Effect of nitrite concentration and pH on most probable number enumerations of non-growing Nitrobacter spp.

Abstract Enumerations of nitrite-oxidizing bacteria in soil samples by a Most Probable Number technique, often showed relatively high cell numbers at a low nitrite concentration compared with the numbers of ammonium-oxidizing bacteria. It was hypothesized that the high numbers enumerated at low nitrite concentration would represent non-growing or organotrophically growing cells of nitrite-oxidizing species. In this paper, the sensitivity of non-growing Nitrobacter species to high nitrite concentrations as well as to low pH was examined. Different Nitrobacter species were pre-cultured at 0.5 mM nitrite. Non-growing cells differing in age were enumerated at different nitrite concentrations and pH values. The incubation period lasted for 5 months at 20°C. However, during the incubation periods of the older non-growing cells, it appeared that a period of 5 months might have been too short for reaching constant numbers. Early stationary cells of all species that were studied appeared not to be affected by high nitrite concentrations or low pH. Eight- and 18-month-old non-growing cells of Nitrobacter hamburgensis were also insensitive to 5 mM nitrite. The numbers of 8- and 18-month-old resting cells of N. vulgaris were only repressed by a combination of 5 mM nitrite and a low pH. Eight-month-old non-growing cells of N. winogradskyi were sensitive to 5 mM irrespective of pH, but 18-month-old cells only to 5 mM nitrate at low pH. The numbers of 8- and 18-month-old resting cells of N. winogradskyi serotype agilis were repressed by low pH rather than high nitrite concentration. Hence, it was concluded that the large differences in numbers of nitrite-oxidizing bacteria obtained with low and high nitrite concentrations in the incubation medium, was not likely to be due to the presence of non-growing Nitrobacter species in soil samples, but rather to the existence of organotrophically growing Nitrobacter cells.     

Effective and robust partial nitrification to nitrite by real-time aeration duration control in an SBR treating domestic wastewater

Achieving sustainable partial nitrification to nitrite has been proven difficult in treating low strength nitrogenous wastewater. Real-time aeration duration control was used to achieve efficient partial nitrification to nitrite in a sequencing batch reactor (SBR) to treat low strength domestic wastewater. Above 90% nitrite accumulation ratio was maintained for long-term operation at normal condition, or even lower water temperature in winter. Partial nitrification established by controlling aeration duration showed good performance and robustness even though encountering long-term extended aeration and starvation period. Process control enhanced the successful accumulation of ammonia oxidizing bacteria (AOB) and washout of nitrite oxidizing bacteria (NOB). Scanning electron microscope observations indicated that the microbial morphology showed a shift towards small rod-shaped clusters. Fluorescence in situ hybridization (FISH) results demonstrated AOB were the dominant nitrifying bacteria, up to 8.3 ± 1.1% of the total bacteria; on the contrary, the density of NOB decreased to be negligible after 135 days operation since adopting process control.     

好氧和缺氧条件下游离亚硝酸对氨氧化菌和亚硝酸盐氧化菌的选择性抑制

【背景】稳定短程硝化是实现城市污水厌氧氨氧化技术的瓶颈,目前国内外关于游离亚硝酸(Free nitrous acid,FNA)对硝化菌活性的影响大多是在曝气条件下进行研究,鲜有关于缺氧条件下FNA对硝化菌活性影响的报道。【目的】探究好氧和缺氧下FNA对氨氧化菌(Ammonia oxidizing bacteria,AOB)和亚硝酸盐氧化菌(Nitrite oxidizing bacteria,NOB:Nitrospira和Nitrobacter)活性的抑制影响。【方法】采用序批式反应器(Sequencing batch reactor,SBR),基于混合液悬浮固体浓度(Mixed liquid suspended solids,MLSS)为8 300 mg/L的全程硝化污泥条件,通过批次试验分别考察好氧和缺氧下FNA(初始浓度为1.16 mg/L)处理48 h后,AOB和NOB活性的变化情况。【结果】好氧FNA处理活性污泥48 h后,FNA浓度维持在1.16-1.17 mg/L,游离氨(Free ammonia,FA)浓度小于0.017 mg/L,AOB、Nitrospira、Nitrobacter丰度均未发生明显变化;过曝气至99 h时,与空白组相比,比氨氮氧化速率(r~+_(NH4-N))、比亚硝酸盐氮氧化速率(r_(NO2-N))均出现小幅下降,分别由3.5、4.828 mg N/(g VSS·h)降至3.3、4.668 mg N/(g VSS·h),且亚硝酸盐氮累积率(Nitrite accumulation rate,NAR)始终低于33.2%。缺氧FNA处理活性污泥48 h后,FNA浓度维持在0.64-1.16 mg/L,FA浓度低于0.039 mg/L,AOB丰度变化较小,而Nitrospira、Nitrobacter丰度均明显下降,分别由3.002 9×10~9、4.245×10~8 copies/g VSS降至1.666 5×10~8、5.163 8×10~7 copies/g VSS;过曝气至99 h时,与空白组相比,r~+_(NH4-N)值下降幅度较小,而r_(NO2-N)值明显降低,由4.828 mg N/(g VSS·h)降至0.007 mg N/(g VSS·h),且在过曝气0-292 h内,NAR均大于94%。【结论】好氧FNA处理活性污泥48 h后对AOB和NOB无明显抑制作用,但缺氧FNA处理活性污泥48 h后对AOB具有轻微抑制作用,而对NOB具有强烈的抑制作用,可以实现稳定的短程硝化。     

Optimization of free ammonia concentration for nitrite accumulation in shortcut biological nitrogen removal process

A shortcut biological nitrogen removal (SBNR) utilizes the concept of a direct conversion of ammonium to nitrite and then to nitrogen gas. A successful SBNR requires accumulation of nitrite in the system and inhibition of the activity of nitrite oxidizers. A high concentration of free ammonia (FA) inhibits nitrite oxidizers, but unfortunately decreases the ammonium removal rate as well. Therefore, the optimal range of FA concentration is necessary not only to stabilize nitrite accumulation but also to achieve maximum ammonium removal. In order to derive such optimal FA concentrations, the specific substrate utilization rates of ammonium and nitrite oxidizers were measured. The optimal FA concentration range appeared to be 5–10 mg/L for the adapted sludge. The simulated results from the modified inhibition model expressed by FA and ammonium/nitrite concentrations were shown very similar to the experimental results.     

Effect of salicylic acid on nitrite reductase and glutamate dehydrogenase activities in maize roots

The effects of 0.01 to 5 m M salicyclic acid on the increase in nitrite reductase or glutamate dehydrogenase activities in maize roots by nitrate or ammonium respectively, were examined. Nitrite reductase activity was inhibited by the highest concentration of the acid. The activity of NADH-glutamate dehydrogenase was stimulated slightly (but consistently) by the lowest concentration and was inhibited by higher concentrations. Total protein content was also inhibited at high concentrations. When the crude enzyme extract was stored at 25°C in light, the glutamate dehydrogenase activity in the control decreased after 4 h of incubation. Low concentrations of the acid had no effect on this decrease but higher concentration accelerated the process. The divalent cations Caz²⁺, Mn²⁺, Mg²⁺ and Zn²⁺ protected against loss of enzyme activity during storage, both in the absence and presence of the acid. The inhibitory effect of 5 m M salicylic acid on glutamate dehydrogenase activity is apparent due to interference with the activity of the enzyme rather than with its synthesis.     

Effect of nitrate and nitrite on sulfide production by two thermophilic,sulfate-reducing enrichments from an oil field in the North Sea

Thermophilic sulfate-reducing bacteria (tSRB) can be major contributors to the production of H₂S (souring) in oil reservoirs. Two tSRB enrichments from a North Sea oil field, NS-tSRB1 and NS-tSRB2, were obtained at 58°C with acetate-propionate-butyrate and with lactate as the electron donor, respectively. Analysis by rDNA sequencing indicated the presence of Thermodesulforhabdus norvegicus in NS-tSRB1 and of Archaeoglobus fulgidus in NS-tSRB2. Nitrate (10 mM) had no effect on H₂S production by mid-log phase cultures of NS-tSRB1 and NS-tSRB2, whereas nitrite (0.25 mM or higher) inhibited sulfate reduction. NS-tSRB1 did not recover from inhibition, whereas sulfate reduction activity of NS-tSRB2 recovered after 500 h. Nitrite was also effective in souring inhibition and H₂S removal in upflow bioreactors, whereas nitrate was similarly ineffective. Hence, nitrite may be preferable for souring prevention in some high-temperature oil fields because it reacts directly with sulfide and provides long-lasting inhibition of sulfate reduction.     

Effect of varying salinity, temperature, ammonia and nitrous acid concentrations on nitrification of saline wastewater in fixed-bed reactors

Nitrification under changing salinities (0-9%), temperatures (6-50 °C), ammonia (0-5 g N L⁻¹) and nitrite concentrations (0-0.4 g N L⁻¹) was investigated in fixed-bed reactors. For all conditions ammonia oxidation rates (AOR) were lower than nitrite oxidation rates (NOR). AORs and NORs increased from 12.5 to 40 °C and were very low at 6 °C and almost zero at 50 °C. No recovery of nitrification was obtained after incubation at 50 °C, whereas nitrification was restorable after incubation at 6 °C. Ammonia concentrations of 5 g N L⁻¹ or nitrite concentrations up to 0.125 g N L⁻¹ decreased AOR to almost zero. AORs and NORs recovered if ammonia or nitrite was removed. At concentrations of 1 and 5 g N L⁻¹ ammonia AOR and NOR were inhibited by 50%, whereas 27 mg N/L nitrite inhibited AOR by 50%.     

Importance of the operating pH in maintaining the stability of anoxic ammonium oxidation (anammox) activity in moving bed biofilm reactors

Two bench-scale parallel moving bed biofilm reactors (MBBR) were operated to assess pH-associated anammox activity changes during long term treatment of anaerobically digested sludge centrate pre-treated in a suspended growth partial nitrification reactor. The pH was maintained at 6.5 in reactor R1, while it was allowed to vary naturally between 7.5 and 8.1 in reactor R2. At high nitrogen loads reactor R2 had a 61% lower volumetric specific nitrogen removal rate than reactor R1. The low pH and the associated low free ammonia (FA) concentrations were found to be critical to stable anammox activity in the MBBR. Nitrite enhanced the nitrogen removal rate in the conditions of low pH, all the way up to the investigated level of 50 mg NO₂-N/L. At low FA levels nitrite concentrations up to 250 mg NO₂-N/L did not cause inactivation of anammox consortia over a 2-days exposure time.     

Modelling of inhibition of nitrite oxidation in biological nitritation processes by free ammonia

To understand nitrite build-up in the nitritation process, the inhibition kinetics of free ammonia (FA) on nitrite oxidation were investigated. FA inhibition on nitrite oxidation was the key factor of nitrite build-up and its type was mixed inhibition. Therefore, the mixed inhibition model should be adapted to a Michaelis–Menten equation to set up a process rate equation for nitrite oxidation in the nitritation process.     

Effect of pH on domoic acid toxicity in mice

Domoic acid is a shellfish toxin which produces gastrointestinal distress, followed by neurological symptoms such as headache, confusion, disorientation and severe deficits in short-term memory. Domoic acid is an amino acid which contains three carboxylic groups, and one imino group, and its solubility, rate of absorption, and elimination would vary depending on the protonation of these groups at different pH's. We propose that domoic acid toxicity varies with pH of administered domoic acid solution. Domoic acid toxicity was measured in mice as the onset times for scratching behaviour, seizure activity, and death, after the intraperitoneal administration of domoic acid at different pH's. Results of the present study show that the scratching behaviour, seizure activity, and death, occurred at 12, 40, and 55 min, after intraperitoneal administration of domoic acid at pH 3.7. Apparently, the onset times for three types of behaviours were relatively long, and well separated from each ot her. Domoic acid toxicity was lowest at pH 3.7, and highest at pH 7.4, with intermediate toxicity at other pH's. The onset time of scratching behaviour was not influenced by pH of domoic acid solution at three different doses. In contrast, the onset times for seizure activity, and death were significantly affected by pH of domoic acid, toxicity being higher at pH 7.4 than at pH 3.7. The pH effect on domoic acid toxicity diminished as the dose of domoic acid was increased. In fact, at 14.5 mg/kg domoic acid toxicity was similar at both pH's of 3.7 and 7.4. It is concluded that in vivo toxicity of domoic acid varies depending on pH of the administered solution. The differential toxicity of domoic acid at different pH may be related to its solubility, rate of absorption, and elimination, depending on the degree of protonation of domoic acid molecule. Domoic acid toxicity would also vary depending on the age of animal, receptor sensitivity and density in different regions of brain. (Mol Ce ll Biochem 167: 179-185, 1997)     

Effects of medium glucose concentration and pH on docosahexaenoic acid content of heterotrophic Crypthecodinium cohnii

The effects of medium glucose concentration and pH on growth and docosahexaenoic acid (DHA, C22:6 ω-3) content of Crypthecodinium cohnii were investigated. Over a range of glucose concentrations (5–40 g l⁻¹) investigated, the highest specific growth rate (0.12 h⁻¹), highest cell dry weight concentration (3.13 g l⁻¹) and highest growth yield on glucose (0.6 g g⁻¹) were obtained at 20 g l⁻¹ glucose. However, the highest degree of fatty acid unsaturation (3.2) and highest DHA proportion (53.4% of total fatty acids) were achieved at 5 g l⁻¹ glucose. Low glucose concentrations enhanced the degree of fatty acid unsaturation and DHA formation. Medium pH also affected cell growth, fatty acid unsaturation and DHA proportion. When medium pH was 7.2, the highest specific growth rate (0.089 h⁻¹), highest cell dry weight concentration (2.73 g l⁻¹), highest growth yield on glucose (0.564 g g⁻¹), highest degree of fatty acid unsaturation (3.4) and highest DHA proportion (56.8% of total fatty acids) were obtained. Results suggest that glucose concentration and pH value could be effectively manipulated to achieve maximum DHA production by C. cohnii.     

Effect of ATP concentration and pH on rigor tension development and dissociation of rigor complex in glycerinated rabbit psoas muscle fiber

Isometric rigor tension development of glycerinated rabbit psoas muscle fibers in a medium, due to the formation of rigor complexes, was estimated at varying ATP concentrations from 0 to 2.5 mM and pH values from 6.75 to 8.20. The dissociation of rigor complexes was also estimated under the same conditions. When muscle fibers developed rigor tension from the relaxed and rigor states, the magnitude of rigor tension increased with increasing concentration of ATP. Transition between rigor and relaxation in single fibers occurred discontinuously at constant levels (critical levels) of ATP which were determined by pH. The critical concentrations of ATP necessary for inducing the transitions between rigor and relaxed states were also increased exponentially with increased pH. Incomplete repetition of tension development by the same fiber was also observed. This incomplete reversibility was divided into two types: one which showed a decay in rigor tension and another which showed no decay. The reason for the incomplete reversibility was discussed.     

Influence of pH, temperature, and inoculum composition on mixed cultures of Streptococcus thermophilus 404 and Lactobacillus bulgaricus 398

The effects of pH, temperature, and inoculum composition on Streptococcus thermophilus 404 and Lactobacillus bulgaricus 398 mixed cultures were studied. Linear quadratic models according to these three variables were proposed to express the growth and acidification characteristics, as well as the final proportion between the two strains. Initial and operating conditions, allowing maximization of these characteristics were determined. Finally, a graphic method to predict both final total concentration and final proportion of the two bacteria as a function of pH, temperature and inoculum composition is presented.     

Effect of pH on the extraction characteristics of succinic and formic acids with Tri-n-octylamine dissolved in 1-octanol

A study was made on the extraction equilibria of succinic and formic acids from aqueous solutions using tri-n-octylamine (TOA) in 1-octanol. It was shown that the loading values of TOA decreased with increasing pH values. The apparent equilibrium constants for each acid-amine complex were determined by an equilibrium model. In the case of succinic acid, the formation of a bisuccinate anion played an important role in the stoichiometry of the acid-amine complex.     

Current perspectives and challenges in understanding the role of nitrite as an integral player in nitric oxide biology and therapy

Beyond an inert oxidation product of nitric oxide (NO) metabolism, current thinking posits a key role for nitrite as a mediator of NO signaling, especially during hypoxia. This concept has been discussed in the context of nitrite serving a role as an endogenous modulator of NO homeostasis, but also from a novel clinical perspective whereby nitrite therapy may replenish NO signaling and prevent ischemic tissue injury. Indeed, the relatively rapid translation of studies delineating mechanisms of action to ongoing and planned clinical trials has been critical in fuelling interest in nitrite biology, and several excellent reviews have been written on this topic. In this article we limit our discussions to current concepts and what we feel are questions that remain unanswered within the paradigm of nitrite being a mediator of NO biology.     

Effect of substrate concentration,inorganic nitrogen,O2 concentration,temperature and pH on dehydrogenase activity in soil

Summary Dehydrogenase activity was measured in a sandy loam soil under a variety of incubation conditions using the reduction of 2-(p-iodophenyl-3-(p-nitrophenyl)-5-phenyl tetrazolium chloride (INT) to iodonitrotetrazolium formazan (INT-formazan). There was a high positive correlation between dehydrogenase activity and substrate concentration, incubation temperature, and soil pH. Dehydrogenase activity also displayed a high negative correlation with O₂ concentrations. Ammonium sulfate at concentrations from 40 to 120 g/g soil had no significant effect on dehydrogenase activity. However, at concentrations of 160 and 200 g/g, dehydrogenase activity was significantly reduced. Potassium nitrate at concentrations ranging from 40 to 200 g/g had no significant effect on soil dehydrogenase activity, whereas sodium nitrite significantly inhibited activity at concentrations of 120 and 160 g/g soil.     

Influence of nutrient solution pH on N2O and N2 emissions from a soilless culture system

The influence of nutrient solution pH on the emission of N2O and N2 was investigated during cultivation of cucumbers in a closed-loop rockwool system. Between pH 4 and 7 these gaseous nitrogen losses increased from 1.6 to 21.1% of the N fertilizer input. This was equivalent to average flux rates of 0.06 and 0.85 kg nitrogen per hectare greenhouse area and day, respectively. The N2O/N2 ratio was inversely related to the total gaseous nitrogen losses. At neutral pH dinitrogen was the main emission product, whereas more acidic conditions favoured the emission of nitrous oxide. The pH effects were probably not indirectly affected by root respiration or exudation as much as by a direct inhibition of the activity of denitrifying microorganisms due to high H+ concentrations since similar results were obtained in unplanted nutrient solution systems with the addition of glucose as carbon source. Despite the low microbial denitrification activity under acidic conditions, nitrogen balance deficits of up to one-fifth of the N input still occurred. It is suggested these losses were predominantly caused by chemodenitrification.     

The effect of pH on the rate of relaxation of isolated barnacle myofibrillar bundles

CO₂-induced acidosis in barnacle muscle fibres prolongs the relaxation phase of the electrically stimulated contraction (Ashley, C.C., Franciolini, F., Lea, T.J. and Lignon, J. (1979) J. Physiol. 296, 71P). In order to test if this effect is due to a direct action of H⁺ on the relaxation kinetics of the myofilaments, isolated myofibrillar bundles were contracted and relaxed in Ca²⁺ buffer solutions at pH 6.0 and 7.1, in the presence of 20 mM caffeine to inactivate the sarcoplasmic reticulum. At pH 7.1, the relaxation half-time was reduced from 1.5 to 0.3 s as the EGTA concentration in the relaxing solution was progressively increased from 0.3 to 50 mM. The resulting curve was shifted in the direction of increasing EGTA concentration by lowering the pH to 6.0. This effect could be explained by the reduction in affinity of Ca²⁺ for EGTA at pH 6.0, since relaxation half-times for a given relaxing pCa (calculated from the contaminating Ca²⁺ concentrations in the relaxing solutions) were shorter (by about 40%) at pH 6.0 compared with 7.1. However, similar experiments using the new Ca²⁺-chelating agent 1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid (BAPTA), which is much less pH sensitive than EGTA, indicated that there was no significant difference between relaxation half-times at pH 6.0 and 7.1 for a given relaxing pCa. It is concluded that because no prolongation of relaxation of the myofibrils was observed on lowering the pH from 7.1 to 6.0, the effect of CO₂ on the relaxation of intact muscle fibres is probably due to a modification of sarcoplasmic reticulum activity.     

The effect of the pH level on the communities of anoxygenic phototrophic bacteria of soda lakes of the southeastern Transbaikal Region

The effect of pH on the structure of the communities of anoxygenic phototrophic bacteria (APB) was studied under laboratory conditions. Samples of natural APB communities were inoculated into media that differed in pH values, which were 7, 9.5, or 10.5. The structure of the APB communities in the obtained enrichment cultures at all pH values depended also on the mineralization levels of the media, which were the same as in the lakes from which samples were taken. The same dependence of the community structure on salinity was observed as in the case of the natural communities that had been described previously. APB were most diverse in the enrichment cultures grown at pH 9.5. The shift of the pH to either neutral or extremely alkaline values restricted the species diversity within the APB community, resulting in marked predominance of the most adapted forms. It was shown that the status of Ectothiorhodospira species within the community could serve not only as an indicator of salinity but also as an indicator of pH in soda lakes with a water mineralization of higher than 5 g/l. The statuses of various APB groups in the community as dependent on pH and salinity are discussed, as well as possible changes in these statuses due to changes in the water level and other environmental parameters in the studied lakes.