Nitrogen mineralization, nitrification and denitrification potential in contrasting lowland rain forest types in Central Kalimantan, Indonesia

ARTICLE IN PRESS

Soil Biology & Biochemistry 39 (2007) 2992–3003 www.elsevier.com/locate/soilbio

Nitrogen mineralization, nitrification and denitrification potential in contrasting lowland rain forest types in Central Kalimantan, Indonesia R.R.E. Vernimmena,, H.A. Verhoefa, J.M. Verstratenb, L.A. Bruijnzeela, N.S. Klompa, H.R. Zoomera, P.E. Wartenberghb a Faculty of Earth and Life Sciences, Vrije Universiteit, Amsterdam, The Netherlands Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam, Amsterdam, The Netherlands

b

Received 5 January 2007; received in revised form 5 June 2007; accepted 7 June 2007 Available online 27 July 2007

Abstract Nitrogen mineralization and denitrification potential in litter were measured during a dry and a wet period in a Bornean Lowland Evergreen Rain Forest (LERF) and two nearby Heath Forests (HF) of contrasting stature. Nitrification was very low or non-existent in all forest types and ammonification was the major constituent of nitrogen (N) mineralization. Rates of net N mineralization in the HFs on infertile sandy soils were lower than in the LERF on a more nutrient-rich clay soil or other LERFs, both during dry and wet conditions. We attribute the differences to the lower litter quality in the HFs compared to LERF. When dissolved organic nitrogen (DON-N) was included, N uptake was the same (15–17 mg g1 d1) in all three forest types. We conclude that N availability is the same in all three forest types and that N deficiency is not the reason for the reduced stature of Heath Forests compared to LERF. All three-forest types had denitrifiers present in the ectorganic layers but denitrification will only play a minor role in the N-cycle as nitrification rates were very low. r 2007 Elsevier Ltd. All rights reserved. Keywords: Nitrogen mineralization; Denitrification potential; Heath forest; Lowland Evergreen Rain Forest; Borneo; Dissolved organic nitrogen

1. Introduction The microbial mineralization of ammonium (NH4-N) from soil organic matter is the principal source of plantavailable nitrogen (N) in most forest ecosystems and rates of N mineralization can regulate the productivity of many forests (Nadelhoffer et al., 1983; Pastor et al., 1984; Miller et al. 2003). The rate of ammonification (the transformation of organic N to NH4-N) and nitrification (the oxidation of NH4-N to nitrate (NO3-N)) play a key role in the N cycle by making N available for plants and microbes, and by making N susceptible to leaching and denitrification losses. Factors affecting nitrification in soils Corresponding author. Department of Hydrology and Geo-Environmental Sciences, Faculty of Earth and Life Sciences, Vrije Universiteit, De Boelelaan 1085, 1081 HV, Amsterdam, The Netherlands. Tel.: +31 (0)20 5987377; fax: +31 (0)20 5989940. E-mail addresses: [email protected], [email protected] (R.R.E. Vernimmen).

0038-0717/$ - see front matter r 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.soilbio.2007.06.005

are pH, moisture, temperature, C:N ratio of the litter, the presence of plant-produced allelochemicals, and the supply of essential nutrients, notably phosphorus (Robertson, 1982; Tietema et al., 1992; Nugroho et al., 2007). N-conserving mechanisms may be better developed in nutrient-poor than in nutrient-rich environments (Jordan et al., 1979; Pugnaire and Chapin, 1993). For example, Jordan et al. (1979) have shown how low pH and high tannin content of the root humus layer in an undisturbed rain forest on Oxisol of low nutrient content at San Carlos, Venezuela is effective in conserving nitrogen through depressing populations of denitrifying bacteria. Heath forests (HF), a distinctive lowland rainforest type, are known for their occurrence on highly acidic and infertile soils that are also allegedly deficient in N. HF is distinctly shorter in stature and poorer in species (but high in endemism), simpler in structure, and of lesser biomass than Lowland Evergreen Rain Forest (LERF) on clay soils (Whitmore, 1998). One of the reasons for the distinctive

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features of HF has been hypothesized to be nutrient deficiency, particularly N (Proctor, 1999). As in similarly structured wet montane forests the mineral nutrient supply in HF is thought to be restricted because a large proportion of nutrients (particularly N and P) is bound in undecayed litter or unmineralized humus (Coomes and Grubb, 1996; Hafkenscheid, 2000). Low N concentrations in HF leaves and litterfall in Borneo and Amazonia (Proctor et al., 1983; Coomes, 1997) and the abundance of insectivorous plants (Whitmore, 1998) suggest that N may indeed be in short supply in HF. Studies of nitrification in undisturbed tropical forests (top) soils have reported a wide range in nitrification rates (o0.1–6 mg N g1 d1) (reviewed by Robertson, 1989; Hafkenscheid, 2000; Van Dam, 2001) with sometimes negative values (Matson and Vitousek, 1987). Low to very low nitrification and nitrogen mineralization rates have been reported for HF soils (Marrs and Proctor, unpublished) and in wet montane tropical soils (Marrs et al., 1988; Hafkenscheid, 2000) while high rates have been found in fertile volcanic soils (Marrs et al., 1988) and high to intermediate rates in lowland Oxisols and Ultisols (Vitousek and Matson, 1988; Montagnini and Buschbacher, 1989). Low nitrification and mineralization rates are expected in HFs with their low pH and high concentration of tannins in leaf material (Proctor et al., 1983; Vernimmen

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et al., in preparation). Jordan et al. (1979) attributed the low number of nitrifying bacteria in a Venezuelan HF to low soil pH and high concentrations of tannins in the root mat. However, there are conflicting reports on the effects of tannins on nitrification (Kraus et al., 2003). Furthermore, relatively high nitrification rates are known to occur in a variety of soils at pH values of less than 4.0 (Robertson, 1982; Schmidt, 1982, Kowalchuk and Stephen, 2001). This paper aims to test the hypothesis that low rates of nitrogen turnover may play an important part in the reduced stature of HF. Rates of nitrification, ammonification and net nitrogen mineralization on the forest floor plus potential denitrification rates in the ectorganic layers were measured in a tall LERF and in two nearby HFs of contrasting stature in Central Kalimantan, Indonesia. In addition, the effect of drought on nitrogen-transformation rates was assessed by comparing measurements made during a dry and a wet period. 2. Study area The present study was conducted in the remote 450 ha Barito Ulu research area in Central Kalimantan (01030 4100 S, 114100 1000 E) near the confluence of the Rekut and Busang rivers in the Sumber Barito District (Fig. 1). The entirely forested area belongs to the lower parts of the Muller Mountain range and has a rolling topography with

Fig. 1. Location of the Barito Ulu research area in Central Kalimantan, Indonesia. Site map courtesy of Mr. S.R.G. Ridgeway; LERF ¼ Lowland Evergreen Rain Forest, THF ¼ tall Heath Forest, and SHF ¼ stunted Heath Forest.

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plateaus at ca. 180 m a.s.l., and steeply incised rivers. Measurements were carried out in three plots covered with Lowland Evergreen Rain Forest (LERF, 0.25 ha), tall Heath Forest (THF, 0.28 ha) and stunted Heath Forest (SHF, 0.13 ha), respectively. The two HF plots were located next to each other at ca. 180 m a.s.l. on a gently sloping (1–51) ridge of northerly to northwesterly orientation, whereas the LERF was located on a much steeper (25–351) and dissected slope approximately 400 m from the HF plots (Fig. 1). Average tree height was estimated at ca. 40 m in the LERF and was measured in the tall (20.276.3 m) and in the stunted HF (15.475.3 m). The two HF plots were underlain by haplic Podzols whereas the LERF plot was on a haplic Acrisol (FAO, 2006). pHH2O (1 : 2.5 w/v) ranged from 4.3 in the LERF to 3.9 in the THF and 3.3 in the SHF. At the HF sites, the mineral soil was covered by a mat of roots growing on top of, and attaching themselves to freshly fallen litter. The root mat was generally between 5 and 15 cm thick in the HF but very thin (max. 3 cm) in the LERF. Central Kalimantan has a humid tropical climate (type Af in the Ko¨ppen system). Temperatures vary little during the year (25.470.8 1C) while diurnal variations typically range from 23 to 30 1C. Mean annual rainfall as measured at 130 m a.s.l. at the Barito Ulu Base Camp (800 m from the HF plots) for the period 1994–2002 was 36257560 mm distributed over an average of 208733 rain days per year. The wettest year on record (1995) had 4400 mm of rain (226 rain days) and the driest (1997) 2575 mm and 135 rain days. Rainfall is distributed fairly evenly within the year and is mostly convective in origin. In general, January and March are the wettest months with about 400 mm each, while the July–September period is relatively dry although still receiving ca. 215 mm per month on average. Despite the humid climate dry months (Po100 mm, sensu Bru¨nig, 1969) are known to occur on Borneo (Bru¨nig, 1969; Vernimmen et al., 2007). Sliding 30-d precipitation totals revealed even more dry periods (Vernimmen et al., 2007). 3. Methods 3.1. Ammonification, nitrification, and net nitrogen mineralization 3.1.1. Field The experiments were conducted under dry (19 August–2 September 2002) and wet conditions (30 June–14 July 2003). In the week preceding the experiment under dry conditions, total measured rainfall was 0.6 mm, while this was 49.9 mm for the experiment under wet conditions. One to two weeks prior to the experiments ten PVC cylinders with a diameter of 15.9 cm and a height of approximately 20 cm were installed randomly in each of the plots (see for description: Hafkenscheid, 2000). Due to the thick surface root mat, the cylinders could not be inserted into the soil without severely damaging the root mat which would affect

the results. Therefore, the cylinders were installed on top of the root mat, while assuring good contact between the cylinder and the root mat using moulding clay, to prevent nitrogen loss by nitrate leaching. At the start of the experiment each cylinder was covered with a tilted plastic sheet (positioned approximately 5 cm above the rim) to keep out any dry and wet deposition. Reference litter samples were taken near the cylinders and transported to the Base Camp in plastic bags for further processing. After an incubation period of 14 days, litter samples were collected from inside the cylinders. 3.1.2. Laboratory For the litter samples that had been incubated under dry conditions, 150 ml of 1 M KCl was added to 10 g of fresh weight of each sample, while for the experiment under wet conditions, 150 ml of 0.5 M K2SO4 was used as an extractant, enabling analysis of dissolved organic nitrogen (DON) as well. To allow comparing of dry and wet conditions, KCl was also used as extractant for half the samples from the experiment conducted under wet conditions. All solutions were prepared with fresh water from a nearby stream which was filtered through a Fison ion exchange column producing an electric conductivity value of 3 mS cm1. All samples were extracted within 32 h of collection. Blanks were run in order to ensure quality control. The samples were shaken for 2 h after which they were filtered, first over a Whatman filter (0.80 mm) and secondly over a Millipore filter (0.45 mm). A total of 5 ml of concentrated H2SO4 was added to keep pH below 2 to prevent bacterial growth and N-transformations. The samples were stored at temperatures below 8 1C until analysis in The Netherlands. Additionally, remaining fresh litter was oven-dried at 80 1C to constant weight for gravimetric determination of field moisture conditions and subsequently dry weight, thereby allowing calculation of concentrations of NO3 and NH4 per gram dry weight. NH4-N, NO3-N and NO2-N in both extractants were analyzed colorimetrically with a Skalar Auto Analyzer at the Faculty of Earth and Life Sciences, Vrije Universiteit Amsterdam (FELS-VUA) for samples extracted with KCl and at the Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam (IBED-UoA) for samples extracted with K2SO4. No nitrite could be detected in any sample. Total nitrogen (Nt) was determined after oxidation of organic nitrogen, and NH4 and NO2 were measured in the K2SO4/H2SO4 extract after oxidation to nitrate (Lowry and Mancy, 1978). After oxidation was completed, Nt was measured colorimetrically as nitrite after reduction on a Cd-column (Lowry and Mancy, 1978) with a Skalar Auto Analyzer. Subsequently, DON was calculated as NtNO3NH4. To determine the effect of the different analyzers used in the two laboratories, samples with known concentrations of NO3 and NH4 were analyzed in both KCl and K2SO4 and with both analyzers. The comparative analyzer test

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3.2. Potential denitrification 3.2.1. Field Potential denitrification rates were measured in six to nine replicates from the litter (L), fragmentation (F) and topsoil (organic) horizons (H) in the SHF and THF, as sampled on 1 August 2002 (2002 experiment). No organic horizon was sampled in the THF. Between 12 and 19 May 2003 (2003 experiment), the experiment was repeated with nine replicates at each location, including the LERF. The organic horizon in the THF was sampled but not in the LERF as it was too thin. In the week preceding the 2002 experiment total measured rainfall was 83.4 mm vs. 82.0 mm for the 2003 experiment.

3.2.2. Laboratory A total of 100 ml of a 0.2 mM KH2PO4/0.2 mM CaCl2/ 0.2 mM MgSO4-solution (De Boer et al., 1992) was added to 20 g of fresh weight sample and pre-incubated for 24 h at ambient temperature (23–30 1C) in open bottles in the field laboratory at the Base Camp. All samples were preincubated within 32 h of collection. To determine the potential denitrification rates of the samples, a NO3solution was added after pre-incubation, to a final concentration of 2 mM (initial concentration was between 0.032 and 0.113 mM NO3-N g1 dry weight) (Laverman, 2000). Samples were shaken thoroughly and a 5 ml subsample was taken to determine initial concentrations of mineral N. A total of 5 ml of concentrated H2SO4 was added to keep pH below 2 so as to prevent bacterial growth and N-transformations and the sample was kept at temperatures below 8 1C until analysis at FELS-VUA. A total of 5 ml of the NO3-solution was added to the remaining sample to keep the volume constant after subsampling. Samples were shaken gently in a refrigerator basket for 24 h in a nearby stream at approximately 25 1C, before another 5 ml subsample was taken. A final 5 ml subsample was taken after 48 h. Subsamples were treated the same way as the subsamples taken after pre-incubation. For the 2003 experiment an extra subsample was taken after 72 h. Remaining fresh litter was oven-dried at 80 1C to constant weight for gravimetric determination of moisture content. Potential denitrification rates were calculated from the differences in the NO3 concentrations.

3.3. Statistics Because criteria for ANOVA were not always met for the mineralization data (normality and homogeneity of variance) the non-parametric Kruskal–Wallis test was used instead (only if the results consisted of more than two groups, otherwise the Mann–Whitney test was used). If the Kruskal–Wallis test revealed a significant difference (Po0.05) between the results of the three forest types the Mann–Whitney test was used for pairwise group comparisons with a Bonferroni correction for the P value. Because three groups were compared P became 0.016 (0.05/3 comparisons). Gravimetric litter moisture contents were regressed against initial NH4-N, NO3-N concentrations, ammonification, nitrification and net N mineralization rates using Pearson correlation (Po0.05). The results of the denitrification experiment were tested using a single ANOVA. If the ANOVA noted a significant difference (Po0.05) between the results of an experiment a Tukey HSD test was used as a post hoc (only if the results consisted of more than two groups). All statistical analyses were performed using SPSS 12.0.1. 4. Results 4.1. Ammonification, nitrification, and net nitrogen mineralization 4.1.1. Moisture condition effects Moisture conditions were different between the two experiments except in the SHF (Fig. 2). Average gravimetric moisture contents of the litter samples ranged from 50% to 76% during the dry period and between 100% and 150 gravimetric moisture content [%]

showed no significant effect of analyzer on measured concentrations of NO3 and NH4. Also the extractant used (KCl or K2SO4) had no effect on the measured concentrations. Net N mineralization rates were calculated as the sum of net ammonification and net nitrification divided by incubation time, with net ammonification defined as final NH4-N minus initial NH4-N, and net nitrification as final NO3-N minus initial NO3-N (Robertson and Vitousek, 1981).

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a

b c

100

50

0

LERF

THF forest type

SHF

Fig. 2. Average gravimetric moisture contents (%) (with standard errors) of litter samples used in the nitrification experiments during dry (light grey bars) and wet (dark grey bars) conditions (n ¼ 10) in Lowland Evergreen Rain Forest (LERF), tall Heath Forest (THF), and stunted Heath Forest (SHF) at Barito Ulu, Central Kalimantan. Bars carrying different superscripts are different (Po0.05) between forest types.

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139% during the wet period. Moisture contents did not differ significantly at the 0.05 level between plots during the dry period but moisture content was higher in LERF litter during the wet period compared to the litter of the two HFs. Significant correlations (Po0.05) between net nitrogen mineralization and gravimetric moisture content were only found in LERF litter under wet conditions and in the SHF under dry conditions (Table 1). However, whilst the correlation was positive in LERF, it was negative in the SHF. Except for a significant negative correlation between nitrification rate and gravimetric litter moisture content under wet conditions, no significant correlations were found between nitrogen transformation rates and gravimetric moisture contents in the THF. 4.1.2. Nitrogen transformations in litter: comparisons between different forest types Initial NH4 concentrations in LERF litter during dry conditions were higher than those in the two HFs, whereas

there was no difference between forest types under wet conditions (Table 2). Initial NH4 concentrations in LERF litter during dry conditions were roughly five times higher than those during wet conditions. In the two HFs initial NH4 concentrations were approximately two times higher during dry conditions than during wet conditions. Initial NO3-N concentrations in litter were not different in the respective forest types, both during wet and dry conditions. Initial NO3-N concentrations during wet conditions were eight times higher in LERF and THF litter and four times in SHF litter compared to their dry condition equivalents. Ammonification was the major constituent of net N mineralization and was six times higher in LERF litter compared to HF litter for both moisture conditions. Average ammonification rate in SHF litter under dry conditions was very low compared to the other two forest types but because of the broad range in observed rates it was not significantly different from the ammonification rate in the THF.

Table 1 Pearson correlation coefficients between initial NH4-N and NO3-N concentrations, ammonification, nitrification and net N-mineralization in litter and gravimetric litter moisture contents during the mineralization experiment in Lowland Evergreen Rain Forest (LERF), tall Heath Forest (THF), and stunted Heath Forest (SHF) at Barito Ulu, Central Kalimantan LERF

THF

Dry NH4-N initial NO3-N initial Ammonification rate Nitrification rate Net N-mineralization rate *

P o0.05;

Wet *

0.717 0.231 0.169 0.369 0.187

0.360 0.309 0.979** 0.865 0.980**

Dry 0.310 0.601 0.01 0.592 0.028

SHF Wet 0.852 0.996** 0.297 0.970** 0.427

Dry

Wet **

0.889 0.656* 0.929** 0.230 0.938**

0.228 0.331 0.311 0.073 0.310

P o0.01.

**

Table 2 Average values and ranges (in parentheses) of initial inorganic nitrogen concentrations (mg g1) and rates of transformation (mg g1 d1) in litter of Lowland Evergreen Rain Forest (LERF), tall Heath Forest (THF), and stunted Heath Forest (SHF) at Barito Ulu, Central Kalimantan extracted with KCl. Incubations were performed between August 19 and September 2, 2002 (dry conditions) and between June 30 and July 14, 2003 (wet conditions) with number of replicates in parentheses Site

Period Initial concentration (mg g1)

Final concentration (mg g1)

Net NH4-N production (ammonification) (mg g1 d1)

Net NO3-N production (nitrification) (mg g1 d1)

Net N mineralization (mg g1 d1)

NH4-N

NO3-N

NH4-N

NO3-N

LERF Dry (10) Wet (5)

35.0 (18.660.1)a# 6.67 (0–14.0)a#

0.30 (0–0.76)a# 2.55 (1.65–3.69)a#

78.4 (21.4–139.6)a 81.0 (32.8–127)a

1.26 (0–4.95)a 0.93 (0–1.67)a

3.10 (0.09,7.78)a

0.07 (0.01,0.30)a#

3.17 (0.09,7.77)a

5.31 (2.34–8.84)a

0.12 (0.16,0.08)a#

5.20 (2.21–8.76)a

THF Dry (9) Wet (5)

10.2 (2.81–26.3)b 4.35 (0–8.14)a

0.27 (0–0.93)a# 2.50 (0.89–5.17)a#

17.4 (8.49–35.7)b 16.6 (3.10–25.7)b

0.10 (0–0.90)bc 0.21 (0–0.62)a

0.51 (0.29,1.72)b

0.01 (0.07,0.03)ac#

0.50 (0.29,1.69)b

0.88 (–0.08,1.84)b

0.16 (0.34,0.02)a#

0.71 (0.21,1.82)b

SHF

8.19 (0–51.1)b 3.58 (0–17.9)a

0.54 (0–1.19)a# 1.96 (1.13–3.74)a#

8.33 (3.44–20.7)c 15.0 (7.1726.2)b

0.25 0.01 (2.86,0.82)b (0–0.80)ac# 0.93 0.82 (0.63,1.87)b (0.381.70)a#

0.02 (0.09,0)bc

0.01 (2.87,0.77)b

0.07 (0.20,0.01)a

0.74 (0.71,1.71)b

Dry (10) Wet (5)

Values carrying different superscripts are different (Po0.016) between plots during either dry or wet conditions. Values carrying a # are different (Po0.05) between the dry and wet conditions.

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4.2.2. Potential denitrification rates in the ectorganic layers of different forest types The average potential denitrification rates for LERF, THF and SHF litter during the two experiments are given in Table 4. Potential denitrification rates on a dry weight basis during the 2002 experiment were always two to four times higher in L-horizon material compared with the F- or H-horizon, whereas there was no difference between F- and H-horizons. Neither was there a difference during the 2002 experiment between forest types when the same ectorganic horizons were compared. During the 2003 experiment, no differences were found in potential denitrification rates between the L- and F-horizons of all forest types. Potential denitrification rates differed however between the L- and H-horizons in the two HFs, where the rates were up to two times higher in the L-horizon. Potential denitrification rates were three to four times higher in the L- and F-horizons of the LERF compared with the THF and SHF, whereas the rates in the

Nitrification rates in the two HFs were low or nonexistent, and were especially low under dry conditions in SHF litter. Nitrification rates during dry conditions were higher in LERF- and THF litter (i.e. less negative) compared to wet conditions. Net N mineralization rates in LERF litter were higher compared to rates in the two HFs both under wet and dry conditions whereas there was no difference in net N mineralization rates between the two HFs. Net N mineralization rates were six to seven times higher in LERF litter compared to HFs litter (leaving the negative value found for dry conditions in the SHF aside; Table 2). 4.1.3. Nitrogen partitioning under wet conditions Average initial and final inorganic nitrogen and DON-N fractions of Nt in K2SO4 extractions of litter during the wet period are listed in Table 3. Representing 73–89% of total extractable N, initial DON-N dominated the extract with NO3-N contributing only 1–3%. On average, approximately 15–17 mg g1 N was taken up by roots or immobilized by bacteria during the incubation period (Table 3), with no significant differences between forest types. After the two week incubation period most DON-N was immobilized in LERF litter (3 mg g1 d1) followed by THF- (1.8 mg g1 d1) and SHF litter (1.2 mg g1 d1). However, these differences were not significant (Mann–Whitney, P40.05). DON-N and NH4-N production were not correlated.

250 gravimetric moisture content [%]

F

4.2. Potential denitrification 4.2.1. Moisture conditions Gravimetric moisture contents of the ectorganic samples of the two denitrification experiments are shown in Fig. 3. Average gravimetric moisture contents of the L- and F-horizons ranged between 99–115% and 135–167%, respectively, during the 2002 experiment. During the 2003 experiment, average gravimetric moisture contents ranged from 70% to 114%, 132% to 197% and 166% to196% in the L-, F- and H-horizons, respectively. Because approximately the same amount of rainfall had been received by the forest floor one week before the start of both experiments no significant differences (P40.05) in gravimetric moisture content were found, except for the F-horizon in the THF and the H-horizon in the SHF.

H∗

∗

F

200

F H

150

F

L F∗ L

L L

L

100

H

∗

L

50

0

LERF

THF forest type

SHF

Fig. 3. Average gravimetric moisture content (%) with standard errors indicated by the error bars for the litter- (L), fragmentation- (F) and topsoil (organic) horizon (H) samples from the denitrification experiment in 2002 (light grey bars) and 2003 (dark grey bars) in Lowland Evergreen Rain Forest (LERF), tall Heath Forest (THF), and stunted Heath Forest (SHF) at Barito Ulu, Central Kalimantan; n ¼ 9 except for 2002 experiment in SHF, n ¼ 6. Bars of the same horizon within a forest type carrying an asterisk are different (Po0.05) between the two experiments.

Table 3 Average of initial and final inorganic nitrogen and DON fractions of Nt, in samples extracted with K2SO4 of litter of Lowland Evergreen Rain Forest (LERF), tall Heath Forest (THF), and stunted Heath Forest (SHF) at Barito Ulu, Central Kalimantan under wet conditions Site

LERF THF SHF

n

9 10 10

Nt (mg g1)

91.8 36.4 37.6

Nt (mg g1)

Initial (% of Nt) NH4-N

NO3-N

DON-N

24.3a 8.5bc 11.3ac

2.5a 2.1a 1.4a

73.2a 89.4bc 87.3ac

Values carrying different superscripts are different (Po0.016) between plots.

74.4 20.2 22.9

Final (% of Nt) NH4-N

NO3-N

DON-N

64.0a 62.4ab 27.1ac

2.2a 0b 0.8b

33.9a 37.6a 72.1b

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Table 4 Average potential denitrification rates (mg N g dw1 d1) with ranges in parentheses for Lowland Evergreen Rain Forest (LERF), tall Heath Forest (THF), and stunted Heath Forest (SHF) at Barito Ulu, Central Kalimantan in 2002 and 2003 Forest type

Ectorganic layer

Potential denitrification rate (mg N g dw1 d1) n

n

2002 48 h

LERF THF

SHF

L F L F H L F H

9 9 6 6 6

n.d. n.d. 0.12 0.05 n.d. 0.09 0.04 0.02

2003 48 h

(0.28,0.02)a1 (0.08,0.02)b1 (0.12,0.07)a1 (0.04,0.00)b1* (0.05,0.03)b*

9 9 9 9 9 9 9 9

0.43 0.41 0.14 0.11 0.08 0.11 0.08 0.05

72 h (0.63,0.33)a1# (0.54,0.21)a1# (0.21,0.10)a2 (0.24,0.02)ac2 (0.15,0.04)bc1 (0.22,0.04)a2 (0.12,0.01)ac2* (0.07,0.04)bc1*

No more substrate No more substrate 0.12 (0.17,0.08)a2 0.13 (0.26,0.03)a2 0.08 (0.13,0.04)a1 0.14 (0.23,0.09)a2 0.10 (0.15,0.05)ac2 0.07 (0.10,0.05)bc1

# rates after 24 h of incubation, because no substrate was left after 48 h of incubation (except for 12 samples). Values carrying different lowercase letters are different between layers of a forest type in 2002 (Po0.05) or 2003 after 48 h (Po0.016) and 72 h (Po0.05) of incubation. Values carrying different numbers are different (Po0.05) between forest types for the same ectorganic layers after 48 h of incubation. Values carrying an asterisk are different between the two experiments after 48 h of incubation; n.d. ¼ not determined.

respective ectorganic horizons of the two HFs did not differ from one another. Potential denitrification rates were higher during the 2003 experiment compared to the 2002 experiment for the F- and H-materials of the SHF. Fractions of potentially denitrified NO3-N after 24, 48 and 72 h of incubation for both experiments are shown in Fig. 4. At the start of both experiments average NO3 concentrations ranged between 2.8 and 4.9 mg g1 dry weight. After 48 h, on average, almost no NO3 was left in the LERF ectorganic horizons whereas there was still 70–90% left in the HFs depending on ectorganic horizon and moisture conditions. Potential denitrification rates accelerated in SHF material during the last 24 h of the experiment but rates remained more or less the same in the THF. 5. Discussion 5.1. Ammonification, nitrification, and net nitrogen mineralization 5.1.1. Nitrogen pools at the start of the experiments NH4-N was the predominant form of inorganic-N in the litter from all three forest types, accounting on average for 64–72% and 94–99% of inorganic-N during dry and wet conditions, respectively. Higher initial NH4-N concentrations in LERF litter compared to HF litter suggest a higher availability of inorganic N, most likely due to a higher overall fertility level in the LERF. This higher fertility is also reflected in the concentrations of N and other nutrients in leaf litterfall, which were all higher in the LERF compared to the two HFs (Vernimmen et al., in preparation). Higher initial NO3-N concentrations in the litter layer during wet conditions were to be expected because of the higher activity of bacteria during moister conditions, while

NH4-N concentrations will be lowered accordingly due to nitrification or low ammonification rates. Because there was no effect of analyzer type used on measured concentrations, any differences between the analyses using KCl or K2SO4 as extractant can be attributed to either the extractant or the implicitly high spatial variability of the litter material in species-rich tropical rain forests. Because this cannot be ascertained, further discussion will focus on the comparison of net N mineralization between wet and dry periods using KCl as an extractant, and on DON-N concentrations in the K2SO4 extracts. 5.1.2. Nitrification Nitrification was low or slightly negative in both LERF and HF litter (Table 2). The low nitrification rates in the HFs may have been exacerbated by the combined negative effects of low pH and high concentrations of tannins in leaf material on the numbers of nitrifying bacteria (Jordan et al., 1979; Tietema et al., 1992). Another reason for reduced nitrification may be decreased microbial NH4-N availability due to occurrence of ectomycorrhizae (Chandler, 1985). Tree families that form ectomycorrhizae (notably Dipterocarpaceae and most Myrtaceae; Malloch et al., 1980) constituted 40–60% (in terms of basal area) of the identified species in the respective forest types at Barito Ulu. 5.1.3. Net nitrogen mineralization Table 5 provides a comparison of nitrification, ammonification and net nitrogen mineralization rates in ectorganic layers at selected tropical sites. Negative nitrification rates in organic layers were only found in the forests from the present study, elsewhere nitrification contributed 30–50% of net N mineralization. The high nitrification rates found in other studies probably reflect higher NH4-N

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Fig. 4. Fractions of potentially denitrified NO3 with time for (a) Lowland Evergreen Rain Forest (LERF), (b) tall Heath Forest (THF), and (c) stunted Heath Forest (SHF) at Barito Ulu, Central Kalimantan in 2002 (open markers) and 2003 (filled markers) for L-horizon (circles), F-horizon (squares) and H-horizon (diamonds) materials.

availability. Initial NH4-N concentrations in the organic layers of Spodosols and Oxisols/Ultisols (USDA, 1951) were 10–12 times and 2–3 times higher, respectively compared to those of litter layers under dry conditions in the forests of this study. Ammonification rates in LERF litter fell within the range found by other studies on Oxisols and Ultisols. Net N mineralization in litter of the LERF at Barito Ulu was significantly higher compared to rates in the two HFs which were not significantly different from each other. The higher net N mineralization rates in LERF litter compared to HF litter were reflected by the higher initial concentrations of NH4-N and DON-N in LERF litter, which, together with its higher quality (lower C:N ratio and phenol concentrations), provide more favourable conditions for the microbial community (Anderson et al., 1983).

5.1.4. The role of DON-N While there is a difference in net production of inorganic nitrogen during wet conditions between the LERF and the two HF sites (Table 2), approximately the same amount of nitrogen was immobilized in each forest during the incubation period of 14 days, when DON-N was included (between 15 and 17 mg N g1, Table 3). Jones et al. (2005) stated that the direct uptake of DON-N by plants, without the prior need f or microbial mineralization, has the potential to be a primary factor in ecosystem functioning, particularly in N-limiting environments. However, experimental evidence for such a mechanism is still lacking. Our results seem to provide at least some circumstantial evidence of the suggested importance of this alternative N-pathway.

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Table 5 Concentrations of mineral nitrogen (NO3-N and NH4-N, mg g1), total nitrogen (%) plus net rates of ammonification, nitrification and net N mineralization in ectorganic layers of selected tropical forest sites Site

NO3-N (mg g1)

NH4-N (mg g1)

Nt (%)

Ammonification (mg N g1 d1)

Nitrification (mg N g1 d1)

N mineralization (mg N g1 d1)

THF, litter on Spodosola SHF, litter on Spodosola Brazil, organic layer on Spodosolc,d LERF, litter on Ultisola Brazil, organic layer on Oxisol/Ultisolc,d San Carlos, Venezuela, rootmat on Oxisol/Ultisole

0.27–2.5 0.54–2.0 0 0.30–2.6 0.7–2.0 5.9–9.0

4.4–10.2 3.6–8.2 102 6.7–35.0 93.5–94.4 28.8–80.8

0.55b 0.40b 1.14 0.96 0.84–0.97 1.0–1.1

0.73 0.50 2.4 4.4 4.8–5.4 0.47–3.0

0.10 0.05 1.5 0.04 2.0–2.6 0.37–3.1

0.63 0.44 3.9 4.4 6.8–8.0 0.84–6.1

a

This study. Vernimmen et al. (in preparation). c Livingston et al. (1988). d Vitousek and Matson (1988). e Montagnini and Buschbacher (1989). b

5.2. Potential denitrification Denitrifying bacteria were present in the ectorganic layers of all three forest types as was demonstrated by the two experiments and as such they affect the availability of NO3-N. Potential denitrification rates were three to four times higher in the L-horizon and four to five times higher in the F-horizon materials in the LERF compared to the two HFs. A higher availability of mineral N in litter in the LERF (Table 2) may be one of the reasons for the higher potential denitrification rates encountered in the LERF ectorganic layers. Griffiths et al. (1993) showed that potential denitrification rates were strongly correlated with levels of mineralizable N in volcanic soils in Costa Rica. A second, perhaps more important reason for the higher potential denitrification rates in the ectorganic layers in the LERF is the better quality of the litter in terms of carbon sources that the denitrifiers require to reduce NO2-N or NO3-N to N2O (Griffiths et al., 1993). As indicated earlier, the quality of litter in the LERF is better, and probably consists of more easily decomposable material, because of its lower C:N ratio (547SD 3.7) and lower concentration of phenols (62.3711.8 mg tannic acid equivalents (TAE) g1) compared to the litter in the two HFs (C:N ratio of 102710.2 and 14179.7, phenol concentration of 98.87 17.6 and 108.7716.4 mg TAE g1 for THF and SHF, respectively, Vernimmen et al., in preparation). In addition, concentrations of DOC in litter percolate were much higher in the LERF (529 kg C ha1 yr1) compared to the THF (356 kg C ha1 yr1) and SHF (285 kg C ha1 yr1) (Vernimmen, unpublished data). Matson et al. (1987) reported that glucose (but not NO3) increased denitrification rates in Costa Rica, which further illustrates the importance of the carbon source. Significant differences in denitrification rates between ectorganic layers in the two HFs were only found between the L- and F-horizons during the 2002 experiment and between the F- and H-horizons during the 2003 experiment. Laverman et al. (2000) and Raat (2005) also found

that denitrification rates were higher in the L-horizon of an acid coniferous forest. Higher pH values and the presence of fungi were suggested by Laverman et al. (2000) as being responsible for the higher denitrification rates in the L horizon. Because pH was not measured separately in the respective ectorganic layers at Barito Ulu one cannot be certain whether this could be a reason for the observed differences. Notwithstanding the fact that denitrifiers were present in the ectorganic layers, it is not likely that the denitrification process will contribute much to the overall loss of N from the HFs. Although nitrification rates were negative in the two HFs, 70–90% of available substrate was left after 48 h of incubation and the concentrations of the added substrate (0.6–1.1 mg NO3-N g1 dry weight) were much higher than the concentrations of NO3-N in litter sampled in the field (0.3–2.6 mg NO3 N g1 dry weight; Table 2). Despite the higher denitrification rates in the LERF compared to the two HFs, nitrification rates were not different under wet conditions (Table 2). As such, denitrification will only play a minor role in the availability of NO3-N in the LERF. 5.3. Effect of litter moisture content on nitrogen transformations Gravimetric moisture contents are considered to be poorly correlated with physiological availability of water or microbial activity (Tietema et al., 1992), which seems to be supported by the present findings when considering correlations for the different forests studied. There was only a significant positive correlation between N mineralization and moisture content in LERF litter during wet conditions whereas a negative correlation was found for SHF litter during dry conditions (Table 1). However, considering all data points from all three forest types together (Fig. 5), there seems to be a positive effect of gravimetric moisture content (when 4 c. 100%) on both ammonification and N mineralization, and a negative effect

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on nitrification throughout the measured moisture range, which is probably related to denitrification. Whilst studies in Costa Rica (Vitousek and Denslow, 1986; Matson et al., 1987) found some small seasonal fluctuations in N mineralization, Marrs et al. (1991) reported greater nitrogen transfers in the rainy season and lower in LERF in Brazil, with maximum mineralization during the transition period between the dry and wet seasons. Marrs et al. (1991) related these greater fluctuations to a pronounced wet and dry season cycle, something that is largely absent at the present study site in Central Kalimantan (cf. Bru¨nig, 1969). 6. Conclusion

N transformation rate [μg N g−1 d−1]

The current findings support the hypothesis that N mineralization rates are lower in HF litter than in LERF

litter, both during dry and wet conditions. Results for the two HFs of contrasting stature were not significantly different from one another. Net nitrification was very low or non-existent in all forest types and ammonification was the major constituent of nitrogen mineralization. We attribute the differences in N mineralization rates to the lower litter quality in the HFs compared to LERF litter as indicated by the much higher C:N ratio and concentrations of phenols in HF leaf litterfall. However, when DON-N was included, uptake of N was approximately the same (15–17 mg N g1 d1), suggesting that N availability is the same in all three forest studied and hence cannot be a reason for the reduced stature of HFs. All three forest types had denitrifiers present in their ectorganic horizons. However, denitrification will only play a minor role in the N-cycle because nitrification rates were very low. Gravimetric moisture conditions of litter in all

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Fig. 5. (a) Ammonification, (b) nitrification, and (c) net N-mineralization rates (mg g1 d1) against gravimetric litter moisture content in Lowland Evergreen Rain Forest (circles), tall Heath Forest (diamonds), and stunted Heath Forest (squares) at Barito Ulu, Central Kalimantan during dry (open markers) and wet (closed markers) conditions for litter samples extracted with KCl.

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