Morio Miyahara

Nitrous oxide emission from nitrifying activated sludge dependent on denitrification by ammonia-oxidizing bacteria.

Nitrous oxide (N(2)O) is emitted during the aerated nitrification process of wastewater treatment, but its mechanism is not understood. In this study, we employed a model system to clarify the mechanism of N(2)O emission, utilizing the activated sludge derived from a piggery effluent. Aerated incubation of the sludge with ammonium (NH(4)(+)) or hydroxylamine (NH(2)OH) resulted in the emission of a significant amount of N(2)O. The emission stopped when the nitrification substrate (NH(4)(+) or NH(2)OH) was exhausted. When NH(4)(+) was replaced with nitrate (NO(3)(-)) and nitrite (NO(2)(-)), no N(2)O was emitted. This result suggests that the N(2)O emission under nitrifying conditions did not depend on the oxidation of NO(2)(-) by nitrite-oxidizing bacteria (NOB) or denitrification by heterotrophic denitrifiers but depended on the oxidation of NH(4)(+) by ammonia-oxidizing bacteria (AOB). When NO(2)(-), the product of nitrification by AOB, was added to the NH(4)(+)-oxidizing system, N(2)O emission was enormously enhanced, suggesting that N(2)O was formed via denitrification. Diethyldithiocarbamate (DCD), an inhibitor of copper-containing nitrite reductase (NirK), strongly blocked N(2)O emission from NH(2)OH. Furthermore, the expression of the gene (nirK) encoding NirK of AOB was detected in the sludge exposed to the nitrifying conditions. The results showed that N(2)O emission during the nitrification process depends on denitrification by AOB that reside in the activated sludge. This study provides direct evidence for the cause of N(2)O emission from activated sludge (non-pure culture).

Potential of Aerobic Denitrification by Pseudomonas stutzeri TR2 To Reduce Nitrous Oxide Emissions from Wastewater Treatment Plants

ABSTRACT In contrast to most denitrifiers studied so far, Pseudomonas stutzeri TR2 produces low levels of nitrous oxide (N2O) even under aerobic conditions. We compared the denitrification activity of strain TR2 with those of various denitrifiers in an artificial medium that was derived from piggery wastewater. Strain TR2 exhibited strong denitrification activity and produced little N2O under all conditions tested. Its growth rate under denitrifying conditions was near comparable to that under aerobic conditions, showing a sharp contrast to the lower growth rates of other denitrifiers under denitrifying conditions. Strain TR2 was tolerant to toxic nitrite, even utilizing it as a good denitrification substrate. When both nitrite and N2O were present, strain TR2 reduced N2O in preference to nitrite as the denitrification substrate. This bacterial strain was readily able to adapt to denitrifying conditions by expressing the denitrification genes for cytochrome cd1 nitrite reductase (NiR) (nirS) and nitrous oxide reductase (NoS) (nosZ). Interestingly, nosZ was constitutively expressed even under nondenitrifying, aerobic conditions, consistent with our finding that strain TR2 preferred N2O to nitrite. These properties of strain TR2 concerning denitrification are in sharp contrast to those of well-characterized denitrifiers. These results demonstrate that some bacterial species, such as strain TR2, have adopted a strategy for survival by preferring denitrification to oxygen respiration. The bacterium was also shown to contain the potential to reduce N2O emissions when applied to sewage disposal fields.

Electricity generation from cattle manure slurry by cassette-electrode microbial fuel cells.

Cassette-electrode microbial fuel cells (CE-MFCs) are efficient and scalable devices for electricity production from organic waste. Previous studies have demonstrated that CE-MFCs are capable of generating electricity from artificial wastewater at relatively high efficiencies. In this study, a single-cassette CE-MFC was constructed, and its capacity for electricity generation from cattle manure suspended in water (solid to water ratio of 1:50) was examined. The CE-MFC reactor was operated in batch mode for 49 days; electricity generation became stable 2 weeks after initiating the operation. The maximum power density was measured at 16.3 W m⁻³ on day 26. Sequencing analysis of PCR-amplified 16S rRNA gene fragments obtained from the original manure and from anode biofilms suggested that Chloroflexi and Geobacteraceae were abundant in the anode biofilm (29% and 18%, respectively), whereas no Geobacteraceae sequences were detected in the original manure sample. The results of this study suggest that CE-MFCs can be used to generate electricity from water-suspended cattle manure in a scalable MFC system.

Bioaugmentation of a wastewater bioreactor system with the nitrous oxide-reducing denitrifier Pseudomonas stutzeri strain TR2

In bioaugmentation technology, survival of inoculant in the treatment system is prerequisite but remains to be a crucial hurdle. In this study, we bioaugmented the denitrification tank of a piggery wastewater treatment system with the denitrifying bacterium Pseudomonas stutzeri strain TR2 in two pilot-scale experiments, with the aim of reducing nitrous oxide (N(2)O), a gas of environmental concern. In the laboratory, strain TR2 grew well and survived with high concentrations of nitrite (5-10 mM) at a wide range of temperatures (28-40°C). In the first augmentation of the pilot-scale experiment, strain TR2 inoculated into the denitrification tank with conditions (30°C, ~0.1 mM nitrite) survived only 2-5 days. In contrast, in the second augmentation with conditions determined to be favorable for the growth of the bacterium in the laboratory (40-45°C, 2-5 mM nitrite), strain TR2 survived longer than 32 days. During the time when the presence of strain TR2 was confirmed by quantitative real-time PCR, N(2)O emission was maintained at a low level even under nitrite-accumulating conditions in the denitrification and nitrification tanks, which provided indirect evidence that strain TR2 can reduce N(2)O in the pilot-scale system. Our results documented the effective application of growth conditions favorable for strain TR2 determined in the laboratory to maintain growth and performance of this strain in the pilot-scale reactor system and the decrease of N(2)O emission as the consequence.

Advenella faeciporci sp. nov., a nitrite-denitrifying bacterium isolated from nitrifying–denitrifying activated sludge collected from a laboratory-scale bioreactor treating piggery wastewater

Strain M-07(T) was isolated from nitrifying-denitrifying activated sludge treating piggery wastewater. Phylogenetic analysis based on 16S rRNA gene sequences demonstrated that strain M-07(T) belonged to the genus Advenella. 16S rRNA gene sequence similarity between M-07(T) and Advenella incenata CCUG 45225(T), Advenella mimigardefordensis DPN7(T) and Advenella kashmirensis WT001(T) was 96.5, 97.3 and 96.9%, respectively. The DNA G+C content of strain M-07(T) was 49.5 mol%, which was approximately 5 mol% lower than the range for the genus Advenella (53.5-58.0 mol%). The predominant cellular fatty acids of strain M-07(T) were C(16:0), summed feature 3 (comprising C(16:1)ω7c and/or iso-C(15:0) 2-OH), C(17:0) cyclo and summed feature 2 (comprising one or more of C(14:0) 3-OH, iso-C(16:1) I, an unidentified fatty acid with an equivalent chain-length of 10.928 and C(12:0) alde). The isoprenoid quinone was Q-8. On the basis of phenotypic characteristics, phylogenetic analysis and DNA-DNA relatedness, strain M-07(T) should be classified as a novel species of the genus Advenella, for which the name Advenella faeciporci sp. nov. is proposed. The type strain is M-07(T) ( = JCM 17746(T)  = KCTC 23732(T)).

Electricity generation from rice bran in microbial fuel cells

BackgroundRice bran is a by-product of the rice milling process and mostly discarded in Japan. Although many studies have shown that microbial fuel cells (MFCs) are able to generate electricity from organic wastes, limited studies have examined MFCs for generating electricity from rice bran.FindingsLaboratory-scale single-chamber MFCs were inoculated with paddy field soil and supplied with rice bran for examining electricity generation. Power outputs and microbiome compositions were compared between MFCs containing pure water as the liquid phase (MFC-W) and those containing mineral solution (MFC-M). Polarization analyses showed that both MFCs successfully generated electricity with the maximum power densities of 360 and 520 mW m−2 (based on the projected area of anode) for MFC-W and MFC-M, respectively. Amplicon-sequencing analyses revealed that Trichococcus and Geobacter specifically occurred in anode biofilms in MFC-W and MFC-M, respectively.ConclusionsThe results suggest that rice bran is a feasible fuel by itself for generating electricity in MFCs.

Sodium chloride concentration determines exoelectrogens in anode biofilms occurring from mangrove-grown brackish sediment

Single-chamber microbial fuel cells (MFCs) were inoculated with mangrove-grown brackish sediment (MBS) and continuously supplied with an acetate medium containing different concentrations of NaCl (0-1.8M). Different from MFCs inoculated with paddy-field soil (high power outputs were observed between 0.05 and 0.1M), power outputs from MBS-MFCs were high at NaCl concentrations from 0 to 0.6M. Amplicon-sequence analyses of anode biofilms suggest that different exoelectrogens occurred from MBS depending on NaCl concentrations; Geobacter occurred abundantly below 0.1M, whereas Desulfuromonas was abundant from 0.3M to 0.6M. These results suggest that NaCl concentration is the major determinant of exoelectrogens that occur in anode biofilms from MBS. It is also suggested that MBS is a potent source of microbes for MFCs to be operated in a wide range of NaCl concentrations.

Effectiveness of heat treatment to protect introduced denitrifying bacteria from eukaryotic predatory microorganisms in a pilot-scale bioreactor.

Bioaugmentation of bioreactor systems with pre-cultured bacteria has proven difficult because inoculated bacteria are easily eliminated by predatory eukaryotic-microorganisms. Here, we demonstrated an intermediate thermal treatment was effective for protecting introduced denitrifying bacteria from eukaryotic predators and consequently allowed the inoculated bacteria to survive longer in a denitrification reactor.

Streptomyces griseus Enhances Denitrification by Ralstonia pickettii K50, Which Is Possibly Mediated by Histidine Produced during Co-Culture

Ralstonia pickettii K50 (strain K50) is a denitrifying bacterium that produces low levels of N2O under aerobic conditions. In this study, we found that co-culturing of strain K50 with Streptomyces griseus significantly enhanced the denitrification activity of strain K50 in an artificial wastewater (AWW) system. Most factors that enhance denitrification activity were in the high molecular weight fraction of the cell-free broth of S. griseus, and were suggested to be extracellular proteases. Further investigation revealed that the cultivation of strain K50 in protease-treated AWW medium fully enhanced denitrification, and that a shortage of amino acids in the medium limited it. Among the 20 standard amino acids tested, only histidine had a significant effect in inducing denitrification by strain K50. Our results indicatate that histidine is a novel inducer of bacterial denitrification.

Anode macrostructures influence electricity generation in microbial fuel cells for wastewater treatment.

Microbial fuel cells (MFCs) are devices that exploit microbes for generating electricity from organic substrates, including waste biomass and wastewater pollutants. MFCs have the potential to treat wastewater and simultaneously generate electricity. The present study examined how anode macrostructure influences wastewater treatment, electricity generation and microbial communities in MFCs. Cassette-electrode MFCs were equipped with graphite-felt anodes with three different macrostructures, flat-plate (FP), vertical-fin (VF), and horizontal-fin (HF) structures (these were composed of a same amount of graphite felt), and were continuously supplied with artificial wastewater containing starch as the major organic constituent. Polarization analyses revealed that MFCs equipped with VF and HF anodes generated 33% and 21% higher volumetric power densities, respectively, than that of MFCs equipped with FP anodes. Organics were also more efficiently removed from wastewater in MFCs with VF and HF anodes compared to reactors containing FP anodes. In addition, pyrosequencing of PCR-amplified 16S rRNA gene fragments from microbial samples collected from the anodes showed that the presence of fins also affected the bacterial compositions in anode biofilms. Taken together, the findings presented here suggest that the modification of anodes with fins improves organics removal and electricity generation in MFCs. The optimization of anode macrostructure therefore appears to be a promising strategy for improving MFC performance without additional material costs.