Effects of nitrogen and phosphorus addit (1)
Effects of nitrogen and phosphorus
addition on soil microbial community in a
secondary tropical forest of China
Jian Li, Zhian Li, Faming Wang, Bi Zou,
Yao Chen, Jie Zhao, Qifeng Mo, Yingwen
Li, Xiaobo Li & Hanping Xia
Biology and Fertility of Soils
Cooperating Journal of International
Society of Soil Science
ISSN 0178-2762
Biol Fertil Soils
DOI 10.1007/s00374-014-0964-1
1 23
Your article is protected by copyright and
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to self-archive your article, please use the
accepted manuscript version for posting on
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the accepted manuscript version in any
repository, provided it is only made publicly
available 12 months after official publication
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on Springer's website. The link must be
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publication is available at link.springer.com”.
1 23
Author's personal copy
Biol Fertil Soils
DOI 10.1007/s00374-014-0964-1
ORIGINAL PAPER
Effects of nitrogen and phosphorus addition on soil microbial
community in a secondary tropical forest of China
Jian Li & Zhian Li & Faming Wang & Bi Zou & Yao Chen &
Jie Zhao & Qifeng Mo & Yingwen Li & Xiaobo Li &
Hanping Xia
Received: 23 April 2014 / Revised: 17 September 2014 / Accepted: 23 September 2014
# Springer-Verlag Berlin Heidelberg 2014
Abstract Nutrient availability greatly regulates soil microbial
processes and functions in tropical forests. However, few
studies have explored the impacts of nitrogen (N) addition
(100 kg P ha − 1 year − 1 ), phosphorus (P) addition
(100 kg N ha−1 year−1), and N×P interaction on soil microbial
biomass and microbial community composition in tropical
forests. We established a field nutrient manipulation experiment in a secondary tropical forest of South China. Soil
physicochemical properties and microbial community composition were measured. Analysis of phospholipid fatty acids
(PLFAs) was used to determine soil microbial biomass and
composition, and both were related to environmental factors
by the redundancy analysis (RDA) and principal response
curves (PRC). We demonstrated that N addition usually did
not affect microbial biomass, which was increased by P addition over 3 years of fertilization. Nitrogen addition decreased
soil bacterial biomass but did not affect soil fungal biomass
after 3 years of fertilization. After P addition, soil fungal
biomass increased faster than soil bacterial biomass, indicating a more sensitive response of soil fungi to P addition than
bacteria. Phosphorus addition increased fungi/bacteria ratio
(F/B) ratios after 3 years of fertilization. Both N and P
J. Li : Z. Li : F. Wang (*) : B. Zou : Y. Chen : Q. Mo : Y. Li :
X. Li : H. Xia
Key Laboratory of Vegetation Restoration and Management of
Degraded Ecosystems, South China Botanical Garden, Chinese
Academy of Sciences, 510650 Guangzhou, People’s Republic
of China
e-mail: wangfm@scbg.ac.cn
J. Li : Y. Chen : Q. Mo
University of Chinese Academy of Sciences, 10000 Beijing, People’s
Republic of China
J. Zhao
Key Laboratory of Agro-ecological Processes in Subtropical Region,
Institute of Subtropical Agriculture, Chinese Academy of Sciences,
410125 Changsha, People’s Republic of China
additions had different effects on soil microbial community
in this tropical forest and, thus, probably altered ecosystem
functioning.
Keywords Nutrients . Fertilization . Biomass . Soil microbial
community . Tropical forest
Introduction
Nitrogen (N) and phosphorus (P) play an important role in
plant growth since they are essential nutrients for plants (Elser
et al. 2007). It is generally believed that in temperate and high
latitude ecosystems, N frequently limits plant growth and
organic matter storage (Vitousek and Howarth 1991).
However, most tropical forests occur on highly weathered
soils, where much of the original P-rich parent material has
been lost, and most of the remaining P is occluded on iron and
aluminum oxides (Miller et al. 2001). Therefore, it is generally
believed that plant growth is limited by P in tropical forests
(Walker and Syers 1976; Vitousek 1984; Vitousek and
Farrington 1997; Cleveland et al. 2011). However, a metaanalysis study suggested that N limitation was equally strong
for temperate and tropical forests (LeBauer and Treseder
2008). Recent studies also have demonstrated that both N
and P limit tropical forest plant growth (Wright et al. 2011).
These conflicting results reflect an uncertainty in the nutrient
limitation of tropical forest.
Soil microbial community plays a key role in terrestrial
ecosystem by carrying out almost all soil biochemical processes (Kennedy 1999; Artursson et al. 2006). They mineralize organic matters to simple inorganic compounds and recycle growth-limiting nutrients for autotrophs (Griffiths et al.
2012). These processes are essential for soil fertility and plant
growth. Soil microbe therefore acts as both a sink and a source
of available nutrients. The availability of essential soil
Author's personal copy
Biol Fertil Soils
nutrients (e.g., N and P) could influence the activities, biomass, and compositions of soil microbial communities
(Treseder and Allen 2002; He et al. 2008). Furthermore, recent
researches have shown that anthropogenic N deposition is
increasing dramatically in tropical regions (Hietz et al. 2011)
and P deposition may already play an important role as a
source of P in tropical regions (Sayer et al. 2012).
Understanding the effect of N and P addition on soil microbial
activities can thus improve our capacity to predict how the soil
microbial community will respond to environmental change
in tropical forest ecosystems.
The effect of N fertilization or deposition on soil microbial
biomass and community composition has been well studied
(Waldrop et al. 2004; Gallo et al. 2004; Frey et al. 2004;
Chung et al. 2007). Treseder (2008) analyzed 82 field experiment data and concluded that negative effects of N addition
on soil microbial biomass were widespread in terrestrial
ecosystems. However, the effects of N addition or deposition
on microbial diversity were inconsistent among various
studies. For example, Balser (2001) reported that alteration
of microbial community composition by N addition varied in
three Hawaiian tropical forest soils. Gallo et al. (2004) also
found that the response of functions and structure of microbial
decomposer communities to N amendment changed with
forest types. DeForest et al. (2004) reported that N addition
decreased microbial biomass without any evidence that N
addition altered microbial community composition. These
inconsistent results indicated that the response of soil microbial communities to N fertilization is highly variable among
different environments.
Compared with studies on N addition, only a few studies
focused on the effects of P addition on soil microbial biomass
and microbial community, and most of these studies were
conducted in agriculture ecosystems (Beauregard et al. 2010;
Shi et al. 2013; Tan et al. 2013). In an agriculture soil, Shi et al.
(2012) reported no significant changes on soil microbial biomass by 17 years of P fertilization. Bünemann et al. (2004)
also observed no significant difference in microbial, bacterial,
and fungal biomass under maize-crotalaria fallow rotation
with or without P fertilization. The response of soil microbial
biomass and community composition to P fertilization in
tropical forests is poorly documented and cannot get a consistent conclusion (Wang et al. 2008; Liu et al. 2012). Recent
work indicates that P addition in tropical forest can increase
soil microbial biomass and alter the composition of soil microbial community (Liu et al. 2012), but this effect has been
transient and disappearing over 4 years of fertilization (Liu
et al. 2013). It remains unclear if these changes are consistent
across various forests in tropical areas.
Although previous studies have studied well the response
of soil microbial community to N addition, only few studies
have explored impacts of N addition, P addition, and N×P
interaction on soil microbial biomass and community
composition in tropical forests. In this study, we designed a
long-term (>3 years) N and P addition experiment in a secondary tropical forest, which is facing high atmospheric N
deposition (over 40 kg N ha−1 year−1), to test the following
two hypotheses: (1) N fertilization would have negative effects on microbial biomass and composition of soil microbial
community; and (2) P fertilization would have positive effects
on the two microbial variables.
Materials and methods
Site description
This study was carried out at the Xiaoliang Tropical Coastal
Ecosystem Research Station of the Chinese Academy of
Sciences (CAS) (21°27′ N, 110° 54′ E), southwest of
Guangdong Province, China. This region is characterized by
tropical monsoon climate with a mean annual temperature of
23 °C. Annual rainfall ranges from 1400 to 1700 mm with a
variation of dry and wet seasons. The wet season is from April
to October and the dry season from November to March. The
soil is latosol developed from granite (Yu and Pi 1985).
The study was conducted in a restored mixed forest. The
forest started as Eucalyptus exserta plantation in 1959, and
then 312 plant species were introduced between 1964 and
1975 (Ding et al. 1992; Ren et al. 2007). Later, the forest
succeeded and naturally colonized species displaced almost
all planted species by the 1990s. Now, the most common tree
species are as follows: Castanopsis fissa, Cinnamomum
camphora, Carallia brachiata, Aphanamixis polystachya,
Ternstroemia pseudoverticillata, Acacia auriculaiformis,
Cassia siamea, Albizia procera, Albizia odoratissima,
Leucaena leucocephala, Aquilaria sinensis, Chakrasia
tabularis, Syzygium levinei, Schefflera heptaphylla,
Syzygium hancei, Psychotria rubra, and Aporusa dioica.
The forest is considered as typical tropical secondary forest
in regard to biodiversity and structure complexity of the forest
community.
Experimental design
An N and P addition experiment was designed as a randomized complete block (n=5) and established in the secondary
tropical forest in September 2009. Each block was located in a
site more than 50 m apart in the forest. Within each block, four
10×10 m plots were established and each plot was surrounded
by a 2 m wide buffer strip in each site. Four treatments, N
addition (+N), P addition (+P), N addition with P addition (+
NP), and control (CK) with no addition of mineral nutrients,
were carried out randomly with four plots within each block.
Both N and P were applied at 100 kg ha−1 year−1. Briefly,
476.6 g NH4NO3 (equal to 166.6 g N) and/or 808 g Na2HPO4
Author's personal copy
Biol Fertil Soils
Table 1 Soil physical and chemical characteristics of the tropical
forest before the start of fertilization in 2009
Data are expressed as means±SE
(n=5). Values are not significant
at P
addition on soil microbial community in a
secondary tropical forest of China
Jian Li, Zhian Li, Faming Wang, Bi Zou,
Yao Chen, Jie Zhao, Qifeng Mo, Yingwen
Li, Xiaobo Li & Hanping Xia
Biology and Fertility of Soils
Cooperating Journal of International
Society of Soil Science
ISSN 0178-2762
Biol Fertil Soils
DOI 10.1007/s00374-014-0964-1
1 23
Your article is protected by copyright and
all rights are held exclusively by SpringerVerlag Berlin Heidelberg. This e-offprint is
for personal use only and shall not be selfarchived in electronic repositories. If you wish
to self-archive your article, please use the
accepted manuscript version for posting on
your own website. You may further deposit
the accepted manuscript version in any
repository, provided it is only made publicly
available 12 months after official publication
or later and provided acknowledgement is
given to the original source of publication
and a link is inserted to the published article
on Springer's website. The link must be
accompanied by the following text: "The final
publication is available at link.springer.com”.
1 23
Author's personal copy
Biol Fertil Soils
DOI 10.1007/s00374-014-0964-1
ORIGINAL PAPER
Effects of nitrogen and phosphorus addition on soil microbial
community in a secondary tropical forest of China
Jian Li & Zhian Li & Faming Wang & Bi Zou & Yao Chen &
Jie Zhao & Qifeng Mo & Yingwen Li & Xiaobo Li &
Hanping Xia
Received: 23 April 2014 / Revised: 17 September 2014 / Accepted: 23 September 2014
# Springer-Verlag Berlin Heidelberg 2014
Abstract Nutrient availability greatly regulates soil microbial
processes and functions in tropical forests. However, few
studies have explored the impacts of nitrogen (N) addition
(100 kg P ha − 1 year − 1 ), phosphorus (P) addition
(100 kg N ha−1 year−1), and N×P interaction on soil microbial
biomass and microbial community composition in tropical
forests. We established a field nutrient manipulation experiment in a secondary tropical forest of South China. Soil
physicochemical properties and microbial community composition were measured. Analysis of phospholipid fatty acids
(PLFAs) was used to determine soil microbial biomass and
composition, and both were related to environmental factors
by the redundancy analysis (RDA) and principal response
curves (PRC). We demonstrated that N addition usually did
not affect microbial biomass, which was increased by P addition over 3 years of fertilization. Nitrogen addition decreased
soil bacterial biomass but did not affect soil fungal biomass
after 3 years of fertilization. After P addition, soil fungal
biomass increased faster than soil bacterial biomass, indicating a more sensitive response of soil fungi to P addition than
bacteria. Phosphorus addition increased fungi/bacteria ratio
(F/B) ratios after 3 years of fertilization. Both N and P
J. Li : Z. Li : F. Wang (*) : B. Zou : Y. Chen : Q. Mo : Y. Li :
X. Li : H. Xia
Key Laboratory of Vegetation Restoration and Management of
Degraded Ecosystems, South China Botanical Garden, Chinese
Academy of Sciences, 510650 Guangzhou, People’s Republic
of China
e-mail: wangfm@scbg.ac.cn
J. Li : Y. Chen : Q. Mo
University of Chinese Academy of Sciences, 10000 Beijing, People’s
Republic of China
J. Zhao
Key Laboratory of Agro-ecological Processes in Subtropical Region,
Institute of Subtropical Agriculture, Chinese Academy of Sciences,
410125 Changsha, People’s Republic of China
additions had different effects on soil microbial community
in this tropical forest and, thus, probably altered ecosystem
functioning.
Keywords Nutrients . Fertilization . Biomass . Soil microbial
community . Tropical forest
Introduction
Nitrogen (N) and phosphorus (P) play an important role in
plant growth since they are essential nutrients for plants (Elser
et al. 2007). It is generally believed that in temperate and high
latitude ecosystems, N frequently limits plant growth and
organic matter storage (Vitousek and Howarth 1991).
However, most tropical forests occur on highly weathered
soils, where much of the original P-rich parent material has
been lost, and most of the remaining P is occluded on iron and
aluminum oxides (Miller et al. 2001). Therefore, it is generally
believed that plant growth is limited by P in tropical forests
(Walker and Syers 1976; Vitousek 1984; Vitousek and
Farrington 1997; Cleveland et al. 2011). However, a metaanalysis study suggested that N limitation was equally strong
for temperate and tropical forests (LeBauer and Treseder
2008). Recent studies also have demonstrated that both N
and P limit tropical forest plant growth (Wright et al. 2011).
These conflicting results reflect an uncertainty in the nutrient
limitation of tropical forest.
Soil microbial community plays a key role in terrestrial
ecosystem by carrying out almost all soil biochemical processes (Kennedy 1999; Artursson et al. 2006). They mineralize organic matters to simple inorganic compounds and recycle growth-limiting nutrients for autotrophs (Griffiths et al.
2012). These processes are essential for soil fertility and plant
growth. Soil microbe therefore acts as both a sink and a source
of available nutrients. The availability of essential soil
Author's personal copy
Biol Fertil Soils
nutrients (e.g., N and P) could influence the activities, biomass, and compositions of soil microbial communities
(Treseder and Allen 2002; He et al. 2008). Furthermore, recent
researches have shown that anthropogenic N deposition is
increasing dramatically in tropical regions (Hietz et al. 2011)
and P deposition may already play an important role as a
source of P in tropical regions (Sayer et al. 2012).
Understanding the effect of N and P addition on soil microbial
activities can thus improve our capacity to predict how the soil
microbial community will respond to environmental change
in tropical forest ecosystems.
The effect of N fertilization or deposition on soil microbial
biomass and community composition has been well studied
(Waldrop et al. 2004; Gallo et al. 2004; Frey et al. 2004;
Chung et al. 2007). Treseder (2008) analyzed 82 field experiment data and concluded that negative effects of N addition
on soil microbial biomass were widespread in terrestrial
ecosystems. However, the effects of N addition or deposition
on microbial diversity were inconsistent among various
studies. For example, Balser (2001) reported that alteration
of microbial community composition by N addition varied in
three Hawaiian tropical forest soils. Gallo et al. (2004) also
found that the response of functions and structure of microbial
decomposer communities to N amendment changed with
forest types. DeForest et al. (2004) reported that N addition
decreased microbial biomass without any evidence that N
addition altered microbial community composition. These
inconsistent results indicated that the response of soil microbial communities to N fertilization is highly variable among
different environments.
Compared with studies on N addition, only a few studies
focused on the effects of P addition on soil microbial biomass
and microbial community, and most of these studies were
conducted in agriculture ecosystems (Beauregard et al. 2010;
Shi et al. 2013; Tan et al. 2013). In an agriculture soil, Shi et al.
(2012) reported no significant changes on soil microbial biomass by 17 years of P fertilization. Bünemann et al. (2004)
also observed no significant difference in microbial, bacterial,
and fungal biomass under maize-crotalaria fallow rotation
with or without P fertilization. The response of soil microbial
biomass and community composition to P fertilization in
tropical forests is poorly documented and cannot get a consistent conclusion (Wang et al. 2008; Liu et al. 2012). Recent
work indicates that P addition in tropical forest can increase
soil microbial biomass and alter the composition of soil microbial community (Liu et al. 2012), but this effect has been
transient and disappearing over 4 years of fertilization (Liu
et al. 2013). It remains unclear if these changes are consistent
across various forests in tropical areas.
Although previous studies have studied well the response
of soil microbial community to N addition, only few studies
have explored impacts of N addition, P addition, and N×P
interaction on soil microbial biomass and community
composition in tropical forests. In this study, we designed a
long-term (>3 years) N and P addition experiment in a secondary tropical forest, which is facing high atmospheric N
deposition (over 40 kg N ha−1 year−1), to test the following
two hypotheses: (1) N fertilization would have negative effects on microbial biomass and composition of soil microbial
community; and (2) P fertilization would have positive effects
on the two microbial variables.
Materials and methods
Site description
This study was carried out at the Xiaoliang Tropical Coastal
Ecosystem Research Station of the Chinese Academy of
Sciences (CAS) (21°27′ N, 110° 54′ E), southwest of
Guangdong Province, China. This region is characterized by
tropical monsoon climate with a mean annual temperature of
23 °C. Annual rainfall ranges from 1400 to 1700 mm with a
variation of dry and wet seasons. The wet season is from April
to October and the dry season from November to March. The
soil is latosol developed from granite (Yu and Pi 1985).
The study was conducted in a restored mixed forest. The
forest started as Eucalyptus exserta plantation in 1959, and
then 312 plant species were introduced between 1964 and
1975 (Ding et al. 1992; Ren et al. 2007). Later, the forest
succeeded and naturally colonized species displaced almost
all planted species by the 1990s. Now, the most common tree
species are as follows: Castanopsis fissa, Cinnamomum
camphora, Carallia brachiata, Aphanamixis polystachya,
Ternstroemia pseudoverticillata, Acacia auriculaiformis,
Cassia siamea, Albizia procera, Albizia odoratissima,
Leucaena leucocephala, Aquilaria sinensis, Chakrasia
tabularis, Syzygium levinei, Schefflera heptaphylla,
Syzygium hancei, Psychotria rubra, and Aporusa dioica.
The forest is considered as typical tropical secondary forest
in regard to biodiversity and structure complexity of the forest
community.
Experimental design
An N and P addition experiment was designed as a randomized complete block (n=5) and established in the secondary
tropical forest in September 2009. Each block was located in a
site more than 50 m apart in the forest. Within each block, four
10×10 m plots were established and each plot was surrounded
by a 2 m wide buffer strip in each site. Four treatments, N
addition (+N), P addition (+P), N addition with P addition (+
NP), and control (CK) with no addition of mineral nutrients,
were carried out randomly with four plots within each block.
Both N and P were applied at 100 kg ha−1 year−1. Briefly,
476.6 g NH4NO3 (equal to 166.6 g N) and/or 808 g Na2HPO4
Author's personal copy
Biol Fertil Soils
Table 1 Soil physical and chemical characteristics of the tropical
forest before the start of fertilization in 2009
Data are expressed as means±SE
(n=5). Values are not significant
at P