Methylamine,an inhibitor of ammonium oxidation and chemoautotrophic growth in the marine nitrifying bacteriumNitrosococcus oceanus

Methylamine (CH₃NH ₃ ⁺ ) appeared to utilize the same transport mechanism as ammonium (NH ₄ ⁺ ) to enter cells ofNitrosococcus oceanus. Methylamine uptake did not show clear evidence of saturable kinetics and was not fully saturated at 20 mM. Assimilated CH₃NH ₃ ⁺ was not incorporated into macromolecular constituents, but inhibited rates of nitrification, chemoautotrophic CO₂ fixation and growth. The degree of inhibition was dependent on the relative concentrations of NH ₄ ⁺ and CH₃NH ₃ ⁺ . Rates of CO₂ fixation and growth were inhibited four times more than the rate of nitrification.     

Ammonia oxidation by the methane oxidising bacterium Methylococcus capsulatus strain bath

Soluble extracts of Methylococcus capsulatus (Bath) that readily oxidise methane to methanol will also oxidise ammonia to nitrite via hydroxylamine. The ammonia oxidising activity requires O₂, NADH and is readily inhibited by methane and specific inhibitors of methane mono-oxygenase activity. Hydroxylamine is oxidised to nitrite via an enzyme system that uses phenazine methosulphate (PMS) as an electron acceptor. The estimated K_mvalue for the ammonia hydroxylase activity was 87 mM but the kinetics of the oxidation were complex and may involve negative cooperativity.Abbreviations PMS Phenazine methosulphate - NADH nicotinamide adenine dinucleotide, reduced form - K_m Michaelis constant - NO₂^- nitrite - NH₂OH hydroxylamine     

Significance of gaseous NO for ammonia oxidation by Nitrosomonas eutropha

Nitrification by the obligately lithoautotrophic ammonia oxidizer Nitrosomonas eutropha was significantly inhibited when nitric oxide was removed from the culture medium by means of intensive aeration and turbulence. Nearly complete recovery of ammonia oxidation could be achieved by adding 100 ppm NO to the supplied air. Inhibition of ammonia oxidation occurred also upon addition of the NO binding agens 2,3-Dimercapto-1-propane-sulfonic acid (DMPS). Recovery of ammonia oxidation occurred within 3 h in the presence of 100 ppm NO and within 76 h in the absence of externally added NO. In co-cultures of N. eutropha and the NO detoxifying bacterium Pseudomonas PS88, hardly any nitrification was detectable and release of NO was extremely low when the heterotroph was provided with an organic substrate. When cells of Pseudomonas PS88 were added to a mixotrophically nitrifying culture of N. eutropha the release of NO decreased drastically upon the addition and ammonia oxidation ceased. These results confirm for the first time the significance of NO in the course of ammonia oxidation by N. eutropha.     

Description of a new species of Nitrosococcus

A new species of Nitrosococcus is described. It resembles Nitrosococcus oceanus in shape, size, and ultrastructure of the cells. However, the new species has a more pronounced salt requirement, corresponding to its natural habitats. Two strains were isolated from a salt lake in Saudi Arabia and a salt lagoon in the Mediterranean Sea, respectively. In contrast to N. oceanus, both isolates of the new species were unable to utilize urea as ammonia source. Both species also differed in gelelectrophoretic cell protein patterns. The name N. halophilus is proposed.     

Effect of metal-binding agents and other compounds on methane oxidation by two strains of Methylococcus capsulatus

Inhibition studies of methane mono-oxygenase activity in whole cell suspensions of Methylococcus capsulatus (Texas) and M. capsulatus (Bath) were performed and the results compared. The inhibition pattern for M. capsulatus (Bath) was not only substantially different from the pattern obtained with M. capsulatus (Texas) but also very limited in the number of potent inhibitors specific for methane oxidation. To confirm the whole cell results of M. capsulatus (Bath) similar experiments were done using cell-free extracts. It was found that only acetylene (100% inhibition) and 8-hydroxyquinoline (71%) significantly inhibited methane oxidation, verifying the restricted inhibition pattern found with the whole cell suspensions. Eight acetylenic compounds were tested for specific inhibition of methane oxidation by whole cells and cell-free extracts of M. capsulatus (Bath). Only two compounds (acetylene and propyne) gave 100% inhibition in both cases with three other compounds (but-1-yne, but-2-yne and propyn-1-ol) giving weaker inhibitions. The inhibition pattern of methane oxidation by whole cell suspensions and cell-free extracts of M. capsulatus (Bath) is discussed and reasons for the prominent results are suggested.     

Isolation of a moderate halophilic ammonia-oxidizing bacterium,Nitrosococcus mobilis nov. sp.

An ammonia-oxidizing bacterium was isolated from a sample of brackish water (North Sea, Harbour of Husum). It is a motile large coccus 1.5–1.7 m in diameter. The extensive cytomembrane system occurring as flattened vesicles in the peripheral region of the cytoplasm and as intrusions into the center of the cytoplasm is to be emphasized as a characteristic mark of identification. The lithoauto-trophically growing bacterium turned out to be an obligate halophile. Because of its physiological and morphological properties, we assigned it to the genus Nitrosoccus and propose the name Nitrosococcus mobilis.     

Gaseous NO2 as a regulator for ammonia oxidation of Nitrosomonas eutropha

Cells of Nitrosomonas eutropha strain N904 that were denitrifying under anoxic conditions with hydrogen as electron donor and nitrite as electron acceptor were unable to utilize ammonium (ammonia) as an energy source. The recovery of ammonia oxidation activity was dependent on the presence of NO₂. Anaerobic ammonia oxidation activity was observed in a helium atmosphere supplemented with 25 ppm NO₂ after 20 h. Ammonia oxidation activity was detected after 2–3 days using an oxic atmosphere with 25 ppm NO₂. In contrast, ammonia consumption started after 8–9 days under oxic conditions without the addition of NO₂; in this case, small amounts of NO and NO₂ were detected and their concentrations increased with increasing ammonia oxidation activities. Hardly any ammonia oxidation was detected when nitrogen oxides were removed by intensive aeration. It would seem, therefore, that NO₂ is the master regulatory signal for ammonia oxidation in Nitrosomonas eutropha. Anaerobic ammonia oxidation activity was inhibited by the addition of NO. This inhibition was partly compensated by either increasing the NO₂ concentration or by using 2,3-dimercapto-1-propane-sulfonic acid as a NO binding substrate. DMPS was inhibitory to nitrification under oxic conditions, while increased amounts of NO or NO₂ led to increased oxidation activities.     

Thermophilic methane production and oxidation in compost

Methane cycling within compost heaps has not yet been investigated in detail. We show that thermophilic methane oxidation occurred after a lag phase of up to one day in 4-week old, 8-week old and mature (>10-week old) compost material. The potential rate of methane oxidation was between 2.6 and 4.1 micromol CH4(gdw)(-1)h(-1). Profiles of methane concentrations within heaps of different ages indicated that 46-98% of the methane produced was oxidised by methanotrophic bacteria. The population size of thermophilic methanotrophs was estimated at 10(9) cells (gdw)(-1), based on methane oxidation rates. A methanotroph (strain KTM-1) was isolated from the highest positive step of a serial dilution series. This strain belonged to the genus Methylocaldum, which contains thermotolerant and thermophilic methanotrophs. The closest relative organism on the basis of 16S rRNA gene sequence identity was M. szegediense (>99%), a species originally isolated from hot springs. The temperature optimum (45-55 degrees C) for methane oxidation within the compost material was identical to that of strain KTM-1, suggesting that this strain was well adapted to the conditions in the compost material. The temperatures measured in the upper layer (0-40 cm) of the compost heaps were also in this range, so we assume that these organisms are capable of effectively reducing the potential methane emissions from compost.     

The relationship between inorganic nitrogen oxidation and organic carbon production in batch and chemostat cultures of marine nitrifying bacteria

Rates of nitrification and organic C production were determined in batch and chemostat cultures of marine nitrifying bacteria; two NH ₄ ⁺ -oxidizing species and one NO ₂ ^– -oxidizing spezies. With increasing age in batch cultures and with decreasing flow rates in chemostats, cellular organic C and N concentrations declined while the intracellular ratio of C:N remained constant. With decreasing flow rates in chemostats, there was a reduction in (a) carboxylating enzyme activity per unit of cellular organic C (the potential for chemoautotrophic CO₂ fixation), and (b) the yield of organic C. For both NH ₄ ⁺ and NO ₂ ^– oxidizers, rates of nitrification and C yield were lowest at very slow chemostat growth rates, when compared with optimal growth rates in batch cultures. For both NH ₄ ⁺ and NO ₂ ^– -oxidizing species, the stoichiometric relationship between nitrification and organic C production did not remain constant and appeared to be dependent on the availability of the inorganic N substrate. The organic C yield from NH ₄ ⁺ oxidation and hence the free energy efficiency declined with increasing age in batch cultures and with decreasing flow rates in chemostats. The C yield from NO ₂ ^– oxidation and the free energy efficiency at slow chemostat growth rates was also lower than that at the optimal growth rate in batch culture.     

The methane monooxygenase gene cluster of Methylosinus trichosporium: cloning and sequencing of the mmoC gene

Methane monooxygenase (MMO) is the enzyme responsible for the conversion of methane to methanol in methanotrophic bacteria. The soluble MMO enzyme complex from Methylosinus trichosporium also oxidizes a wide range of aliphatic and aromatic compounds in a number of potentially useful biotransformations. In this study we have used heterologous DNA probes from the type X methanotroph Methylococcus capsulatus (Bath) to isolate mmo genes from the type II methanotroph M. trichosporium. We report here that the gene encoding the reductase component, Protein C of MMO, lies adjacent to the genes encoding the other components of soluble MMO in M. trichosporium but is separated by an open reading frame of unknown function, orfY. The complete nucleotide sequence of these genes is presented. Sequence analysis of mmoC indicates that the N-terminus of Protein C has significant homology with 2Fe2S ferredoxins from a wide range of organisms.Abbreviations MMO methane monooxygenase     

Diel methane emissions in stands of Spartina alterniflora and Suaeda salsa from a coastal salt marsh

In this study, we aimed to understand the influence of plant type on the monthly variations of diel CH₄ fluxes from Spartina alterniflora and Suaeda salsa of coastal salt marshes at three growth stages (July, August and September). Dissolved CH₄ concentrations in porewater and sediment redox potentials were monitored, as were aboveground plant biomass and stem densities. CH₄ fluxes exhibited clear monthly variations and peaked in September in the S. alterniflora and S. salsa mesocosms. However, no discernible diel variation was observed in the CH₄ flux in the S. salsa mesocosm, probably due to its weak gas transport capacity. By contrast, notable diel variations of CH₄ flux with the peak of 1.42 and 3.67 mg CH₄ m⁻² h⁻¹ at 12:00 and the lowest of 0.75 and 2.11 mg CH₄ m⁻² h⁻¹ at 3:00 or 6:00 were observed in the S. alterniflora mesocosm on 11 August and 11 September, respectively, but not in July mainly due to low plant biomass masking diel variations in the porewater CH₄ concentration. The ratios of the maximum flux to minimum flux over the course of the day in the S. alterniflora mesocosm on 10 July, 11 August and 11 September were 1.28, 1.89 and 1.76, respectively, and corresponding values for porewater CH₄ concentration were 1.31, 1.39 and 1.17, respectively. CH₄ flux significantly correlated with CH₄ concentration in porewater, and both were significantly related to air temperature. These findings indicate that CH₄ production and CH₄ flux at the middle growth stage (August) exhibited greater responses to changes in air temperature, which in turn induced the higher diel variation. The higher diel cycle for CH₄ flux in August than in September was likely due to the higher proportion of CH₄ oxidized during diffusion within the aerenchyma system. Our results suggest that the extent of diel variations in CH₄ flux may have depended on the gas transport capacity of plants, and the highest diel variation occurred at the middle growth stage.     

Microbial desulfurization of coal and oxidation of pure pyrite byThiobacillus ferrooxidans andAcidianus brierleyi

Summary Thiobacillus ferrooxidans andAcidianus brierleyi were capable of oxidizing pure pyrite as well as oxidizing sulfur in coal. First order reactions were assumed in the kinetic analysis performed. For oxidation of pure pyrite the rate constant was higher forA. brierleyi than forT. ferrooxidans. For sulfur removal from coal the values of the rate constants were comparable for the two microorganisms.     

Anaerobic ammonia oxidation by cell-free extracts of Nitrosomonas eutropha

Cell-free extracts of Nitrosomonas eutropha oxidized ammonia to nitrite with NO₂ (N₂O₄) as electron acceptor. The ammonia oxidation activity was shown to be sensitive against oxygen. In the absence of oxygen ammonia and NO₂ were consumed in a ratio of approximately 1:2 and hydroxylamine occurred as an intermediate. NO was released in amounts equimolar to the consumption of NO₂. After passing the cell suspension through a French pressure cell and fractionating it by density gradient centrifugation using a linear sucrose gradient, two soluble and two membrane fractions were detectable. Highest ammonia oxidation activity was measured in the membrane fractions and highest hydroxylamine oxidation activity in the soluble fractions. The K_S values of the ammonia oxidizing system in cell-free extracts was about 20 m NH₃ and remained unchanged between pH 7.25 to 8.25.     

Isolation of mutants of the obligate methanotroph Methylomonas albus defective in growth on methane

A strain of Methylomonas albus BG8WM, a type 1 obligate methanotroph, which had been maintained for 2 ycars by serial passage on solid medium containing methanol as a carbon source was found to mutate at a frequency of 10^-5-10^-6 to resistance to dichloromethane (DCM^R), the parental strain BG8 did not give rise to DCM^R colonies. DCM^R strains were no longer capable of growth on methane as a carbon cource and exhibited greatly reduced or undetectable methane mono-oxygenase activity. The mutants fell into three groups on the basis of SDS-PAGE analysis of the polypeptide profiles of the particulate fraction of cell extracts. One or more of four polypeptides of Mr 70,000, 50,000, 25,000 and 23,000 were implicated as being components of the methane mono-oxygenase. Spontaneous reversion to growth on methane and sensitivity to dichloromethane occurred at a frequency of about 10^-8. The loss of methane mono-oxygenase activity was not associated with loss of the resident 55 kb plasmid.Abbreviations DCM^R dichloromethane-resistant - SDS-PAGE sodium dodecyl sulphate polyacrylamide gel electrophoresis - NMS nitrate minimal salts medium     

Emission and oxidation of methane in Equisetum fluviatile stands growing on organic sediment and sand bottoms

Methane emission and rhizospheric CH₄ oxidation were studied in stands of Equisetum fluviatile, a common cryptogam in boreal lakes. The experiment was performed in mesocosms with organic sediment or sand bottoms under natural variation of temperature and light using the light-oxic – dark-anoxic chamber (LO/DA) technique. Net CH₄ emission from the organic sediment during the growing season varied between 3.4 and 19.0 mg m^–2 h^–1, but from sand the net CH₄ emission was only 3–10% of that measured from the organic sediment. In the organic sediment net CH₄ emission was very significantly correlated with sediment temperature (r² = 0.92). In the sand mesocosms the variation of net CH₄ emission was better correlated with the shoot biomass than with sediment temperature variation during the growing season, indicating that methanogens were severely limited by substrate availability and were probably dependent on substrates produced by E. fluviatile. The proportion of the methane oxidized of the potential CH₄ emission in summer did not differ significantly between the bottom types. The net CH₄ emission during the growing season as a proportion of the seasonal maximum of the shoot biomass was significantly higher in the organic sediment mesocosms (6.5%) than in sand (1.7%). The high CH₄ emissions observed from dense well-established E. fluviatile stands in the field appear to be more related to temperature-regulated turnover of detritus in the anaerobic sediment and less to CH₄ oxidation and seasonal variation in plant growth dynamics