nitrogen transformations in two nitrogen saturated forest ecosystems subjected to an experimental...

Plant and Soil 175: 205-218, 1995. 205 @ 1995 Kluwer Academic Publishers. Printed in the Netherlands.

Nitrogen transformations in two nitrogen saturated forest ecosystems subjected to an experimental decrease in nitrogen deposition

C.J. Koopmans, W.C. Lubrecht and A. Tieterna Department of Physical Geography and Soil Science, University of Amsterdam, Nieuwe Prinsengracht 130, 1018 VZ Amsterdam, The Netherlands*

Received 16 December 1994• Accepted in revised form 22 March 1995

Key words: atmospheric deposition, forest, incubation, mineralization, nitrification, nitrogen

Abstract

Nitrogen transformations were studied in the forest floor and mineral soil (0-5 cm) of a Douglas fir forest (Pseudot- suga menziesii (Mirb.) Franco.) and a Scots pine forest (Pinus sylvestris L.) in the Netherlands. Current nitrogen depositions (40 and 56 kg N ha-1 yr-I , respectively) were reduced to natural background levels (1-2 kg N ha-1 yr- ~) by a roof construction. The study concentrated on rates and dynamic properties of nitrogen transformations and their link with the leaching pattern and nitrogen uptake of the vegetation under high and reduced nitrogen deposition levels. Results of an in situ field incubation experiment and laboratory incubations were compared. No effect of the reduced N deposition on nitrogen transformations was found in the Douglas fir forest. In the Scots pine forest, however, during some periods of the year nitrogen transformations were significantly decreased under the low nitrogen deposition level. At low nitrogen inputs a net immobilization occurred during most of the year leading to a very small net mineralization for the whole year. In laboratory and in individual field plots nitrogen transformations were negatively correlated with initial inorganic nitrogen concentrations. Nitrogen budget estimates showed that nitrogen transformations were probably underestimated by the in situ incubation technique. Nevertheless less nitrogen was available for plant uptake and leaching at the low deposition plots.

Introduction

For many years the effects of increased N input via atmospheric deposition in forest ecosystems have been a matter of research (Van Breemen et al., 1982) and discussion (Abet et al., 1989; Van Breemen et al., 1984). The effects, such as large scale forest decline and pollution of groundwater reserves with nitrate, are a serious problem in many European countries and in north America.

Most studies investigate the effects of increased N input (Aber et al., 1989; Dise and Wright, 1992). This question, however, seems to be too limited for the Netherlands, as in many sites in this country increased nitrate leaching to the groundwater does occur already (Van Breemen and Verstraten, 1991). Consequently we investigated whether nitrogen saturation and its effects on ecosystem functioning are reversible as a

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result of decreased nitrogen input, in order to support the discussion of possible abatement policies. To this end, a field-scale manipulation project was started in 1988 in two coniferous forest ecosystems (Van Dijk et al., 1992). Nitrogen and sulphur inputs were reduced to natural background levels by a roof construction and the effects of this decreased atmospheric input on ecosystem functioning was studied. This project was part of the EU project NITREX, a consortion of comparable field manipulation experiments with N deposition in coniferous forest ecosystems. Within three months after the start of these experiments, soil solution mon- itoring at both sites has shown a drastic decrease in NH4 and NO3 concentrations due to the removal of N and S in throughfall water (Boxman et al., 1995). Beside the N deposition level, the cause of this quick response may include changes in relative N source-sink strengths in the soil. Net mineralization is the main process responsible for the availability of nitrogen in the

206

soil. Net nitrification is seen as an important process contributing to soil acidification and nitrogen losses due to leaching and possibly to gaseous emissions of nitrogen.

In our study we concentrated on the nitrogen transformation processes in the upper soil layers, in the fourth year after the nitrogen deposition was reduced. The study aimed at gathering information about the rates and dynamic properties of the transformations in the different plots in two coniferous ecosystems, Speuld and Ysselsteyn. Various incubation techniques in the field and in the laboratory were used, as the measured N transformation rates are affected by the method used (Adams et al., 1989; Ht~bner et al., 1991; Tietema et al., 1992). The results of these different methods were compared and the possible artefacts dis- cussed.

Materials and methods

Site description

This study was carried out at two sites (Table 1) in the Netherlands with relatively high N and S deposition levels. The Speuld site is situated in the central part of the Netherlands (52°13'N, 5 ° 39'E). It is a plantation of Douglas fir (Pseudotsuga menziesii (Mirb.) Fran- co.). The well-drained loamy sand soil was classified as an Haplic Podzol (FAO/UNESCO, 1988). Nitrogen deposition (Table 2) of inorganic N in throughfall water during the last 4 years was 40 kg ha- 1 yr- 1, mainly in the form of NH4-N (Boxman et al., 1995). The Yssel- steyn site is located in the south eastern part of the Netherlands (51 ° 30'N, 5 ° 55'E). The site is dominated by 45-year old Scots pine (Pinus sylvestris L.). The soil, classified as an Haplic Podzol (FAO/UNESCO, 1988) has a relatively organic rich mineral top layer (50 cm) as a result of ploughing before tree plan- ting. N deposition in throughfall amounted up to 56 kg ha-l y r - l (Table 2) also mainly in the form of NH4-N (Boxman et al., 1995). The climate in the Netherlands is temperate with a mean annual air temperature of 9-9.5 °C and a mean annual precipitation of approxi- mately 750 mm.

Experimental set up

During the winter of 1988-1989 transparent roofs, co- vering 14 × 26 meter, were built underneath the canopy at the two sites. Two (10 × 10 m) plots were established

underneath the roof, leaving a bufferzone of 2 m around each plot.

In the first plot, the low deposition plot, N and S depositions were reduced to natural background levels. The second plot under the roof, the high deposition plot, received the unaltered N and S deposition. A third plot, the ambient plot, was established outside the roof. The effects of the reduction in N and S deposition can be studied by comparing the high and the low deposition plot. By comparing the high deposition plot and the ambient plot the effect of the roof, the watering regime and other environmental influences of the roof on the ecosystem processes (Gundersen et al., 1994) can be investigated.

The water is applied to the plots under the roof by means of an automated sprinkling system with a real time watering regime. The low deposition plot receives artificial throughfall water. Sea salts were added to deionized water to reach concentrations comparable with throughfall water. Nitrogen and S are excluded, leading to a strongly reduced deposition level (1-2 kg ha-l yr-1). The throughfall water intercepted by the roof is sprinkled to the high deposition plot under the roof. The ambient plot outside the roof, receives unaltered throughfall water.

Field incubations

After four years of N manipulations in the field, net nitrification and net mineralization rates were measured using an in situ incubation technique. At both sites for a one-year period (May 1992 - May 1993) a sequential in situ incubation technique was used with intact soil cores (Adams and Attiwil, 1986; Raison et al., 1987; Tietema et al., 1992). The individual incubation periods lasted six weeks, which was considered to be a compromise between the minimum time neces- sary to measure differences in ammonium and nitrate concentrations and a maximum period in which artificial conditions (e.g. soil moisture content) within the soil cores would influence microbial activity. In each plot two intact soil cores were taken within 10 cm from each other in 5 replicates. PVC columns of 25 cm height and 7 cm internal diameter were used. Aera- tion was possible through small holes in the top of the columns (diameter= 13 ram). One column of each pair was taken directly to the laboratory for initial NH4- N and NO3-N determinations. The other column was closed on top and bottom, and placed back in the hole for a period of 6 weeks. After this period these samples too were analyzed for inorganic N. After storage

Table 1. Vegetation and soil characteristics of the Speuld and Ysselsteyn site

Speuld Ysselsteyn

Tree Douglas fir Scots Pine Pseudotsuga menziesii Pinus sylvestris

Tree age 35 45

Stem density (stems ha -1 ) 800 650

Soil type Haplic Podzol Haplic Podzol

Bulk density (g cm -3) 1.01 (0-5) cm Ah) 0.95 (0-5 cm Ah + Ae)

pH(IMKCX) LF: 2.86 Ah: 3.17 LF: 2.91 Ah+Ae: 3.04

% N L: 2.13 F: 2.02 Ah: 0.15 L: 1.75 F: 2.3 Ah+Ae: 0.2

C/N (by weight) LF: 21 Ah: 16 LF: 19 Ah: 10

207

Table 2. Nitrogen cycling fluxes for the Speuld and Ysselsteyn site. Litter fluxes are averages from the period 1990-1992. Nitrogen fluxes in the soil from Ysselsteyn are averages from the period 1990-1992, for Speuld from 1991-1992. (Partly adapted from Boxman et al, 1995)

Speuld Ysselsteyn

Plot Low High Ambient Low High Ambient

Litter production (kg dry matter ha - t yr - l )

2021 2699 2699 3470 3465 3793

N flux in litter (kg ha - I y r - l ) 26 34 38 43 49 56

Yhroughfall flux (kg h a - J)

NH4-N 0.7 26.2 29.7

NO3-N 0.4 8.3 10.5

Total 1.1 34.5 40.2

0.7 29.0 44.1

0.3 6.2 11.1

1.0 35.2 55.5

Leaching l0 cm depth (kg ha -1 )

NH4-N 1.4 2. l 12,6

NO3-N 4.2 15.8 65.6

Total 5.6 17.9 78.2

3.5 13.8 20.8

16.6 33.1 45.7

21.1 46.9 66.5

Leaching 90 cm depth (kg ha - l )

NH4-N 0 0 0.9

NO3-N 0 7.9 84.0

Total 0 7.9 84.9

1.4 0 1.4

10.8 33.1 63.8

12.2 33.1 65.2

at 2 °C for maximally 48 hours the soil cores were split into the forest floor and the upper 5 cm of the mineral soil. Roots (> 1 mm) were removed from the samples. The fresh samples were extracted with 1 M KC1 in a 1:20 and 1:10 extraction ratio for the forest floor and mineral soil respectively. After pHKcb measurement the extract was filtered (0.45 ,urn), and NH4-N and NO3-N concentrations were measured on a continuous flow autoanalyzer.

The N transformation rates were calculated as the difference in concentrations between the two paired soil cores. The difference in nitrate concentrations was used to calculate the net nitrification rate, while the difference in total inorganic N was used to determine the net N mineralization rate. To convert the transformation rates in mg N k g - 1 6 w e e k s - l to an areal estimate, the mean dry weights of each layer in the cores at the various sites were used.

208

percolation water

soil column

glass- I filter funnel pressure

sample ~ ~ vial data logger

Fig. 1.

valve

Percolation system. See text for more details.

Laboratory incubations

In December 1993, 96 intact soil cores (diameter=-5 cm, height=10 cm) were taken from the plots under the roof of the Ysselsteyn site to determine rates of net mineralization and net nitrification with three different incubation methods in the laboratory.

The first method involved the incubation of homogenized soil samples in petri dishes. Eight pairs of soil cores from each plot were separated in the organic and upper 5 cm of the mineral horizon and homogenized and pooled per plot. Initial NH4-N and NO3-N concentrations were determined in 1 M KCI extracts of four replicates taken from the pooled sample. Another four samples of the organic and mineral soil were incubated for 6 weeks at 12°C followed by the analysis of the final NH4-N and NO3-N concentrations.

The second method involved the incubation of intact soil cores in the laboratory. Eight pairs of soil cores from each plot were used. Initial inorganic N concentrations were determined, separately for the organic and mineral soil, in one core of each pair. The other soil core was incubated at 12 °C, at field moisture content, for 6 weeks.

The third incubation method involved a percolation system (Fig. 1). Intact soil cores were placed on top of glassfilter-funnels (1.6/tm pores). Each funnel was connected to a system keeping the soil core at a pressure of - 1 5 0 cm (pF 2.18) through a small pump connected to a data logger. This prevented anaerobic conditions and the system allowed drainage to a sample vial. Soil cores were incubated on this system at 12 °C

for six weeks. Three times a week the soil cores were watered, corresponding to 5 mm day- I. Previously the system was tested for denitrification, measuring rates of N20 emissions from soil cores placed in a closed chamber immediately after water addition. N20 emis- sion were always less than 2% of the total amount of nitrogen nitrified and was therefore assumed to be neglectable during the time of incubation. Except for N and S which was applied at different levels, an artificial throughfall solution comparable to the throughfall water in the field was added. Eight replicate soil cores from the low deposition plot were watered with a no-N rain solution (no N addition). Eight replicate samples from the high deposition plot also received the no-N rain solution, whereas eight other replicate soil cores from this plot received a high-N rain solution, containing (NH4)2SO4 at a concentration comparable to that in ambient throughfall water (710/~mol N L-~), Leachates were pooled for one week and analyzed for the inorganic nitrogen species. In addition after six weeks NH4-N and NO3-N concentrations in the soil cores were determined in 1 M KC1 extracts.

Statistical analysis

The lack of plot replication allows to test only between plots and not between treatments. Samples from each plot were considered as replications. Differences between plots and sampling dates of net nitrification, net mineralization and inorganic N (NH4 and NO3) concentrations in the field and the percolation experiment were evaluated with a repeated measurement ANOVA of the GLM (General Linear Model) proce- dure of the SAS statistical package (SAS, 1988). All other differences were tested through a student t-test. Regulating factors were investigated by multiple linear regression analysis using the regression (REG) proce- dure of SAS. In this study effects with probabilities of p < 0.05 were assumed to be significant unless other probabilities are indicated.

Results

The Douglas fir forest in Speuld

In situ field incubations Ammonium concentrations, measured in KC1 extracts of the forest floor, were lower in the low deposition plot compared to the high deposition plot during almost

209

SPEULD

700

600 - A -+- LOW AMMONWM

-O- High LF 500 -- T cl J

’ ‘A1 ’ Ambient

P z 2

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time (date)

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AMMONIUM

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Urn. (da)

6 NITRATE

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urns ("ate)

6,

D NITRATE

6 Ah T

1 l/5/92 22/I 51/7 11/O 25/10 4112 14/l/93 26/2 1 Z/4

time (date)

Fig. 2. Initial concentrations of ammonium (A) and nitrate (B) in the forest floor and O-5 cm of the mineral soil (C, D) for the three plots at the Speuld site. Error bars (n=5) and significant differences between the low and high deposition plot (T = treatment effect) and/or between the high deposition plot and the ambient plot (R = roof effect) at p= 0.05 are indicated.

the whole year (Fig. 2). This was more pronounced in the mineral soil were NHb-N levels for the high deposition plot amounted to a maximum value of 26 mgkg-‘, whereas NHs-N levels for the low deposition plot were always below 5 mg kg- ‘. The low deposition plot tended also towards lower nitrate levels compared to the other plots, especially in the mineral soil (Fig. 2).

The net nitrification rate of the forest floor at the Speuld site was highest in spring and autumn (Fig. 3). The net nitrification did not differ between the plots and treatments, except for two periods with small differences between the high deposition plot and the ambient plot, probably due to a roof effect. A roof effect for the nitrification rate occurred also in the mineral soil in spring and summer. Nitrogen transformation rates were calculated for the whale year period (Table 3). Differences in the nitrification rate between the low and high deposition plot were small. However, a strong roof effect was observed for the net nitrification rate in the mineral soil. In the forest floor of Speuld almost no

differences in the net mineralization rate between the plots were found for the whole year (Fig. 3). In the mineral soil net mineralization rates in the low deposition plot were somewhat less than in the high deposition plot, resulting in a small decrease in total net mineralization on a whole year basis under decreased N deposition (Table 3).

The Scats pine forest in Ysselsteyn

In situfield incubations The Ysselsteyn site showed relatively larger variations in inorganic nitrogen levels and nitrogen transformation rates within the plots and during the year (Fig. 4 and 5) than the Speuld site (Fig. 2 and 3). In the high deposition plot, NH4-N levels were significantly higher compared to the low deposition plot, especially in spring 1992. In this period also a strong roof effect was observed. In the mineral soil average NH4-N levels of the low deposition plot (17 mg kg-‘) were significantly lower than in the high deposition plot (32 mg kg-‘).

210

Table 3. Nitrogen transformation rates (kg N ha - l yr -1) at the Speuld and Ysselsteyn site• Values are means calculated from the in situ field incubation experiment with standard deviations between parentheses. LF - forest floor, M - top 5 cm of the mineral soil

Horizon Net mineralization Net nitrification

Low High Ambient Low High Ambient

Speuld LF 28.6 (7.4) 32.6 (8.3) 29.1 (12.1) 7.9 (2.2) 8.4 (1.9

M 13.7 (4.1) 27.5 (9.2) 22.1 (4.1) 3.5 (2.0) 2.7 (2.3)

Total 42.3 (8.5) 60.1 (12•4) 51.2 (12.8) 11•4 (3.6) 11.1 (3.0)

9.9 (1.4)

11.1 (2.3) 21.0 (5.3)

Ysselsteyn LF 3.0 (16.2) 35•8 (24.9) 81.0 (16.8) 7.4 (5.4) 10.8 (4.0) 15.0 (3.5) M 0.7 (14.7) 39.7 (25.7) 90•3 (32.3) 6.1 (2.7) 6.0 (4.6) 9.2 (3.7)

Total 3.7 (21.9) 75•5 (35•7) 171•3 (36.4) 13.5 (6.0) 16.8 (6•1) 24•2 (5.1)

SPEULD

z

z m E

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M I N E R A L I Z A T I O N

A h

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t i m e (date ) t i m e ( d a t e ) 1214

Fig. 3. Net nitrification (A) and net mineralization (B) in the forest floor and 0-5 cm of the mineral soil (C, D) for the three plots at the Speuld site. Error bars (n=5) and significant differences between the low and high deposition plot (T = treatment effect) and/or between the high deposition plot and the ambient plot (R=roof effect) at p= 0.05 are indicated.

Ni t r a t e l eve l s s h o w e d a seasona l pat tern , w i th h ig h es t

l eve l s in the fores t f loor espec ia l ly in s u m m e r w h e n N t r a n s f o r m a t i o n ra tes were re la t ive ly low. S t r ik ing were

the h i g h N O 3 - N c o n c e n t r a t i o n s o f the low depos i t i on

plot . In the mine ra l soil these h i g h NO3 leve ls were

absent•

In Ysse l s t eyn the net n i t r i f ica t ion rates in the am-

b ien t fores t soil (Fig. 5) s h o w e d a s t rong seasonal effect

Y S S E L S T E Y N

211

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time (date) time (date)

Fig. 4. Initial concentrations ammonium (A) and nitrate (B) in the forest floor and 0-5 cm of the mineral soil (C, D) for the three plots at the Ysselsteyn site Error bars (n-5) and significant differences between the low and high deposition plot (T - treatment effect) and/or between the high deposition plot and the ambient plot (R - roof effect) atp- 0.05 are indicated.

varying from negative net nitrification during dry summer periods up to 75 mg N kg -1 6 weeks -1 in October 1992. The net nitrification rate at the Ysselsteyn site (Fig. 5) was slightly higher than at the Speuld site (Fig. 3) with highest values in spring and autumn. The low deposition plot showed during two periods in summer and autumn a significant lower net nitrification than the high deposition plot. For the mineral soil significant differences between the plots were found only for the high deposition plot and the ambient plot in spring 1992. Net nitrification for the whole year (Table 3) showed a lower net nitrification at the plots under the roof and a somewhat reduced net nitrification in the low deposition plot compared to the high deposition plot.

A seasonal pattern with rates up to 405 mg N kg-1 6 weeks- 1 in autumn was observed for the net mineralization rates in the forest floor of the ambient plot. This seasonal pattern was less pronounced under the roof. In both plots under the roof the net mineralization was lower compared to the ambient plot during the summer

and autumn period. This was especially pronounced in the low deposition plot where we found a small net immobilization during spring and summer, resulting in a very low net mineralization for the whole year (Table 3). The results from the mineral soil confirm the strong variation within and between the plots (Fig. 5). The overall net mineralization in the forest soil of the low deposition plot (Table 3) was very low.

Laboratory incubations Table 4 shows initial soil conditions and N transformation rates in the forest floor of the intact soil cores and the homogenized forest floor incubated in petri dishes in December 1993. In the intact soil cores, the initial NHa-N concentrations of the forest floor were significantly lower in the low deposition plot compared to the high deposition plot. In the homogenized forest floor smaller differences were observed. NO3-N concentrations were highest in the low deposition plot, although differences were only significant in the homogenized litter. These initial conditions and differences between

212

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Fig. 5. Net nitrification (A) and net mineralization (B) in the forest floor and 0-5 cm of the mineral soil (C, D) for the three plots at the Ysselsteyn site. Error bars (n=5) and significant differences between the low and high deposition plot (T = treatment effect) and/or between the high deposition plot and the ambient plot (R = roof effect) at p= 0.05 are indicated•

Table 4. Characteristics and N transformations of the forest floor of the Ysselsteyn site, incubated in the laboratory (12 °C) as intact soil cores and after homogenization. Standard errors are given in brackets and significance levels (*** p< 0. 001; ** p<0. 01; * p<0.1) are indicated. See text for details

Intact soil cores (n=8) Homogenized soil (n-4) Soil Low High p Low High p characteristic deposition deposition low- deposition deposition low-

plot plot high plot plot high

Moisture (%) 136 (12) 147 (8)

NH4-N (mg kg -x) 100 (17) 195 (24)

NO3-N (rag kg -1) 115 (28) 60 (6) Nitrification 47 (25) 49 (12) (rag k g - x 6 weeks- l)

Mineralization 121 (28) 92 (29) (mgkg -1 6 weeks - l )

128 (2) 164 (1) ***

165 (2) 158 (1) * 82 (3) 57 (1) ***

29 (5) 51 (4) *

131 (9) 139 (8)

the plots are comparable to the conditions found in the in situ incubations. Rates of net nitrification and net mineralization were in the same range for both laboratory methods, under the controlled temperature

conditions and in the same order of magnitude as field rates. Nitrogen transformations did not differ significantly between the plots except for the net nitrification rate in the homogenized litter.

o

-6 E :L

=-

o" z

- - + - - LDP, no-N - - ~ - - LOP, no-N • 0 ' HDP, high-N

60

50

40

30

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t ime (week)

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Net nitrate fluxes (A) and ammonium fluxes (B) of the percolation system during the six weeks incubation. An artificial rain solution was percolated through the soil of the low deposition plot (LDP) and the high deposition plot (HDP) containing no-N (LDP and HDP) or high N (HDP) nitrogen concentrations.

(mg N/soil core. 6 weeks)

S

4

3

2

1

0 LDP, no-N HDP, no-N HDP, high-N

I~.o, n~tr~,cat,o. [] opt m,oora,zat)o. I

Fig. 7. Calculated rates of nitrogen transformations in incubated soil samples of the Ysselsteyn site. Soil cores of the low deposition (LDP) and high deposition (HDP) plot were percolated with an artificial rain solution containing 710 ~mol L - 1 of nitrogen (high N treatment) or no nitrogen (no-N treatment).

213

(mg Nlsoi~ core.6 weeks)

intact percolat ion intact percolation

LDP HDP

[ [ ~ net nRrif ication E ] n e t m, . . . . I lzation J

Fig. 8. Nitrogen transformations in the soil of the low deposition plot (LDP) and high deposition plot (HDP) of Ysselsteyn incubated either as intact soil cores or percolated with an artificial rain solution without nitrogen.

In the percolation experiment, the net nitrate flux (output minus input) did not significantly differ between the plots or between the two N addition rates (Fig. 6). A steady state of the net nitrate output was reached after three weeks. A similar net ammonium flux was found for the no-N treatments. In the high- N treatment the net ammonium flux fluctuated around zero (Fig. 6). Calculation of N transformation rates were based on the last three weeks of incubation with constant nitrate effluxes. No significant differences were found in the net nitrification rate between the plots (Fig. 7). Neither were any differences in net nitrification observed between the no-N and high-N treatments. The net mineralization rate was highest in the no-N treatments (significant different from the high-N treatment at p=0.05). Highest net mineralization was observed in the low deposition plot receiving the no-N rain solution. In the high deposition plot almost all N net mineralized was also nitrified.

If the two laboratory incubations of intact soil cores are compared, the incubation including percolation showed much higher net nitrogen transformation rates compared to the incubation of the intact soil cores without percolation (Fig. 8). Although differences in net nitrification between the plots were small for both methods, the percolation increased the net nitrification rate considerably. For both methods, net mineralization was higher in the low deposition plot and highest in the percolated treatments.

Correlations between nitrogen transformations and environmental factors Figure 9 shows soil temperature and moisture during the incubation periods. Temperatures are averages

214

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Fig. 9. Initial soil moistures (forest floor) and temperatures (6 weeks averages, 1 cm below soil surface) for the plots at the Speuld (A) and Ysselsteyn (B) site during the in situ field incubation.

from under the roof during the six weeks period. Dif- ferences in daily mean temperature between the plots outside and under the roof were small (<0.1 o C, 1 cm below soil surface) (Boxman et al., 1995). Soil moisture at the start of the incubations differed sometimes between the plots although a completely automated sprinkling system was used. At both sites soil moisture was generally highest outside the roof. In Yssel- steyn these differences occurred especially in autumn, in Speuld during the summer period.

In a multiple linear regression analysis rates of nitrification and net mineralization were correlated per plot with the factors initial moisture, temperature, pH, initial NH4-1evels and initial NO3-1evels (p<0.05). These factors could explain 0.15--0.86% of the variation in the net nitrification and net mineralization in the Speuld and Ysselsteyn forests (Table 5). In Speuld net nitrification was significant (negatively) correlated with the initial NO3-N concentration except for the forest floor in the high deposition plot and in the mineral soil of the ambient plot. Significant (negatively) correlations between the net mineralization and initial NH4-N concentrations were found in the forest floor of the low

deposition plot, the ambient plot and in the mineral soil of the high deposition plot. In all three plots of Speuld in the forest floor and the mineral soil a significant (p<0.0001) positive correlation between net nitrification and net mineralization was found (R 2 = 0.44 and 0.41 for the low and the high deposition plot respectively).

In the plots of Ysselsteyn net nitrification of the forest floor was (negatively) correlated with the initial NO3 concentration. Net mineralization in the forest floor of the low deposition plots of Ysselsteyn showed a negative correlation with the initial NH4-N level. Whereas a positive correlation was found between the net mineralization and the initial NH4-N level in the mineral soil of the high deposition and ambient plot. Correlations of the transformation rates with the abiotic factors soil moisture and temperature, at both sites, were limited (Table 5) and could be only little improved by the distinction of a summer and winter period.

Discussion

Although nitrogen transformations in forests subjected to increased nitrogen depositions have been studied before (Tietema et al., 1992), the knowledge of the effects of a decrease in atmospheric nitrogen deposition on nitrogen transformation rates in the soil is limited. The budget of the unaltered forests in this study showed both forests are nitrogen saturated according to the definition of Aber (1989). B oxman et al. (1995) suggested a quick response of nitrogen concentrations in the soil water, especially a reduction of the NO3-N leaching in the low deposition plots. The belowground N cycling patterns were investigated in this study as they may play a critical role in regulating the NO3-N leaching pattern (Van Miegroet et al., 1990) and the turnover rate of the organic matter.

Table 6 shows the nitrogen budget for the forest floor and upper part of the mineral soil (10 cm) of the Speuld and Ysselsteyn site. The results of the N transformations determined with the in situ incubation were combined with measured N fluxes from Boxman et al. (1995). The balance for Speuld shows that less nitrogen is available for plant uptake in the low deposition plot compared to the high deposition plot. If plant uptake is assumed to be at least the amount of the N flux in the above-ground litter production (minimum plant uptake), in the low deposition plot of Speuld a minimum amount of 11 kg N ha- 1 yr- 1 remains available for plant growth (surplus). In the high deposition plot

215

Table 5. Adjusted R 2 of a multiple linear regression analysis between the rate of net nitrification and net mineralization and the initial NO3-N and NH4-N concentrations, pH, moisture and temperature of the Speuld and Ysselsteyn forests

Horizon Low deposition plot High deposition plot Ambient plot R 2 R 2 R 2

Speuld Nitrification LF 0.52 0.53 0.45

M 0.86 0.21 0.72

Mineralization LF 0.58 0.52 0.39 M 0.80 0.46 0.77

Ysselsteyn Nitrification LF 0.38 0,46 0.34

M 0.47 0.15 0.32

Mineralization LF 0.56 0.15 0.15 M 0.41 0.25 0.30

Table 6. Calculated N budget for the forest floor and mineral soil for the three plots at the Speuld and Ysselsteyn site, all fluxes (kg N ha - l yr -1)

Budget Flux Speuld Ysselsteyn Low High Ambient Low High Ambient

In Deposition I 35 40 1 35 56 In Mineralization 42 60 51 4 76 171 Out Leaching 6 18 78 20 47 67 Balance Total 37 77 13 ( - 15) 64 160 Minimum Litter plant uptake production 26 34 38 43 49 56 Surplus Plant growth 11 43 (-25) (-58) 15 104

more nitrogen remains available. In the ambient plot a gap of 25 kg N h a - 1 y r - 1 remains. In this ambient plot nitrogen leaching from the rooting zone excee- ded input via throughfall water by more than 44 kg N ha - l . This is rather high compared to results from Van der Maas (1990) measured a few years earlier in the same forest (other plots). Calculation of the nitrogen budget for Ysselsteyn leads to the conclusion that there is hardly any nitrogen available for plant uptake and leaching in the low deposition plot, mainly due to the very low deposit ion and the low net mineralization rate (Table 6). It should be realized that this simple balance does not include N uptake through the canopy which may decrease soil nitrogen uptake in the balance,

however, the nitrogen fluxes of the belowground litter production were not included which may increase the minimum plant uptake in the balance.

The in situ field method probably underestimates the nitrogen transformations in the soil (Popovic, 1980; Tietema et al., 1992; Williams, 1992). Nitrogen transformations may be underestimated due to the use of closed incubation columns. The percolation study showed that water running through the soil enhances the microbial processes. A removal of toxic plant and microbial components by water flow and a optimal distribution of substrates may result in increased transformations rates.

216

The soil water as sampled with suction cups for the determination of N fluxes (Boxman et al., 1985) might be the mobile water with a relatively short residence time in the soil. As a results nitrogen transformation rates measured the year round in situ may show a dis- crepancy with soil water nitrogen fluxes. But the main N transformations do not take place only in the soil water but also on soil surfaces and inside soil aggre- gates. It was assumed the KCI extracts represent the nitrogen available for the microorganisms, transformations and plant uptake.

At the Speuld site a quick response of the nitrogen concentrations in the soil water was not underlined by the initial inorganic N concentrations measured in KC1 extracts the year round. Only small differences were found in initial NH4-N and NO3-N levels if the low deposition and the high deposition plot were compared. In the mineral soil differences in inorganic N levels were significantly reduced in the low deposition plot. Variation in the nitrogen transformation rates within the plots were small at the Speuld site if compared to the Ysselsteyn site; no major changes in N transformation rates were detectable. The nitrogen transformations are not affected by the decrease in nitrogen input, due to the stage (Aber et al., 1989) of nitrogen saturation of this ecosystem. In an evaluation of nitrogen cycling in European forests Tietema and Beier (1995) showed the intermediate position of the Speuld forest between Scandinavian sites receiving low nitrogen depositions and the very high deposition level of the Ysselsteyn site. The site shows not only an intermediate position in nitrogen deposition level but also in response of the vegetation, soil and soil water. A limited response of the nitrogen transformation processes was confirmed by a decomposition study which showed no significant response in the rate of litter decomposition due to the decrease in nitrogen deposition (Boxman et al., 1995). Also the nitrogen concentrations in the needles did not show a significant response after four years of nitrogen manipulation (Boxman et al., 1995). At present, the nitrogen transformations and probably the microbial activity was not seriously affected by the reduction in nitrogen depositions. A new equilibrium of the soil- vegetation system, adapted to the new N deposition level may be reached in the long run.

At the Ysselsteyn site ammonium concentrations were highest in the high deposition plot. Nitrate concentrations were highest in the forest floor of the low deposition plot during the year. Soil samples used for the laboratory experiments confirmed these difference as well as H20 extracts of forest floor samples of the

same plots (Tietema and Van Dam, 1995). The resulting change in NH4/NO3 ratio in the soil was not expected but may result from differences in the microbial composition and nutrient uptake in the soil.

In the Ysselsteyn forest the year round net mineralization rate was reduced in the low deposition plot. In the intact soil cores incubated in the laboratory no significant differences in the net mineralization between the plots could be observed. In the percolated system a decrease in net mineralization was found in the cores treated continuously with a high nitrogen concentration solution. A decrease in net mineralization due to high continuous NH4 additions was reported earlier for sev- eral soils however, and may result from pH effects or increased immobilization of N (Houdijk, 1993; Kre- itinger et al., 1985; White and Gosz, 1987). Results did not indicate a pH effect so probably an increased N immobilization of the microorganisms would be responsible (White and Gosz, 1987). This hypotheses was not tested within this experiment. A decrease in the rate of nitrification may be expected if the nitrogen availability decreases (Aber et al., 1989). Although this effect was observed during some periods of the year, it did not lead to a significant overall year round decrease in the net nitrification in the low deposition plot of Ysselsteyn. Differences in the chemical composition of the sprinkling water in the field may explain some of the discrepancies observed between nitrogen transformations observed in situ and in the laboratory. In the laboratory, soil cores from both plots received an artificial rain solution, whereas in the field the low deposition plot receiving an artificial rain solution is compared with the high deposition plot receiving origi- nal throughfall water. The effects of organic substances in the throughfall water on nitrogen transformations in these soils are unknown.

The net nitrification rates of the incubated soil cores in the laboratory, intact and homogenized, were in the same order as those for the field. In the percolation system an overall increase in the rate of nitrification rates was observed but no effect of either the plot or the ammonium treatment on net nitrification rates was found. It must be kept in mind that these laboratory experiments do not include any fluctuations of the nitrogen availability and transformation throughout the year.

In the Ysselsteyn forest we found higher nitrogen transformations compared to the Speuld site. This was confirmed by higher decomposition rates at this site (Boxman et al., 1995) and a higher substrate quality (lower lignin to nitrogen ratio, Koopmans, in prep.).

In the previous years, the nitrogen depositions in the area around Ysselsteyn decreased already considerably (Houdijk, 1993). Together with the high organic matter decomposition and high leaching, nitrogen availability may have decreased significantly and more than in the Speuld forest. The needles in the low deposition plot too, at this site showed a significant response in the arginine level, the sink for an excess of nitrogen (Box- man et al., 1995). This may confirm that less nitrogen becomes available in the soil in the low deposition plot in Ysselsteyn.

The use of the tracer ~SN, applied during a one year period to the manipulated plots in the Ysselsteyn and Speuld site (Kj~naas et al., 1992) in combination with the use of NIICCE, a model for cycling of N and C istopes in coniferous forest ecosystems (Van Dam and Van Breemen, 1995) will hopefully increase our insight into the role of the deposition level on the nitrogen transformations in the soil of the Speuld and Ysselsteyn site.

Conclusions

From this study it may be concluded that there is no direct relationship between the N leaching pattern of the plots and the net nitrogen transformation rates for the Speuld site. The limited response of the nitrogen concentrations in the needles under the decreased input level confirms that nitrogen transformations are hardly affected at this site after four years of N manipulations. Nitrogen is still available for microorganisms and plant uptake even under the reduced nitrogen deposition level.

The reduction in N deposition at the Ysselsteyn site leads to a decrease in nitrogen dynamics in the upper soil layers during certain periods of the year in this highly N saturated site. Differences in N leaching could be partly explained from differences in N dynamics in the upper soil horizons. At the low nitrogen deposition plot a net immobilization occurred during most of the year leading to a very low net mineralization for the year round period. Decreased net mineralization rates measured in the low deposition plot in the Ysselsteyn site may lead to the reduced nitrogen uptake by the vegetation, smaller leaching losses and relatively higher immobilization rates of the nitrogen deposited.

217

Acknowledgements

This research was financially supported by the Dutch Priority Program on Acidifcation and by the EU- Environmental Program (STEP).

We wish to thank P F Wartenberg for his contribu- tion in the analytical work, Dr Ir G Heuvelink for his advise on statistics and Prof Dr J M Verstraten, Dr W de Boer and Dr D van Dam for reviewing an earlier draft of the manuscript.

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