Chie Katsuyama

Complementary cooperation between two syntrophic bacteria in pesticide degradation.

Interactions between microbial species, including competition and mutualism, influence the abundance and distribution of the related species. For example, metabolic cooperation among multiple bacteria plays a major role in the maintenance of consortia. This study aims to clarify how two bacterial species coexist in a syntrophic association involving the degradation of the pesticide fenitrothion. To elucidate essential mechanisms for maintaining a syntrophic association, we employed a mathematical model based on an experimental study, because experiment cannot elucidate various conditions for two bacterial coexistence. We isolated fenitrothion-degrading Sphingomonas sp. TFEE and its metabolite of 3-methyl-4-nitrophenol (3M4N)-degrading Burkholderia sp. MN1 from a fenitrothion-treated soil microcosm. Neither bacterium can completely degrade fenitrothion alone, but they can utilize the second intermediate, methylhydroquinone (MHQ). Burkholderia sp. MN1 excretes a portion of MHQ during the degradation of 3M4N, from which Sphingomonas sp. TFEE carries out degradation to obtain carbon and energy. Based on experimental findings, we developed mathematical models that represent the syntrophic association involving the two bacteria. We found that the two bacteria are characterized by the mutualistic degradation of fenitrothion. Dynamics of two bacteria are determined by the degree of cooperation between two bacteria (i.e., supply of 3M4N by Sphingomonas sp. TFEE and excretion of MHQ by Burkholderia sp. MN1) and the initial population sizes. The syntrophic association mediates the coexistence of the two bacteria under the possibility of resource competition for MHQ, and robustly facilitates the maintenance of ecosystem function in terms of degrading xenobiotics. Thus, the mathematical analysis and numerical computations based on the experiment indicate the key mechanisms for coexistence of Sphingomonas sp. TFEE and Burkholderia sp. MN1 in syntrophic association involving fenitrothion degradation.

N2O Emission from Degraded Soybean Nodules Depends on Denitrification by Bradyrhizobium japonicum and Other Microbes in the Rhizosphere

A model system developed to produce N2O emissions from degrading soybean nodules in the laboratory was used to clarify the mechanism of N2O emission from soybean fields. Soybean plants inoculated with nosZ-defective strains of Bradyrhizobium japonicum USDA110 (ΔnosZ, lacking N2O reductase) were grown in aseptic jars. After 30 days, shoot decapitation (D, to promote nodule degradation), soil addition (S, to supply soil microbes), or both (DS) were applied. N2O was emitted only with DS treatment. Thus, both soil microbes and nodule degradation are required for the emission of N2O from the soybean rhizosphere. The N2O flux peaked 15 days after DS treatment. Nitrate addition markedly enhanced N2O emission. A 15N tracer experiment indicated that N2O was derived from N fixed in the nodules. To evaluate the contribution of bradyrhizobia, N2O emission was compared between a nirK mutant (ΔnirKΔnosZ, lacking nitrite reductase) and ΔnosZ. The N2O flux from the ΔnirKΔnosZ rhizosphere was significantly lower than that from ΔnosZ, but was still 40% to 60% of that of ΔnosZ, suggesting that N2O emission is due to both B. japonicum and other soil microorganisms. Only nosZ-competent B. japonicum (nosZ+ strain) could take up N2O. Therefore, during nodule degradation, both B. japonicum and other soil microorganisms release N2O from nodule N via their denitrification processes (N2O source), whereas nosZ-competent B. japonicum exclusively takes up N2O (N2O sink). Net N2O flux from soybean rhizosphere is likely determined by the balance of N2O source and sink.

Hybrid Nitrous Oxide Production from a Partial Nitrifying Bioreactor: Hydroxylamine Interactions with Nitrite

The goal of this study was to elucidate the mechanisms of nitrous oxide (N2O) production from a bioreactor for partial nitrification (PN). Ammonia-oxidizing bacteria (AOB) enriched from a sequencing batch reactor (SBR) were subjected to N2O production pathway tests. The N2O pathway test was initiated by supplying an inorganic medium to ensure an initial NH4+-N concentration of 160 mg-N/L, followed by 15NO2- (20 mg-N/L) and dual 15NH2OH (each 17 mg-N/L) spikings to quantify isotopologs of gaseous N2O (44N2O, 45N2O, and 46N2O). N2O production was boosted by 15NH2OH spiking, causing exponential increases in mRNA transcription levels of AOB functional genes encoding hydroxylamine oxidoreductase (haoA), nitrite reductase (nirK), and nitric oxide reductase (norB) genes. Predominant production of 45N2O among N2O isotopologs (46% of total produced N2O) indicated that coupling of 15NH2OH with 14NO2- produced N2O via N-nitrosation hybrid reaction as a predominant pathway. Abiotic hybrid N2O production was also observed in the absence of the AOB-enriched biomass, indicating multiple pathways for N2O production in a PN bioreactor. The additional N2O pathway test, where 15NH4+ was spiked into 400 mg-N/L of NO2- concentration, confirmed that the hybrid N2O production was a dominant pathway, accounting for approximately 51% of the total N2O production.

Unique distribution of deep groundwater bacteria constrained by geological setting

We collected groundwater samples at depths of up to 482 m from three boreholes drilled into sedimentary rock within two formations in Hokkaido, Japan. The prokaryotic community in each subsurface groundwater sample was analysed by microscopic counts and cloning-sequencing the 16S rRNA genes. On total direct counts, there were between 4.61 × 10(4) and 5.06 × 10(6) prokaryote cells ml(-1) in the samples, which is similar to the numbers observed at the marine subsurface. However, the vertical distribution of the prokaryotes did not show a simple decrease in abundance with increasing depth. A high abundance of cells with significant amounts of RNA was identified in the domain Bacteria using fluorescence in situ hybridization, with a high frequency of dividing cells at the transition zone between the two sedimentary rock formations. Cloning-sequencing analysis showed the predominance of γ-Proteobacteria at this transition zone at 281-312 m. The horizontal heterogeneity of the microbial distribution in the subsurface environment was also demonstrated by a relatively high density of members of the domain Archaea in borehole HDB-4, drilled only 1.5 km northeast of HDB-6 and in the same formation.