Introduction of Dalbergia Odorifera Enhances Nitrogen Absorption on Eucalyputs Through Stimulating Microbially Mediated Soil Nitrogen-cycling

doi:10.21203/rs.3.rs-683628/v1

Download PDF

Research Article

Introduction of Dalbergia Odorifera Enhances Nitrogen Absorption on Eucalyputs Through Stimulating Microbially Mediated Soil Nitrogen-cycling

https://doi.org/10.21203/rs.3.rs-683628/v1

This work is licensed under a CC BY 4.0 License

You are reading this latest preprint version

Background: There is the substantial evidence that Eucalyptus for nitrogen (N) absorption and increasing the growth benefit from the introduction of N-fixing species, but the underlying mechanisms for microbially mediated soil N cycling remains unclear.

Methods: We investigated the changes of soil pH, soil water content (SWC), soil organic carbon (SOC), total N (TN), inorganic N (NH₄⁺-N and NO₃^--N), microbial biomass and three N-degrading enzyme activities as well as the biomass and N accumulation of Eucalyptus between a pure Eucalyptus urophylla × grandis plantation (PP) and a mixed Dalbergia odorifera and Eucalyptus plantation (MP) in Guangxi Zhuang Autonomous Region, China.

Results: Compared with the PP site, soil pH, SWC, SOC and TN in both seasons were significantly higher at the MP site, which in turn enhanced microbial biomass and the activities of soil N-degrading enzymes. The stimulated microbial activity at the MP site likely accelerated soil N mineralization, providing more available N (NH₄⁺-N in both seasons and NO₃^--N in the wet-hot season) for Eucalyptus absorption. Overall, the N accumulation of Eucalyptus at the MP site was increased by 19.7% and 21.9%, promoting the biomass increases of 15.1% and 19.2% in the dry-cold season and wet-hot season, respectively.

Conclusion: Our results reveal the importance of microbially mediated soil N cycling in the N absorption on Eucalyputs. Introduction of D. odorifera can enhance N absorption and growth on Eucalyputs, improve soil N availability and increased soil C sequestration, which hence can be considered to be an effective sustainable management option of Eucalyptus plantations.

Forestry

Eucalyptus plantations

Soil physical-chemical properties

Microbial biomass

Soil enzyme activities

Nitrogen availability

Eucalyptus is one of the most extensively planted commercial plantation timber genera in the world (Lino et al. 2016). Currently, most Eucalyptus plantations are grown as monocultures and have been intensively managed in short-rotation with continuous cropping (Huang et al. 2017). However, due to high nitrogen (N) consumption with successive rotations, the monoculture plantations of Eucalyptus have rapidly depleted soil nutrients (especially N) and water (Liu et al. 1998; Sicardi et al. 2004). The massive application of N fertilizer is common to ensure high and sustainable stand production (Laclau et al. 2005), but this not only increases economic costs but also leads to soil acidification, compaction, contamination of groundwater, and other negative impacts on the environment (Goncalves et al. 1997). Consequently, seeking an optimal silvicultural practice to achieve N sustainable management of Eucalyptus plantations is in urgent need.

The N-fixing trees have been widely considered as being important in balancing N losses due to timber harvesting and in reducing the demand for fertilizer application in Eucalyptus plantations (May and Attiwill 2003). The growth of N-fixing plants relies largely on the fixed atmospheric-N that accounts for 10% − 90% of the N used by the N-fixing species (Nygren and Leblanc 2015). Therefore, introduction of N-fixing species into Eucalyptus plantations would decrease the N uptake of trees from soil, in turn increasing soil N retention (Bouillet et al. 2008; Koutika et al. 2019). In addition, the N can be transferred from N₂-fixing species to Eucalyptus, which allows Eucalyptus to benefit directly from symbiotic N fixation (Paula et al. 2015; Yao et al. 2019, 2021). In recent years, introducing N-fixing species to improve N absorption and productivity on Eucalyptus has been a popular focus of research, but the experimental results remain contradictory depending on the selected N-fixing species and stand conditions (Forrester et al. 2006; Firn et al. 2007; Bouillet et al. 2013). Moreover, because of lack of understanding the underlying mechanisms of soil N cycling, it is still hard to predict how combinations of the N-fixing species and specific sites can lead to the best benefits, making the extension of this silvicultural practice remains difficult around the world.

The advantages of Eucalyptus mixed with N₂-fixing tree species are usually attributed to the increase in soil organic N source (Paula et al. 2015; Yao et al. 2019). However, it is important to note that most of these increased organic N sources due to the introduction of N-fixing species becoming available to Eucalyptus have to be transformed to inorganic N by the microbial decomposition of the plant tissues and soil organic matter (Versini et al. 2016). Shift of microbial activity under the mixed plantation of Eucalyptus and N-fixing species would alter soil N cycling and availability, which ultimately affect the N absorption on Eucalyputs (Huang et al. 2014). Microorganisms involve soil biogeochemical cycling mainly through producing specific extracellular enzymes, which has been confirmed by extensive researches (Huang et al. 2017). Soil N-degrading enzymes including leucine aminopeptidase (LAP), β-1,4-N-acetylglucosaminidase (NAG) and Urease can serve as indicators of energy N demand (Schimel et al. 2017), which catalyze terminal reactions to depolymerize organic N (Sinsabaugh et al., 2008). Therefore, detecting the change of microbial biomass and soil N-degrading enzyme activities will help us to better understand the underlying mechanisms for microbially mediated soil N cycling under the mixed plantations of Eucalyptus and N-fixing species.

Eucalyptus has been introduced and cultivated in south China since the 1970s, and the area of Eucalyptus plantations has reached 4.6 million ha by 2014, of which almost half in Guangxi Zhuang Autonomous Region, China (Zhao et al. 2018). In recent years, a variety of N-fixing species such as Acacia mangium and D. odorifera were introduced to improve N absorption and productivity on Eucalyptus planations in Guangxi (Huang et al. 2014; Yao et al. 2019). Here we conducted a mixed plantation of Eucalyptus (E. urophylla × E. grandis) and D. odorifera (MP) and a Eucalyptus pure plantation (PP) in a long-term field experiment. Previous work in pot experiment has observed a percentage of 6.6–13.6% N transfer from D. odorifera to Eucalyptus and an increase of 20.4–33.2% in the dry matter yields of Eucalyptus compared to the monoculture plantation (Yao et al. 2019). In this study, we investigated the changes of soil pH, soil water content (SWC), soil organic carbon (SOC), total N (TN), inorganic N (NH₄⁺-N and NO₃⁻-N), microbial biomass and three N-degrading enzyme activities as well as the biomass and N accumulation of Eucalyptus between PP and MP sites. We would focus on the role of microbial function in driving soil N availability in the dry-cold and wet-hot seasons. Specifically, we hypothesized that the introduction of D. odorifera would increase soil organic matter (SOC and TN) and improve soil environment (pH and SWC), which in turn stimulated microbial activities (biomass and N-degrading enzyme activities) and improved N availability (NH₄⁺-N and NO₃⁻-N), ultimately enhancing Eucalyptus N absorption and productivity.

Experimental design

The study site was located at the Experimental Center of Tropical Forestry, Chinese Academy of Forestry (22°07′ N, 106°93′ E), Pingxiang City, Guangxi Zhuang Autonomous Region, China. The mean annual precipitation is around 1400 mm, falling mainly from April through September (Wang et al., 2010) and mean annual temperature is 21°C. The soils were formed from granite, classified as red soil in Chinese soil classification, equivalent to oxisol in USDA Soil Taxonomy.

In March 2015, a complete randomized block design with three replicates was established to compare a pure E. urophylla × grandis plantation (PP) and a mixed species plantation of D. odorifera and E. urophylla × grandis (MP), the proportion in the mixed plantation stands was 1:1 (50E:50D). Three sampling plots (15 m × 15 m) were randomly established within each of the stand type, a 200 m buffer separates the PP and MP plantations. The two species were planted alternately at 2 m spacing in the row, with 2.5 m between rows, giving a total stocking density of 2000 trees ha^− 1. The fertilizers applied on planting were 140 g N plant^− 1 (urea: CO(¹⁴NH₂)₂, buried at 20 cm from the each plant), as well as 18 g plant^− 1 K (KCl), 56 g P plant^− 1 (CaH₂PO₄).

Sample of plants collected and N analysis

Five plants in each plot were harvested on March 20 and August 25, 2019, which including below-ground (i.e., roots) and above-ground. The trees of above-ground were separated into components: leaves, branches (living branches and dead branches), stem wood and stem bark. The stem of each tree was sawn into 2 m sections according to Monsic’s stratified clip method (You et al. 2018). However, much of the root growth was shallow and lateral, it was difficult to distinguish the fine roots of one tree from those of another. To address this problem, a large soil pit of 1.5 m diameter and 1.0 m depth was excavated around each target tree. All the materials were collected and weighed up fresh weight immediately, and then 500 g of every composition sample was taken in laboratory.

The harvested material was dried at 65°C until constant dry weight period for the biomass analyses. The dried plant material was ground in a ball mill (< 0.1 mm) for the N concentration analyses with a continuous-flow chemical analyzer (AA3) after the degradation using 10 ml H₂SO₄. The biomass and N accumulation of E. urophylla × grandis in the pot was the sum of leaves, stem, branches, bark and roots.

Sample of soils collected and analysis

On March 19 and August 24, representing the dry-cool season and the wet-hot seasons of the year 2019, respectively, five soil samples were randomly collected from each plot at 0–20 cm depth. During sampling within a plot, the corer was wiped clean of obvious soil particles with paper towel. All samples were stored at -20℃and sieved < 2 mm to remove visible stones, soil animals, roots and plant materials prior to analysis pH, ammonium (NH₄⁺-N) and nitrate nitrogen (NO₃⁻-N). Some of the air dried and ball milled sample (sieved < 0.2 mm) were used for the concentration of TN, soil microbial biomass carbon (MBC) and nitrogen (MBN) and SOC analyses.

The pH of the soil samples was measured in a 1:2.5 soil/water suspension. SOC was determined by dichromate oxidation and titration with ferrous ammonium sulfate, the TN concentration was determined by a continuous flow chemical analyzer (AA3) and followed by detection of NH₄⁺-N and NO₃⁻-N. Data was also collected on C/N ratio of soil (C/Nsoil).

MBC and MBN were measured by fumigation-extraction, using 0.5 M K₂SO₄ as the extraction agent (Vance et al. 1987), with a total organic carbon analyzer (1020A; OI, College Station, TX, USA), and were calculated by (Vance et al. 1987).

MBC = E_C/k_EC (1)

Where E_C= (organic C extracted from fumigated soils) - (organic C extracted from non-fumigated soils), and k_EC= 0.38.

MBN = E_N/k_EN (2)

Where E_N= (organic N extracted from fumigated soils) - (organic N extracted from non-fumigated soils), and k_EN= 0.45.

In addition, soil microbial function was expressed by the soil extracellular enzyme activities, involved NAG, LAP and Urease, respectively. We determined the soil enzyme activities using the conventional p-nitrophenol (pNP) assays (Baldrian, 2009) and we took the following specific operation steps: (1) weight 1.25 g fresh soil and add 12.5 mL of 50 mM Sodium acetate buffer, then extract it with using low-energy sonication for 2 min as the soil sample to be tested; (2) take 200 ul soil sample into place microplates and added the 50 ml stock solution to analysis the activity of LAP (stock solution: 200 µM of the L-Leucine-7-amido-4-Methylcoumarin hydrochloride), NAG (stock solution: 200 µM of the 4-MethylumbelliferylN-7-acetyl-β-D-glucosaminide) and Urease (20 mM of the urea), respectively; (3) and then place microplates were incubated for 4 hours in the darkness at 25℃; (4) enzyme assays were performed with a fully automatic full-wavelength enzyme labeling apparatus for an excitation wavelength of 355 nm and an emission wavelength of 460 nm, slit width of 25 nm. Enzyme activities were assayed in duplicate with one control, to which substrate was added after incubation and subtracted from the sample value. The enzyme activities were measured by 3 replicates for each soil sample and expressed as µmol per gram dry soil and incubation time.

Statistical analyses

Statistical analyses were performed using SPSS software (SPSS Inc., Chicago, IL, USA). Analysis of Variance (ANOVA) was used to determine the statistical significance (α = 0.05) of tand type, season and their interactive effect on N accumulation and biomass of Eucalyptus as well as soil physico-chemical properties and microbial activity. Tukey's multiple comparison test (HSD) was conducted if significant effects of stand type or season was found. Pairwise relationships between plant N accumulation, soil physico-chemical properties and microbial activities were tested using Pearson correlation coefficients.

A structural equation modelling (SEM) approach was also used to test a conceptual model for microbially mediated soil N cycling in the dry-cold and the wet-hot seasons. The SEM analysis was performed with the IBM SPSS Amos 20.0 using the maximum likelihood estimation method. Several tests were used to assess model fit: the Chi-square (c2)-test, comparative fit index (CFI) and root square mean error of approximation (RMSEM).

The biomass and the N content of E. urophylla × grandis

The biomass of Eucalyptus was significantly by stand type, with greater at the MP than the PP sites in both seasons (P < 0.05). At both PP and MP sites, the biomass of Eucalyptus in stems was greater than that of the other organs in both seasons (Fig. 1a and b). The N accumulation of the whole plant was significantly affected by season, stand type and their interaction (Table S1). Similar to biomass, the N accumulation of Eucalyptus was also significantly greater at the MP than the PP sites in the both seasons (Fig. 1c and d). However, the distribution of N accumulation in different organs was different from that of biomass. Generally, the N accumulation of branch was greater than that of the other organs except for the stems in the both seasons.

Soil physico-chemical properties

Soil pH had no significant difference between the dry-cold and wet-hot season, but was significantly higher at the MP site than at the PP site (Table 1). The concentrations of SOC, TN, NO₃⁻-N and NH₄⁺-N as well as the ratio of SOC to TN (C/N) and SWC were significantly affected by stand type and season. SOC, TN, NH₄⁺-N and SWC were significantly higher at the MP than the PP sites and in the wet-hot season than dry-cold season (p < 0.05 for all, Table 1). The soil NO₃⁻-N concentration at the MP site was higher than that at the PP site in the wet-hot season only (Table 1). Compared with the PP site, soil C/N ratio was significantly decreased at the MP site in both seasons (Table 1).

Table 1

Soil Physical chemical properties as affected by introduction N₂-fixing trees and seasons in *Eucalyptus* plantations. PP = pure *Eucalyptus* plantation; MP = mixed with N₂-fixing species and *Eucalyptus* plantation.
Source of variation		pH	SWC	SOC	TN	NO₃⁻-N	NH₄⁺-N	C/N
Dry-cold	MP	4.83 ± 0.12 a	18.63 ± 1.43 b	21.49 ± 3.25 b	1.34 ± 0.13 b	4.22 ± 0.46 b	16.99 ± 3.17 b	16.09 ± 2.03 b
Dry-cold	PP	4.56 ± 0.27 b	16.83 ± 0.86 c	18.96 ± 2.81 c	1.04 ± 0.15 c	4.09 ± 0.36 b	13.55 ± 2.01c	18.78 ± 3.14 a
Wet-hot	MP	4.91 ± 0.16 a	21.48 ± 1.63 a	24.70 ± 3.62 a	1.56 ± 0.15 a	5.94 ± 0.88 a	17.46 ± 1.48 a	16.29 ± 3.69 b
Wet-hot	PP	4.60 ± 0.14 b	19.99 ± 1.11 b	20.85 ± 2.57 b	1.12 ± 0.10 c	4.25 ± 0.27 b	13.59 ± 1.56 c	18.15 ± 0.94 a
Season (S)		ns	64.832^***	25.48^***	100.852^***	87.659^***	144.709^***	ns
Stand type (T)		7.844^*	19.332^***	18.631^***	9.330^**	48.121^***	24.905^***	7.844^*
S × T		ns	ns	ns	8.522^**	50.178^***	13.667^***	ns

Microbial biomass and enzymes activities

Two-way ANOVAs showed that MBC and MBN were significantly affected by stand type and season (p < 0.05, Fig. 2), but no significant interactive effect was found (p > 0.05, Fig. 2). The MBC and MBN were significantly higher at the MP than the PP sites and in the wet-hot season than dry-cold season (Fig. 2a and b). However, the ratio of MBC/MBN did not significantly differ between the PP and MP sites in the both seasons (Fig. 2c). All of the soil N-degrading enzyme activities were also significantly affected by stand type and season (p < 0.05, Fig. 3). Specifically, only the activities of NAG and Urea in the wet-hot season were significantly higher (p < 0.05) at the MP than the PP sites (Fig. 3).

Relationships of soil microbial biomass and enzymes activities with soil and plant N pools

Soil pH did not correlated with any other properties in the dry-cold season, while in the wet-hot season it was positively correlated with SWC, SOC, TN, NAG activity and NO₃⁻-N (Table 2). SWC was positively correlated with MBC, the activities of NAG and Urea, and the N accumulation of Eucalyptus in the dry-cold season, and with SOC, TN, NAG activity, NH₄⁺-N, NO₃⁻-N and the N accumulation of Eucalyptus in the wet-hot season, respectively (Table 2). SOC also did not correlated with any other properties in the dry-cold season, but in the wet-hot season it was positively correlated with TN, MBC, all of N-degrading enzyme activities, NH₄⁺-N, NO₃⁻-N and the N accumulation of Eucalyptus in the wet-hot season. Soil TN content was positively correlated with MBN and the N accumulation of Eucalyptus in the dry-cool season, and with MBC, the activities of NAG and Urea, NO₃⁻-N and the N accumulation of Eucalyptus in the wet-hot season, respectively (Table 2). MBC and MBN were positively correlated with all of the N-degrading enzyme activities in the wet-hot season expect for the relationship between MBN and LAP activity (Table 2). MBC was also positively correlated with NH₄⁺-N, NO₃⁻-N and the N accumulation of Eucalyptus in the wet-hot season (Table 2). NAG activity was positively correlated with NH₄⁺-N, NO₃⁻-N and the N accumulation of Eucalyptus in both seasons (Table 2). LAP activity was positively correlated with NH₄⁺-N and the N accumulation of Eucalyptus in the wet-hot season but not in the dry-cold season, while Urea activity was positively correlated with NH₄⁺-N and the N accumulation of Eucalyptus in the dry-cold season but not in the wet-hot season (Table 2). Both NH₄⁺-N and NO₃⁻-N were positively correlated with the N accumulation of Eucalyptus in the wet-hot season, while in the dry-cold season only the NH₄⁺-N was positively correlated with the N accumulation of Eucalyptus (Table 2).

Table 2

The collection among the soil physical-chemical properties, soil enzymes, soil microbial biomass and N content in the plants
Source of variation	pH	SWC	SOC	TN	MBC	MBN	LAP	NAG	Urease	NH₄⁺-N	NO₃⁻-N	PN
pH		ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns
SWC	0.976^**		ns	ns	0.813^*	ns	ns	0.895^*	0.899^*	ns	ns	0.946^**
SOC	0.824^*	0.857^*		ns	ns	ns	ns	ns	ns	ns	ns	ns
TN	0.836^*	0.87^*	0.835^*		ns	0.864^*	ns	ns	ns	ns	ns	0.818^*
MBC	ns	ns	0.888^*	0.968^**		ns	ns	ns	ns	ns	ns	ns
MBN	ns	ns	ns	ns	ns		ns	ns	ns	ns	ns	ns
LAP	ns	ns	0.866^*	ns	0.812^*	ns		ns	ns	ns	ns	ns
NAG	0.889^*	0.947^**	0.931^**	0.908^*	0.978^**	0.825^*	0.908^**		0.994^**	0.96^**	0.858^*	0.962^**
Urease	ns	ns	0.850^*	0.855^*	0.866^*	0.859^*	ns	0.861^**		0.935^**	ns	0.967^**
NH₄⁺-N	ns	0.859^*	0.883^*	ns	0.894^*	ns	0.94^**	0.946^**	ns		0.897^*	0.877^*
NO₃⁻-N	0.902^*	0.910^*	0.947^**	0.951^**	0.952^**	ns	ns	0.928^*	ns	0.831^*		ns
PN	ns	0.883^*	0.873^*	0.909^**	0.976^**	ns	0.894^*	0.984^**	ns	0.960^**	0.893^*
SOC = soil organic carbon, TN = soil total nitrogen, NH₄⁺-N = ammonium nitrogen, NO₃⁻-N = nitrate nitrogen, PN = plant nitrogen content, MBC = microbial biomass carbon, MBN = microbial biomass nitrogen, LAP = Leucine aminopeptidase, NAG = β-1, 4-N-acetylglucosaminidase. The values in the blue area are in the wet-hot season and in the yellow areas are in the dry-cool season. ns, ^* and ^** = F-value not significant, significant at P > 0.05 and P < 0.05 and P < 0.01, respectively.

Pathway analysis for Eucalyptus to enhance N absorption

The structural equation model on the regulatory pathway of soil N dynamics well passed all the statistical tests on adequacy of microbial biomass and enzyme activities (Fig. 4a and 4b). Overall, soil pH, SWC, SOC and TN in both seasons were significantly higher at the MP than PP sites, which together stimulated microbial biomass and most of soil N-related enzyme activities although the control mechanisms are different between seasons. The enhanced microbial activity likely accelerated soil N mineralization, providing more available N (NH₄⁺-N and NO₃⁻-N) for Eucalyptus absorption. Path analysis pointed to direct and positive controls of soil N availability (NH₄⁺-N and NO₃⁻-N) by the activity of soil N-related enzymes in both seasons but the control by microbial biomass in the wet-hot season only. Soil environment positively corrected with microbial biomass in the dry-cold season only. SOC and TN positively corrected with both microbial biomass and enzyme activity in the dry-cold season and there was also a positive relationship between microbial biomass and enzyme activity. In the wet-hot season, SOC and TN did not directly affect soil enzyme activity, but have an indirect effect via influencing microbial biomass.

Mixed plantations of Eucalyptus and N-fixing species have been recognized as one of effective silvicultural practices to increase Eucalyptus productivity while maintaining soil fertility, compared to Eucalyptus monocultures (Epron et al. 2013; Tchichelle et al. 2017). For example, introducting Acacia mangium into E. grandis provides better conditions for restoring the soil fertility and soil biodiversity, hence providing better sustainability of the cropping systems than pure E. grandis plantations (Garay et al. 2004). However, the results reported previously remain contradictory depending on the selected N-fixing species and stand conditions (Forrester et al. 2006; Firn et al. 2007; Bouillet et al. 2013). In this study, we discovered that the N absorption and productivity of Eucalyputs as well as soil TN and N availability were significantly enhanced by introducting D. odorifera into the Eucalyptus plantations. This suggests that mixed plantations of Eucalyptus with D. odorifera could be considered as an effective sustainable management option in Guangxi Zhuang Autonomous Region, southern China.

The enhanced N absorption on Eucalyputs with introduction of D. odorifera was likely attributed to the improved soil N availability, as N accumulation of Eucalyputs was directly correlated with both soil NH₄⁺-N and NO₃⁻-N concentrations (Table 2). Using structural equation model, we found that the higher soil organic matter (SOC and TN) and improved soil environment (pH and SWC) could indirectly enhance soil N availability via stimulating microbial activity (microbial biomass and enzyme activity) (Fig. 4). Increased soil N storage under the mixed plantation of Eucalyptus and N₂-fixing species is common, as the growth of N-fixing plants relies largely on the fixed atmospheric-N (Paula et al. 2018). Therefore, introduction of N-fixing species into Eucalyptus plantations would decrease the N uptake of trees from soil, in turn increasing soil N retention (Bouillet et al. 2008). Faster growth of N-fixing tree species and Eucalyptus due to higher N availability at the MP sites could also increase the input of plant residuals and hence enhance SOC and TN. Similar results were also found in other studies with a mixed plantation such as E. globulus and A. mearnsii in Australia (Forrester et al. 2005), Eucalyptus grandis and Acacia mangium in Brazil (Bini et al. 2013a, 2013b) and in Congo (Koutika et al. 2019, 2020), or Eucalyptus regnans and Acacia dealbata in southeastern Australia (Pfautsch et al. 2009). Taken together, the higher SOC ,TN and inorganic N at the MP sites (Table 1) indicate that mixed-species plantation not only improved soil N availability, aiming at preventing soil degradation; but also increased soil C sequestration, contributing to mitigating climate change.

The greater soil microbial biomass at the MP than PP sites (Fig. 2) suggests that the presence of leguminous trees in the system could improve stand conditions for microbial proliferation (Bini et al. 2013a), while the driving factors differed between the seasons in this study. In the dry-cold season, soil microorganisms are usually subject to water limitation. Thus, increased SWC under the MP site could stimulate microbial biomass. This was supported by the positive relationship between MBC and SWC in the dry-cold season observed in this study (Table 2). The positive relationship between TN and MBN in the dry-cold season (Table 2), suggests that increased TN under the MP site could alleviate microbial N limitation and enhance microbial N assimilation. This in turn may explain why enhanced microbial biomass under the MP site did not directly contribute to higher N availability in the dry-cold season. In the wet-hot season, both temperature and SWC are relatively higher at our study sites, thus increased SWC and pH under the MP site did not significantly influence microbial biomass (Table 2). Alternatively, we found direct and positive effects of SOC and TN on microbial biomass, suggesting that microbial biomass in the wet-hot season was mainly limited by substrate availability in this study. Introduction of D. odorifera into Eucalyputs plantation significantly increased MBC but not MBN in the wet-hot season, thus having a direct and strong contribution to soil N availability. This was supported by the positive relationships of MBC with NH₄⁺-N or NO₃⁻-N in the wet-hot season observed in this study (Table 2).

Microbial communities involve soil biogeochemical cycling primarily through producing specific extracellular enzyme (Bowles et al. 2014). Increased microbial biomass usually enhanced N-degrading extracellular enzyme production in the soil. Increased TN could also stimulate the activity of N-degrading enzymes as providing more N resource for microbial decomposition. We have observed positive relationships of MBC with the activities of all N-degrading extracellular enzymes in the wet-hot season, while in the dry-cold season the enzyme activity was controlled by both TN and microbial biomass (Fig. 4). Previous studies have also reported that hydrolytic enzyme activities increased (Huang et al. 2017) with increasing soil TN and MBC since N-fixing trees were introduced into the Eucalyputs plantations (Bini et al. 2013b; Huang et al. 2017). It is well known that increasing in N-degrading enzyme activities reflect a higher rate of N mineralization and a higher level of N availability (Tabatabai et al. 2010). Although we have not determined the N mineral processes in this study, previous studies have frequently found that introducing N-fixing trees into Eucalyptus plantations can stimulate microbial activity (Pereira et al. 2018, 2019) to increase the rate of N mineralization (Voigtlaender et al. 2012, 2019). Alternatively, we indeed found NH₄⁺-N was positively correlated with the N accumulation of Eucalyptus in both seasons, while the similar relationship was only occurs in the wet-hot season for NO₃⁻-N. This result is consistent with the observation that the growth of many forest plant species depend on ammonium (Eiter 1972).

Compared with the Eucalyptus monoculture, we found that the N absorption and productivity of Eucalyptus were significantly increased by introduction of D. odorifera into the Eucalyptus plantation. Introduction of D. odorifera into the Eucalyptus plantation could also improve soil N availability and increase soil C and N storage. Moreover, our results highlight the importance of microbially mediated soil N cycling in the N absorption on Eucalyputs. These findings in turn can provide government and policy makers with useful tools to achieve N sustainable management in Eucalyptus plantations.

TN: total nitrogen, SOC: soil organic carbon, SWC: soil water content, PP: pure E. urophylla × grandis plantation, MP: a mixed species plantation of D. odorifera and E. urophylla × grandis, MBC: soil microbial biomass carbon, MBN: soil microbial biomass nitrogen, LAP: leucine aminopeptidase, NAG: β-1,4-N-acetylglucosaminidase.

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Availability of data and materials

Data available on request from the authors.

Competing interests

The authors declare that they have no competing interests.

Funding

This work was supported by the National Natural Science Foundation of China (No. 31460196 and 31870461), the Innovation Project of Guangxi Graduate Education (NO. YCBZ2018012), the “Hundred Talent Program” of South China Botanical Garden at the Chinese Academy of Sciences (No. Y761031001), and the “Young Top-notch Talent” in Pearl River talent plan of Guangdong Province, and by the Science (No. 2019QN01L763).

Acknowledgements

All who contributed towards the article who does not meet the criteria for authorship including anyone who provided professional writing services or materials have acknowledged.

The authors are grateful to Junfei Xiong at Chinese Academy of Forestry, Guangxi, China, for collecting the soils, harvesting and measuring of the plants in the field. We are also grateful to Dafeng Hui at Tennessee State University, Nashville, TN, USA, for his valuable comments and English improvements.

Authors’ contributions

All the authors designed the experiments; Xianyu Yao, Zhi Nong and Qianchun Zhang carried out the experiments; Xianyu Yao, Shaoming Ye and Qi Deng analyzed the experimental results, Xianyu Yao and Haiju Zhou analyzed the data and developed analysis tools; Xianyu Yao, Shaoming Ye and Qi Deng wrote the manuscript, and all authors read and approved the final manuscript.

Author details

¹Key Laboratory of Vegetation Restoration and Management of Degraded Ecosystems, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou, Guangdong 510650, China. ²Center for Plant Ecology, Core Botanical Gardens, Chinese Academy of Sciences, Guangzhou 510650, China. ³State Key Laboratory of Conservation and Utilization of Subtropical Agro-bioresources, College of Forestry, Guangxi University, Daxue East Road 100, Nanning, Guangxi 530004, China. ⁴Experimental Center of Topical Forestry, Chinese Academy of Forestry, Pingxiang, Guangxi 532600,

China.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Baldrian P (2009) Microbial enzyme-catalyzed processes in soils and their analysis. Plant Soil Environ 55:370–378
Bini D, Figueiredo AF, da Silva MCP, Vasconcellos RLdeF, Cardoso EJBN (2013b) Microbial biomass and activity in litter during the initial development of pure and mixed plantations of Eucalyptus grandis and Acacia mangium. Rev Bras Ciência do Solo 37:76–85
https://doi.org/10.1590/S0100-06832013000100008
Bini D, Santos CAd, Bouillet J-P, Goncalves JLM, Cardoso EJBN (2013a) Eucalyptus grandis and Acacia mangium in monoculture and intercropped plantations: Evolution of soil and litter microbial and chemical attributes during early stages of plant development. Appl Soil Ecol 63: 57–66. https://doi.org/10.1016/j.apsoil.2012.09.012
Bouillet JP, Laclau JP, Goncalves JLM, Moreira MZ, Trivelin P, Jourdan C, Galiana A (2008) Mixed-species plantations of Acacia mangium and Eucalyptus grandis in Brazil 2. N accumulation in the stands and N₂ biological fixation. Forest Ecol Manag 255(12):3918–4393. https://doi:10.1016/j.foreco.2007.10.050
Bouillet J-P, Laclau J-P, Gonçalves JLM, Voigtlaender M, Gava JL, Leite FP, Hakamada R, Mareschal L, Mabiala A, Tardy F, Levillain J, Deleporte P, Epron D, Nouvellon Y (2013) Eucalypt and Acacia tree growth over entire rotation in single-and mixed-species plantations across five sites in Brazil and Congo. Forest Ecol Manag 301:89–101. https://doi.org/10.1016/j.foreco.2012.09.019
Bowles TM, Acosta-Martínez V, Calderón F, Jackson LE (2014) Soil enzyme activities, microbial communities, and carbon and nitrogen availability in organic agroecosystems across an intensively-managed agricultural landscape. Soil Biol Biochem 68:252–262
http://dx.doi.org/10.1016/j.soilbio.2013.10.004
Eiter HM (1972) Effect of nitrogen nutrition upon sugar content and dry weight of juvenile lodgepole pine and white spruce. Can J For Res 2:434–440
Epron D, Nouvellon Y, Mareschal L, Moreira RM, Koutika L-S, Geneste B, Delgado-Rojas JS, Laclau J-P, Sola G, Gonçalves JLM, Bouillet J-P (2013) Partitioning of net primary production in Eucalyptus and Acacia stands and in mixed-species plantations: Two case-studies in contrasting tropical environments. Forest Ecol Manag 301:101–111
https://dx.doi.org/10.1016/j.foreco.2012.10.034
Firn J, Erskine PD, Lamb D (2007) Woody species diversity influences productivity and soil nutrient availability in tropical plantations. Oecologia 154:521–533. https://10.1007/s00442-007
-0850-8
Forrester DI, Bauhus J, Cowie AL, Vanclay JK (2006) Mixed-species plantations of Eucalyptus with N-fixing trees: A review. Forest Ecol Manag 233(2):211–230
https://doi.org/10.1016/j.foreco.2006.05.012
Forrester DI, Bauhus J, Cowie AL (2005) On the success and failure of mixed-species tree plantations: lessons learned from a model system of Eucalyptus globulus and Acacia mearnsii. Forest Ecol Manag 209:147–155. https://doi:10.1016/j.foreco.2005.01.012
Garay I, Pellens R, Kindel A, Barros E, Franco AA (2004) Evaluation of soil conditions in fast-growing plantations of Eucalyptus grandis and Acacia mangium in Brazil: a contribution to the study of sustainable land use. Appl Soil Ecol 27:177–187. https://doi.org/10.1016/j.apsoil
03.007
Goncalves JLM, Barros NF, Nambiar EKS, Novais RF (1997) Soil and stand management for short-rotation plantations. In: Nambiar, E.K.S., Brown, A.G. (Eds.), Management of Soil, Nutrients and Water in Tropical Plantation Forests. Australian Centre for International Agricultural Research, Canberra 379–418
Huang XM, Liu SR, Wang H, Hu Z, Li ZG, You YM (2014) Changes of soil microbial biomass carbon and community composition through mixing N-fixing species with Eucalyptus Urophylla in subtropical China. Soil Biol Biochem 73:42–48. http://dx.doi.org/10.1016/j.soilbio.2014.01.021
Huang X, Liu S, You Y, Wen YG, Wang H, Wang JX (2017) Microbial community and associated enzymes activity influence soil carbon chemical composition in Eucalyptus urophylla plantation with mixing N₂-fixing species in subtropical China. Plant Soil 414:199–212
https://10.1007/s11104-016-3117-5
Koutika L-S, Cafiero L, Bevivino A, Merino A (2020) Organic matter quality of forest floor as a driver of C and P dynamics in acacia and eucalypt plantations established on a Ferralic Arenosols, Congo. Forest Ecosyst 7:40. https://doi.org/10.1186/s40663-020-00249-w
Koutika L-S, Ngoyi S, Cafiero L, Bevivino A (2019) Soil organic matter quality along rotations in acacia and eucalypt plantations in the Congolese coastal plains. Forest Ecosyst 6:39
https://doi.org/10.1186/s40663-019-0197-8
Laclau J-P, Ranger J, Deleporte P, Nouvellon Y, Saint-André L, Marlet S, Bouillet J-P (2005) Nutrient cycling in a clonal stand of Eucalypt and an adjacent savanna ecosystem in Congo 3. Input–output budgets and consequences for the sustainability of the plantations. Forest Ecol Manag 210:375–391
Lino IAN, Santos VM, Escobar IEC, Silva DKA, Araújo ASF, Maia LC (2016) Soil enzymatic activity in Eucalyptus grandis plantations of different ages. Land Degrad Develop 27:77–82
https://doi.10.1002/ldr.2454
Liu SR, Li XM, Niu LM (1998) The degradation of soil fertility in pure larch plantation in the northeastern part of China. Ecol Eng 10:75–86
May BM, Attiwill PM (2003) Nitrogen-fixation by Acacia dealbata and changes in soil properties 5 years after mechanical disturbance or slash-burning following timber harvest. Forest Ecol Manag 181: 339–355. https://doi.org/10.1016/S0378-1127(03)00006-9
Nygren P, Leblanc HA (2015) Dinitrogen fixation by legume shade trees and direct transfer of fixed N to associated cacao in a tropical agroforestry system. Tree Physiol 35:134–137
https://doi.org/10.1093/treephys/tpu116
Paula RR, Bouillet J-P, Gonçalves JLM, Trivelin PCO, Balieiro FC, Nouvellon Y, Oliveira JC, Júnior JCD, Bordron B, Laclau J-P (2018) Nitrogen fixation rate of Acacia mangium Wild at mid rotation in Brazil is higher in mixed plantations with Eucalyptus grandis Hill ex Maiden than in monocultures. Ann Forest Sci 75:14. https://doi.org/10.1007/s13595-018-0695-9
Paula RR, Bouillet J-P, Trivelin PCO, Zeller B, Gonçalves JLM, Nouvellon Y, Bouvet J-M, Plassard C, Laclau J-P (2015) Evidence of short-term belowground transfer of nitrogen from Acacia mangium to Eucalyptus grandis trees in a tropical planted forest. Soil Biol Biochem 91: 99–108. https:// dx.doi.org/10.1016/j.soilbio.2015.08.017
Pereira APA, Durrer A, Gumiere T, Gonçalves José LM, Robin A, Bouillet JP, Wang J, Verma JP, Singh BK, Cardoso EJBN (2019) Mixed Eucalyptus plantations induce changes in microbial communities and increase biological functions in the soil and litter layers. Forest Ecol Manag 433:332–342. https://doi.org/10.1016/j.foreco.2018.11.018
Pereira APA, Zagatto MRG, Brandani CB (2018) Acacia changes microbial indicators and increases C and N in soil organic fractions in intercropped Eucalyptus plantations. Front Microbiol 9:1–13. https://doi.org/10.3389/fmicb.2018.00655
Pfautsch S, Rennenberg H, Bell TL, Adams MA (2009) Nitrogen uptake by Eucalyptus regnans and Acacia spp. - preferences, resource overlap and energetic costs. Tree Physiol 29:389–399. https://doi.org/10.1093/treephys/tpn033
Schimel J, Becerra CA, Blankinship J (2017) Estimating decay dynamics for enzyme activities in soils from different ecosystems. Soil Biol Biochem 114:5–11
https://doi.org/10.1016/j.soilbio.2017.06.023
Sicardi M, García-Pérchac F, Frioni L (2004) Soil microbial indicators sensitive to land use conversion from pastures to commercial Eucalyptus grandis (hill ex maiden) plantations in Uruguay. Appl Soil Ecol 27(2):125–133. https://doi.org/10.1016/j.apsoil.2004.05.004
Sinsabaugh RL, Lauber CL, Weintraub MN, Ahmed B, Allison SD, Crenshaw C, Contosta AR, Causack D, Frey S, Gallo ME, Gartner TB, Hobbie SE, Holland K, Keeler BL, Powers JS, Stursova M, Takacs-Vesbach C, Wallenstein MD, Zak DR, Zeglin LH (2008) Stoichiometry of soil enzyme activity at global scale. Ecol Lett 11:1252–1264
https://doi.org/10.1111/j.1461-0248.2008.01245.x
Tabatabai MA, Ekenler M, Senwo ZN (2010) Significance of enzyme activities in soil nitrogen mineralization. Commun Soil Sci Plan 41(5):595–605
https://doi.org/10.1080/00103620903531177
Tchichelle S, Mareschal L, Koutika L-S, Epron D (2017) Biomass production, nitrogen accumulation and symbiotic nitrogen fixation in a mixed species plantation of eucalypt and Acacia on a nutrient-poor tropical soil. Forest Ecol Manag 403:103–111.
https://dx.doi.org/10.1016/j
foreco.2017.07.041
Vance ED, Brookes PC, Jenkinson DS (1987) An extraction method for measuring soil microbial biomass C. Soil Biol Biochem 19:703–707. https://doi.org/10.1016/0038-0717(87)90052-6
Versini A, Laclau J-P, Mareschal L, Plassard C, Diamesso LA, Ranger J, Zeller B (2016) Nitrogen dynamics within and between decomposing leaves, bark and branches in eucalyptus planted forests. Soil Biol Biochem 101:55–64. http://dx.doi.org/10.1016/j.soilbio.2016.06.034
Voigtlaender M, Brandani CB, Caldeira DRM, Tardy F, Bouillet JP, Gonçalves JLM, Moreira MZ, Leite FP, Brunet D, Paula RR, Laclau J-P (2019) Nitrogen cycling in monospecific and mixed-species plantations of Acacia mangium and Eucalyptus at 4 sites in Brazil. Forest Ecol Manag 436: 56–67. https://doi.org/10.1016/j.foreco.2018.12.055
Voigtlaender M, Laclau JP, Gonalves JLDM, Piccolo MDC, Moreira MZ, Nouvellon Y, Ranger J, Bouillet J-P (2012) Introducing Acacia mangium trees in Eucalyptus grandis plantations: consequences for soil organic matter stocks and nitrogen mineralization. Plant Soil 352(1–2): 99–111. https://doi.10.1007/s11104-011-0982-9
Wang H, Liu SR, Mo JM, Wang JX, Makeschin F, Wolff M (2010) Soil organic carbon stock and chemical composition in four plantations of indigenous tree species in subtropical China. Ecol Res 25: 1071–1079. https://doi.10.1007/s11284-010-0730-2
Yao XY, Liao LN, Huang YZ, Fan G, Yang M, Ye SM (2021) The physiological and molecular mechanisms of N transfer in Eucalyptus and Dalbergia odorifera intercropping systems using root proteomics. BMC Plant Biol 21:201
https://doi.org/10.1186/s12870-021-02969-9
Yao XY, Li YF, Liao LN, Sun G, Wang HX, Ye SM (2019) Enhancement of nutrient absorption and interspecific nitrogen transfer in a Eucalyptus urophylla × eucalyptus grandis and Dalbergia odorifera mixed plantation. Forest Ecol Manag 449:117465
https://doi.org/10.1016/j.foreco.2019.117465
You Y, Huang X, Zhu H, Liu S, Liang H (2018) Positive interactions between Pinus massoniana and Castanopsis hystrix species in the uneven-aged mixed plantations can produce more ecosystem carbon in subtropical China. Forest Ecol Manag 410:193–200
http://dx.doi.org/10.1016/j.foreco.2017.08.025
Zhao BC, Xu JD, Chen BY, Cao XF, Yuan TQ, Wang SF, Charlton A, Sun R-C (2018) Selective precipitation and characterization of lignin-carbohydrate complexes (LCCs) from Eucalyptus. Planta 247(5):1077–1087. https://doi.org/10.1007/s00425-018-2842-9

Supplementaryfile.docx

Download PDF

Editorial decision: Major Revision
23 Jul, 2021
Review #2 received at journal
22 Jul, 2021
Review #1 received at journal
20 Jul, 2021
Reviewer #2 agreed at journal
11 Jul, 2021
Reviews received at journal
08 Jul, 2021
Reviewers invited by journal
08 Jul, 2021
Editor invited by journal
08 Jul, 2021
Editor assigned by journal
08 Jul, 2021
Reviewer #1 agreed at journal
07 Jul, 2021
Submission checks completed at journal
07 Jul, 2021
First submitted to journal
02 Jul, 2021

You are reading this latest preprint version

Introduction of Dalbergia Odorifera Enhances Nitrogen Absorption on Eucalyputs Through Stimulating Microbially Mediated Soil Nitrogen-cycling

Status:

Version 1

Abstract

Figures

Background

Materials And Methods

Experimental design

Sample of plants collected and N analysis

Sample of soils collected and analysis

Statistical analyses

Results

Soil physico-chemical properties

Microbial biomass and enzymes activities

Relationships of soil microbial biomass and enzymes activities with soil and plant N pools

Pathway analysis for Eucalyptus to enhance N absorption

Discussion

Conclusion

Abbreviations

Declarations

Ethics approval and consent to participate

Consent for publication

Availability of data and materials

Competing interests

Funding

Acknowledgements

Authors’ contributions

Author details

Declaration of competing interest

References

Supplementary Files

Status:

Version 1