Dynamics of soil nitrate after forest fertilization as monitored by the plant nitrate reductase assay

Forest Ecology and Management, 44 ( 1991 ) 223-238

223

Elsevier Science Publishers B.V., Amsterdam

Dynamics of soil nitrate after forest fertilization as monitored by the plant nitrate reductase assay L. H6gbom and P. Htigberg Department of Forest Site Research, Swedish University of Agricultural Sciences, S-901 83 Ume& Sweden (Accepted 4 September 1990 )

ABSTRACT H6ghom, L. and H6gherg, P., 1991. Dynamics of soil nitrate after forest fertilization as monitored by the plant nitrate reductase assay. For. Ecol. Manage., 44: 223-238. Nitrate reductase activity (NRA) of the grass Deschampsiaflexuosa (L.) Trin., was used to monitor available soil nitrate after fertilization of coniferous forests in northern Sweden with ammonium nitrate or urea (300 or 150 kg N ha- t ). Small plots (9 m 2), at three sites of different quality class, were followed for 2-3 years. The poorest site is a groundwater recharge area, whereas discharges occur temporarily at the intermediate site and frequently at the richest site; these variations in transport of solutes explain the differences in productivity. Applications of ammonium nitrate gave, as expected, rapid and substantial increases in NRA. In the case of urea, slow and small increases in NRA indicated that nitrification took place. Treatment with ammonium chloride at the poorest site (pH 4.0) produced an increase in NRA similar to that given by urea application. On ammonium nitrate treated plots in discharge areas, there was a decline from peak NRA to levels near those of the controls within 2 weeks, in connection with a period of rainfall and high groundwater level. On plots treated with ammonium nitrate in the recharge area, soil nitrate remained at a high level throughout the first growing period. We suggest that leaching of nitrate may be a common phenomenon in discharge areas. We further suggest that ~SN fertilizer budgets, which have been conducted only in recharge areas, should also be carried out at sites with discharge.

INTRODUCTION

Nitrogen is the most limiting nutrient for forest production in temperate forests (e.g. Tamm, 1964). Nitrogen fertilization is, therefore, a common practice to increase forest production. Detailed investigations using 15N-labelled fertilizers have shown that 1095% of the fertilizer nitrogen can be recovered in the vegetation and the soil a few years after fertilization (e.g. Overrein, 1972; Heilman et al., 1982; Melin and N6mmik, 1988 ). The rest is thought to have been leached, denitrified, or, in the case of urea, volatilized. Elevated concentrations of nitrate are often found in forest streams after 0378-1127/91/$03.50

© 1991 Elsevier Science Publishers B.V. All rights reserved.

224

L. HOGBOM AND P. HOGBERG

nitrogen fertilization (e.g. Gonzalez and Plamondon, 1977-1978; Grip, 1982). Part of this effect is caused by fertilizer falling directly into stream channels, but leaching must also be important because fertilizer falling directly into stream channels cannot alone explain the observed changes in nitrate concentration (Grip, 1982). Denitrification rates have so far been found to be low in free-draining forest soils (Robertson and Tiedje, 1984; Goodroad and Keeney, 1984; Nohrstedt, 1988 ). Losses by leaching or denitrification could follow not only directly from application of nitrate, but also as a result of increased nitrification after nitrogen fertilization. Fertilization with urea has been shown to increase nitrification (Heilman et al., 1982; Adams and Attiwill, 1984; Popovic, 1985; H6gberg et al., 1986). Nitrification was often believed to be of little importance in acid forest soils, but recent research has clearly shown that nitrification can occur also at low pH in conifer forests (Robertson, 1982; Runge, 1983 ). Although the ~SN labelling technique has been applied only in groundwater recharge areas or lysimeter studies, studies of stream-water quality have been conducted also in broken terrain with heterogeneous soils and varying hydrology. Ideally, however, these methods should be combined to obtain a detailed picture of the fate of fertilizer nitrogen. Although highly desirable, it would be costly to apply the ~SN labelling technique on a large number of plots. Tension-free lysimeters have often been used to monitor nutrient flows in soils. The great drawback with these lysimeters in groundwater discharge areas especially is that the water flow is mainly lateral or upward in such areas

1

_L_ Fig. 1. A tension-freelysimeterin a groundwaterdischargearea. The arrows indicate flows of water and solutes.

SOIL NITRATE AFTER FOREST FERTILIZATION

225

(Eriksson, 1985). Downward leaching, which is the only possibility in tension-free lysimeters, is negligible (Fig. 1 ). Thus, the major water flow in the soil at discharge areas does not enter these lysimeters. On the other hand, tension lysimeters are fairly expensive, and, like tension-free lysimeters, cause some soil disturbance during installation and will not measure the nitrate available for plants. In this paper we report the application of the plant nitrate reductase activity (NRA) assay (H6gberg et al., 1986) as a tool for studies of the temporal variation in soil nitrate after forest fertilization. The NRA assay is influenced not only by the availability of nitrate (which includes a soil moisture component ) but also partly by temperature and light. However, these factors may be neglected when small plots under the same or similar tree canopies in a restricted area are compared (H6gberg et al., 199 lb). We were especially interested in ( 1 ) differences caused by ammonium nitrate and urea fertilizer, and (2) differences in nitrate dynamics between sites of different quality class and hydrology. MATERIALS AND METHODS

Site descriptions and weather conditions Our studies were made within the Kulb~icksliden and Svartberget Experimental Forests, 60 km NW of U m e L northern Sweden. The bedrock is dominated by veined gneisses. All sites are under the highest coastline (altitude 250-260 m ). The mean annual rainfall is 530 mm, and the mean annual temperature is 1.1°C. The wet deposition of nitrate is generally less than 2 kg NO3-N h a - l year- l, and usually comprises about a third of the total nitrogen deposition (Degermark, 1987, 1988, 1989). Our experiments were conducted at three sites, described below in order of increasing site quality class. The Aheden site (64 ° 13'N, 19 ° 30' E) is situated at 175 m altitude on level and thick delta/lacustrine sediments, which form a recharge area for groundwater (Table 1 ). The soil textural class is coarse silt, and the soil is an Orthic Podzol (FAO-UNESCO, 1974). The field layer is dominated by Faccinium spp. (mainly V. vitis-idaea) and Deschampsia flexuosa. (Nomenclature of vascular species follows Turin et al. ( 1964-1980). ) The tree layer is dominated by Pinus sylvestris (Table 2 ). The Renberget site (64°15'N, 19 ° 49'E) is located on a gentle slope on wave-washed gravelly sand, at 255 m altitude just below the highest coastline (Table 1 ). This is a discharge area for groundwater for fairly short periods, as indicated by patches of Sphagnum spp. moss (outside the plots). The soil is an Orthic Podzol. The field layer is dominated by V. myrtillus, but this site has a richer herbaceous flora than/~heden. The tree layer is totally dominated by Picea abies (Table 2 ).

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L. HOGBOM AND P. HOGBERG

TABLE I Soils of the sites studied Soil properly

Site

Soil type Thickness of Ao horizon (cm) Thickness of A2 horizon (cm)

Aheden

Renberget

Kulb~icksliden

Orthic Podzol 3-5 2-5

Orthic Podzol 8-11 12-13

Humic Podzol 5-9 7-9

0.0 0.0 31.3 65.0 4.0

15.1 19.9 56.7 7.5 1.5

6.5 11.7 44.1 34.5 3.3

Mechanical composition (%) of C horizon Textural class ( m m ) Stone > 20 Gravel 20-2 Sand 2-0.06 Silt 0.06-0.002 Clay <0.002

Chemical properties of Ao horizon pH(H20) NO3-N (mg g- J dry mass) NH4-N (mgg - l dry mass) Total N (% dry mass) C/N

4.1 0.0 a 0.09 1.16 36.2

(0.04) a (0.01)a (0.05) a (0.99) ~

4.5 0.0 a 0.09 1.23 36.7

(0.10) b

(0.01)" (0.05) a (1.03) a

4.9 0.0 a 0.11 1.42 28.3

(0.11) c (0.01)~ (0.07) b (0.86) b

Figures not followed by the same letter (row by row) are not significantly different at the P < 0 . 0 5 level. Figures in parenthesis are standard errors.

TABLE 2 Properties of the stands studied ( 1988 ) Stand property

Dominant tree species Site index (H~oo) a Basal area (m 2 ha -~ ) Stand age (year) Stand height ( m ) b Volume (m s ha-~ )

Site ~,heden

Renberget

Kulb~icksliden

Pinus sylvestris

Picea abies

Picea abies

T19 18 75 16 220

G22 26 140 22 350

G24 14 150 22 320

aBased on site properties (H~igglund and Lundmark, 1977 ). bHeight of tree of mean basal area.

228

L. HOGBOMAND P. HOGBERG

The Kulb~icksliden site (64 ° 11 'N, 19°38'E) is situated at 225 m altitude on a moderate slope on wave-washed sand, just below the bare rocks of the highest coastline (Table 1 ). Outflows of groundwater occur for long periods, as indicated by the dominance of Sphagnum spp. and liverworts (hepatics) in the bottom layer. Hardpan formation is frequent and provides further evidence of substantial lateral groundwater movement on the plots. The soil is a Humic Podzol. There is a ditch below two of the blocks but above the third; this ditch should not affect the groundwater movement on the plots (compare with the pit in Fig. 1 ). The field layer consists of a diverse flora, among which Lactuca alpina and Equisetum silvestris dominate. The tree layer is dominated by Picea abies (Table 2 ). 1987

1988

1.0/

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Fig. 3. NRA on plots fertilized in early summer 1987. (a) Aheden (Experiments 2 and 3); (b) Renberget (Experiment 2); (c) Kulbiicksliden (Experiment 2). Each symbol is the mean of three samples (except before treatments when n = 9-12): O, control; V, 150 kg N as AN ha- ~; A, 150 kg N as U ha- ]; C2, 150 kg N as AC ha- ~. In the lower part of the figure, solid bars indicate daily precipitation and the continuous line daily mean temperatures. Small arrows indicate minimum temperatures below zero, and large arrows show when fertilizers were applied.

SOIL NITRATE AFTER FOREST FERTILIZATION

229

Climatic data are recorded regularly at the Svartberget Research Station, which is within 2 km of Renberget and/~heden, and 12 km from Kulb~icksliden. Late summer and early autumn in 1986 were cooler and wetter than usual. The growing period in 1987 was also cool and wet, with a temperature sum of 575 (threshold value + 5 ° C ) and a rainfall of 350 mm, whereas the temperature and precipitation in 1988 were closer to the average, with a temperature sum of 950 and a rainfall of 310 mm. The groundwater level was very high throughout summer and autumn in 1987. Details of ambient weather conditions can be found in Figs. 2 and 3 and in reports from the research station, which also include observations on hydrological conditions and atmospheric deposition (Degermark, 1987, 1988, 1989).

Experiments The experiments had a randomized block design with three blocks less than 50 m apart at each site. Plots were quadratic, with an area of 9 m 2 (3 m × 3 m), and there was a buffer strip of 1 m between plots. Each treatment was applied to one plot per block. Experiment 1 was conducted at Aheden. Fertilization was carried out in early August 1986 using two doses of nitrogen fertilizer (150 and 300 kg N h a - l ) , in two forms, ammonium nitrate (AN) and urea (U) as granules. Control (C) plots were left untreated. On 25 September 1986, a test of the maximal induced NRA was made to indicate the level of NRA when the availability of nitrate was not limiting. Five 0.25 m 2 plots outside the blocks were fertilized with 1000 kg of NO3-N (KNO3) in solution 3 days before sampling, and sampled at the same time as a regular sampling for the experiment (see H6gberg et al., 1991 ). Experiment 2 was carried out at all three sites. At/~heden, the plots were interspersed within the blocks used in Experiment 1, and the control plots were thus the same for the two experiments on this site. Fertilizer was applied in early June 1987. One dose of nitrogen (150 kg N ha -1 ) as AN or U was used. Experiment 3 was started at/kheden at the same time as Experiment 2, and used the same blocks and control plots as Experiments 1 and 2. Plots were treated with ammonium chloride (AC, 150 kg of N h a - l ) , which is not a conventional fertilizer. This experiment was done to provide a comparison with the U treatment, i.e. to test the application of ammonium without enhancing soil pH.

Analysis of NRA Leaves of Deschampsiaflexuosa were sampled for NRA assays during periods without snow cover from before treatments to mid-September 1988.

230

L. HOGBOM AND P. HOGBERG

Samples were collected between 0900 and 1200 h, and analysed within 3 h for in vivo NRA (H6gberg et al., 1986). Approximately 0.5-1.0 g (fresh mass) of leaf tissue was taken from each plot at each sampling.

Soil analyses Soil samples for chemical analyses were taken from the Ao horizon with a soil auger of 10 cm width. Five samples were taken around each block, i.e. 15 samples per site. No sampling was made on plots. The pH was measured in a suspension of soil in deionized water (the volume ratio of soil: water was 1 : 3 ). NO~- and NH + were extracted with 2 N KC1 for l h. The filtrates were analysed largely according to Henriksen and Selmer-Olsen (1970) and Koroleff (1976), respectively. Carbon and total nitrogen were determined on a Perkin-Elmer CHN-analyser, which determines C as CO2 after combustion and N as N2 after combustion and subsequent reduction on a thermal conductivity detector.

Statistical analysis The statistical analysis of the material was performed by using one-way ANOVA, one for each sampling date. In case of significance, Tukey's range test was applied. Two-way ANOVA, with time as one factor, could not be used as the NRA values are not independent on single plots over time. In addition, time and treatment as variables interact strongly. RESULTS

General The seasonal pattern in NRA was the same on all three sites: NRA was highest in the spring and early summer, and low in the summer, but sometimes also showed an increase in the autumn. There was very little difference between sites, the blocks at a site and the plots within a block before treatment.

Experiment 1 NRA increased rapidly after fertilization with AN (Fig. 2). Both dosages of AN gave significantly higher NRA than the control in the first year, but,in the second year only the higher dose gave higher NRA throughout the whole season (Table 3). The increases in NRA after U treatment were small. The higher dose gave a significantly higher NRA than the control occasionally in all 3 years, whereas

231

SOIL NITRATE AFTER FOREST FERTILIZATION T ABLE 3 Statistical a n a l y s i s o f E x p e r i m e n t 1 (one-way A N O V A for every s a m p l i n g d a t e ) Date

F

P

Experiment 1:1986/~heden 1 231.9 0.0001 2 234.2 0.0001 3 381.0 0.0001 4 128.5 0.0001 5 341.5 0.0001 6 214.3 0.0001

Test result

300AN ~ 300AN a 300AN a 300AN a 300AN a 300AN ~

150AN b 150AN b 150AN a 150AN a 150AN a 150AN a

300U c 300U c 300U b 300U b 300U b 300U b

150U c 150U ¢ 150U b 150U b 150U b 150U bc

C~ Cc Cb Cb Cc Cc

Experiment 1:1987 ~,heden 1 2 3 4 5 6 7 8 9 10 11 12 Experiment 1 2 3 4

2.3 8.1 5.5 2.4 11.4 6.3 10.3 7.1 1.0 4.6 3.4 1.8

0.1184 0.0017 0.0080 0.0996 0.0007 0.0048 0.0005 0.0030 0.4419 0.0155 0.0420 0.1958

...................................................... 300AN b 150AN ab 300U a 150U b Cb 300AN a 150AN ab 300U ab 150U b Cb ......................................................... 300AN a 150AN ab 300U t~ 150U t~ C t~ 300AN a 150AN a 300U ~ 150U b Cb 300AN a 150AN ab 300U be 150U t~ Cc 300AN a 1 5 0 A N ab 300U b 150U b Cb .............................................................. 300AN a 150AN ab 300U ab 150U b Cb 300AN ~ 150AN ab 300U "b 150U ab Cb ................................................................

1:1988/~heden 1.3 3.3 0.4 3.3

0.3145 0.0470 0.8041 0.518

3 0 0 A N "b

150AN ~b

300U"

150U ~b

Cb

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

In case of significance, T u k e y ' s test was used. T r e a t m e n t s not followed by the s a m e letter (row by r o w ) are s i g n i f i c a n t l y different ( P < 0.05 ). C, control.

the lower dose gave higher NRA only once in the first year. AN gave higher NRA than U during the first year irrespective of dosage of nitrogen. The m a x i m u m induced activity on non-treated plots outside the blocks at the end of September was 1.4_+ 0.1 #mol NO~- g - l h - l as compared with 0.1 + 0.0/~mol NO~- g- l h - l on control plots.

Experiment 2 Aheden The NRA was higher on plots treated with AN than on control plots during the first year only (Table 4 and Fig. 3 ). The U treatment had no effect on the NRA compared with the control during the first year. Fertilization with AN also gave higher NRA than the U treatment during the whole of the first season.

232

L. HOGBOM AND P. HOGBERG

TABLE4 Statistical analysis of Experiments 2 and 3 (one-way ANOVA for every sampling date) Date

F

P

Test r e s u l t

Experiments 2 and 3:1987 ,~heden 1 2 3 4 5 6

11.1 12.9 10.1 26.6 13.3 42.5

0.0012 0.0006 0.0023 0.0001 0.0006 0.0001

7

16.4

0.0002

AN"

8 9

10.5 8.3

0.0015 0.0036

AN" AN"

10 1.5 0.2645 E x p e r i m e n t s 2 and 3:1988 ~heden 1 1.0 0.4343 2 4.3 0.0302 3 1.4 4 0.7 E x p e r i m e n t 2:1987 1 24.0 2 20.5 3 11.4 4 8.8 5 6 7 8 9

26.0 1.8 1.1 6.5 8.8

0.2881 0.5837

AN" AN" AN a AN a AN a AN"

A C ab AC b AC b AC b AC b AC b

Ub Ub Ub Ub Ub Ub

Cb Cb Cb Cb Cb Cb

AC °

U°

Cb

AC ° A C ab

U° Ub

C° Cb

......................................... ......................................... A N ab AC" U ~° Cb ......................................... .........................................

Renberget 0.001 0.002 0.009 0.016

AN a AN a AN a AN"

Ub Ub Ub

C° C° C°

0.001 0.246 0.399 0.032 0.016

AN ~ U° C° .............................. .............................. A N a° Ua Cb AN" U" Cb

U ab

C0

Experiment 2:1988 Renberget 1 3.6 2 1.0 3 6.1 4 3.1 5 4.3 E x p e r i m e n t 2:1987 1 19.2 2 61.7 3 6.1 4 4.5 5 18.6 6 7 8 9

1.2 4.0 0.8 9.4

0.096 0.418 0.035 0.120 0.081

.............................. .............................. AN" U" Ca .............................. ..............................

Kulbiieksliden 0.002 0.000 0.036 0.063 0.005

AN a Ub Cb AN a Ub Cb AN a U "b Cb .............................. AN ~ Ub Cc

0.367 0.078 0.494 0.014

.............................. .............................. .............................. A N ab Ua Cb

Experiment 2:1988 Kulbficksliden 1 2 3 4

0.9 1.2 0.6 1.2

0.468 0.356 0.570 0.365

In c a s e o f s i g n i f i c a n c e , T u k e y ' s t e s t w a s u s e d . T r e a t m e n t s r o w ) a r e s i g n i f i c a n t l y d i f f e r e n t ( P < 0 . 0 5 ). C, c o n t r o l .

not followed by the same letter (row by

SOIL NITRATE AFTER FOREST FERTILIZATION

233

Renberget A rapid increase in NRA after the AN treatment was followed by a rapid decrease down to levels only slightly above the control (Table 4 and Fig. 3 ), from which it did not differ in the second season. The U treatment produced a small increase in NRA only at the end of the first year. AN gave higher NRA than U in the first year after fertilization.

Kulbiicksliden The results at this site were similar to those at Renberget (Table 4 and Fig. 3 ). After the rapid initial increase in NRA after AN treatment, there was a rapid decline. The U treatment also gave higher NRA only at the end of the first year. AN gave higher NRA than U only directly after fertilization.

Experiment 3 Both the AC and U treatments gave a slightly but not significantly higher NRA than on control plots (Table 4 and Fig. 3). There was no difference between the AC and U treatments. DISCUSSION

General The plots studied could in some respects be considered as small. Trees and other plants outside fertilized plots could have responded in a compensatory manner with enhanced root growth and nitrate uptake on the plots (see Drew and Saker, 1975 ). This would have led to a larger drain of nitrogen from the plots than would be the case if these were localized in a fertilized stand. However, changes in the field layer vegetation, e.g. at/~heden, were abrupt at the boundary of fertilized plots. Theoretically, the differences encountered shouid, nevertheless, be considered as conservative estimates. The use of small plots, on the other hand, allows monitoring of nitrate losses from discharge areas on slopes. The seasonal variation in NRA, i.e. high NRA in spring and autumn, is partly a result of changes in available soil nitrate, and partly an effect of ambient temperature on the level of nitrate reductase (HSgberg et al., 1991 ). More enzyme is induced at lower than at higher temperatures. For example, the sudden increases in NRA at the end of June 1987 are simultaneous with a decrease in temperature (Figs. 2 and 3 ). High levels of nitrate in the soil in the spring and a u t u m n are the result of production in excess of uptake and immobilization. Stream water in the study area has its highest concentrations o f nitrate during winter and spring (Degermark, 1987, 1988, 1989 ).

234

L. HOGBOM AND P. HOGBERG

Influence of form and dosage of nitrogen Application of a fertilizer containing nitrate, i.e. AN, gave rapid increases in NRA, as expected (Figs. 2 and 3 ). The rate of the subsequent decline varied between sites (see below). In the case of U, there were small increases in NRA, indicating an increase in nitrate availability depending on the rate of nitrification. Hence, both forms of fertilizer gave temporary pulses of nitrate through the soil system, but the form and duration of the pulses varied. This will influence the patterns of leaching. Grip ( 1982 ) found that the total loss of nitrate, as measured in forest streams over 3 years, was larger after fertilization with U than with AN, although there was a large initial loss after AN fertilization. The relation between nitrate supply and NRA follows a saturation curve (H6gberg et al., 1986 ). This explains why there was no particular difference between the two doses of AN applied at/l, heden (Fig. 2 ), which gave an NRA close to the maximal induced NRA at that time of the year at that site.

Recharge and discharge areas The soils at all sites are derived from gneisses, and the composition of the underlying bedrock cannot account for the differences in soil chemical properties, such as pH (Table I ). These differences can only be explained by variations in water flow. Outflows of base-rich groundwater, for example, can explain the relatively high pH of the mor at the two better sites. This mass flow also includes nitrogen compounds. Also, higher base saturation, particularly a high concentration of calcium, which is thought to promote nitrogen turnover, is correlated with a high total concentration of nitrogen in the mor of Nordic boreal forests (Dahl et al., 1967). There is a strong correlation between the concentrations of calcium and nitrogen and the vegetation of the field layer and forest productivity (Dahl et al., 1967; Lahti and V~iis~inen, 1987 ). In the Swedish site index classification system, which is based on empirical data, the hydrology (length of slopes) and field layer vegetation are important components (H/igglund and Lundmark, 1977; see also H6gberg et al., 1990). Our data from soil chemical analyses showed that pH was significantly higher on sites with groundwater discharge (Table 1 ). The nitrogen concentration and C / N ratio of the mor was also highest and lowest, respectively, at Kulb~icksliden. There were, however, no differences in the concentrations of extractable ammonium and nitrate, which demonstrates the difficulties in accounting for the supposed differences in the fluxes of these ions. The rapid decline in NRA after the first positive response to AN fertilization at Renberget and Kulb~icksliden was striking. The rate of the decrease was close to the half-life of nitrate reductase in D. flexuosa (Lee and Stewart,

SOIL NITRATE AFTER FOREST FERTILIZATION

235

1978 ), which may suggest a total disappearance of the substrate. Such a rapid decrease could, theoretically, be caused by a repression of NRA by ammonium or its assimilation products; a phenomenon found in some higher plants (e.g. Srivastava, 1980) and in Sphagnum moss (Woodin and Lee, 1987). However, two facts suggest that this is not the case. First, the nitrogen concentration of D. flexuosa on the control plots varied only between 1.4 and 1.7% total N between the sites. Second, Hrgberg et al. (1986) found highly elevated NRA in populations of D. flexuosa that had been fertilized with high dosages of AN and U for 15 consecutive years. It also seems highly unlikely that plant uptake, microbial immobilization and denitrification could remove the 75 kg N h a - l applied directly as nitrate from the available pool in about 2 weeks. It should be stressed in this context that the NRA method is particularly sensitive to small amounts of nitrate (H/Sgberg et al., 1986 ). We can only explain the major part of this decline as a loss of nitrate from the plots during the period of massive outflows of groundwater, which coincidentally followed fertilization. Unsaturated topsoils with prevailing vertical transport of water are found immediately below the plots. The nitrate lost should reappear in discharge areas downslope (see Eriksson, 1985 ). Grip ( 1982 ) first found elevated concentrations of nitrate directly in connection with AN fertilization, of which some had fallen in the stream channels, and then again a month later during a period of high rainfall and runoff. Heilman et al. (1982) also found a significant loss of nitrogen under conditions of heavy rainfall shortly after fertilization. Grip ( 1982 ) calculated that the leakage took place within a zone of a few metres along stream channels, provided that all the nitrate in that zone was lost. Our results suggest that leakage may be a more widespread phenomenon in discharge areas. That nitrate is retained for longer periods in recharge areas is shown not only by data from Aheden (Figs. 2 and 3), but also by other observations in the field (HSgberg et al., 1986, unpublished data, 1991 ). The question arises, however, as to whether the sites chosen are representative. The soils at Renberget in particular, and at Kulb~icksliden, are fairly coarse textured (Table 1 ). Water flows through the more typical fine-textured till soils are slower. Hence, leakage should also be slower. Detailed studies of retention of fertilizer nitrogen using 15N labelling have, as far as we know, been made only in recharge areas or in lysimeters which exclude lateral flows (Overrein, 1972; Melin et al., 1983; Melin and NiSmmik, 1988). Rodhe ( 1987 ) estimated the extent of discharge areas in l 0 forested basins in Sweden ranging from 3 to 660 ha in size. During snowmelt and rainfall events, the discharge areas were estimated at 2-60% and 0.2-17%, respectively (with median values of 22% and 3%, respectively). However, individual stands likely to be selected for fertilization, e.g. Renberget, may comprise a large fraction of discharge areas. In Sweden it is advised to avoid using fertilizer close to stream channels and on soils with coarse texture, but the point raised here,

236

L. HOGBOM AND P. HOGBERG

i.e. that perhaps discharge areas in general should be avoided, has not been considered. This hypothesis should be tested in ~SN studies.

Nitrification Nitrification increased as compared with the control after fertilization with U, irrespective of the initial soil pH (Figs. 2 and 3 ). Nitrification in acid soils has often been said to be dependent on microsites with favourable pH (e.g. Runge, 1983); for example, around U granules. This need not be so, as indicated by the increased nitrification after application of AC, which instead acidifies the soil solution through cation exchange. Thus, our results add some support to the statement that the correlation between pH and relative nitrifi -~ cation is poor (e.g. Robertson, 1982; Runge, 1983). We find it particularly interesting that our results are obtained from measurements of undisturbed (with the exception of the a m m o n i u m addition ) soil studied in situ. ACKNOWLEDGEMENTS This project was funded by The Swedish Forestry Research Foundation. We would like to thank Professor N. Nykvist, and Drs. H. Grip. H.-O. Nohrstedt, H. Ivarsson and S. Holm, for helpful suggestions, a Westerbergh for field and laboratory assistance, E. Sinclair for laboratory assistance and Nigel Rollison for linguistic revision.

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