodegradation. Anaerobic, unacclimated sludge and glucose were used as seed and primary substrate, respectively. TCE-degrading granules developed after 1.5 months of start-up. They grew at an accelerated pace for 7 months. The TCE-degrading granules had a maximum diameter of 2.5 mm and specific methanogenic activity of 1.32 g chemical oxygen demand (COD) g^–1 total suspended solid (TSS) day^–1. 94% COD and 90% TCE removal efficiencies were achieved when the reactor was operating at loading rates as high as 160 mg TCE l^–1 day^–1 and 14 g COD l^–1 day^–1, after 230 days of continuous operation.     

Aerobic granulation in sequencing batch reactors with different reactor height/diameter ratios

Effects of reactor height/diameter ratios ranged from 24 to 4 corresponding to reactor settling velocities from 12 to 2 m h^?1 on aerobic granulation were investigated. It was found that granules appeared after 1-week operation and granule volume percentages exceeded 50% after 2–3 weeks in four reactors. In addition, similar granule fraction of 94–96% was found at steady state in all four reactors. Sludge volume index (SVI), average sludge size, biomass density and granule settling velocity at steady state were around 50 ml g^?1, 1800 μm, 53 g l^?1 and 40 m h^?1, respectively, in four reactors. Extracellular polymeric substances (EPS) and specific oxygen uptake rate (SOUR) were around 38 mg g^?1 VSS and 40 mg O₂ g^?1 VSS h^?1, respectively. Denaturing gradient gel electrophoresis (DGGE) fingerprint of sludge in four reactors showed the same microbial population shift during the start-up period and same microbial community structure during steady-state period. These results recommended strongly that reactor height/diameter ratio or reactor setting velocity in the used range in this study did not affect granule formation, physical characteristics, microbial community structure of granules and stable operation of granular sludge reactor. Reactor height/diameter ratio thus can be very flexible in the practice, which is important for the application of aerobic granule technology.     

Effect of temperature and long-term operation on passively immobilizedZymomonas mobilis for continuous ethanol production

Summary A fibrous support was used forZ. mobilis immobilization. The system showed a broad optimum temperature range (25–35°C) for highest ethanol productivity, ethanol yield and glucose conversion during continuous fermentation of a 100 g/L glucose medium. Ethanol production and glucose conversion kept steady during two months of continuous operation at D=1h^–1.     

Mass cultivation of Nannochloropsis sp. in annular reactors

A study was made on the mass cultivation of Nannochloropsis sp. in newly designed annular reactors operated under natural, artificial or combined illumination. The annular reactor consists of two 2-m-high Plexiglas cylinders of different diameter placed vertically one inside the other so as to form an annular culture chamber. Artificial illumination is supplied by lamps placed inside the inner cylinder. Two annular reactors of different diameter (50 and 91 cm), light path (4.5 and 3.0 cm) and illuminated surface area (5.3 and 9.3 m²) were experimented with. The effect of two different artificial light sources (fluorescent tubes and metal halide lamps) on culture productivity was investigated in both systems. The highest productivity on a per reactor basis (about 34 g (d. wt) reactor^–1 24 h^–1) was achieved with the larger reactor illuminated by a 400-W metal halide lamp. From February to May a 91-cm reactor illuminated only with natural light was operated in parallel with a 91-cm reactor subjected to combined illumination. Under natural illumination productivity increased from 16.6 g (d. wt) reactor^–1 d^–1 in February to 34.1 g (d. wt) reactor^–1 d^–1 in May. Under combined illumination productivity was 41.3 g (d. wt) reactor^–1 d^–1 in February and increased up to 48.3 g (d. wt) reactor^–1 d^–1 in May. Although the culture exposed to combined illumination always attained higher yields, the productivity gap between the two cultures decreased gradually along the season as solar radiation and minimum night temperatures increased. A 1200-L plant made of ten 50-cm annular reactors was set up and operated for two years with combined illumination yielding an average of 270 g of dry Nannochloropsis sp. biomass per day. More than 2000 L of concentrate suspension (50 g (d. wt) L^–1) of Nannochloropsis sp. were produced and successfully used by fish hatcheries as live feed for rotifers and for rearing seabream larvae with the green-water technique. This study indicates that the annular reactor can be profitably used for long-term cultivation of Nannochloropsis in temperate climates. Besides reliability and ease of operation, the main advantage of the system is that it can be used under natural illumination, yet artificial light can be also supplied to maintain high productivity levels in winter or on cloudy days.     

Outdoor cultivation ofArthrospira platensis during autumn and winter in temperate climates

Arthrospira (Spirulina) platensis M2 was grown outdoors in 50-mm diameter tubular reactors under the climatic conditions of central Italy (Florence) from September to December 1995 and in March 1996. Except for September, the cultures temperature was regulated. Mean productivities of 0.83, 0.44 and 0.61 g dry wt L^–1 d^–1 were achieved in autumn (September–October), winter (November–December) and March, respectively. In autumn and winter, the photosynthetic efficiency of the cultures and the degree of correlation between productivity and solar irradiance were significantly greater than in summer. The effect of cell density and aeration rate on productivity was evaluated in September. The productivity of cultures operated at high supra-optimal population density was about 30% less at high aeration rate (1.0 LL^–1 min^–1), and 50% less at standard aeration rate (0.17 LL^–1 min^–1), than that of control cultures kept at optimal population density and standard aeration rate. The reduction of productivity in high-density cultures was due to lower daylight output rates and higher night biomass losses (the latter were particularly relevant under standard aeration conditions). The main factor limiting productivity in closed reactors during autumn was the night temperature. Heating the cultures during daylight hours on sunny days did not cause any significant increase of the yields, since under sunlight the unheated cultures also reached the optimal temperature for growth early in the morning. On cloudy days, the day-time temperature of the unheated cultures remained well below the optimum, however this had only a limited effect on productivity since algal growth was mainly light-limited.     

Biomass and icosapentaenoic acid productivities from an outdoor batch culture of Phaeodactylum tricornutum UTEX 640 in an airlift tubular photobioreactor

An outdoor tubular photobioreactor, based on an external-loop airlift, has been used to cultivate the diatom Phaeodactylum tricornutum UTEX 640, rich in icosapentaenoic acid (EPA). The system was operated in batch mode. The specific growth rate in the exponential phase, averaged over the whole day, was 0.254 day^–1 · A 25 gl^–1 maximum biomass concentration was obtained by the end of the culture period. The dissolved oxygen generated never rose over 210% of air saturation nor was there impediment to biomass generation or inhibition of photosynthesis. Furthermore, around 2 × 10¹⁷ quanta cm^–2 s^–1 light intensity seemed to saturate the photosynthetic activity. Biomass losses during the night could be reduced to less than 5% by maintaining the culture at 16°C at night. The maximum concentration and productivity of EPA were 423 mgl^–1 and 13 mgl^–1 day^–1, respectively, at the end of the culture, although a 21-mgl^–1 day^–1 net EPA generation rate was obtained at the linear growth phase.     

Method for measuring microbial degradation and mineralization of14C-labeled chitin obtained from the blue crab,Callinectes sapidus

A method for measuring microbial degradation and mineralization of radiolabeled native chitin is described.¹⁴C-labeled chitin was synthesized in vivo by injecting shed blue crabs (Callinectes sapidus) with N-acetyl-D-[1-¹⁴C]-glucosamine and allowing for its incorporation into the exoskeleton. The cuticle had a total organic carbon content of 0.48 mg C mg^–1 with a specific radioactivity of 6,356 CPM mg^–1. Glucosamine, i.e., chitin content as determined colorimetrically, was 22% (w/w). Microbial degradation and mineralization rates were assessed in batch culture using¹⁴C-chitin as substrate and York River water as inoculum. Replicate flasks were sampled daily for enumeration of chitinoclastic bacteria and the radiolabel recovered as particulate¹⁴C-chitin or¹⁴CO₂. The amount of¹⁴CO₂ generated was directly proportional to the loss of particulate¹⁴C-chitin, with 96% of the added label recovered as the sum of both phases. The maximum rate of mineralization was 207 mg day^–1 g^–1 seeded¹⁴C-chitin at 20°C. Highest chitinoclastic bacterial counts corresponded to the period of maximum rate of chitinolysis. It is suggested that the rate of chitin mineralization is limited by exoenzymatic depolymerization and not by chitin concentration.VIMS Contribution no. 1215.     

Pyrolytic characteristics of heterotrophic Chlorella protothecoides for renewable bio-fuel production

Heterotrophic Chlorella protothecoides cells were pyrolyzed in athermogravimetric analyzer to investigate the pyrolytic characteristics anddetermine the kinetic parameters. Heating rates of 15, 40, 60 and 80 ^°C^-1 up to a final temperature of 800 ^°C wereused. The pyrolysis reactions mainly took place between 160–520^°C with a volatile yield of about 80%. The devolatilization stageconsisted of two main temperature zones (I and II) with a transition at300–320 ^°C. Crude lipid in cells decomposed at Zone II whileother main components at Zone I. The increase of heating rate caused alateral shift to higher temperatures in the thermograms, a decrease ofactivation energies for the devolatilization stage and an increase of both theinstantaneous maximum and average reaction rates. The difference ofactivation energies between two zones implied that more energy input forlipid pyrolysis seems needed in comparison with other main components.These data are useful for the design, operation, and modeling of thepyrolysis systems for microalgae.     

Continuous butanol production from whey permeate with immobilized Clostridium beyerinckii LMD 27.6

Summary The aim of this study was to find the conditions necessary for the continuous butanol production from whey permeate with Clostridium beyerinckii LMD 27.6, immobilized in calcium alginate beads. The influence of three parameters on the butanol production was investigated: the fermentation temperature, the dilution rate (during start-up and at steady state) and the concentration of calcium ions in the fermentation broth. It was found that both a fermentation temperature of 30° C and a dilution rate of 0.1 h^-1 or less during the start-up phase are required to achieve continuous butanol production from whey permeate. Butanol can be produced continuously from whey permeate in reactor productivities sixteen times higher than those found in batch cultures with free C. beyerinckii cells on whey media.     

Hypertonic cryohemolysis of human red blood cells

Summary Hypertonic cryohemolysis of human erythrocytes is caused by incubation of the cells in hypertonic medium at a temperature of 20–50°C (stage 1), with subsequent cooling to 0°C (stage 2). In 0.86m sucrose hemolysis increases, with increasing stage 1 temperature, whereas in 1m NaCl cryohemolysis has a temperature optimum at a stage 1 temperature of about 30°C.Cryohemolysis is inhibited by preceding ATP depletion of the cells and bypreincubation of the cells in hypertonic medium at 0°C. In general, anesthetics inhibit cryohemolysis strongly. Only in 1m NaCl at stage 1 temperatures in the range of 40–50°C is cryohemolysis stimulated by these drugs, if present during the entire incubation period. This effect is abolished, however, when the anesthetic is added after piror incubation of the cells at 40–50°C in 1m NaCl.Ghost-bound ANS fluorescence indicates complicated conformation changes in the membrane structure during the various experimental stages leading to cryohemolysis.Some of the experimental results can be considered as examples of molecular hysteresis, thus indicating several different metastable structures of the membrane, under various experimental conditions.The described results support the working hypothesis of Green and Jung that the experimental procedure results in membrane protein damage, preventing normal adaptation of the membrane during cooling.     

Stimulation of artemisinin production in Artemisia annua hairy roots by the elicitor from the endophytic Colletotrichum sp.

Artemisinin content in hairy roots of Artemisia annua was increased from 0.8 mg g^–1 dry wt to 1 mg g^–1 dry wt by using elicitor treatment of mycelial extracts from the endophytic fungus Colletotrichum sp. The increase of artemisinin was dependent on the growth stage of hairy roots as well as on the dose of the elicitor applied. When hairy roots of 23-day-old cultures (later growth phase) were exposed to the elicitor at 0.4 mg total sugar ml^–1 for 4 days, the maximum production of artemisinin reached to 13 mg l^–1, a 44% increase over the control. This is the first report on the stimulation of artemisinin production in hairy roots by the elicitor from an endophytic fungus of A. annua.     

Continuous acetonitrile degradation in a packed-bed bioreactor

A 20-l packed-bed reactor filled with foamed glass beads was tested for the treatment of acetonitrile HPLC wastes. Aeration was provided by recirculating a portion of the reactor liquid phase through an aeration tank, where the dissolved oxygen concentration was kept at 6 mg/l. At a feeding rate of 0.77 g acetonitrile l^–1 reactor day^–1, 99% of the acetonitrile was removed; and 86% of the nitrogen present in acetonitrile was released as NH₃, confirming that acetonitrile volatilization was not significant. Increasing the acetonitrile loading resulted in lower removal efficiencies, but a maximum removal capacity of 1.0 g acetonitrile l^–1 reactor day^–1 was achieved at a feeding rate of 1.6 g acetonitrile l^–1 reactor day^–1. The removal capacity of the system was well correlated with the oxygenation capacity, showing that acetonitrile removal was likely to be limited by oxygen supply. Microbial characterization of the biofilm resulted in the isolation of a Comamonas sp. able to mineralize acetonitrile as sole carbon, nitrogen and energy source. This organism was closely related to C. testosteroni (91.2%) and might represent a new species in the Comamonas genus. This study confirms the potential of packed-bed reactors for the treatment of a concentrated mixture of volatile pollutants.     

Erratum to: Bacterial diversity in toxic<Emphasis Type="Italic"> Alexandrium tamarense</Emphasis> blooms off the Orkney Isles and the Firth of Forth

The genetic diversity of the bacterial community associated with Alexandrium tamarense blooms was studied in blooms of the toxic dinoflagellates in the waters around the Orkney Isles and the Firth of Forth (Scotland). For toxin and molecular analysis of the bacterial communities associated with the toxic bloom, water samples were taken in 1998 and 1999 from A. tamarense blooms. The bacterial community structure, as determined by DGGE (denaturing gradient gel electrophoresis) showed clear differences between all three investigated size fractions (dinoflagellate-associated bacteria, attached bacteria and free-living bacteria), with high diversity within each sample. DNA sequence analysis of the dominant and most frequent DGGE bands revealed the dominance of Proteobacteria, mainly of the Roseobacter clade, with similarities of 91–99%. Moreover, DGGE bands occurring at the same position in the gel throughout in most samples corroborate the presence of several specific Proteobacteria of the Roseobacter clade. Overall, 500 bacteria were isolated from the bloom and partly phylogenetically analysed. They were members of two prokaryotic phyla, the Proteobacteria and the Bacteroidetes, related to Proteobacteria of the and subdivisions (Alteromonas, Pseudoalteromonas and Colwellia). All bacteria were tested for the production of sodium channel blocking (SCB) toxins using mouse neuroblastoma assay. No production of SCB toxins was found and high performance liquid chromatography (HPLC) analysis confirmed these results. The content of total paralytic shellfish poisoning (PSP) toxin in the water samples, as measured within the toxic dinoflagellate blooms using HPLC, ranged from 53 to 2191 ng PSP l^–1 in 1998 and from 0 to 478 ng PSP l^–1 in 1999. Changes in PSP toxin content were not accompanied by changes of DGGE band patterns. We therefore presume that the bacterial groups identified in this study were not exclusively associated with toxic A. tamarense, but were generally associated with the phytoplankton.An erratum to this article can be found at Communicated by H.-D. Franke