Competition between Ammonia-Oxidizing Archaea and Bacteria from Freshwater Environments.

In the environment, nutrients are rarely available in a constant supply. Therefore, microorganisms require strategies to compete for limiting nutrients. In freshwater systems, ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) compete with heterotrophic bacteria, photosynthetic microorganisms, and each other for ammonium, which AOA and AOB utilize as their sole source of energy and nitrogen. We investigated the competition between highly enriched cultures of AOA (AOA-AC1) and AOB (AOB-G5-7) for ammonium. Based on the amoA gene, the newly enriched archaeal ammonia oxidizer in AOA-AC1 was closely related to Nitrosotenuis spp., and the bacterial ammonia oxidizer in AOB-G5-7, Nitrosomonas sp. strain Is79, belonged to the Nitrosomonas oligotropha group (Nitrosomonas cluster 6a). Growth experiments in batch cultures showed that AOB-G5-7 had higher growth rates than AOA-AC1 at higher ammonium concentrations. During chemostat competition experiments under ammonium-limiting conditions, AOA-AC1 dominated the cultures, while AOB-G5-7 decreased in abundance. In batch cultures, the outcome of the competition between AOA and AOB was determined by the initial ammonium concentrations. AOA-AC1 was the dominant ammonia oxidizer at an initial ammonium concentration of 50 µM, and AOB-G5-7 was dominant at 500 µM. These findings indicate that during direct competition, AOA-AC1 was able to use ammonium that was unavailable to AOB-G5-7, while AOB-G5-7 dominated at higher ammonium concentrations. The results are in strong accordance with environmental survey data suggesting that AOA are mainly responsible for ammonia oxidation under more oligotrophic conditions, whereas AOB dominate under eutrophic conditions. IMPORTANCE Nitrification is an important process in the global nitrogen cycle. The first step, ammonia oxidation to nitrite, can be carried out by ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB). In many natural environments, these ammonia oxidizers coexist. Therefore, it is important to understand the population dynamics in response to increasing ammonium concentrations. Here, we study the competition between AOA and AOB enriched from freshwater systems. The results demonstrate that AOA are more abundant in systems with low ammonium availabilities and that AOB are more abundant when the ammonium availability increases. These results will help to predict potential shifts in the community composition of ammonia oxidizers in the environment due to changes in ammonium availability.

Impact of Heavy Metal Pollution on Ammonia Oxidizers in Soils in the Vicinity of a Tailings Dam, Baotou, China.

Soil heavy metal pollution has received increasing attention due to their toxicity to soil microorganisms. We have analyzed the effects of heavy metal pollution on ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in soils in the vicinity of a tailings dam of Baotou region, China. Results showed that AOB were dominated with Nitrosomonas-like clusters, while AOA was dominated by group1.1b (Nitrososphaera cluster). Single Cd and Cr contents, as well as compound heavy metal pollution levels, had a significant negative impact on soil potential nitrification rate and both diversities of AOA and AOB. No clear relationship was found between any single heavy metal and abundance of AOA or AOB. But compound pollution could significantly decrease AOA abundance. The results indicated that heavy metal pollution had an obviously deleterious effect on the abundance, diversity, activity and composition of ammonia oxidizers in natural soils.

Start-up and bacterial community compositions of partial nitrification in moving bed biofilm reactor.

Partial nitrification (PN) has been considered as one of the promising processes for pretreatment of ammonium-rich wastewater. In this study, a kind of novel carriers with enhanced hydrophilicity and electrophilicity was implemented in a moving bed biofilm reactor (MBBR) to start up PN process. Results indicated that biofilm formation rate was higher on modified carriers. In comparison with the reactor filled with traditional carriers (start-up period of 21 days), it took only 14 days to start up PN successfully with ammonia removal efficiency and nitrite accumulation rate of 90 and 91%, respectively, in the reactor filled with modified carriers. Evident changes of spatial distributions and community structures had been detected during the start-up. Free-floating cells existed in planktonic sludge, while these microorganisms trended to form flocs in the biofilm. High-throughput pyrosequencing results indicated that Nitrosomonas was the predominant ammonia-oxidizing bacterium (AOB) in the PN system, while Comamonas might also play a vital role for nitrogen oxidation. Additionally, some other bacteria such as Ferruginibacter, Ottowia, Saprospiraceae, and Rhizobacter were selected to establish stable footholds. This study would be potentially significant for better understanding the microbial features and developing efficient strategies accordingly for MBBR-based PN operation.

Effects of Bacterial Community Members on the Proteome of the Ammonia-Oxidizing Bacterium Nitrosomonas sp. Strain Is79.

UNLABELLED: Microorganisms in the environment do not exist as the often-studied pure cultures but as members of complex microbial communities. Characterizing the interactions within microbial communities is essential to understand their function in both natural and engineered environments. In this study, we investigated how the presence of a nitrite-oxidizing bacterium (NOB) and heterotrophic bacteria affect the growth and proteome of the chemolithoautotrophic ammonia-oxidizing bacterium (AOB) Nitrosomonas sp. strain Is79. We investigated Nitrosomonas sp. Is79 in co-culture with Nitrobacter winogradskyi, in co-cultures with selected heterotrophic bacteria, and as a member of the nitrifying enrichment culture G5-7. In batch culture, N. winogradskyi and heterotrophic bacteria had positive effects on the growth of Nitrosomonas sp. Is79. An isobaric tag for relative and absolute quantification (iTRAQ) liquid chromatography-tandem mass spectrometry (LC-MS/MS) proteomics approach was used to investigate the effect of N. winogradskyi and the co-cultured heterotrophic bacteria from G5-7 on the proteome of Nitrosomonas sp. Is79. In co-culture with N. winogradskyi, several Nitrosomonas sp. Is79 oxidative stress response proteins changed in abundance, with periplasmic proteins increasing and cytoplasmic proteins decreasing in abundance. In the presence of heterotrophic bacteria, the abundance of proteins directly related to the ammonia oxidation pathway increased, while the abundance of proteins related to amino acid synthesis and metabolism decreased. In summary, the proteome of Nitrosomonas sp. Is79 was differentially influenced by the presence of either N. winogradskyi or heterotrophic bacteria. Together, N. winogradskyi and heterotrophic bacteria reduced the oxidative stress for Nitrosomonas sp. Is79, which resulted in more efficient metabolism. IMPORTANCE: Aerobic ammonia-oxidizing microorganisms play an important role in the global nitrogen cycle, converting ammonia to nitrite. In their natural environment, they coexist and interact with nitrite oxidizers, which convert nitrite to nitrate, and with heterotrophic microorganisms. The presence of nitrite oxidizers and heterotrophic bacteria has a positive influence on the growth of the ammonia oxidizers. Here, we present a study investigating the effect of nitrite oxidizers and heterotrophic bacteria on the proteome of a selected ammonia oxidizer in a defined culture to elucidate how these two groups improve the performance of the ammonia oxidizer. The results show that the presence of a nitrite oxidizer and heterotrophic bacteria reduced the stress for the ammonia oxidizer and resulted in more efficient energy generation. This study contributes to our understanding of microbe-microbe interactions, in particular between ammonia oxidizers and their neighboring microbial community.

[Seasonal changes of ammonia-oxidizing bacterial communities during tropical forest restoration]. / 热带森林恢复过程中氨氧化细菌群落的季节变化.

In this study, we used a high-throughput sequencing technology to survey the dry-wet seasonal change characteristics of soil ammonia-oxidizing bacteria (AOB) communities in the three restoration stages [i.e., Mallotus paniculatus community (early stage), Millettia leptobotrya community (middle stage), and Syzygium oblatum community (later stage)] of Xishuangbanna tropical forest ecosystems. We analyzed the effects of soil physicochemical characteristics on AOB community composition and diversity during tropical forest restoration. The results showed that tropical forest restoration significantly affected the relative abundance of dominant AOB phyla and their dry-wet seasonal variation. The maximum relative abundance of Proteobacteria (71.3%) was found in the early recovery stage, while that of Actinobacteria was found in the late recovery stage (1.0%). The abundances of Proteobacteria and Actinobacteria had the maximum ranges of dry-wet seasonal variation in the early and late stages, respectively. The abundance of dominant AOB genera and its dry-wet seasonal variation varied across tropical forest restoration stages. The maximum average relative abundance of Nitrosospira and Nitrosomonas in the late recovery stage was 66.2% and 1.5%, respectively. In contrast, the abundance of Nitrosovibrio reached its maximum (25.6%) in the early recovery stage. The maximum dry-wet seasonal variation in relative abundance of Nitrosospira and Nitrosomonas occurred in the early recovery stage, while that of Nitrosovibrio occurred in the middle recovery stage. The Chao1, Shannon, and Simpson diversity indices of AOB communities increased along the restoration stages, which were significantly higher in the wet season than in the dry season. The results of canonical correspondence analysis showed that soil easily oxidized carbon was the main factor controlling AOB community diversity and Actinobacteria abundance. Soil bulk density and temperature were the main factors affecting Proteobacteria abundance. Soil pH, microbial biomass carbon, water content, ammonium nitrogen, bulk density, and temperature were the main factors controlling the abundances of Nitrosospira, Nitrosomonas, and Nitrosovibrio. Therefore, tropical forest restoration can regulate the change of relative abundance of dominant AOB taxa via mediating the changes of soil temperature, bulk density, and readily oxidized carbon, leading to an increase in soil AOB community diversity.

Dynamics of specific ammonia-oxidizing bacterial populations and nitrification in response to controlled shifts of ammonium concentrations in wastewater.

Ammonia-oxidizing bacteria (AOB) are essential for the nitrification process in wastewater treatment. To retain these slow-growing bacteria in wastewater treatment plants (WWTPs), they are often grown as biofilms, e.g., on nitrifying trickling filters (NTFs) or on carriers in moving bed biofilm reactors (MBBRs). On NTFs, a decreasing ammonium gradient is formed because of the AOB activity, resulting in low ammonium concentrations at the bottom and reduced biomass with depth. To optimize the NTF process, different ammonium feed strategies may be designed. This, however, requires knowledge about AOB population dynamics. Using fluorescence in situ hybridization (FISH) and confocal laser scanning microscopy, we followed biomass changes during 6 months, of three AOB populations on biofilm carriers. These were immersed in aerated MBBR tanks in a pilot plant receiving full-scale wastewater. Tanks were arranged in series, forming a wastewater ammonium gradient mimicking an NTF ammonium gradient. The biomass of one of the dominating Nitrosomonas oligotropha-like populations increased after an ammonium upshift, reaching levels comparable to the high ammonium control in 28 days, whereas a Nitrosomonas europaea-like population increased relatively slowly. The MBBR results, together with competition studies in NTF systems fed with wastewater under controlled ammonium regimes, suggest a differentiation between the two N. oligotropha populations, which may be important for WWTP nitrification.

Analysis of microbial population dynamics in a partial nitrifying SBR at ambient temperature.

In this study, a lab-scale partial nitrifying sequencing batch reactor (SBR) was developed to investigate partial nitrification at ambient temperature (16-22 °C). Techniques of denaturing gradient gel electrophoresis (DGGE), cloning, and fluorescence in situ hybridization (FISH) were utilized simultaneously to study microbial population dynamics. Partial nitrification was effectively achieved in response to shifts of influent ammonium concentrations. DGGE results showed that higher ammonia concentration referred to lower ammonia-oxidizing bacteria (AOB) diversity in the SBR. Phylogenetic analysis revealed that all the predominant AOB was affiliated with Nitrosomonas genus. FISH analysis illustrated AOB was the predominant nitrifying bacteria of microbial compositions when SBR achieved partial nitrification (PN) at ambient temperature.

Use of real-time QPCR in biokinetics and modeling of two different ammonia-oxidizing bacteria growing simultaneously.

A real-time quantitative polymerase chain reaction (QPCR) was used to evaluate biokinetic coefficients of Nitrosomonas nitrosa and N. cryotolerans clusters growing simultaneously in a batch mode of ammonia oxidation. The mathematical models based on Monod equation were employed to describe the competitive relationship between these clusters and were fitted to experimental data to obtain biokinetic values. The maximum growth rates (µ(m)), half-saturation coefficients (K(S)), microbial yields (Y) and decay coefficients (k(d)) of N. nitrosa and N. cryotolerans were 1.77 and 1.21 day(-1), 23.25 and 23.06 mg N·L(-1), 16 × 10(8) and 1 × 10(8) copies·mg N(-1), 0.26 and 0.20 day(-1), respectively. The estimated coefficients were applied for modeling continuous operations at various hydraulic retention times (HRTs) with an influent ammonia concentration of 300 mg N·L(-1). Modeling results revealed that ammonia oxidation efficiencies were achieved 55-98 % at 0.8-10 days HRTs and that the system was predicted to be washed out at HRT of 0.7 days. Overall, use of QPCR for estimating biokinetic coefficients of the two AOB cluster growing simultaneously by use of ammonia were successful. This idea may open a new direction towards biokinetics of ammonia oxidation in which respirometry tests are usually employed.

Effects of nitrogen dioxide and anoxia on global gene and protein expression in long-term continuous cultures of Nitrosomonas eutropha C91.

Nitrosomonas eutropha is an ammonia-oxidizing betaproteobacterium found in environments with high ammonium levels, such as wastewater treatment plants. The effects of NO(2) on gene and protein expression under oxic and anoxic conditions were determined by maintaining N. eutropha strain C91 in a chemostat fed with ammonium under oxic, oxic-plus-NO(2), and anoxic-plus-NO(2) culture conditions. Cells remained viable but ceased growing under anoxia; hence, the chemostat was switched from continuous to batch cultivation to retain biomass. After several weeks under each condition, biomass was harvested for total mRNA and protein isolation. Exposure of N. eutropha C91 to NO(2) under either oxic or anoxic conditions led to a decrease in proteins involved in N and C assimilation and storage and an increase in proteins involved in energy conservation, including ammonia monooxygenase (AmoCAB). Exposure to anoxia plus NO(2) resulted in increased representation of proteins and transcripts reflective of an energy-deprived state. Several proteins implicated in N-oxide metabolism were expressed and remained unchanged throughout the experiment, except for NorCB nitric oxide reductase, which was not detected in the proteome. Rather, NorY nitric oxide reductase was expressed under oxic-plus-NO(2) and anoxic-plus-NO(2) conditions. The results indicate that exposure to NO(2) results in an energy-deprived state of N. eutropha C91 and that anaerobic growth could not be supported with NO(2) as an oxidant.

Growth of ammonia-oxidizing archaea and bacteria in cattle manure compost under various temperatures and ammonia concentrations.

A recent study showed that ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) coexist in the process of cattle manure composting. To investigate their physiological characteristics, liquid cultures seeded with fermenting cattle manure compost were incubated at various temperatures (37°C, 46°C, or 60°C) and ammonium concentrations (0.5, 1, 4, or 10 mM NH (4) (+) -N). The growth rates of the AOB and AOA were monitored using real-time polymerase chain reaction analysis targeting the bacterial and archaeal ammonia monooxygenase subunit A genes. AOB grew at 37°C and 4 or 10 mM NH (4) (+) -N, whereas AOA grew at 46°C and 10 mM NH (4) (+) -N. Incubation with allylthiourea indicated that the AOB and AOA grew by oxidizing ammonia. Denaturing gradient gel electrophoresis and subsequent sequencing analyses revealed that a bacterium related to Nitrosomonas halophila and an archaeon related to Candidatus Nitrososphaera gargensis were the predominant AOB and AOA, respectively, in the seed compost and in cultures after incubation. This is the first report to demonstrate that the predominant AOA in cattle manure compost can grow and can probably oxidize ammonia under moderately thermophilic conditions.

Influence of growth manner on nitrifying bacterial communities and nitrification kinetics in three lab-scale bioreactors.

The effects of growth type, including attached growth, suspended growth, and combined growth, on the characteristics of communities of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) were studied in three lab-scale Anaerobic/Anoxic(m)-Oxic(n) (AmOn) systems. These systems amplified activated sludge, biofilms, and a mixture of activated sludge and biofilm (AS-BF). Identical inocula were adopted to analyze the selective effects of mixed growth patterns on nitrifying bacteria. Fluctuations in the concentration of nitrifying bacteria over the 120 days of system operation were analyzed, as was the composition of nitrifying bacterial community in the stabilized stage. Analysis was conducted using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and real-time PCR. According to the DGGE patterns, the primary AOB lineages were Nitrosomonas europaea (six sequences), Nitrosomonas oligotropha (two sequences), and Nitrosospira (one sequence). The primary subclass of NOB community was Nitrospira, in which all identified sequences belonged to Nitrospira moscoviensis (14 sequences). Nitrobacter consisted of two lineages, namely Nitrobacter vulgaris (three sequences) and Nitrobacter alkalicus (two sequences). Under identical operating conditions, the composition of nitrifying bacterial communities in the AS-BF system demonstrated significant differences from those in the activated sludge system and those in the biofilm system. Major varieties included several new, dominant bacterial sequences in the AS-BF system, such as N. europaea and Nitrosospira and a higher concentration of AOB relative to the activated sludge system. However, no similar differences were discovered for the concentration of the NOB population. A kinetic study of nitrification demonstrated a higher maximum specific growth rate of mixed sludge and a lower half-saturation constant of mixed biofilm, indicating that the AS-BF system maintained relatively good nitrifying ability.

Inoculum effects on community composition and nitritation performance of autotrophic nitrifying biofilm reactors with counter-diffusion geometry.

The link between nitritation success in a membrane-aerated biofilm reactor (MABR) and the composition of the initial ammonia- and nitrite-oxidizing bacterial (AOB and NOB) population was investigated. Four identically operated flat-sheet type MABRs were initiated with two different inocula: from an autotrophic nitrifying bioreactor (Inoculum A) or from a municipal wastewater treatment plant (Inoculum B). Higher nitritation efficiencies (NO(2)(-)-N/NH(4)(+)-N) were obtained in the Inoculum B- (55.2-56.4%) versus the Inoculum A- (20.2-22.1%) initiated reactors. The biofilms had similar oxygen penetration depths (100-150 µm), but the AOB profiles [based on 16S rRNA gene targeted real-time quantitative PCR (qPCR)] revealed different peak densities at or distant from the membrane surface in the Inoculum B- versus A-initiated reactors, respectively. Quantitative fluorescence in situ hybridization (FISH) revealed that the predominant AOB in the Inoculum A- and B-initiated reactors were Nitrosospira spp. (48.9-61.2%) versus halophilic and halotolerant Nitrosomonas spp. (54.8-63.7%), respectively. The latter biofilm displayed a higher specific AOB activity than the former biofilm (1.65 fmol cell(-1) h(-1) versus 0.79 fmol cell(-1) h(-1) ). These observations suggest that the AOB and NOB population compositions of the inoculum may determine dominant AOB in the MABR biofilm, which in turn affects the degree of attainable nitritation in an MABR.

Nitrification in fixed-bed reactors treating saline wastewater.

Halophilic nitrifiers belonging to the genus Nitrosomonas and Nitrospira were enriched from seawater and marine sediment samples of the North Sea. The maximal ammonia oxidation rate (AOR) in batch enrichments with seawater was 15.1 mg N L(-1) day(-1). An intermediate nitrite accumulation was observed. Two fixed-bed reactors for continuous nitrification with either polyethylene/claysinter lamellas (FBR A) or porous ceramic rings (FBR B)were run at two different ammonia concentrations, three different ammonia loading rates (ALRs), + or - pH adjustment,and at an increased upflow velocity. A better overall nitrification without nitrite accumulation was observed in FBR B. However, FBR A revealed a higher AOR and nitrite oxidation rate of 6 and 7 mg N L(-1) h(-1), compared to FBR B with 5 and 5.9 mg N L(-1) h(-1), respectively. AORs in the FBRs were at least ten times higher than in suspended enrichment cultures. Whereas a shift within the ammoniaoxidizing population in the genus Nitrosomonas at the subspecies level occurred in FBR B with synthetic seawater at an increasing ALR and a decreasing pH, the nitrite oxidizing Nitrospira population apparently did not change.

Transcriptome Analysis of the Ammonia-Oxidizing Bacterium Nitrosomonas mobilis Ms1 Reveals Division of Labor between Aggregates and Free-living Cells.

Bacteria change their metabolic states to increase survival by forming aggregates. Ammonia-oxidizing bacteria also form aggregates in response to environmental stresses. Nitrosomonas mobilis, an ammonia-oxidizing bacterium with high stress tolerance, often forms aggregates mainly in wastewater treatment systems. Despite the high frequency of aggregate formation by N. mobilis, its relationship with survival currently remains unclear. In the present study, aggregates were formed in the late stage of culture with the accumulation of nitrite as a growth inhibitor. To clarify the significance of aggregate formation in N. mobilis Ms1, a transcriptome analysis was performed. Comparisons of the early and late stages of culture revealed that the expression of stress response genes (chaperones and proteases) increased in the early stage. Aggregate formation may lead to stress avoidance because stress response genes were not up-regulated in the late stage of culture during which aggregates formed. Furthermore, comparisons of free-living cells with aggregates in the early stage of culture showed differences in gene expression related to biosynthesis (ATP synthase and ribosomal proteins) and motility and adhesion (flagella, pilus, and chemotaxis). Biosynthesis genes for growth were up-regulated in free-living cells, while motility and adhesion genes for adaptation were up-regulated in aggregates. These results indicate that N. mobilis Ms1 cells adapt to an unfavorable environment and grow through the division of labor between aggregates and free-living cells.

Chemoorganoheterotrophic growth of Nitrosomonas europaea and Nitrosomonas eutropha.

The ammonia oxidizers Nitrosomonas europaea and Nitrosomonas eutropha are able to grow chemoorganotrophically under anoxic conditions with pyruvate, lactate, acetate, serine, succinate, alpha-ketoglutarate, or fructose as substrate and nitrite as terminal electron acceptor. The growth yield of both bacteria is about 3.5 mg protein (mmol pyruvate)(-1) and the maximum growth rates of N. europaea and N. eutropha are 0.094 d(-1) and 0.175 d(-1), respectively. In the presence of pyruvate and CO2 about 80% of the incorporated carbon derives from pyruvate and about 20% from CO2. Pyruvate is used as energy and only carbon source in the absence of CO2 (chemoorganoheterotrophic growth). CO2 stimulates the chemoorganotrophic growth of both ammonia oxidizers and the expression of ribulose bisphosphate carboxylase/oxygenase is down-regulated at increasing CO2 concentration. Ammonium, although required as nitrogen source, is inhibitory for the chemoorganotrophic metabolism of N. europaea and N. eutropha. In the presence of ammonium pyruvate consumption and the expression of the genes aceE, ppc, gltA, odhA, and ppsA (energy conservation) as well as nirK, norB, and nsc (denitrification) are reduced.

Stability of partial nitrification and microbial population dynamics in a bioaugmented membrane bioreactor.

Bioaugmentation of bioreactors focuses on the removal of numerous organics, with little attention typically paid to the maintenance of high and stable nitrite accumulation in partial nitrification. In this study, a bioaugmented membrane bioreactor (MBR) inoculated with enriched ammonia-oxidizing bacteria (AOB) was developed, and the effects of dissolved oxygen (DO) and temperature on stability of partial nitrification and microbial community structure, in particular on nitrifying community were evaluated. The results showed that DO and temperature played the most important roles in the stability of partial nitrification in the bioaugmented MBR. The optimal operation conditions were found at 2-3 mgDO/L and 30 degrees C, achieving 95% ammonia oxidization efficiency and 0.95 of nitrite ratio (NO2-/NOx-). High-DO (5-6 mg/L) and low-temperature (20 degrees C) had negative impacts on nitrite accumulation, leading to its drop to 0.6. However, the nitrite ratio achieved in the bioaugmented MBR was higher than that in most previous literatures. Denaturing gradient gel electrophoresis (DGGE) and fluorescence in situ hybridization (FISH) were used to provide an insight into the microbial community. It showed that Nitrosomonas-like species as the only detected AOB remained predominant in the bioaugmented MBR all the time, which coexisted with numerous heterotrophic bacteria. The heterotrophic bacteria responsible for mineralizing soluble microbial products (SMP) produced by nitrifiers belonged to Cytophaga-Flavobacterium-Bacteroides (CFB) group, alpha-, beta-, and gamma- Proteobacteria. The fraction of AOB ranging from 77% to 54% was much higher than that of NOB (0.4-0.9%), which might be the primary cause for the high and stable nitrite accumulation in the bioaugmented MBR.

The drought and high wet soil condition impact on PAH (phenanthrene) toxicity towards nitrifying bacteria.

A few previous studies showed that the low soil moisture could interact with the toxic effect of the polycyclic aromatic hydrocarbons (PAHs) towards animals (mostly invertebrates). In the present research the impact of the soil moisture in the wide range (from the drought to high moisture conditions) in three different soil materials on toxic effect of the PAH (phenanthrene) towards soil microorganisms (nitrifying bacteria activity) was evaluated. The three dry soil materials were artificially contaminated with phenanthrene (0, 1, 10, 100 and 1000 mg kg-1 dry mass of soil) and moistened to the varied levels of the soil moisture (30% WHC (dry), 55% WHC (optimal) and 80% WHC (highly wet conditions)). After 7 days incubation, the nitrification potential was measured. The results of the proposed ANCOVA multiple regression model (adjusted R2 = 0.91), showed that the increase of soil moisture enhanced the toxicity of the phenanthrene towards nitrification potential and this combined moisture-phenanthrene effect was soil dependent. Therefore, the effect of the soil moisture in combination with the soil diversity should not be missed in the ecotoxicological risk assessment of the PAHs.

Mixed nitrifying bacteria culture under different temperature dropping strategies: Nitrification performance, activity, and community.

In this study, the nitrification performance, metabolic activity, antioxidant enzyme activity as well as bacterial community of mixed nitrifying bacteria culture under different temperature dropping strategies [(#1) growth temperature kept at 20 °C; (#2) sharp1 decreased from 20 °C to 10 °C; (#3) growth at 20 °C for 6 days followed by sharp decrease to 10 °C; and (#4) gradual decreased from 20 °C to 10 °C] were evaluated. It was shown that acclimation at 20 °C for 6 days allowed to maintain better nitrification activity at 10 °C. The nitrite oxidation capacity of nitrifiers was significantly correlated with the relative light unit (RLU) (p < .05) and the fluctuation of superoxide dismutase (SOD) enzyme activity (p < .01). With serial #3 showed the highest RLU levels and the least SOD enzyme fluctuation as compared to serials #2 and #4. Throughout the experimental period, Nitrosospira and Nitrosomonas as well as Nitrospira were identified as the predominant ammonia-oxidizing bacteria (AOB) and nitrate-oxidizing bacteria (NOB). The dynamic change of AOB/NOB ratios and nitrification activity in serials #2-#4 demonstrated that AOB recovered better than NOB with long-term 10 °C exposure, and the nitrification performance was mainly limited by the nitrite oxidation capacity of NOB. Applying 6 days acclimation at 20 °C was beneficial for the mixed nitrifying bacteria culture to cope with low temperature (10 °C) stress, possibly due to the maintenance of metabolic activity, antioxidant enzyme activity stability as well as appropriate AOB/NOB ratio.

Effects of ammonium and nitrite on communities and populations of ammonia-oxidizing bacteria in laboratory-scale continuous-flow reactors.

This study investigated the effects of ammonium and nitrite on ammonia-oxidizing bacteria (AOB) from an activated sludge process in laboratory-scale continuous-flow reactors. AOB communities were analyzed using specific PCR followed by denaturing gel gradient electrophoresis, cloning and sequencing of the 16S rRNA gene, and AOB populations were quantified using real-time PCR. To study the effect of ammonium, activated sludge from a sewage treatment system was enriched in four reactors receiving inorganic medium containing four different ammonium concentrations (2, 5, 10 and 30 mM NH(4) (+)-N). One of several sequence types of the Nitrosomonas oligotropha cluster predominated in the reactors with lower ammonium loads (2, 5 and 10 mM NH(4) (+)-N), whereas Nitrosomonas europaea was the dominant AOB in the reactor with the highest ammonium load (30 mM NH(4) (+)-N). The effect of nitrite was studied by enriching the enriched culture possessing both N. oligotropha and N. europaea in four reactors receiving 10-mM-ammonium inorganic medium containing four different nitrite concentrations (0, 2, 12 and 22 mM NO(2) (-)-N). Nitrosomonas oligotropha comprised the majority of AOB populations in the reactors without nitrite accumulation (0 and 2 mM NO(2) (-)-N), whereas N. europaea was in the majority in the 12- and 22-mM NO(2) (-)-N reactors, in which nitrite concentrations were 2.1-5.7 mM (30-80 mg N L(-1)).

Determination of growth rate and yield of nitrifying bacteria by measuring carbon dioxide uptake rate.

Nitrifier growth parameters--the maximum growth rate (microAmax) and yield (YA)--were estimated by measuring the rate of carbon dioxide uptake and additional rates of oxygen uptake and ammonia (or nitrite) use. Batch tests in a combined titrimetric and offgas analyzer with enriched Nitrobacter and Nitrosomonas cultures and an activated sludge sample were performed. The measured microAmax values for the Nitrobacter and Nitrosomonas cultures were 0.67 +/- 0.03 day(-1) and 0.54 +/- 0.09 day(-1), while the YA values were 0.072 +/- 0.01 g volatile suspended solids (VSS) x g nitrogen (N)(-1) and 0.14 +/- 0.02 gVSS x gN(-1), respectively. For the activated sludge sample, microAmax was observed to increase with pH (microAmax = 0.72 x 3.3(pH-7.1)) over the range 6.8 to 7.1. All microAmax and YA values determined by this method were similar to those previously reported. Compared with other microAmax and YA estimation methods, this method allows for unique microAmax and YA estimations for given conditions from a single experiment.