Noriatsu Ozaki

In situ activity and spatial organization of anaerobic ammonium-oxidizing (anammox) bacteria in biofilms.

ABSTRACT We investigated autotrophic anaerobic ammonium-oxidizing (anammox) biofilms for their spatial organization, community composition, and in situ activities by using molecular biological techniques combined with microelectrodes. Results of phylogenetic analysis and fluorescence in situ hybridization (FISH) revealed that “Brocadia”-like anammox bacteria that hybridized with the Amx820 probe dominated, with 60 to 92% of total bacteria in the upper part (<1,000 μm) of the biofilm, where high anammox activity was mainly detected with microelectrodes. The relative abundance of anammox bacteria decreased along the flow direction of the reactor. FISH results also indicated that Nitrosomonas-, Nitrosospira-, and Nitrosococcus-like aerobic ammonia-oxidizing bacteria (AOB) and Nitrospira-like nitrite-oxidizing bacteria (NOB) coexisted with anammox bacteria and accounted for 13 to 21% of total bacteria in the biofilms. Microelectrode measurements at three points along the anammox reactor revealed that the NH4+ and NO2− consumption rates decreased from 0.68 and 0.64 μmol cm−2 h−1 at P2 (the second port, 170 mm from the inlet port) to 0.30 and 0.35 μmol cm−2 h−1 at P3 (the third port, 205 mm from the inlet port), respectively. No anammox activity was detected at P4 (the fourth port, 240 mm from the inlet port), even though sufficient amounts of NH4+ and NO2− and a high abundance of anammox bacteria were still present. This result could be explained by the inhibitory effect of organic compounds derived from biomass decay and/or produced by anammox and coexisting bacteria in the upper parts of the biofilm and in the upstream part of the reactor. The anammox activities in the biofilm determined by microelectrodes reflected the overall reactor performance. The several groups of aerobic AOB lineages, Nitrospira-like NOB, and Betaproteobacteria coexisting in the anammox biofilm might consume a trace amount of O2 or organic compounds, which consequently established suitable microenvironments for anammox bacteria.

Influences of air temperature change on leisure industries – case study on ski activities –

To evaluate the influences of air temperature change on ski activities, thechanges in the numbers of skiers visiting seven ski areas in Japan were predicted in conjunction with climate change. First, having built a model forpredicting snow depth based on the budgets of water and heat using the air temperature and precipitation data collected nationwide, we demonstrated goodagreement between the predicted and observed snow depths (p < 0.01 and the ratios for more than 81% cases ranged from 0.5 to 2). Second, the relationshipbetween the number of skiers and the depth of snow at one of the seven ski areaswas analyzed statistically on a daily basis. In addition, we did the same on amonthly basis at six other ski areas and compared the observed and predictednumbers of skiers (p < 0.01 and the ratios for more than 94% cases ranged from 0.5 to 2). Using this model and the relationship between daily snow depth andnumber of skiers, the changes in skier numbers in the seven ski areas werepredicted for several scenarios with respect to air temperature changes; e.g. a more than 30% drop in visiting skiers was forecast in almost all ski areas in Japan except northern region (Hokkaido) and/or high altitude regions (center of the Main Island) under the condition of a 3 °C increase in air temperature. The vulnerability of the ski industry and its adaptationto climate change are discussed.

Ecophysiological Role and Function of Uncultured Chloroflexi in an Anammox Reactor

The coexistence of uncultured heterotrophic bacteria belonging to the phylum Chloroflexi has often been observed in anaerobic ammonium oxidation (anammox) reactors fed with synthetic nutrient medium without organic carbon compounds. To determine if coexisting Chloroflexi in anammox reactors scavenge organic matter derived from anammox bacterial cells, the present study was conducted to investigate the substrate uptake pattern of the uncultured Chloroflexi present in an anammox reactor and to clarify if they take up microbial products derived from anammox bacterial cells. To accomplish this, combined microautoradiography and fluorescence in situ hybridization (MAR-FISH) was conducted. Phylogenetic analysis revealed that 36% of the clones analyzed in this study were affiliated with Chloroflexi. The sequence similarities to Anaerolinea thermophila and Caldilinea aerophila within the phylum Chloroflexi were only 81.0-88.7% and 80.3-83.8%, respectively. The uncultured Chloroflexi were found to incorporate sucrose, glucose, and N-acetyl-glucosamine. The (14)C-tracing experiment revealed that the uncultured Chloroflexi were clearly MAR-positive, indicating the utilization of decaying anammox bacterial cell materials. Taken together, these results indicate that coexisting uncultured Chloroflexi in anammox reactors scavenge organic compounds derived from anammox bacterial cells.

Biological oxidation of dissolved methane in effluents from anaerobic reactors using a down-flow hanging sponge reactor

Anaerobic wastewater treatment plants discharge dissolved methane, which is usually not recovered. To prevent emission of methane, which is a greenhouse gas, we utilized an encapsulated down-flow hanging sponge reactor as a post-treatment to biologically oxidize dissolved methane. Within 3 weeks after reactor start-up, methane removal efficiency of up to 95% was achieved with a methane removal rate of 0.8 kg COD m(-3) day(-1) at an HRT of 2 h. After increasing the methane-loading rate, the maximum methane removal rate reached 2.2 kg COD m(-3) day(-1) at an HRT of 0.5 h. On the other hand, only about 10% of influent ammonium was oxidized to nitrate during the first period, but as airflow was increased to 2.5 L day(-1), nitrification efficiency increased to approximately 70%. However, the ammonia oxidation rate then decreased with an increase in the methane-loading rate. These results indicate that methane oxidation occurred preferentially over ammonium oxidation in the reactor. Cloning of the 16S rRNA and pmoA genes as well as phylogenetic and T-RFLP analyses revealed that type I methanotrophs were the dominant methane oxidizers, whereas type II methanotrophs were detected only in minor portion of the reactor.

Physiological Characterization of an Anaerobic Ammonium-Oxidizing Bacterium Belonging to the “Candidatus Scalindua” Group

ABSTRACT The phylogenetic affiliation and physiological characteristics (e.g., Ks and maximum specific growth rate [μmax]) of an anaerobic ammonium oxidation (anammox) bacterium, “Candidatus Scalindua sp.,” enriched from the marine sediment of Hiroshima Bay, Japan, were investigated. “Candidatus Scalindua sp.” exhibits higher affinity for nitrite and a lower growth rate and yield than the known anammox species.

Hydraulic effects on sludge accumulation on membrane surface in crossflow filtration.

Membrane filtration technology for application of wastewater treatment has been developing recently. In the application to wastewater treatment, it is major concern to remove cake layer on membrane surface effectively with crossflow shear stress. Hydraulic effect of sludge accumulation process on membrane surface in bubble and non-bubble driven crossflow filtration was studied. Maximum sludge accumulation. sludge accumulation rate, and lag phase were introduced to describe sludge accumulation process, and the effects of hydraulic conditions were clarified experimentally. Maximum sludge accumulation and sludge accumulation rate were dependent on aeration intensity, and were less depend on flow channel width and MLSS concentration. Their tendencies were explained by shear stress. Shear stress was thought to be the major hydraulic factor that influences them. Lag phase was dependent on aeration intensity, flow channel width, and MLSS concentration. A non-dimensional equation was proposed to explain dependencies of flow channel width based on consideration of hydraulic behavior of MLSS particles and shear stress.

Dissolved methane oxidation and competition for oxygen in down-flow hanging sponge reactor for post-treatment of anaerobic wastewater treatment

Post-treatment of anaerobic wastewater was undertaken to biologically oxidize dissolved methane, with the aim of preventing methane emission. The performance of dissolved methane oxidation and competition for oxygen among methane, ammonium, organic matter, and sulfide oxidizing bacteria were investigated using a lab-scale closed-type down-flow hanging sponge (DHS) reactor. Under the oxygen abundant condition of a hydraulic retention time of 2h and volumetric air supply rate of 12.95m(3)-airm(-3)day(-1), greater than 90% oxidation of dissolved methane, ammonium, sulfide, and organic matter was achieved. With reduction in the air supply rate, ammonium oxidation first ceased, after which methane oxidation deteriorated. Sulfide oxidation was disrupted in the final step, indicating that COD and sulfide oxidation occurred prior to methane oxidation. A microbial community analysis revealed that peculiar methanotrophic communities dominating the Methylocaldum species were formed in the DHS reactor operation.

Forecasting the changes in lake water quality in response to climate changes, using past relationships between meteorological conditions and water quality

In order to forecast the effects of global warming on the water environment, the relationship between meteorological conditions and lake water quality was investigated statistically using 17 years of monitoring data obtained from a shallow eutrophic lake, Lake Kasumigaura. The usefulness of the DPY (difference from the previous year) method was confirmed for removing the watershed change (e.g. land cover, population, etc.). From the analysis of the relationships between air temperature and water temperature on a monthly basis, the delay of time was seen to be negligible, but the gain was reduced due to the high frequency of change. As the amount of precipitation affected their relationship on a yearly basis, the slopes of 1·0–1·2 °C water temperature/ °C air temperature were determined with the DPY method by excluding the combinations of the years having large difference in annual precipitation. The deterioration of lake water quality, such as increases in COD (chemical oxygen demand) and decreases in transparency, was quantitatively assessed as corresponding to an increase in air temperature. In addition, we found that higher precipitation led to high nitrogen concentrations on a monthly basis, as well as on a yearly basis, probably induced by both the runoff of soilwater having high concentrations and the lowering of residence times of lake water. Copyright

Phosphate recovery as concentrated solution from treated wastewater by a PAO-enriched biofilm reactor

Phosphorus recovery from wastewaters and its recycling are of importance for sustaining agricultural production. During the conventional enhanced biological phosphorus removal process, phosphorus is removed by withdrawing excess sludge from wastewater. However, excess sludge disposal is costly and energy intensive. A proposed novel process for phosphorus recovery from sewage treatment will result in no excess sludge if a polyphosphate accumulating organisms (PAOs) enrichment biofilm can be applied to effluents containing phosphate. This process allows the recovery of phosphate as phosphate-concentrated solutions by controlling PAOs to absorb and release phosphate. A reactor consisting of a modified trickling filter with a synthetic substrate (5 mg P L⁻¹) was operated to form a PAO-enriched biofilm. As a result of the enrichment, the concentration of phosphate of >100 mg P L⁻¹ was successfully achieved. During this experiment, no sludge withdrawal was carried out over the duration of the operation of 255 days. To highlight the new process, the principle of enriching PAOs on biofilm and concentrating phosphate from treated sewage is explained, and a discussion on phosphate recovery performance is given.

Phylogenetic diversity and ecophysiology of Candidate phylum Saccharibacteria in activated sludge

Candidate phylum Saccharibacteria (former TM7) are abundant and widespread in nature, but little is known about their ecophysiology and detailed phylogeny. In this study phylogeny, morphology and ecophysiology of Saccharibacteria were investigated in activated sludge from nine wastewater treatment plants (WWTPs) from Japan and Denmark using the full-cycle 16S rRNA approach in combination with microautoradiography (MAR) and fluorescence in situ hybridization (FISH). Phylogenetic analysis showed that Saccharibacteria from all WWTPs were evenly distributed within subdivision 1 and 3 and in a distinct phylogenetic clade. Three probes were designed for the distinct saccharibacterial groups, and revealed morphotypes representing thin filaments, thick filaments and rods/cocci. MAR-FISH results showed that most probe-defined Saccharibacteria utilized glucose under aerobic-, nitrate reducing- and anaerobic conditions. Some Saccharibacteria also utilized N-acetylglucosamine, oleic acid, amino acids and butyrate, which are not predicted from available genomes so far. In addition, some filamentous Saccharibacteria exhibited β-galactosidase and lipase activities determined using a combination of enzyme-labeled fluorescence and FISH (ELF-FISH). No uptake of acetate, propionate, pyruvate, glycerol and ethanol was observed. These results indicate that Saccharibacteria is a phylogenetically diverse group and play a role in the degradation of various organic compounds as well as sugar compounds under aerobic-, nitrate reducing- and anaerobic conditions.