Biol Fertil Soils (1997) 24:323328

Springer-Verlag 1997

O R I G I N A L PA P E R

T. Mahmood R. Ali K. A. Malik S. R. A. Shamsi

Denitrification with and without maize plants (Zea mays L.) under irrigated field conditions

Received: 15 April 1996

Abstract The study was conducted under irrigated field conditions to examine the effect of maize plants on denitrification. Both planted and unplanted field plots received 150 kg N ha1 as urea. In a third treatment, which was also planted and received urea at 150 kg N ha1, the soil nitrate N content was brought up to equal to that in the unplanted plots by applying additional doses of N as calcium nitrate. Soil cores were collected 24 and 72 h after irrigation and the denitrification rate was measured by the acetylene inhibition method. Nitrate-N content, aerobically mineralizable C, microbial biomass carrying capacity and denitrification potential were also studied on field-moist soil. Maize plants grown under field conditions always had the potential to increase denitrification in conditions of both high and low water-filled porosity. When nitrate-N content of the planted soil decreased due to plant uptake, denitrification was reduced in the planted soils. However, when nitrate-N uptake by plants was compensated through additional doses of nitrate fertilizer, denitrification was always higher in planted than unplanted soil. The stimulatory effect of plants on denitrification was observed at both high and low soil nitrate-N concentrations, though it was more pronounced at high nitrate-N levels. The effect of plants on denitrification and related parameters was confined to the root zone. Key words Denitrification Denitrification potential Irrigated field Mineralizable carbon Effect of maize plants

Introduction
It is well established that roots of growing plants release metabolizable C compounds into the surrounding soil (Rovira and McDougall 1967). Plant-derived C in soil also comprises sloughed off root cells, root hairs, small rootlets and products of autolysis and microbial breakdown (Rovira et al. 1979; Rovira 1981). The contribution of root exudates to decomposable substrate varies with plant species and growth stage and may amount to 625% of the net photosynthates (Barber and Martin 1976; Haller and Stolp 1985; Milchunas et al. 1985; Biondini et al. 1988). Root-derived C in soil under cereal crops may be as high as 15002040 kg C ha1 (Martin and Puckridge 1982; Haider et al. 1987). However, the extent to which this root-derived C is utilized by soil heterotrophs is controversial. Shamoot et al. (1968) found that only 30% of the fresh rhizodeposits was released as CO2 during 14 weeks. On the other hand, Sauerbeck and Johnen (1976, 1977) reported that 80% of the CO2 from planted soil originated from rhizodeposits. Utilization of root-derived C by microbes under denitrifying conditions is also less well understood. Haider et al. (1985) found a higher denitrification capacity in wheatplanted than unplanted soil, but the increase in denitrification due to plants was proportionally lower if compared with the increase in soluble C in the soil. In another study, these authors again did not find and increase in denitrification due to actively growing corn plants and concluded that the root exudates were not able to fuel the denitrification process (Haider et al. 1987). In contrast, Haller and Stolp (1985) demonstrated utilization of corn root exudates by Pseudomonas aeruginosa under both aerobic and denitrifying conditions. Wollersheim et al. (1987) studied the interaction of bulk density and soil water tension with denitrification in the rhizosphere of spring wheat. These authors found a lower root biomass when high bulk density accompanied low water tension but the denitrification intensity (N2O g1 root) increased due to higher root exudation under such stress conditions. Klemedtsson et al. (1987) attributed stimulation of denitrifying bacteria by

T. Mahmood (u) R. Ali Soil Biology Division, Nuclear Institute for Agriculture and Biology, PO Box 128, Faisalabad, Pakistan K. A. Malik National Institute for Biotechnology and Genetic Engineering, PO Box 577, Faisalabad, Pakistan S. R. A. Shamsi Department of Botany, University of the Punjab, Lahore, Pakistan

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roots mainly to an effect of decreased O2 in the planted soil. Prade and Trolldenier (1988) reported an increase in the rhizosphere effect on denitrification with increasing soil organic matter content due to enhanced rhizosphere O2-depletion. Plant nutritional and soil factors in relation to rhizosphere denitrification have been discussed by Trolldenier (1989). Little information is available on the effect of plants on denitrification under field conditions. Vinther (1984) reported 2.5 and 5 times higher denitrification due to barley plants in field plots receiving 30 and 150 kg N ha1, respectively. The present study was carried out to examine under irrigated field conditions the effect of maize plants on denitrification and on some factors governing its rate.

capture detector. Analysis of CO2 was carried out on a Gasukuro Kogyo 370 gas chromatograph equipped with a thermal conductivity detector (TCD). Nitrous oxide production was adjusted for N2O dissolved in water using Bunsen absorption coefficients (Moraghan and Buresh 1977).

Analysis Soil sampling for other analyses was done in the same way as for denitrification, but the soil (four cores) from each replicate plot was pooled and mixed, and visible roots manually removed and passed through a 2-mm screen. The soil was processed within 4 h of collection and analysed. We did not apply an electrostatic treatment to remove root hairs and fine root material as done by Haider et al. (1985). Therefore, in our study the effect of rhizodeposits and root exudates could not be distinguished. Soil nitrate-N was determined by the micro-Kjeldahl method as described by Keeney and Nelson (1982). The chloroform-fumigation method as described by Groffman and Tiedje (1989) was used for measurement of microbial biomass carrying capacity (MBCC). A 10-g portion of the field-moist soil was subjected to CHCl3 fumigation at 30 C for 24 h, followed by the removal of CHCl3 vapours and inoculation with fresh soil. Soil was incubated at 30 C for 10 days. Incubations were carried out in 100-ml serum vials sealed with a silicone rubber septum. At the end of incubation, the headspace was analysed for CO2-C by gas chromatography using a TCD. A kc factor of 0.45 was used to calculate the MBCC. The incubation and analytical procedure for measuring aerobically mineralizable C (AMC) of soil was similar to that for MBCC, except that soil was unfumigated and no kc factor used. For denitrification potential (DNP), 10-g portions of the field-moist soil in 100-ml serum vials were treated with 10 ml KNO3 solution to establish a 1 NO 3 level equivalent to 200 lg N g . After sealing with a silicone rubber septum, the vials were made anaerobic by evacuating and flushing 3 times with O2-free N2. The headspace was then replaced by acid-washed C2H2 (0.1 atm.) and the vials were incubated at 30 C. After 48 h of incubation, contents in the vials were vigorously shaken by hand and the headspace analysed for N2O by gas chromatography using a TCD. Water-filled pore space (WFPS) was calculated as WFPS = [(gravimetric water contentsoil bulk density)/total soil porosity], where soil porosity= [1 soil bulk density/particle density].

Materials and methods

Field experiments The field site was located at the Nuclear Institute for Agriculture and Biology, Faisalabad. The soil, which belongs to the Hafizabad series, is a deep, moderately well drained sandy-clay loam developed in mixed calcareous medium-textured alluvium derived from the Himalayas. The site had been under a wheat-maize rotation for the previous 2 years. Properties of the plough layer were as follows: pH, 7.8; bulk density, 1.56 g cm3; particle density, 2.71 g cm3, saturation, 38%; total organic C, 1.14%, total N, 0.07%; sand, 49%; silt, 27.4%; and clay, 23.6%. Twelve experimental plots (48 m) were set out for three treatments which were laid down in a randomized complete block design, each replicated 4 times. All plots received P2O5 at 50 kg ha1 (as single superphosphate) at the time of land preparation and K2O (as K2SO4) at 110 kg ha1:50 kg ha1 at land preparation and 30 kg ha1 at both 37 and 52 days after germination (DAG). All plots also received urea-N at 150 kg ha1:50 kg ha1 at sowing and 40, 30 and 30 kg ha1 at 37, 52 and 63 DAG, respectively. Treatments included planted (T 1), unplanted (T 2) and planted+nitrate (T 3). To T 3, calcium nitrate was applied at the time of irrigation to equalize its soil nitrate-N content with that of T 2. Total calcium nitrate applied to T 3 during the 88-day study period was equivalent to 173 kg N ha1. After land preparation and fertilizer application, the T 1 and T 3 plots were seeded to maize (Zea mays L. var. Composit-17) with plant-to-plant and row-to-row distances of 20 and 30 cm, respectively. However, to study the effect of distance from plants on denitrification, the row-to-row distance in some portions of T 1 plots was increased to 60 cm. To control insect pests, diazenon was applied at 15 kg ha1 to all plots with the first irrigation. Soil was sampled at 24 and 7296 h after each irrigation. We assumed an active layer of 15 cm, taking no account of processes in the deeper layers. Quantification of soil respiration and denitrification Denitrification and soil respiration rates under field conditions were measured by the soil-core incubation method (Ryden et al. 1987). From each replicate plot, four intact soil cores (315 cm diameterlength) were randomly extracted in PVC sleeves with a sampling device similar to that of Rice and Smith (1982). Sampling for planted treatments (T 1 and T 3) was done at a distance of 1 cm from maize plants. Soil cores from each replicate plot were placed together in a field incubation jar (nominal volume 800 ml). The jars were sealed with silicone rubber stoppers that had a septum port to facilitate gas sampling. After replacing the headspace with acid-washed C2H2 (0.1 atm.), the jars were incubated in holes made within the experimental field. After 4 and 12 h of incubation, the atmosphere in the jars was repeatedly mixed with a 50-ml syringe and gas samples removed for analyses of N2O and CO2. Nitrous oxide was analysed on a Hitachi 26330 gas chromatograph equipped with a 63Ni electron

Statistics The data were subjected to analysis of variance followed by Duncans Multiple Range Test (Steel and Torrie 1980). To satisfy the assumption of variance homogeneity, data for denitrification rate, soil NO 3-N and soil respiration were log-transformed before statistical analyses. Correlation analyses were performed using Lotus 1-2-3 software.

Results
There was no effect of plants on WFPS. Therefore, results of the average WFPS of all treatments are presented (Table 1). Water-filled pore space 24 h after irrigation or rainfall ranged between 64% and 84% and was significantly higher (P<0.05) than that observed at 72 h after irrigation when it ranged between 45% and 57%. The WFPS of soil sampled 24 h after first irrigation was significantly lower than that recorded 24 h after later irrigations (P<0.05). This was probably due to the loose structure of the plough layer and the prolonged dry period (33 days) preceding the first irrigation, which caused rapid movement of the irrigation water down to deeper soil layers.

325 Table 1 Water-filled pore space (WFPS), temperature and denitrification rate in the field soil Days after germination (DAG) 18 (I) 20 38 (I) 40 64 (I) 66 75 (I) 78 85 (I) 88 (R) b Treatment mean
a c b b

WFPS (%) 66 53 77 56 81 57 77 45 77 64

Temperature a (8C) 23.0 18.8 26.8 23.8 27.0 29.0 31.2 31.0 29.5 28.3

Denitrification rate (g N ha1 day1) Planted 454 21 359 75 128 219 171 14 56 126 162 b a c b c a c a c c Cd
c

Unplanted 226 28 740 46 698 104 482 17 751 337 343 c a b b b c b a b b B

Planted+nitrate 849 a 55 a 1202 a 144 a 963 a 148 b 4935 a 43 a 2347 a 862 a 1155 A

Date mean 510 35 767 78 596 157 1863 25 1051 442 Ed I C H D G A I B F

Average soil temperature (5 cm) during the incubation period Sampling 24 h after irrigation (I ) or rainfall (R ) Values in each row followed by different letter (lower case ) are significantly different at P<0.05 with respect to treatment effect at given sampling date (DAG) d Values for date or treatment means followed by a different letter (upper case ) are significantly different at P<0.05 Table 2 Nitrate-N content of the field soil with and without maize plants Days after germination (DAG) Treatment a (lg N g1) Planted Unplanted 16.7 a 13.4 ab 3.9 a 16.1 a 7.4 a 14.3 a 11.7 a 10.6 a 1.1 a 3.9 a 9.4 A Planted+ nitrate b 18.5 a 17.1 a 3.2 a 13.0 a 8.1 a 18.7 a 11.8 a 12.4 a 1.1 a 3.9 a 10.8 A Date mean 15.9 13.3 2.9 13.3 5.2 14.8 8.5 9.3 0.7 3.6 Ad A C A D A B B E C

18 12.3 a c 20 9.6 b 38 1.5 b 40 10.8 a 64 0.1 b 66 11.4 a 75 2.0 b 78 5.0 b 85 0.1 b 88 3.0 a Treatment mean 5.9 B d
a

All treatments received urea at 150 kg N ha1:50 kg at sowing, 40 kg at 37 DAG and 30 kg each on 52 and 63 DAG b Received additional doses of N as calcium nitrate on 17, 37, 63 and 74 DAG to level its nitrate N content with that of the unplanted soil c Values in each row followed by a different letter (lower case ) are significantly different at P<0.05 with respect to treatment effects at the given sampling date (DAG) d Values for date or treatment means followed by a different letter (upper case ) are significantly different at P<0.05

At early stages of crop growth, i.e. up to 20 days after germination (DAG), nitrate-N content of the planted (T 1) soil was not different from that of the unplanted soil (Table 2). However, during the later growth period due to plant uptake, nitrate-N content of the planted (T 1) soil was generally lower than that of the unplanted soil (P<0.05). Averaging all sampling dates, nitrate-N of planted soil (T 1) was significantly lower than that of the unplanted soil (P<0.05). However, addition of calcium nitrate to planted soil (T 3) at different stages balanced the nitrate-N of planted soil with that of the unplanted soil. The spatial variability in denitrification rate was high and gave a mean CV value of 49%, with a range of 7

105%. Nevertheless, differences between treatments were statistically distinguishable. Denitrification was strongly influenced by WFPS as evidenced by a strong correlation between denitrification and WFPS (r = 0.751; P<0.001). The rates recorded after 24 h of irrigation were 375 times higher than those recorded after 72 h (P<0.05). A higher denitrification rate in the planted (T 1) than unplanted soil was recorded at initial stages of growth (18 DAG), when planted soil (T 1) denitrified twice as high as the unplanted soil (Table 1). A reverse trend was observed during all later samplings done 24 h after irrigation or rainfall. At lower WFPS, a stimulatory effect of plants was also observed on one occasion (66 DAG). On this occasion, however, soil nitrate-N of the planted (T 1) and unplanted soils was high. Addition of calcium nitrate to the planted soil (T 3) caused a marked increase in denitrification rate as compared with the unplanted soil and this was observed at all sampling dates. The relative increase in denitrification rate due to plants ranged from 0.4- to 2.8-fold, with an exceptionally high stimulation (9.2-fold) recorded at 75 DAG. The stimulatory effect of plants on denitrification was also observed at low soil nitrate-N concentrations (38, 85 and 88 DAG). Comparison of the unplanted and planted+nitrate treatments also revealed that plants stimulated denitrification at both high (6481%) and low (56 57%) WFPS. On average, plants caused a 2.8- and 1.3fold increase in denitrification rate at high and low WFPS, respectively. However, if the very high stimulation (9.2fold) caused by plants at 75 DAG is not included in the comparison, the average stimulatory effect of plants on denitrification was almost similar at high (1.5-fold) and low (1.3-fold) WFPS. Cumulative denitrification losses during the 88-day study period were 2.6, 5.9 and 21.4 kg N ha1 from planted, unplanted and planted+nitrate treatments, respectively. Results of different indices of C availability in the planted and unplanted soils are presented in Table 3. Irrespective of sampling date, C availability was higher in the

326 Table 3 Carbon availability in the field soil with and without maize plants Days after germination (DAG) Treatment Planted Unplanted Planted+ nitrate 107 82 180 159 89 127 148 89 134 126 124 b b a b c b b b b b B a a b a b a a ab a a A Date mean

Aerobically mineralizable C (lg g1) 96 a 18 111 a a 20 95 a 72 a 38 239 a 125 c 40 158 a 114 b 64 114 a 60 c 66 113 a 83 b 75 123 b 51 c 78 109 a 69 b 85 111 b 66 c 88 121 a 82 b Treatment mean 129 A b 82 B Soil respiration rate (kg C ha1 day1) 18 15.2 a 8.2 20 5.9 a 2.4 38 9.6 a 6.9 40 14.6 a 2.1 64 6.9 b 2.5 66 34.5 a 9.3 75 34.3 a 10.2 78 25.4 a 5.0 85 34.2 a 7.1 88 30.3 a 14.1 Treatment mean 21.1 A 6.8

105 C b 83 D 181 A 144 B 88 D 108 C 107 C 89 D 104 C 110 C

15.4 a 5.8 a 11.1 a 10.2 a 15.5 a 24.2 a 53.5 a 24.9 a 24.6 a 28.7 a 21.4 A a a ab a a ab a a a a A

12.9 4.7 9.2 9.0 8.3 22.7 32.7 18.4 22.0 24.4

D G E F F AB A CD BC AB

Microbial biomass carrying capacity (lg C g1) 18 341 a 324 a 320 20 304 a 280 a 289 38 347 a 260 b 294 40 371 a 263 b 401 64 296 a 202 b 305 66 281 a 214 b 262 75 365 a 226 b 358 78 387 a 271 ab 335 85 379 a 241 b 376 88 316 ab 260 b 363 Treatment mean 339 A 254 B 330 Denitrification potential (ng N g1 h1) 18 358 a 354 a 20 323 a 261 a 38 365 a 164 b 40 522 a 280 b 64 481 a 146 b 66 574 a 328 b 75 917 b 353 c 78 876 a 378 b 85 683 a 196 b 88 459 a 277 b Treatment mean 556 A 274 B
a

328 291 300 345 268 252 316 331 332 313

AB B B A C C AB AB AB AB

391 a 219 a 244 ab 406 ab 424 a 507 a 1136 a 760 a 703 a 454 a 524 A

368 268 258 403 350 470 802 671 527 397

E F F DE EF CD A B C DE

Values in each row followed by a different letter (lower case ) are significantly different at P<0.05 with respect to treatment effects at a given sampling date (DAG) b Values for date or treatment means followed by a different letter (upper case ) are significantly different at P<0.05

planted (T 1) than unplanted soil (P<0.05). A marked temporal pattern was observed in soil respiration rate, DNP and AMC in planted soil. A slightly lower MBCC was observed on two occasions (64 and 66 DAG) probably due to the moisture stress period between 40 and 60 DAG

when irrigation water was not available. Otherwise, the temporal pattern for MBCC was relatively uniform. Temporal changes in different parameters of C availability were also observed in unplanted soil though these were less pronounced than with the planted soil. On almost all sampling occasions, soil respiration rate was higher in the planted (T 1) than unplanted soil (P<0.05), and on average a 2.1-fold increase in soil respiration rate was observed due to plants. The denitrification potential of the planted and unplanted soils was not different during the initial 20 days. At later stages, however, planted soil (T 1) always possessed a higher DNP than the unplanted soil (P<0.05). Regardless of sampling date, the DNP of the planted soil was twice as high as that of the unplanted soil (P<0.05). Aerobically mineralizable C followed similar trends to those observed for DNP, though the average increase in AMC due to plants was 0.6-fold. Like DNP and AMC, the MBCC of the planted and unplanted soils was similar until 20 DAG. However, during the later growth period it was generally higher in planted (T 1) than in the unplanted soil (P<0.05). The stimulatory effect of plants on MBCC was less pronounced than on soil respiration rate, DNP and AMC, producing a 0.3-fold increase over unplanted soil. Comparing planted (T 1) and planted+nitrate (T 3) treatments, though the addition of calcium nitrate caused better plant growth, its effect on C availability in soil was not frequently observed. An increased soil respiration rate was observed on 64 DAG. Aerobically mineralizable C was slightly higher in nitrate-amended planted soil on 75 and 85 DAG whereas the same was observed for DNP on 75 DAG. However, nitrate-amendment to planted soil sometimes caused a decrease in AMC (38 and 64 DAG). Microbial biomass carrying capacity of the planted soil was not affected by nitrate amendment. The relationship between denitrification rate and different C availability indices was evaluated by linear regression. Considering all the data (n = 30), the denitrification rate was significantly correlated with soil respiration (r = 0.514; P<0.01) and DNP (r = 0.444; P<0.05). Regression analyses based on the data of unplanted and planted+nitrate treatments (n = 20) improved the correlation between denitrification rate and soil respiration (r = 0.786; P<0.001), DNP (r = 0.719; P<0.001) and AMC (r = 0.434; P<0.05). These results signify the positive effects of plants on denitrification by the increased O2 consumption in the soil and by supplying C substrate to denitrifiers. Distance from plants had a marked influence on denitrification rate and related parameters (Table 4). Denitrification rate was significantly lower in the soil close to plants than that collected at a distance 15 or 30 cm from maize plants (P<0.05). A similar trend was observed for soil nitrate-N content. Different parameters of C availability, e.g. soil respiration rate, DNP, AMC and MBCC, revealed a significantly higher C availability in the root zone soil than in the distant soil (P<0.05).

327 Table 4 Effect of distance from maize plants on denitrification and related parameters a Parameter Distance from plant (cm) 0 Denitrification rate (g N ha day ) Soil respiration rate (kg C ha1 day1) Soil nitrate N content (lg g1) Denitrification potential (ng N g1 h1) Aerobically mineralizable C (lg g1) Microbial biomass carrying capacity (lg C g1) Water-filled pore space (%)
a b 1 1

15
b

30 1007 a 6.2 b 2.8 a 178 b 154 b 283 b 78 a

330 b 9.6 a 1.5 b 365 a 239 a 373 a 77 a

707 a 6.5 b 2.8 a 216 b 162 b 284 b 79 a

Investigated 24 h after irrigation 38 days after germination Within each row, means followed by a different letter are significantly different at P<0.05

Discussion
At almost all stages of growth, planted soil possessed a higher C availability to soil heterotrophs as indicated by a higher soil respiration rate, AMC and MBCC of planted than unplanted soil. Similarly, under denitrifying conditions, planted soil showed a higher C availability after 38 DAG as indicated by DNP results. Comparing planted (T 1) and unplanted soils for denitrification rate in the field, plants stimulated denitrification only when soil nitrate-N concentration was high. This happened at the first irrigation when plants were 18 days old. The stimulatory effect of plants on denitrification as observed at the early growth stage may be attributed mainly to the higher O2 consumption in planted than in unplanted soil. This is supported by the higher soil respiration rate in planted soils and the almost similar C availability (DNP) of the planted and unplanted soils at 18 DAG. However, when plant uptake decreased the soil nitrate-N content, planted soil (T 1) always denitrified at lower rates than the unplanted soil. The decrease in denitrification in planted soil as observed during the later stages of plant growth is in agreement with the findings of Guenzi et al. (1978) and Kovalenko and Cameron (1978), who attributed this phenomenon to competition between plant roots and denitrifiers for available nitrate-N. The positive effect of plants on denitrification could be demonstrated at all growth stages when unplanted soil was compared with planted soil that received inputs of calcium nitrate (T 3) to equalize its nitrate-N with that of the unplanted soil. The planted (T 3) soil always denitrified at higher rates than the unplanted soil. At 38 DAG and later, C availability for aerobic as well as anaerobic microbial respiration was greater in planted (T 3) than unplanted soil. Thus the stimulatory effect of plants on denitrification during 3888 DAG may partly be due to more rapid O2 depletion and higher C availability to denitrifying population in planted (T 3) than unplanted soil. These results are in agreement with those of Smith and Tiedje (1979), who reported higher denitrification in planted soil under nitrate non-limiting conditions. However, Smith and Tiedje (1979) did not find stimulation of denitrification due to plants when soil nitrate-N was kept low (2 lg g1). In the present study, however, stimulation of denitrification by plants was also recorded at low soil nitrate-N concentrations. This occurred at 38, 85 and 88 DAG when soil ni-

trate-N content ranged between 1 and 4 lg g1. The stimulation of denitrification at low NO 3-N levels is consistent with the findings of Klemedtsson et al. (1987) and may be attributed to increased C availability in the planted soil. Comparison of the unplanted and planted+nitrate (T 3) treatments also revealed that the extent of the stimulatory effect of plants on denitrification varied with soil nitrate-N concentration. At low soil nitrate-N levels (14 lg g1), the presence of plants produced a 1.4-fold increase in denitrification, whereas at higher nitrate-N levels (719 lg g1) the observed increase due to plants averaged 2.5-fold. These results are almost consistent with those of Vinther (1984), who found 2.5- and 5-fold increases in denitrification with barley plants grown in field plots receiving 30 and 150 kg N ha1, respectively. Regarding the effect of distance from the plant, the root zone soil showed a higher C availability than the distant soil. However, owing to the lower nitrate-N content, the root zone soil denitrified at a lower rate than the soil at 15 or 30 cm from the plants. Thus positive or negative effects of plants on denitrification and related parameters were confined to the root zone. These results agree with those of Smith and Tiedje (1979), who reported a higher denitrification potential in the root zone soil than in the soils collected at a distance of 15 or 30 cm from corn plants. The results of the present study suggest that plants have the potential to increase denitrification through decreased O2 in the soil and by supplying C substrate to denitrifiers. However, such a stimulatory effect of plants can only be observed when sufficient nitrate-N is present in the soil to support denitrifying populations, the situation which may exist particularly at early stages of growth when the requirement of the crop is low. During the later growth phase, the rapid uptake by crop plants leaves a little nitrate-N in the soil and thus denitrification may be substantially reduced in the presence of plants. Since distance from plant may also influence the extent of denitrification, soil should be sampled at various distances from plants while quantifying denitrification in cropped fields.
Acknowledgements This research was partly supported by Kernforschungszentrum Karlsruhe, Germany, under the PAEC-KfK joint project Denitrification Studies under Field and Controlled Conditions. The authors also acknowledge Dr. K. Haider for a critical review of the manuscript.

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