Difference in sporogenous bacterial populations in thermophilic (55 degrees C) and mesophilic (35 degrees C) anaerobic sewage digestion

Spores, sporeforming vegetative cells, and asporogenous populations were enumerated in two semicontinuous anaerobic fermentors digesting municipal primary sludge at 35 and 55 degrees C for more than 87 days. In the 35 degrees C fermentor, the anaerobic total population was 312.5 X 10(6)/ml, with 25.0 X 10(6)/ml being sporogenous. The populations that digest casein, starch, pectin, and cellulose were 23.1 X 10(6), 59.2 X 10(6), 26.2 X 10(6), and 7.3 X 10(6)/ml, respectively, with 2.8 X 10(6), 6.7 X 10(6), 3.4 X 10(6), and 1.5 X 10(6)/ml being sporogenous, respectively. The sporeformers accounted for 8.0 to 20.0% of each of the respective populations. In the 55 degrees C fermentor, the anaerobic total population was 512.5 X 10(6)/ml, with 336.6 X 10(6)/ml being sporogenous. The populations that digest casein, starch, pectin, and cellulose were 97.7 X 10(6), 190.7 X 10(6), 75.8 X 10(6), and 11.2 X 10(6)/ml, respectively, with 47.8 X 10(6), 110.6 X 10(6), 43.3 X 10(6), and 5.1 X 10(6)/ml, respectively, being sporogenous. The sporeformers represented 45.5 to 65.7% of each of the respective populations. The numbers of thermophilic sporeforming vegetative cells in the 55 degrees C fermentor were 9.0 to 19.8 times higher than their counterparts in the 35 degrees C fermentor. Most sporeformers were in the vegetative state in the 35 and 55 degrees C fermentors. After 18 days of fermentation at 55 degrees C, sporeformers carried out most of the digestion; however, the digestion was shared by both sporeformers and asporogenous bacteria after 87 days of fermentation. In the 35 degrees C fermentor, asporogenous bacteria digested most of the sludge. During the 18- and 87-day experimental periods, sporeformers were never predominant.     

Methane from cattle waste: Effects of temperature, hydraulic retention time, and influent substrate concentration on kinetic parameter (k)

The effects of temperature (35 and 55 degrees C), influent volatile solids (VS) concentration (S(0) = 43, 64, 82, 100, and 128 kg VS/m(3)) and hydraulic retention time (HRT = 4, 5, 8, 10, 15, and 25 days) on methane (CH(4)) production from cattle waste were evaluated using 3-dm(3) laboratoryscale fermentors. The highest CH(4) production rate achieved was 6.11 m(3) CH(4) m(-3) fermentor day(-1) at 55 degrees C, four days HRT, and S(0) = 100 kg VS/m(3). Batch fermentations showed an ultimate CH(4) yield (B(0)) of 0.42 m(3) CH(4)/kg VS fed. The maximum loading rates for unstressed fermentation were 7 kg VS m(-3) day(-1) at 35 degrees C and 20 kg VS m(-3) day(-1) at 55 degrees C. The kinetic parameter (K, an increasing K indicates inhibition of fermentation) increased exponentially as S(0) increased, and was described by: K = 0.8 + 0.0016 e(0.06S(0) ). Temperature had no significant effect on K for S(0) between 40 and 100 kg VS/m(3). The above equation predicted published K values for cattle waste within a mean standard error of 7%.     

Comparison of two-stage thermophilic (68 degrees C/55 degrees C) anaerobic digestion with one-stage thermophilic (55 degrees C) digestion of cattle manure

A two-stage 68 degrees C/55 degrees C anaerobic degradation process for treatment of cattle manure was studied. In batch experiments, an increase of the specific methane yield, ranging from 24% to 56%, was obtained when cattle manure and its fractions (fibers and liquid) were pretreated at 68 degrees C for periods of 36, 108, and 168 h, and subsequently digested at 55 degrees C. In a lab-scale experiment, the performance of a two-stage reactor system, consisting of a digester operating at 68 degrees C with a hydraulic retention time (HRT) of 3 days, connected to a 55 degrees C reactor with 12-day HRT, was compared with a conventional single-stage reactor running at 55 degrees C with 15-days HRT. When an organic loading of 3 g volatile solids (VS) per liter per day was applied, the two-stage setup had a 6% to 8% higher specific methane yield and a 9% more effective VS-removal than the conventional single-stage reactor. The 68 degrees C reactor generated 7% to 9% of the total amount of methane of the two-stage system and maintained a volatile fatty acids (VFA) concentration of 4.0 to 4.4 g acetate per liter. Population size and activity of aceticlastic methanogens, syntrophic bacteria, and hydrolytic/fermentative bacteria were significantly lower in the 68 degrees C reactor than in the 55 degrees C reactors. The density levels of methanogens utilizing H2/CO2 or formate were, however, in the same range for all reactors, although the degradation of these substrates was significantly lower in the 68 degrees C reactor than in the 55 degrees C reactors. Temporal temperature gradient electrophoresis profiles (TTGE) of the 68 degrees C reactor demonstrated a stable bacterial community along with a less divergent community of archaeal species.     

The transformation and toxicity of anthraquinone dyes during thermophilic (55 degrees C) and mesophilic (30 degrees C) anaerobic treatments

We studied in batch assays the transformation and toxicity of anthraquinone dyes during incubations with anaerobic granular sludge under mesophilic (30 degrees C) and thermophilic (55 degrees C) conditions. Additionally, the electron shuttling capacity of the redox mediator anthraquinone-2-sulfonic acid (AQS) and subsequent increase on decolourisation rates was investigated on anthraquinone dyes. Compared with incubations at 30 degrees C, serum bottles at 55 degrees C presented distinctly higher decolourisation rates not only with an industrial wastewater containing anthraquinone dyes, but also with model compounds. Compared with batch assays at 30 degrees C, the first-order rate constant "k" of the Reactive Blue 5 (RB5) was enhanced 11-fold and 6-fold for bottles at 55 degrees C supplemented and free of AQS, respectively. However, the anthraquinone dye Reactive Blue 19 (RB19) demonstrated a very strong toxic effect on volatile fatty acids (VFA) degradation and methanogenesis at both 30 degrees C and 55 degrees C. The apparent inhibitory concentrations of RB19 exerting 50% reduction in methanogenic activity (IC50-value) were 55 mg l(-1) at 30 degrees C and 45 mg l(-1) at 55 degrees C. Further experiments at both temperatures revealed that RB19 was mainly toxic to methanogens, because the glucose oxidizers including acetogens, propionate-forming, butyrate-forming and ethanol-forming microorganisms were not affected by the dye toxicity.     

Psychrophilic Microorganisms from Areas Associated with the Viking Spacecraft

Microorganisms capable of growth at 7 C were enumerated and isolated from soil samples from the manufacture and assembly areas of the Viking spacecraft. Populations ranging from 4.2 X 10(3) to 7.7 X 10(6)/g of soil were isolated from the 15 soil samples examined. Temperature requirements were determined, and those growing at 3 C, but not at 32 C, were designated as obligate psychrophiles in this investigation. Populations of soil bacteria, including aerobic sporeformers, ranging from 1.5 X 10(2) to 9.8 X 10(5)/g were capable of growth at 3 C, but not at 32 C. Bacterial isolates were identified to major generic groups. No psychrophilic sporeformers were isolated from soil from the manufacture area, but psychrophilic sporeformers ranged from 0 to 6.1 X 10(3)/g from soil from the assembly area.     

Adaptation of mesophilic anaerobic sewage fermentor populations to thermophilic temperatures.

Thermophilic (50 degrees C) and obligately thermophilic (60 degrees C) anaerobic carbohydrate- and protein-digesting and methanogenic bacterial populations were enumerated in a mesophilic (35 degrees C) fermentor anaerobically digesting municipal primary sludge. Of the total bacterial population in the mesophilic fementor, 9% were thermophiles (36 x 10(6)/ml) and 1% were obligate thermophiles (4.5 x 10(6)/ml). Of these 10%, the percentages of bacteria (thermophiles and obligate thermophiles, respectively) able to use specific substrates were further enumerated as follows: bacteria able to digest albumin, casein, starch, and mono- and disaccharides, 30 and 10%; pectin degraders, 10 and 0.2%; cellulose degraders, 2 and 0.06%; methanogens that grow with H2 and CO2, methanol, and dimethylamine, 9 and 1%; methanogens that grow with formate, 8 and 5%; and methanogens that grow with acetate, 25 and less than 0.8%. Shortly after the temperature was elevated from 35 to 50 or 60 degrees C, the digestion of albumin, casein, starch, and mono- and disaccharides was detected, and methane was produced from H2 and CO2. Methane produced from acetate was not delayed at 50 degrees C, but was delayed by 29 days at 60 degrees C. Methane produced from formate was delayed by 3 days, from methanol by 7 days, and from dimethylamine by 5 days at 50 and 60 degrees C. A 10- and 20-day acclimation period was required for hydrolysis of pectin and cellulose, respectively, at 50 degrees C. Digestion of pectin required 20 days and cellulose longer than 85 days when the temperature was elevated abruptly from 35 to 60 degrees C. The acclimation period for the digestion of pectin and cellulose at 60 degrees C was shortened to 3 and 15 days, respectively, by seeding with a small amount of a culture acclimated to 50 degrees C. The data suggest that enrichment of cellulolytic, pectinolytic, and acetate-utilizing bacteria is crucial for the digestion of sewage sludge at 60 degrees C.     

Adaptation of mesophilic anaerobic sewage fermentor populations to thermophilic temperatures

Thermophilic (50 degrees C) and obligately thermophilic (60 degrees C) anaerobic carbohydrate- and protein-digesting and methanogenic bacterial populations were enumerated in a mesophilic (35 degrees C) fermentor anaerobically digesting municipal primary sludge. Of the total bacterial population in the mesophilic fementor, 9% were thermophiles (36 x 10(6)/ml) and 1% were obligate thermophiles (4.5 x 10(6)/ml). Of these 10%, the percentages of bacteria (thermophiles and obligate thermophiles, respectively) able to use specific substrates were further enumerated as follows: bacteria able to digest albumin, casein, starch, and mono- and disaccharides, 30 and 10%; pectin degraders, 10 and 0.2%; cellulose degraders, 2 and 0.06%; methanogens that grow with H2 and CO2, methanol, and dimethylamine, 9 and 1%; methanogens that grow with formate, 8 and 5%; and methanogens that grow with acetate, 25 and less than 0.8%. Shortly after the temperature was elevated from 35 to 50 or 60 degrees C, the digestion of albumin, casein, starch, and mono- and disaccharides was detected, and methane was produced from H2 and CO2. Methane produced from acetate was not delayed at 50 degrees C, but was delayed by 29 days at 60 degrees C. Methane produced from formate was delayed by 3 days, from methanol by 7 days, and from dimethylamine by 5 days at 50 and 60 degrees C. A 10- and 20-day acclimation period was required for hydrolysis of pectin and cellulose, respectively, at 50 degrees C. Digestion of pectin required 20 days and cellulose longer than 85 days when the temperature was elevated abruptly from 35 to 60 degrees C. The acclimation period for the digestion of pectin and cellulose at 60 degrees C was shortened to 3 and 15 days, respectively, by seeding with a small amount of a culture acclimated to 50 degrees C. The data suggest that enrichment of cellulolytic, pectinolytic, and acetate-utilizing bacteria is crucial for the digestion of sewage sludge at 60 degrees C.     

Morphological change of the acrosome on motile bovine spermatozoa due to storage at 4 degrees C

Swelling of the apical ridge and anterior acrosome of motile bovine spermatozoa was observed during in-vitro storage using differential interference-contrast optics. This morphological alteration is different from that described as the false acrosome reaction on immotile spermatozoa, apparent in ageing semen samples and which has been associated with cell death. In this study, transmission electron microscopy revealed that the apical ridge acrosomal matrix was extended into complex folds and/or projections. Acrosomal and plasma membrane integrity was retained. Storing spermatozoa (1500 X 10(6)/ml) in seminal plasma at 4 degrees C for 1 day was most conducive to the swelling of the apical ridge. Replacing seminal plasma with egg yolk-citrate inhibited swelling. However, incubating semen at 37 degrees C in egg yolk-Tris-fructose extender (25 X 10(6) spermatozoa/ml) after storage in egg yolk-citrate at 4 degrees C for greater than or equal to 3 days restored the swelling characteristic.     

Thermophilic (55-65 degrees C) and extreme thermophilic (70-80 degrees C) sulfate reduction in methanol and formate-fed UASB reactors

The feasibility of thermophilic (55-65 degrees C) and extreme thermophilic (70-80 degrees C) sulfate-reducing processes was investigated in three lab-scale upflow anaerobic sludge bed (UASB) reactors fed with either methanol or formate as the sole substrates and inoculated with mesophilic granular sludge previously not exposed to high temperatures. Full methanol and formate degradation at temperatures up to, respectively, 70 and 75 degrees C, were achieved when operating UASB reactors fed with sulfate rich (COD/SO4(2-)=0.5) synthetic wastewater. Methane-producing archaea (MPA) outcompeted sulfate-reducing bacteria (SRB) in the formate-fed UASB reactor at all temperatures tested (65-75 degrees C). In contrast, SRB outcompeted MPA in methanol-fed UASB reactors at temperatures equal to or exceeding 65 degrees C, whereas strong competition between SRB and MPA was observed in these reactors at 55 degrees C. A short-term (5 days) temperature increase from 55 to 65 degrees C was an effective strategy to suppress methanogenesis in methanol-fed sulfidogenic UASB reactors operated at 55 degrees C. Methanol was found to be a suitable electron donor for sulfate-reducing processes at a maximal temperature of 70 degrees C, with sulfide as the sole mineralization product of methanol degradation at that temperature.     

Methane Production by Fermentor Cultures Acclimated to Waste from Cattle Fed Monensin, Lasalocid, Salinomycin, or Avoparcin

The ability of microorganisms to ferment waste from cattle fed monensin, lasalocid, or salinomycin to methane was determined. Continuously mixed anaerobic fermentors with 3-liter working volumes at 55°C were used; fermentors were fed once per day. Initially, all fermentors were fed waste without antibiotics at 6% volatile solids (VSs, organic matter) and a 20-day retention time (RT) for 60 days. Waste from animals fed monensin, lasalocid, or salinomycin at 29, 20, and 16.5 mg per kg of feed, respectively, was added to duplicate fermentors at the above VSs, and RT. Avoparcin (5 to 45 mg/liter) was not fed to animals but was added directly to duplicate fermentors. Lasalocid and salinomycin had minimal effects on the rate of methane production at RTs of 20 days and later at 6.5 days. Avoparcin caused an increase in organic acids from 599 to 1,672 mg/liter (as acetate) after 4 weeks, but by 6 weeks, acid concentrations declined and the rate of methane production was similar to controls at a 6.5-day RT. The monensin fermentors stopped producing methane 3 weeks after antibiotic addition. However, after a 6-month acclimation period, the microorganisms apparently adapted, and methane production rates of 1.65 and 2.51 liters per liter of fermentor volume per day were obtained with 6% VSs, and RTs of 10 and 6.5 days, respectively. This compares with 1.78 and 2.62 liters/liter per day for controls (P > 0.05). All fermentors that were fed waste containing antibiotics had lower pH values and ammonia and alkalinity concentrations, suggesting less buffering capacity and protein catabolism than in controls. Acclimation results obtained with fermentors at 35°C were similar to those for fermentors at 55°C. These studies indicate that waste from cattle fed these selected growth-promoting antibiotics can be thermophilically fermented to methane at RTs of 6.5 days or longer and VS concentrations of 6%, at rates comparable to waste without antibiotics.     

Removal of bacteria from water by adhesion to cross-linked poly(vinylpyridinium halide).

Cross-linked poly(vinylpyridinium halide) was found to have a novel and remarkable ability to remove bacteria from water. For example, when 10 g (wet weight) of cross-linked poly(N-benzyl-4-vinylpyridinium bromide) was contacted with 20 ml of suspensions of Escherichia coli (9.7 X 10(4) to 9.7 X 10(7)/ml), Salmonella typhimurium (8.0 X 10(6) to 1.1 X 10(7)/ml), Streptococcus faecalis (5.0 X 10(7)/ml), Staphylococcus aureus (8.1 X 10(7)/ml), and Pseudomonas aeruginosa (3.2 X 10(5)/ml) under stirring in sterilized physiological saline at 37 degrees C, 99% of the viable cells of these bacteria were removed in 2 to 6 h. When suspensions of these bacteria (10(5) to 10(8) cells per ml) were passed through a column (20 mm by 100 cm) of cross-linked poly(N-benzyl-4-vinylpyridinium bromide) at 37 degrees C with a flow rate of 0.8 to 1.4 bed volumes per h, 97 to 100% of the viable cells were eliminated from the suspensions during the treatment. Mechanistic studies demonstrated that cross-linked poly(vinylpyridinium halide) irreversibly captured these bacteria alive during the treatment. That is, total organic carbon was removed during the treatment, and the bacteria which adhered to the resin proliferated on the bacterial medium. The adhesion capacity was estimated to be 10(10) cells per g (dry weight). Total organic carbon was also removed even when the bacteria were killed by heat treatment before the column studies.     

Bioaugmentation of a two-stage thermophilic (68 degrees C/55 degrees C) anaerobic digestion concept for improvement of the methane yield from cattle manure

The possibility of improving a two-stage (68 degrees C/55 degrees C) anaerobic digestion concept for treatment of cattle manure was studied. In batch experiments, a 10-24% increase of the specific methane yield from cattle manure and its fractions was obtained, when the substrates were inoculated with bacteria of the genus Caldicellusiruptor and Dictyoglomus. In a reactor experiment inoculation of a 68 degrees C pretreatment reactor with Caldicellusiruptor resulted in a 93% increase in the methane yield of the pretreatment reactor for a period of 18 days, but gave only a slight increase in the overall methane yield of the two-stage setup.     

Kinetic study of sulfide leaching by galvanic interaction between chalcopyrite, pyrite, and sphalerite in the presence of T. ferrooxidans (30 degrees C) and a thermophilic microorganism (55 degrees C)

A systematic, kinetic study and comparison of the leaching of mixed metal sulfides by galvanic conversion and in the presence of bacteria has been carried out for the first time using both powder (-100 to -400 mesh) and larger (bulk) specimen systems. The rates of dissolution of copper from chalcopyrite and zinc from sphalerite as single, electrically isolated (separate) systems were compared with electrically contacting (galvanically coupled) systems involving CuFeS(2)/FeS(2) and ZnS/FeS(2), with and without bacteria and at temperatures of 30 and 55 degrees C. The dissolution of Cu was observed to increase by a factor of 4.6 when the galvanic leaching of CuFeS(2)/FeS(2) was compared to CuFeS(2) leaching at 30 degrees C. When bacteria were present, Cu dissolution increased by an additional factor of 2.1 in the CuFeS(2)/FeS(2) system. At 55 degrees C, the corresponding ratios for Cu were 4.3 and 2.7, respectively. The galvanic leaching of Zn in the ZnS/FeS(2) system compared to ZnS leaching increased by a factor of 2 at 30 degrees C; in the presence of bacteria the dissolution of Zn from the ZnS/FeS(2) system increased by an additional factor of 1.3 at the same temperature. By comparison, the ratio of Cu dissolution from CuFeS(2) in acid-bacterial medium and sterile controls (without bacteria) was 5.5. The corresponding ratio for Zn from ZnS was 2.2 at both 30 and 55 degrees C. The order of reaction was found to be essentially first order for the leaching of powder systems at both 30 and 55 degrees C (with T. Ferrooxidans and thermophilic microorganisms, respectively). The corresponding reaction rate constants were observed to be 12.6 and 22.9 for T. ferrooxidans and the thermophilic microorganisms, respectively. Activation energies for the various systems were also determined.     

Thermotolerance induced by heat, sodium arsenite, or puromycin: its inhibition and differences between 43 degrees C and 45 degrees C

When CHO cells were treated either for 10 min at 45-45.5 degrees C or for 1 hr with 100 microM sodium arsenite (ARS) or for 2 hr with 20 micrograms/ml puromycin (PUR-20), they became thermotolerant to a heat treatment at 45-45.5 degrees C administered 4-14 hr later, with thermotolerance ratios at 10(-3) isosurvival of 4-6, 2-3.2, and 1.7, respectively. These treatments caused an increase in synthesis of HSP families (70, 87, and 110 kDa) relative to total protein synthesis. However, for a given amount of thermotolerance, the ARS and PUR-20 treatments induced 4 times more synthesis than the heat treatment. This decreased effectiveness of the ARS treatment may occur because ARS has been reported to stimulate minimal redistribution of HSP-70 to the nucleus and nucleolus. Inhibiting protein synthesis with cycloheximide (CHM, 10 micrograms/ml) or PUR (100 micrograms/ml) after the initial treatments greatly inhibited thermotolerance to 45-45.5 degrees C in all cases. However, for a challenge at 43 degrees C, thermotolerance was inhibited only for the ARS and PUR-20 treatments. CHM did not suppress heat-induced thermotolerance to 43 degrees C, which was the same as heat protection observed when CHM was added before and during heating at 43 degrees C without the initial heat treatment. These differences between the initial treatments and between 43 and 45 degrees C may possibly be explained by reports that show that heat causes more redistribution of HSP-70 to the nucleus and nucleolus than ARS and that redistribution of HSP-70 can occur during heating at 42 degrees C with or without the presence of CHM. Heating cells at 43 degrees C for 5 hr after thermotolerance had developed induced additional thermotolerance, as measured with a challenge at 45 degrees C immediately after heating at 43 degrees C. Compared to the nonthermotolerant cells, thermotolerance ratios were 10 for the ARS treatment and 8.5 for the initial heat treatment. Adding CHM (10 micrograms/ml) or PUR (100 micrograms/ml) to inhibit protein synthesis during heating at 43 degrees C did not greatly reduce this additional thermotolerance. If, however, protein synthesis was inhibited between the initial heat treatment and heating at 43 degrees C, protein synthesis was required during 43 degrees C for the development of additional thermotolerance to 45 degrees C.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effect of Dietary Monensin or Chlortetracycline on Methane Production from Cattle Waste

Wastes from feedlot cattle fed finishing diets containing either monensin, chlortetracycline, or no antibiotic were investigated as substrates for methane production. We used continuously mixed anaerobic fermentors with 3-liter working volumes at 35 and 55°C; these fermentors were fed once per day. Within a few days after waste from animals fed monensin was added, the volume of methane produced began to decrease in the 55°C fermentors. After 9 days of daily feeding, methane production was severely inhibited, the pH dropped from 7.6 to 5.9, and the concentration of volatile acids increased from 543 to 6,300 mg/liter (as acetate). Although additions of waste from cattle fed monensin were discontinued after 9 days, the fermentors did not resume gas production within 8 weeks. The addition of waste from cattle which had been fed chlortetracycline reduced the methane production rate approximately 20%; however, pH and volatile acid values were comparable to control fermentor values after 40 days. Similar effects were observed with the 35°C fermentors. In a batch fermentation experiment in which 50-g portions of volatile solids from waste of animals fed monensin, chlortetracycline, or no antibiotics were added to fermentors, monensin delayed the onset of methane production for about 40 days, but then these fermentors began to produce methane at a rate comparable to the control rate. The ultimate methane yields from the three types of waste after 180 days were not significantly different. These studies indicate that monensin has a detrimental effect on the conversion of feedlot wastes to methane, unless microorganisms can be adapted to the levels that are present in these wastes.     

Methane production in an UASB reactor operated under periodic mesophilic-thermophilic conditions

Methane production was studied in a laboratory-scale 10 L anaerobic upflow sludge bed (UASB) reactor with periodic variations of the reactor temperature. On a daily basis the temperature was varied between 35 and 45 degrees C or 35 and 55 degrees C with a heating period of 6 h. Each temperature increase was accompanied by an increase in methane production and a decrease in the concentration of soluble organic matter in the effluent. In comparison to a reactor operated at 35 degrees C, a net increase in methane production of up to 22% was observed. Batch activity tests demonstrated a tolerance of mesophilic methanogenic populations to short-term, 2-6 h, temperature increases, although activity of acetoclastic methanogens decreased after 6 h exposure to a temperature of 55 degrees C. 16S sequencing of DGGE bands revealed proliferation of temperature-tolerant Methanospirillum hungatii sp. in the reactor.     

Effect of Temperature and Retention Time on Methane Production from Beef Cattle Waste

The effect of temperature and retention time on the rate of methane production from waste of beef cattle fed a finishing diet was investigated by using continuously mixed 3-liter working volume anaerobic fermentors. The temperatures ranged from 30 to 65°C with 5°C increments between fermentors. The fermentors were fed once per day with 6% volatile solids (organic matter). Retention time for each temperature was varied from 18 to 2.5 days. After 3-volume turnovers, samples were obtained on 4 consecutive days. The highest methane production rate (liters/liter of fermentor per day) and methane yield at that rate (liters/gram of volatile solids) were 1.27 and 0.19 at 9 days and 30°C, 1.60 and 0.16 at 6 days and 35°C, 2.28 and 0.23 at 6 days and 40°C, 2.42 and 0.24 at 6 days and 45°C, 2.83 and 0.14 at 3 days and 50°C, 2.75 and 0.14 at 3 days and 55°C, 3.18 and 0.14 at 2.5 days and 60°C, and 1.69 and 0.17 at 6 days and 65°C. Volatile solids degradation at these retention times and temperatures was between 46 and 54%. The concentrations of volatile acids in the 30 to 55°C fermentors were generally below 2,000 mg/liter, with the exception of the 3-day retention time. The 60 and 65°C fermentors were usually above this level for all retention times. These studies indicate potential rates of methane production from the fermentation of untreated waste of beef cattle fed high-grain finishing diets. This information should serve as preliminary guidelines for various kinetic analyses and aid in economic evaluations of the potential feasibility of fermenting beef cattle waste to methane.     

Acetate Degradation at 70 degrees C in Upflow Anaerobic Sludge Blanket Reactors and Temperature Response of Granules Grown at 70 degrees C

Anaerobic acetate degradation at 70 degrees C and at 55 degrees C (as a reference) was studied by running laboratory upflow anaerobic sludge blanket (UASB) reactors inoculated with mesophilic granular sludge. In UASB reactors fed with acetate-containing media (3 g of chemical oxygen demand [COD] per liter, corresponding to 47 mM acetate) approximately 50 days was needed at 70 degrees C and less than 15 days was needed at 55 degrees C to achieve an effluent COD of 500 to 700 mg/liter. In the UASB reactors at both 70 and 55 degrees C up to 90% of the COD was removed. Batch assays showed that sludges from two 70 degrees C UASB reactors, one run at a low effluent acetate concentration and the other run at a high effluent acetate concentration, exhibited slightly different responses to temperatures in the range from 37 to 70 degrees C. Both 70 degrees C sludges, as well as the 55 degrees C sludge, produced methane at temperatures of 37 to 73 degrees C. The 55 degrees C sludge exhibited shorter lag phases than the 70 degrees C sludges and higher specific methane production rates between 37 and 65 degrees C.     

Effects of Temperature on Methanogenesis in a Thermophilic (58 degrees C) Anaerobic Digestor

The short-term effects of temperature on methanogenesis from acetate or CO(2) in a thermophilic (58 degrees C) anaerobic digestor were studied by incubating digestor sludge at different temperatures with C-labeled methane precursors (CH(3)COO or CO(2)). During a period when Methanosarcina sp. was numerous in the sludge, methanogenesis from acetate was optimal at 55 to 60 degrees C and was completely inhibited at 65 degrees C. A Methanosarcina culture isolated from the digestor grew optimally on acetate at 55 to 58 degrees C and did not grow or produce methane at 65 degrees C. An accidental shift of digestor temperature from 58 to 64 degrees C during this period caused a sharp decrease in gas production and a large increase in acetate concentration within 24 h, indicating that the aceticlastic methanogens in the digestor were the population most susceptible to this temperature increase. During a later period when Methanothrix sp. was numerous in the digestor, methanogenesis from CH(3)COO was optimal at 65 degrees C and completely inhibited at 75 degrees C. A partially purified Methanothrix enrichment culture derived from the digestor had a maximum growth temperature near 70 degrees C. Methanogenesis from CO(2) in the sludge was optimal at 65 degrees C and still proceeded at 75 degrees C. A CO(2)-reducing Methanobacterium sp. isolated from the digestor was capable of methanogenesis at 75 degrees C. During the period when Methanothix sp. was apparently dominant, sludge incubated for 24 h at 65 degrees C produced more methane than sludge incubated at 60 degrees C, and no acetate accumulated at 65 degrees C. Methanogenesis was severely inhibited in sludge incubated at 70 degrees C, but since neither acetate nor H(2) accumulated, production of these methanogenic substrates by fermentative bacteria was probably the most temperature-sensitive process. Thus, there was a correlation between digestor performance at different temperatures and responses to temperature by cultures of methanogens believed to play important roles in the digestor.     

Behavior of Bacillus thuringiensis var. kurstaki under continuous phased cultivation in a cyclone fermentor

Summary Growth, sporulation, insecticidal crystalline protein (ICP) production and plasmids of Bacillus thuringiensis var. kurstaki (HD-1) were investigated during batch and continuous phased cultivation using a laboratory scale cyclone fermentor. When grown in batch culture at 28°C, 93% of the cells sporulated and produced ICP within 10 h of commencement of stationary phase. The batch culture runs were completed within 50 h of inoculation. A predominantly sporogenous and crystalliferous cell population was also obtained by second, stage processing of culture harvested from the first 10 to 25 cycles of continuous phased cultivation. In contrast, after 25 or more cycles of cultivation the population in the continuous phased culture shifted towards a predominantly asporogenous and acrystalliferous one. Culture conditions in continuous phased cultivation did not affect the plasmid content of B. thuringiensis (HD-1), yet influenced sporulation and plasmid-coded ICP production.