Search Results (11)

While temperature, pH, DO, and ammonia nitrogen concentration are known to affect nitrous oxide (N₂O) emissions from ammonia-oxidizing bacteria (AOB), the specific responses of individual AOB species to these environmental variables have yet to be fully elucidated. The present study reports the isolation and pure culture of a new AOB strain, designated as N.eA1, from a stable CANON bioreactor. The strain’s denitrification and N₂O emission were systematically evaluated through a comprehensive analysis of growth kinetics, morphological characteristics, genetic composition, and nitrogen transformation under various environmental processes. Our results indicated that N.eA1 shares 95.33% sequence homology with Nitrosomonas europaea H1 AOB3, and exhibited higher nitrite (NO₂⁻-N) conversion efficiency. Morphological examination revealed white, semi-transparent spherical colonies. The bacterial growth kinetics included adaptation phase (0–12 h), exponential growth phase (12–36 h), stationary phase (36–72 h) and decline phase (after 72 h). Under optimal cultivation conditions (30 °C, DO concentration of 7.3 mg∙L⁻¹, pH 8.0, and NH₄⁺-N concentration of 260 mg∙L⁻¹), the culture achieved a maximum growth rate of 0.0723 h⁻¹, a maximum ammonia oxidation rate (AOR) of 10.74 mg∙(MLVSS∙h)⁻¹, and a minimum doubling time of 9.59 h. The peak time of nitrogen conversion was earlier than that of N₂O emission, with a maximum N₂O-N conversion from NH₄⁺-N of 1.039%. Full article

Using a nitrification inhibitor to decrease nitrification rates in soil represents a promising strategy to improve nitrogen fertilizer use efficiency. Nonetheless, rapid screening of nitrification inhibitors remains challenging. In this study, we propose a strategy to screen potential nitrification inhibitors through a structure–activity relationship (SAR) study based on a rapid determination of nitrification inhibition. To demonstrate this, the nitrification inhibition potentials of cinnamic acid derivatives against Nitrosomonas europaea growth were evaluated in a liquid culture. The SAR study showed that hydroxyl and fluoride groups were the favorable substituents on the benzene ring, and the ester group and double bond in the side chain were essential for maintaining high inhibition efficacy. Three compounds with notable inhibitory efficacy (EC₅₀ = 8–25 μM) were further assessed in agricultural soil, and they displayed a noteworthy reduction in nitrification rate and bacterial amoA gene numbers. Based on the results, we identified methyl cinnamate, methyl 4-hydroxycinnamate, and methyl 4-fluorocinnamate as promising candidates for nitrification inhibition. Full article

Bioregenerative life support systems (BLSS) are currently in development to tackle low recovery efficiencies, high energy demands, as well as food, water, and oxygen production challenges through the regeneration of nutrients from waste streams. The MELiSSA pilot plant has been developed as a testbed for regenerative life support system bioreactor operation and characterization. As nitrogen is a vital resource in such systems, we studied the functional composition of a new packed-bed nitrifying bioreactor inoculated with a co-culture of Nitrosomonas europaea (ATCC 25978) and Nitrobacter winogradskyi (ATCC 25391). After 840 days of autotrophic continuous cultivation, the packed-bed was sampled at five vertical positions, each with three horizontal positions, and the biomass at each position was characterized via qPCR, 16S amplicon sequencing, and liquid chromatography tandem mass spectrometry. The total number of cells within the different sections fluctuated around 8.95 ± 5.10 × 10⁷ cells/mL of beads. Based on 16S amplicons and protein content, N. europaea and N. winogradskyi constituted overall 44.07 ± 11.75% and 57.53 ± 12.04% of the nitrifying bioreactor, respectively, indicating the presence of a heterotrophic population that, even after such a long operation time, did not affect the nitrification function of the bioreactor. In addition, DNA-based abundance estimates showed that N. europaea was slightly more abundant than N. winogradskyi, whereas protein-based abundance estimates indicated a much higher abundance of N. europaea. This highlights that single-method approaches need to be carefully interpreted in terms of overall cell abundance and metabolic activity. Full article

We have recently shown that SmbP, the small metal-binding protein of Nitrosomonas europaea, can be employed as a fusion protein to express and purify recombinant proteins and peptides in Escherichia coli. SmbP increases solubility, allows simple, one-step purification through affinity chromatography, and provides superior final yields due to its low molecular weight. In this work, we report for the first time the use of SmbP to produce a recombinant peptide with anticancer activity: the antitumor-analgesic peptide (BmK-AGAP), a neurotoxin isolated from the venom of the Chinese scorpion Buthus martensii Karsch. This peptide was expressed in Escherichia coli SHuffle for correct, cytoplasmic, disulfide bond formation and tagged with SmbP at the N-terminus to improve its solubility and allow purification using immobilized metal affinity chromatography. SmbP_BmK-AGAP was found in the soluble fraction of the cell lysate. After purification and removal of SmbP by digestion with enterokinase, 1.8 mg of pure and highly active rBmK-AGAP was obtained per liter of cell culture. rBmK-AGAP exhibited antiproliferative activity on the MCF-7 cancer cell line, with a half-maximal inhibitory concentration value of 7.24 μM. Based on these results, we considered SmbP to be a suitable carrier protein for the production of recombinant, biologically active BmK-AGAP. We propose that SmbP should be an attractive fusion protein for the expression and purification of additional recombinant proteins or peptides that display anticancer activities. Full article

With the widespread use of chloramines disinfection, nitrification has become a problem that cannot be ignored. In order to control nitrification in the drinking water distribution system (DWDS), the inactivation effect of free chlorine, monochloramine and chlorine dioxide on ammonia-oxidizing bacterium (AOB) was studied under different temperature (8 °C, 26 °C and 35 °C) and pH (6.0, 7.0 and 8.7) conditions. The inactivation effect of Nitrosomonas europaea (a kind of AOB) by the three disinfectants increases with increasing temperature. As for the raised pH, the inactivation effect of free chlorine and monochloramine on AOB decreased, while that of chlorine dioxide increased. Last, but certainly not least, the experimental data of the disinfection were calculated to develop the N. europaea inactivation kinetic model, which was based on the first order Chick–Watson equation. The proposed model in this study took the two variables, pH and temperature, into consideration simultaneously, which were used to evaluate the average Ct value (multiplying the concentration of the residual disinfectant by the time of contact with N. europaea) required for different disinfectants when they produced the ideal inactivation effect on N. europaea. Full article

The stress response of ammonia-oxidizing bacteria (AOB) to oxygen deprivation limits AOB growth and leads to different nitrification pathways that cause the release of greenhouse gases. Measuring the stress response of AOB has proven to be a challenge due to the low growth rates of stressed AOB, making the sample volumes required to monitor the internal stress response of AOB prohibitive to repeated analysis. In a proof-of-concept study, confocal Raman microscopy with excitation resonant to the heme c moiety of cytochrome c was used to compare the cytochrome c content and activity of stressed and unstressed Nitrosomonas europaea (Nm 50), Nitrosomonas eutropha (Nm 57), Nitrosospira briensis (Nsp 10), and Nitrosospira sp. (Nsp 02) in vivo. Each analysis required no more than 1000 individual cells per sampling; thus, the monitoring of cultures with low cell concentrations was possible. The identified spectral marker delivered reproducible results within the signal-to-noise ratio of the underlying Raman spectra. Cytochrome c content was found to be elevated in oxygen-deprived and previously oxygen-deprived samples. In addition, cells with predominantly ferrous cytochrome c content were found in deprived Nitrosomonas eutropha and Nitrosospira samples, which may be indicative of ongoing electron storage at the time of measurement. Full article

Despite the adverse effects of emerging ZnO nanoparticles (nano-ZnO) on wastewater biological nitrogen removal (BNR) systems being widely documented, strategies for mitigating nanoparticle (NP) toxicity impacts on nitrogen removal have not been adequately addressed. Herein, N-acyl-homoserine lactone (AHL)-based quorum sensing (QS) was investigated for its effects against nano-ZnO toxicity to a model nitrifier, Nitrosomonas europaea. The results indicated that AHL-attenuated nano-ZnO toxicity, which was inversely correlated with the increasing dosage of AHL from 0.01 to 1 µM. At 0.01 µM, AHL notably enhanced the tolerance of N. europaea cells to nano-ZnO stress, and the inhibited cell proliferation, membrane integrity, ammonia oxidation rate, ammonia monooxygenase activity and amoA gene expression significantly increased by 18.2 ± 2.1, 2.4 ± 0.9, 58.7 ± 7.1, 32.3 ± 1.7, and 7.3 ± 5.9%, respectively, after 6 h of incubation. However, increasing the AHL dosage compromised the QS-mediated effects and even aggravated the NPs’ toxicity effects. Moreover, AHLs, at all tested concentrations, significantly increased superoxide dismutase activity, indicating the potential of QS regulations to enhance cellular anti-oxidative stress capacities when facing NP invasion. These results provide novel insights into the development of QS regulation strategies to reduce the impact of nanotoxicity on BNR systems. Full article

Hygienic fecal treatment in resource-oriented sanitation (ROS) systems is an important concern. Although the addition of nitrifying microorganisms is a sustainable fecal treatment method in ROS systems, it is essential to examine the cleanliness of this method. In this study, we investigated the fate of fecal indicators in source-separated fecal samples through tracking Escherichia coli and total coliforms. The effects of adding different amounts of Nitrosomonas europaea bio-seed, along with a constant amount of Nitrobacter winogradskyi bio-seed, were studied. In intact feces samples, the pathogen population underwent an initial increase, followed by a slight decrease, and eventually became constant. Although the addition of nitrifying microorganisms initially enhanced the pathogen growth rate, it caused the reduction process to become more efficient in the long-term. In addition to a constant concentration of 10,000 cells of N. winogradskyi per 1 g feces, a minimum amount of 3000 and 7000 cells of N. europaea per 1 g feces could completely remove E. coli and total coliforms, respectively, in less than 25 days. Increasing the amount of bio-seeds added can further reduce the time required for total pathogen removal. Full article

Nitrosomonas europaea carries numerous toxin-antitoxin systems. However, despite the abundant representation in its chromosome, studies have not surveyed the underlying molecular functions in detail, and their biological roles remain enigmatic. In the present study, we found that a chromosomally-encoded MazF family member, predicted at the locus NE1181, is a functional toxin endoribonuclease, and constitutes a toxin-antitoxin system, together with its cognate antitoxin, MazE. Massive parallel sequencing provided strong evidence that this toxin endoribonuclease exhibits RNA cleavage activity, primarily against the AAU triplet. This sequence-specificity was supported by the results of fluorometric assays. Our results indicate that N. europaea alters the translation profile and regulates its growth using the MazF family of endoribonuclease under certain stressful conditions. Full article

Moving bed biofilm reactors (MBBRs) are increasingly used for nitrogen removal with nitritation-anaerobic ammonium oxidation (anammox) processes in wastewater treatment. Carriers provide protected surfaces where ammonia oxidizing bacteria (AOB) and anammox bacteria form complex biofilms. However, the knowledge about the organization of microbial communities in MBBR biofilms is sparse. We used new cryosectioning and imaging methods for fluorescence in situ hybridization (FISH) to study the structure of biofilms retrieved from carriers in a nitritation-anammox MBBR. The dimensions of the carrier compartments and the biofilm cryosections after FISH showed good correlation, indicating little disturbance of biofilm samples by the treatment. FISH showed that Nitrosomonas europaea/eutropha-related cells dominated the AOB and Candidatus Brocadia fulgida-related cells dominated the anammox guild. New carriers were initially colonized by AOB, followed by anammox bacteria proliferating in the deeper biofilm layers, probably in anaerobic microhabitats created by AOB activity. Mature biofilms showed a pronounced three-dimensional stratification where AOB dominated closer to the biofilm-water interface, whereas anammox were dominant deeper into the carrier space and towards the walls. Our results suggest that current mathematical models may be oversimplifying these three-dimensional systems and unless the multidimensionality of these systems is considered, models may result in suboptimal design of MBBR carriers. Full article

A flow biosensor for the detection of toxicity in water using the ammonia-oxidizing bacterium (AOB) Nitrosomonas europaea as a bioreceptor and a polymeric membrane ammonium-selective electrode as a transducer is described. The system is based on the inhibition effects of toxicants on the activity of AOB, which can be evaluated by measuring the ammonium consumption rates with the ammonium-selective membrane electrode. The AOB cells are immobilized on polyethersulfone membranes packed in a holder, while the membrane electrode is placed downstream in the flow cell. Two specific inhibitors of the ammonia oxidation‒allylthiourea and thioacetamide‒have been tested. The IC₅₀ values defined as the concentration of an inhibitor causing a 50% reduction in the ammonia oxidation activity have been measured as 0.17 μM and 0.46 μM for allylthiourea and thioacetamide, respectively. The proposed sensor offers advantages of simplicity, speed and high sensitivity for measuring toxicity in water. Full article