Nitrogen dynamics and microbial food web structure during a summer cyanobacterial bloom in a subtropical,shallow, well-mixed,eutrophic lake (Lake Taihu,China)

Nitrogen dynamics and microbial food web structure were characterized in subtropical, eutrophic, large (2,338 km²), shallow (1.9 m mean depth), and polymictic Lake Taihu (China) in Sept–Oct 2002 during a cyanobacterial bloom. Population growth and industrialization are factors in trophic status deterioration in Lake Taihu. Sites for investigation were selected along a transect from the Liangxihe River discharge into Meiliang Bay to the main lake. Water column nitrogen and microbial food web measurements were combined with sediment–water interface incubations to characterize and identify important processes related to system nitrogen dynamics. Results indicate a gradient from strong phosphorus limitation at the river discharge to nitrogen limitation or co-limitation in the main lake. Denitrification in Meiliang Bay may drive main lake nitrogen limitation by removing excess nitrogen before physical transport to the main lake. Five times higher nutrient mineralization rates in the water column versus sediments indicate that sediment nutrient transformations were not as important as water column processes for fueling primary production. However, sediments provide a site for denitrification, which, along with nitrogen fixation and other processes, can determine available nutrient ratios. Dissimilatory nitrate reduction to ammonium (DNRA) was important, relative to denitrification, only at the river discharge site, and nitrogen fixation was observed only in the main lake. Reflecting nitrogen cycling patterns, microbial food web structure shifted from autotrophic (phytoplankton dominated) at the river discharge to heterotrophic (bacteria dominated) in and near the main lake.     

Denitrification in lucerne nodules is not involved in nitrite detoxification

Dissimilatory reduction of ionic nitrogen oxides to gaseous forms such as nitrous oxide or nitrogen can be carried out by free living or symbiotic forms of some strains of Rhizobium meliloti. In this paper we investigate whether bacteroid denitrification plays a role in the alleviation of the inhibitory effects of nitrate on nitrogen fixation both in bacteroid incubations as in whole nodules. The presence of a constitutive nitrate reductase (NR) activity in isolated bacteroids caused nitrite accumulation in the incubation medium, and acetylene reduction activity in these bacteroids was progressively inhibited, since nitrite reductase (NiR) activity was unable to reduce all the nitrite produced by NR and denitrification occurred slowly. Even nodules infiltrated with nitrate and nitrite failed to increase gaseous forms of nitrogen substantially, indicating that nitrite availability was not limiting denitrification by bacteroids. In spite of the low rates of bacteroidal denitrification, the effect of nodule denitrification on the inhibition of nitrogen fixation by nitrate in whole plants was tested. For that purpose, lucerne plants (Medicago sativa L. cv. Aragon) were inoculated with two Rhizobium meliloti strains: 102-F-65 (non denitrifying) and 102-F-51 (a highly denitrifying strain). After a seven days nitrate treatment, both strains showed the same pattern of inhibition, and it occurred before any nitrate or nitrite accumulation within the nodules could be detected. This observation, together with the lack of alleviation of the ARA inhibition in the denitrifying strain, and the limited activity of dissimilatory nitrogen reduction present in these bacteroids, indicate a role other than nitrite detoxification for denitrification in nodules under natural conditions.     

Denitrification in the river network of a mixed land use watershed: unpacking the complexities

River networks have the potential to permanently remove nitrogen through denitrification. Few studies have measured denitrification rates within an entire river network or assessed how land use affect rates at larger spatial scales. We sampled 108 sites throughout the network of the Fox River watershed, Wisconsin, to determine if land use influence sediment denitrification rates, and to identify zones of elevated sediment denitrification rates (hot spots) within the river network. Partial least squares regression models identified variables from four levels of organization (river bed sediment, water column, riparian zone, and watershed) that best predicted denitrification rates throughout the river network. Nitrate availability was the most important predictor of denitrification rates, while land cover was not always a good predictor of local-scale nitrate concentrations. Thus, land cover and denitrification rate were not strongly related across the Fox River watershed. A direct relationship between denitrification rate and watershed land cover occurred only in the Wolf River sub-watershed, the least anthropogenically disturbed of the sub-watersheds. Denitrification hot spots were located throughout the river network, regardless of watershed land use, with hot spot location being determined primarily by nitrate availability. In the Fox River watershed, when nitrate was abundant, river bed sediment character influenced denitrification rate, with higher denitrification rates at sites with fine, organic sediments. These findings suggest that denitrification occurring throughout an entire river network, from headwater streams to larger rivers, can help reduce nitrogen loads to downstream water bodies.

     

Effects of bioturbation by tube-dwelling chironomid larvae on oxygen uptake and denitrification in eutrophic lake sediments

1. Oxygen uptake and denitrification were determined in two bioturbated sediments from a eutrophic lake in southern Sweden. In laboratory mesocosms, an organic profundal sediment was incubated with Chironomus plumosus L. and a sandy littoral sediment with an organic-rich top layer was incubated with Polypedilum sp. Both species of chironomid are sediment tube-dwelling. 2. Oxygen consumption, expressed per gram of larval dry weight, was enhanced to the same extent by the larvae in both sediments. Measurements of the respiration rate of individual larvae revealed that the respiration per gram dry weight of the smaller Polypedilum sp. was more than three times higher than that of C. plumosus. 3. Denitrification was measured using the ‘nitrogen isotope pairing’ technique. In the organic sediment, denitrification of nitrate from the water phase (dw) and denitrification of nitrate from coupled nitrification (dn) were each correlated with the biomass of C. plumosus. In the sandy sediment, dw was correlated with the biomass of Polypedilum sp., while dn did not show any correlation with Polypedilum sp. 4. Oxygen uptake in the organic sediment was increased by a factor of 2.5, dw 5-fold and dn 2.5-fold at a biomass of 10 g m^–2 dry weight of C. plumosus. The same biomass of Polypedilum sp. in the sandy sediment resulted in a 2-fold stimulation of oxygen uptake and a 3-fold stimulation of dw, while dn was not affected. These differences in stimulation between oxygen uptake and denitrification by the larvae in the sediments suggest that the stimulation pattern cannot be explained by simple extension of the sediment surface. The burrows evidently reduce the distance between the nitrate source in the water column and the denitrifiers in the anoxic zones. 5. This study indicates that bioturbation by macrofauna elements can have a great impact on denitrification in lake sediments, and that different organisms can influence nitrogen turnover in specific ways.     

Short-term influence of recolonisation by the polycheate worm [2pt] Nereis succinea on oxygen and nitrogen fluxes and [2pt] denitrification: a microcosm simulation

The short-term effects of sediment recolonisation by Nereis succinea on sediment-water column fluxes of oxygen and dissolved inorganic nitrogen, and rates of denitrification, were studied in microcosms of homogenised, sieved sediments. The added worms enhanced oxygen uptake by the sediments, due to the increased surface area provided by the burrow walls and the degree of stimulation was stable with time. Similarly, ammonium fluxes to the water column were stimulated by N. succinea, but declined over the 3 day incubation in all microcosms including the controls. Nitrate fluxes were generally greater in the faunated microcosms, but highly variable with time. Denitrification rates were positively stimulated by N. succinea populations, denitrification of water column nitrate was stimulated 10-fold in comparison to denitrification coupled to nitrification in the sediments. Rates of denitrification of water column nitrate were not significantly different from rates in undisturbed sediment cores with similar densities of N. succinea, whereas rates of coupled nitrification–denitrification were 3-fold lower in the experimental set-up. These results may reflect the relative growth rates of nitrifying and denitrifying bacteria, which allow more rapid colonisation of new burrow surfaces by denitrifier compared to nitrifier populations. The data indicate that recolonisation by burrowing macrofauna of the highly reduced sediments of the Sacca di Goro, Lagoon, Italy, following the annual dystrophic crisis, may play a significant role in the reoxidation and detoxification of the sediments. The increased rates of denitrification associated with the worm burrows, may promote nitrogen losses, but due to the low capacity of nitrifying bacteria to colonise the new burrow structures, these losses would be highly dependent upon water column nitrate concentrations.     

Spatial distribution of denitrifying activity in a stream draining an agricultural catchment

SUMMARY 1. A field study was made of the spatial distribution of denitrification activity in the sediment of the River Dorn, Oxfordshire, England.
2. An assay of denitrifying enzyme activity was used to examine the distribution of denitrification with depth in cores of sediment representative of the types found in the stream. The maximum activity recorded in a predominantly silt sediment core was 5 times greater than that recorded in a sandy gravel core. In both fine sand and silt cores, peaks in denitrifier enzyme activity were shown to correspond to the limit of the nitrate diffusion front. At this depth the redox potential dropped rapidly from + 300 mV to 0 or less. Denitrifying enzyme activity in the stream water was negligible.
3. In situ denitrification activity (I DA) measurements were carried out in an 800 m reach of the Dorn using the acetylene inhibition technique on small sediment cores. Concurrent measurements were also made of stream depth and velocity, nitrate concentration in the interstitial water, and the wet bulk density, loss on ignition, mineraliz- able carbon and total nitrogen contents of the sediment. Mineralizable carbon was the variable which showed the best correlation with I DA. Highest IDAs were associated with accumulations of fine-grained sediment at meander bends. Mean IDAs measured under flood conditions were significantly higher ( P <0.05) than those measured under baseflow. It was estimated that denitrification reduced the nitrate load in the River Dorn by 15% under summer baseflow conditions     

河流沉积物氮循环主要微生物的生态特征

微生物驱动的氮循环过程是全球生物地球化学循环的重要组成部分,由于人类活动的影响,氮循环负荷加剧,氮素的生态平衡和微生物的功能特征也相应地受到干扰。河流生态系统是陆地与海洋联系的纽带,因人类活动过量活性氮的输入导致水体富营养化,明显影响着河流的生态功能以及河口沿岸海洋生态系统的平衡。富含微生物的沉积物对氮素的转化和去除起着至关重要的作用。本文主要介绍河流沉积物氮循环主要功能微生物,包括氨氧化细菌、氨氧化古菌、亚硝酸盐氧化菌、反硝化细菌和厌氧氨氧化细菌的群落特征和生态功能,总结氮相关营养盐、溶氧和季节变化等环境因子,以及河道控制管理措施和污水处理厂扰动等条件下氮循环过程主要功能类群的生态特征和响应关系。指出还需深入全面地研究河流沉积物生态系统氮循环过程的驱动机制和微生物的贡献效率,加强城市河流沉积物微生物功能作用的研究及河道生物修复技术的开发。     

Influence of Potamogeton pectinatus and microphytobenthos on benthic metabolism,nutrient fluxes and denitrification in a freshwater littoral sediment in an agricultural landscape: N assimilation versus N removal

The influence of Potamogeton pectinatus colonisation on benthic nitrogen dynamics was studied in the littoral zone of a lowland pit lake with high nitrate concentration (~200 μM). Our hypothesis was that in aquatic environments where nitrogen availability is not limiting, colonisation by rooted macrophytes changes the dynamics of the benthic nitrogen cycle, stimulating N assimilation and denitrification and increasing the system capacity to take up external nitrogen loads. To test this hypothesis, we quantified and compared seasonal variations of light and dark benthic metabolism, dissolved inorganic nitrogen (DIN) fluxes, denitrification and N assimilation rates in an area colonised by P. pectinatus and a reference site colonised by microphytobenthos. In both areas, the benthic system was net autotrophic and a sink for DIN (2,241–2,644 mmol m^?2 y^?1). Plant colonisation increased nitrogen losses via denitrification by 30% compared to the unvegetated area. In contrast to what is generally observed in coastal marine systems, where the presence of rooted macrophytes limits denitrification rates, under the very high nitrate concentrations in the studied lake, both denitrification (1,237–1,570 mmol m^?2 y^?1) and N assimilation (1,039–1,095 mmol m^?2 y^?1) played important and comparable roles in the removal of DIN from the water column.     

Nitrogen transformations at the sediment–water interface across redox gradients in the Laurentian Great Lakes

The capacity of a lake to remove reactive nitrogen (N) through denitrification has important implications both for the lake and for downstream ecosystems. In large oligotropic lakes such as Lake Superior, where nitrate (NO₃ ^?) concentrations have increased steadily over the past century, deep oxygen penetration into sediments may limit the denitrification rates. We tested the hypothesis that the position of the redox gradient in lake sediments affects denitrification by measuring net N-fluxes across the sediment–water interface for intact sediment cores collected across a range of sediment oxycline values from nearshore and offshore sites in Lake Superior, as well as sites in Lake Huron and Lake Erie. Across this redox gradient, as the thickness of the oxygenated sediment layer increased from Lake Erie to Lake Superior, fluxes of NH₄ ⁺ and N₂ out of the sediment decreased, and sediments shifted from a net sink to a net source of NO₃ ^?. Denitrification of NO₃ ^? from overlying water decreased with thickness of the oxygenated sediment layer. Our results indicate that, unlike sediments from Lake Erie and Lake Huron, Lake Superior sediments do not remove significant amounts of water column NO₃ ^? through denitrification, likely as a result of the thick oxygenated sediment layer.     

Presence of Nitrate-Accumulating Sulfur Bacteria and Their Influence on Nitrogen Cycling in a Shallow Coastal Marine Sediment

Nitrate flux between sediment and water, nitrate concentration profile at the sediment-water interface, and in situ sediment denitrification activity were measured seasonally at the innermost part of Tokyo Bay, Japan. For the determination of sediment nitrate concentration, undisturbed sediment cores were sectioned into 5-mm depth intervals and each segment was stored frozen at −30°C. The nitrate concentration was determined for the supernatants after centrifuging the frozen and thawed sediments. Nitrate in the uppermost sediment showed a remarkable seasonal change, and its seasonal maximum of up to 400 μM was found in October. The directions of the diffusive nitrate fluxes predicted from the interfacial concentration gradients were out of the sediment throughout the year. In contrast, the directions of the total nitrate fluxes measured by the whole-core incubation were into the sediment at all seasons. This contradiction between directions indicates that a large part of the nitrate pool extracted from the frozen surface sediments is not a pore water constituent, and preliminary examinations demonstrated that the nitrate was contained in the intracellular vacuoles of filamentous sulfur bacteria dwelling on or in the surface sediment. Based on the comparison between in situ sediment denitrification activity and total nitrate flux, it is suggested that intracellular nitrate cannot be directly utilized by sediment denitrification, and the probable fate of the intracellular nitrate is hypothesized to be dissimilatory reduction to ammonium. The presence of nitrate-accumulating sulfur bacteria therefore may lower nature's self-purification capacity (denitrification) and exacerbate eutrophication in shallow coastal marine environments.     

The effect of sewage effluent on Grasmere (English Lake District) with particular reference to inorganic nitrogen transformations

SUMMARY. In the summer of 1971 the village of Grasmere was converted from septic tank to mains drainage with sewage treatment at an activated sludge plant. The effluent was discharged into the River Rothay, the main inflow of a nearby small lake (Grasmere). This paper describes some of the effects on the lake. The mean areal hypolimnetic oxygen deficit increased from 274 to 434 mg O₂ m⁻² day⁻¹ with an accompanying marked increase in the degree of deoxygenation in the hypolimnion. Plate counts of bacteria in the surface water increased for 2 years but this increase was not sustained. The mean summer soluble reactive phosphorus concentration did, however, increase significantly ( P = 0.05), but the same was not true of nitrate levels. This paper is particularly concerned with inorganic nitrogen transformations and analyses of the main inflow have shown that 50–98% of the ammonia and 10–40% of the nilrate entering in this river was derived from the sewage effluent. The concentrations in the main body of the lake were usually lower, possibly due to assimilation and denitrification in the shallower reaches of the lake. Seasonal changes in the inorganic nitrogen species in the hypolimnion showed three distinct phases of activity, ammonification, nitrification and denitrification. Nitrification accounted for approximately a quarter of the oxygen uptake in the hypolimnion.     

洱海沉积物中反硝化细菌的分离与反硝化作用研究

自洱海十个点位的沉积物中富集筛选出101株反硝化细菌并从中筛选出1株较强反硝化能力的细菌,命名为EH314。该细菌接触酶(过氧化氢酶)试验、产硫化氢试验和淀粉水解均为阳性,葡萄糖氧化发酵实验结果为氧化菌,产脂酶(Tween 80)试验结果为阴性;初步鉴定该菌为产碱杆菌属细菌;对细菌反硝化能力进行测定发现,菌株EH314能有效地降解水体中的硝酸盐且反硝化可在有氧条件下进行。     

Experimental measurement of sediment nitrification and denitrification in Hamilton Harbour,Canada

This research examines the role of sediment nitrification and denitrification in the nitrogen cycle of Hamilton Harbour. The Harbour is subject to large ammonia and carbon loadings from a waste-water treatment plant and from steel industries. Spring ammonia concentrations rapidly decrease from 4.5 to 0.5 mg 1⁻¹, while spring nitrate concentrations increase from 1 to 2 mg l⁻¹, by mid-summer. A three-layer sediment model was developed. The first layer is aerobic; in it, oxidation of organics and nitrification occurs. The second layer is for denitrification, and the third layer is for anaerobic processes. Ammonia sources for nitrification include diffusion from the water column, sources associated with the oxidation of organics, sources from denitrification and from anaerobic processes. Diffusion of oxygen, ammonia and nitrate across the sediment-water interface occurs. Temperature effects are modelled using the Arrhenius concept. A combination of zero-order kinetics for nitrate or ammonia consumption with diffusion results in a half-order reaction, with respect to the water column loss rate to sediments. From experimental measurement, the rate of nitrification is 200 mg N 1⁻¹ sediment per day, while that of denitrification is 85 mg N 1–1 sediment per day at 20 °C. The Arrhenius activation energy is estimated as 15 000 cal/ mole-K and 17 000 cal/ mole-K for nitrification and denitrification, respectively, between 10 °C and 20 °C. Calculations of the flux of ammonia with the sediments, using the biofilm model, compare favourably with experimental observations. The ammonia flux from the water column is estimated to account for 20% of the observed decrease in water column stocks of ammonia, while the nitrate flux from the water column is estimated to account for 25% of the total nitrogen produced by the sediments.     

Denitrification in deep subsurface sediments

Dissimilatory nitrate reduction (denitrification) in subsurface sediments by indigenous microflora was investigated in samples obtained over a range of depths from 0 to 289 m. Denitrifying activity in sediment samples retrieved from similar stratigraphic horizons at four different sites was determined by measuring the accumulation of N₂O using the acetylene blockage technique. Denitrification was detected in unamended samples which received only prereduced deionized water at almost all depths in all sediments sampled. The surface sediments showed the highest denitrification activity. In the deeper sediments, denitrifying activity was much higher in saturated sandy samples and lower or absent in drier clay samples. Addition of nitrate enhanced denitrification activity in all samples from below the water table down to the maximum depth sampled (289 m), while addition of a carbon (succinate) source in general had no stimulatory effect. These results show that denitrifying microorganisms were present in all of the deep subsurface sediments tested in this study. Furthermore, these results suggest that adequate supplies of metabolizable organic carbon were available to support denitrifying activity. However, denitrification may be limited by inadequate supplies of nitrate in the sediments.     

Denitrification of PVA-immobilized denitrifying photosynthetic bacterium,Rhodobacter sphaeroides

A newly isolated denitrifying strain, Rhodobacter sphaeroides NII2 was immobilized in polyvinyl alcohol (PVA) gel, and the properties of the cells in the gel were examined. The immobilized cells had low or almost no denitrification activity, but the cells were activated by incubation in light with culture medium for denitrification containing 0.5% nitrate and no other nitrogen source. Cells grown in the dark were activated by incubation at an earlier stage and to a higher rate than the light-grown cells. The activation was markedly enhanced in the PVA gel with a low cell concentration. The immobilized cells consumed nitrate with a temporary accumulation of NO₂ and evolved nitrogen gas. The immobilized cells could use various organic compounds as electron donors for denitrification. Thus, the immobilized cells were applied to a continuous treatment of synthetic wastewater using an aparatus devised by this laboratory. The results showed an efficient removal of NO₃-N from the test water.     

Denitrification in Marl and Peat Sediments in the Florida Everglades

The potential for denitrification in marl and peat sediments in the Shark River Slough in the Everglades National Park was determined by the acetylene blockage assay. The influence of nitrate concentration on denitrification rate and N₂O yield from added nitrate was examined. The effects of added glucose and phosphate and of temperature on the denitrification potential were determined. The sediments readily denitrified added nitrate. N₂O was released from the sediments both with and without added acetylene. The marl sediments had higher rates than the peat on every date sampled. Denitrification was nitrate limited; however, the yields of N₂O amounted to only 10 to 34% of the added nitrate when 100 μM nitrate was added. On the basis of measured increases in ammonium concentration, it appears that the balance of added nitrate may be converted to ammonium in the marl sediment. The sediment temperature at the time of sampling greatly influenced the denitrification potential (15-fold rate change) at the marl site, indicating that either the number or the specific activity of the denitrifiers changed in response to temperature fluctuations (9 to 25°C) in the sediment. It is apparent from this study that denitrification in Everglades sediments is not an effective means of removing excess nitrogen which may be introduced as nitrate into the ecosystem with supply water from the South Florida watershed and that sporadic addition of nitrate-rich water may lead to nitrous oxide release from these wetlands.     

Vertical distribution of nitrite reductase genes (nir S) in continental margin sediments of the Gulf of Mexico

Marine sediments account for up to 66% of the loss of nitrogen load to coastal areas. Sedimentary denitrification is the main sink for fixed nitrogen in the global nitrogen budget, and thus it is important to understand the structure and composition of denitrifying communities. To understand the structure and composition of denitrifying communities, the diversity of nitrite reductase (nirS) genes from sediments along the Gulf of Mexico was examined using a PCR-based cloning approach. Sediments were collected at three different depths (0-0.5, 4-5 and 19-21 cm). Geochemical analysis revealed decreasing nitrate and oxygen concentrations with increasing sediment depth. This trend coincided with the decrease in diversity of denitrifying bacteria. LIBSHUFF analysis indicated that the clone library in the shallowest sediment (depth, 0-0.5 cm) was significantly different from that in the deepest sediment (depth, 19-21 cm), and that the deeper sediments (depths of 4-5 and 19-21 cm) were significantly similar. Community structural shifts were evident between the shallowest (oxic zone) and deepest (anoxic zone) sediments. Community changes within the deepest sediments were more subtle, with the presence of different nirS clone sequences gradually becoming dominant or, alternatively, decreasing with depth. The changes in community structure at this depth are possibly driven by nutrient availability, with lower quality sources of carbon and energy leading to the disappearance of nirS sequences common in the top layer. The majority of recovered nirS sequences were phylogenetically divergent relative to known denitrifying bacteria in the database.     

Seasonal effects of zebra mussels on littoral nitrogen transformation rates in Gull Lake, Michigan, U.S.A.

• 1 Zebra mussels (Dreissena polymorpha) are successful colonisers of lake littoral habitats and they interact strongly with littoral benthos. Previous research suggests that localised areas colonised by zebra mussels may be hotspots of nitrogen (N) cycling.
• 2 The effects of zebra mussels on nitrification and denitrification rates were examined approximately every other month for 1 year in Gull Lake, Michigan, U.S.A. Littoral sediment was collected from an area free of zebra mussels and distributed into shallow trays; rocks colonised with zebra mussels were placed in half of the trays, while uncolonised rocks were placed in the remaining trays. After an incubation period of 6–8 weeks in the lake, sediment and zebra mussels were collected from the trays, replaced with new sediment and zebra mussels, and placed in the lake for the next interval. In the laboratory, sediment nitrification and denitrification rates were measured for each tray.
• 3 Sediment nitrification rates did not increase in the presence of zebra mussels; instead nitrification rates were sensitive to changes in water temperature and increased with increasing exchangeable sediment ammonium. In contrast, denitrification rates increased in sediment trays with zebra mussels in the winter when nitrate (NO₃⁻) availability was high and when Chara did not grow in the trays.
• 4 Sediment denitrification was NO₃⁻‐limited in all seasons, regardless of zebra mussel treatment. However, sediment in the presence of zebra mussels responded less to NO₃⁻ addition, suggesting that NO₃⁻ limitation of denitrification can be reduced by zebra mussel activity. Zebra mussels have a seasonally variable impact on sediment denitrification rates, and this may translate into altered seasonal patterns of N cycling in localised areas of lakes where they are particularly abundant.

     

Quantitative selection of denitrifying bacteria in continuous cultures and requirement for organic carbon. I. Starch

A mixed population of bacteria from bottom sludge of nitrogen wastewater reservoir was incubated in continuous culture in medium containing 1000 mg nitrate nitrogen/l and starch. Maximal efficiency of denitrification was 5 mg N/l/h. Marked changes in participation of denitrifying bacteria (16-76%) among total number of bacteria was observed, this being dependent on the ratio of starch concentration (CS) to nitrogen concentration (CN) in the medium. The optimal CS/CN ratio ensuring highest participation of denitrifying bacteria was 3.2. The amount of starch required for the denitrification of a defined quantity of nitrogen is negatively correlated (r = -0.98) with the frequency of the occurrence of denitrifying bacteria (XD) and is: CS = (5.53-0.028XD) CN. The denitrifying bacteria in continuous culture were dominated, depending on CS/CN ratio and flow rate of medium, by Alcaligenes faecalis, Paracoccus denitrificans or Pseudomonas mendocina, that is species unable to hydrolyse starch.