全 文 ：第 50 卷 第 6 期
2 0 1 4 年 6 月
林 业 科 学
SCIENTIA SILVAE SINICAE
Vol. 50，No. 6
Jun．，2 0 1 4
doi:10．11707 / j．1001-7488．20140615
Ｒeceived date: 2014 － 01 － 10; Ｒevised date: 2014 － 04 － 21．
Foundation project: National Sci-Tech Support Plan，China (2012BAD21B04) ．
* Corresponding author: Xing Shiyan．
氮添加对银杏幼林土壤有机碳化学组成
及土壤微生物群落的影响*
张晓文 邢世岩 吴岐奎 刘晓静
(山东农业大学林学院 泰安 271018)
摘 要: 采用磷脂脂肪酸法通过田间试验研究氮添加对银杏人工幼龄林土壤有机碳化学组分、微生物生物量与
群落结构的影响，施氮水平设置为 0 kg·hm － 2 a － 1 N(对照)，50 kg·hm － 2 a － 1 N(低氮)，100 kg·hm － 2 a － 1 N(中氮)及 150
kg·hm － 2 a － 1 N(高氮)，试验周期为 1 年。结果表明:高氮处理显著降低土壤有机碳含量及氧烷基碳组分但增加烷
基碳与氧烷基碳比值。不同施氮水平对土壤微生物量及群落结构的影响不同，总体上中、高氮处理显著影响土壤
微生物量与群落结构组成。中、高氮处理显著降低土壤微生物生物量碳含量但增加土壤微生物生物量氮含量，同
时显著降低土壤总微生物量、真菌生物量及细菌生物量;而且中、高氮处理显著降低革兰氏阳性菌的相对丰度，但
增加真菌细菌比和放线菌的相对丰度。另外，高氮处理显著降低土壤 pH 及土壤呼吸。
关键词: 银杏; 幼龄林; 氮添加; 土壤化学性质; 土壤微生物群落
中图分类号: S714; Q938. 1 文献标识码: A 文章编号: 1001 － 7488(2014)06 － 0115 － 10
Effects of Nitrogen Addition on Chemical Composition of Soil Organic Carbon and
Soil Microbial Community in a Young Ginkgo biloba Plantation
Zhang Xiaowen Xing Shiyan Wu Qikui Liu Xiaojing
( School of Forestry，Shandong Agricultural University Tai’an 271018)
Abstract: To determine the effects of nitrogen deposition on chemical composition of soil organic carbon ( SOC) and
microbial community characteristics in a young subtropical plantation，a field N addition experiment was established in a
young plantation of Ginkgo biloba in China． The experiment was conducted from May 2012 to May 2013 by bimonthly
application of ammonia nitrate solution with four treatments: Control (0 kg·hm － 2 a － 1 N)，Low-N (50 kg·hm － 2 a － 1 N)，
Medium-N (100 kg·hm － 2 a － 1 N) and High-N (150 kg·hm － 2 a － 1 N) ． High-N treatment significantly decreased soil organic
carbon (SOC) concentration and the proportion of O-alkyl but increased Alkyl ∶ O-alkyl ratio． Moreover，N addition
significantly affected soil microbial biomass and altered differentially soil microbial community composition with the amount
of N loaded． Medium-N and High-N treatments significantly decreased microbial biomass carbon ( C) concentration but
increased microbial biomass nitrogen ( N ) concentration． Besides，Medium-N and High-N treatments significantly
decreased soil total microbial biomass，fungal biomass，and bacterial biomass． Additionally，Medium-N and High-N
treatments significantly decreased the relative abundance of Gram-positive bacterial PLFAs and increased fungal: bacterial
ratio as well as the relative abundance of actinomycetes PLFAs． Furthermore，High-N treatment significantly decreased soil
pH，and soil respiration． Our results suggest that increased nitrogen deposition could significantly affect soil microbial
biomass and community composition with the alterations of some soil chemical properties in the studied young plantation of
G． biloba． This study could provide important scientific reference for the mechanism of soil microbial response to nitrogen
deposition in young G． biloba plantations of subtropical China．
Key words: Ginkgo biloba; young plantation; N addition; soil chemical properties; soil microbial community
Nitrogen (N) inputs from anthropogenic sources
are currently estimated to be 30% to 50% greater than
those from natural terrestrial sources and tenfold greater
than anthropogenic inputs from 100 years ago (Fang et
林 业 科 学 50 卷
al．，2011 ) ． In Asia， emissions of reactive N have
increased dramatically，leading to deposition of 30 － 73
kg·hm － 2 a － 1 N in some subtropical forests of southern
China ( Fang et al．，2011 ) ． Increased nitrogen ( N)
deposition caused by human activities has altered
ecosystem functioning and biodiversity． Soil microbial
communities sustain many vital ecosystem processes，
such as nutrient cycling， decomposition of organic
matter and waste，nutrient availability，degradation of
pesticides and contaminants，soil structure，and plant
growth and health ( van Elsas et al．，2006) ． Assessing
the effect of nitrogen deposition on microbial community
structure，diversity and activity in soil is therefore
critical to advancing the understanding of the
functionality，stability，and resilience of managed and
natural ecosystems (Kennedy et al．，1995) ． So better
understanding of microbial compositional and
physiological acclimation mechanisms is critical for
predicting terrestrial ecosystem responses to global
change (Bi et al．，2012) ．
In recent years，numerous studies on the effects of
N addition or simulated N deposition on soil microbes
have been conducted both in the field and laboratory
across different forest or plantation type， however，
most of the related studies focused on the mature
plantation or forest (Wallenstein et al．，2006; Cusack
et al．，2011; Wu et al．，2013; Hu et al．，2010 )，
whereas few studies focused on the young plantations．
In addition，these studies have not shown consistent
effects of N addition on microbial biomass and
community structure， with positive， negative， and
neutral effects being reported． For example， across
temperate forests，Nilsson et al． (2007) reported that
N input had no effect on total fungal biomass in the
soils of oak forests along a natural N deposition
gradient，but Demoling et al． (2008) and Fraterrigo et
al． ( 2006 ) found that fungal biomarkers were
decreased in soils of N-fertilized plots． In contrast，
Gallo et al． ( 2004 ) reported that fungal biomarkers
increased under N addition in north temperate forest
soils． In tropical forest，Cusack et al． (2011) found
that soil microbial biomass increased in response to N
fertilization in both distinct tropical rain forests，but
Liu et al． ( 2013 ) found that nitrogen additions
decreased soil microbial biomass in the short term but
returned to pre-treatment levels over the long term after
four years N addition in a tropical forest of subtropical
China． A common element to these studies is that they
were carried out on mature forests or plantation．
Nitrogen deposition is increasing in non-agricultural
area including forest ecosystem and many previous
studies have proven that nitrogen deposition
significantly affects soil and plants． In addition，
amounts of newly-built plantations were widely
distributed after clear-cutting． Less is known，
however，about how N addition alters on chemical
composition of soil organic carbon ( SOC ) and
microbial community characteristics in a young
subtropical plantation．
Ginkgo biloba ( Ginkgoaceae )， a famous and
widely cultivated plant，is often referred to as a living
fossil ( Yan et al．，2009 ) ． In addition，G． biloba
which is a deciduous gymnosperm species，is the only
remaining species of the once large order Ginkgoales，
with geological records indicating this plant has been
growing on the Earth for 150 － 200 million years，and it
is widely used for street and landscape trees ( He
et al．，2009; Ling et al．，2003) ． However，few studies
focused on the effect of global change such as nitrogen
deposition on G． biloba plantation，especially on soil
microbes． In the present study，to acquire an insight
into the mechanisms of soil microbial response to
nitrogen deposition in subtropical young plantation of
G． biloba， a one-year long field simulated nitrogen
deposition experiment was established． Based on
relevant expectations arising from previously reported
studies on other subtropical plantations， we
hypothesized that: 1) N addition would affect chemical
composition of SOC． 2 ) High-N addition would
significantly decrease soil microbial biomass; 3 ) N
addition would significantly influence soil microbial
community composition regardless of the amount of N
loaded．
1 Materials and methods
1. 1 Study site
The study site was situated in a young G． biloba
(6-year-old) plantation (5 hm2 ) at Xing’an county，
Guangxi province (25°55N，110°24E )，China，and
has a subtropical monsoon and humid climate． The
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第 6 期 张晓文等: 氮添加对银杏幼林土壤有机碳化学组成及土壤微生物群落的影响
mean annual precipitation is 1 842 mm，about 65% of
which occurs from March to September． The annual
average relative humidity is 76% and the mean annual
temperature is 18. 6℃ with the lowest and highest
monthly average temperature of 14. 5 ℃ and 28. 5 ℃ ．
The main ground cover species include Cibotium
barometz，Blechnum orientale， Lophatherum gracile，
Miscanthus floridulu and Ottochloa nodosa． The soil is
classified as Ｒed soil，which predominantly derived
from the granite weathering．
1. 2 Field experimental treatment and sampling
Twelve plots ( 20 m × 20 m ) were randomly
staked out with an at least 15 m buffer in the G． biloba
nursery garden． The experiment included four
treatments with 3 replicates for each treatment: 1 )
Control (without any N addition applied，C); 2) 50
kg·hm － 2 a － 1 N ( Low-N ); 3 ) 100 kg·hm － 2 a － 1 N
(Medium-N); 4) 150 kg·hm － 2 a － 1 N(High-N) ． The
rates of N addition referred to the relevant studies also
conducted in plantations of subtropical China，
especially in Dinghu Mountain Biosphere Ｒeserve
(DHSBＲ ) ( Mo et al．，2007; 2005; Zhang et al．，
2008)，which was relatively near to the studied site in
this paper． N additions ( applied as NH4NO3 ) were
initiated in May 2012 and applied bimonthly over the
course of one year at the rate of 0 g (Control)，952. 38
g (Low-N)，1 904. 77 g (Medium-N) and 2 857. 15 g
(High-N) for each N application ( Liu et al．，2013) ．
Before N addition，NH4NO3 was weighed，dissolved in
20 L distilled water and applied to each plot below the
canopy using a backpack sprayer (Mo et al．，2007) ．
Each control plot received 20 L of water without N
addition as well． During the experiment，nurse and
management were not conducted in order to exclude the
anthropogenic disturbance．
Soil samplings were conducted in May 2013 ．
From each plot， 10 soil cores ( 3. 5 cm inner
diameter) were collected randomly from a 10 cm soil
depth and compounded to one composite sample．
After removing impurities，soils were sieved to 2 mm
mesh size and divided into two parts，one part was
retained for measuring soil chemical properties and the
other was frozen at － 20 ℃ as soon as possible for
analysis of soil microbial biomass and community
structure．
1. 3 Measurements
Soil moisture content was measured by oven-
drying method using 20 g of sampled soil oven-dried at
105 ℃ for 24 h． Soil pH was measured in a 1∶ 2. 5 soil
/ water suspension． SOC was measured by dichromate
oxidation and titration with ferrous ammonium sulfate
( Lu， 2000 ) ． Total N，NH4
+ -N and NO3
－ -N in
filtered 2 mol·L － 1 KCl - extracts of fresh soil sample
were measured with a flow injection auto analyzer
( FIA ) ( Lachat Quik-Chem 8000， Lachat
Instruments，USA ) ． Available P concentration was
analyzed colorimetrically after acidified ammonium
persulfate digestion ( Anderson et al．， 1993 ) ．
Chemical composition of SOC was determined using
solid-state 13 C nuclear magnetic resonance ( NMＲ )
spectroscopy ( Bruker BioSpin Gmbh， Karlsruhe，
Germany) ．
Soil respiration was measured using the static
chamber and gas chromatography techniques， gas
samples were taken with a 100 mL plastic syringe at 0，
10，20，and 30 min after chamber closure (Mo et al．，
2007 ) ． CO2 concentrations were determined using an
Agilent 7890A gas chromatograph．
Soil microbial biomass carbon (MBC) and microbial
biomass N ( MBN ) were estimated by chloroform
fumigation-extraction． MBC and MBN (mg·kg － 1 ) were
calculated according to Wu et al． (1990) and Joergensen
et al．，1990)，respectively．
Soil microbial biomass and community structure
were determined using phospholipid fatty acid (PLFA)
analysis as described by Bossio et al． ( 1998 ) ．
Concentrations of each PLFA were calculated based on
the 19∶ 0 internal standard concentrations． The relative
abundance of individual fatty acid was expressed as the
proportion ( mol% ) of the sum of all fatty acids．
Gram-positive bacteria were identified by the PLFAs:
i14: 0，i15: 0，i16: 0，i17: 0，a15: 0，a17: 0，
Gram-negative bacteria were identified by the PLFAs:
cy17:0，15:0 3OH，16:1 2OH ( Liu et al．，2013;
Frostegrd et al．，1996) ． The fungi were identified by
the PLFAs: 18: 1ω9c ( Myers et al．，2001 )， and
PLFAs 16:1ω5c were used as a marker for arbuscular
mycorrhizal fungi (AMF) (Olsson，1999) ． The ratio
of fungal-to-bacterial PLFAs (18:1ω9c / i14:0，i15:
0，a15:0，i16:0，i17:0，a17:0，cy17:0) was used
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林 业 科 学 50 卷
as an indicator of changes in the relative abundance of
these two microbial groups ( Cao et al．，2010 ) ． The
actinomycetes were identified by the PLFAs 10Me 17:
0 and 10Me 18: 0 (Zak et al．，1996 ) ．
1. 4 Statistical analyses
One-way ANOVAs and Duncan’s test were used
to determine statistical significant differences ( P ＜
0. 05) in soil chemical characteristics，soil microbial
variables among different N addition treatments，which
was performed using SPSS software package 19. 0 for
Windows． Additionally， correlations between soil
chemical properties and microbial variables were
determined using the Pearson’s correlation
coefficients． The composition of soil microbial
community was summarized using a principle
component analysis (PCA) on the relative abundances
(mol% ) of 23 PLFAs in each sample． Furthermore，
the relationship between soil microbial community
composition and soil chemical properties was
determined using Ｒedundancy analysis ( ＲDA ) ． In
addition， soil chemical properties were tested for
significant contribution to the explanation of the
variation in soil microbial community composition with
the Monte Carlo permutation test ( P ＜ 0. 05) ． PCA
and ＲDA were conducted using CANOCO software for
Windows 4. 5 (Microcomputer Power，Inc．，Ithaca，
NY ) ． The figures were plotted by Origin Pro 9. 0
(Origin Lab Corporation，Northampton，MA，USA) ．
2 Ｒesults
2. 1 Soil chemical properties and chemical
compositions of SOC with N additions
By the time of sampling in May 2013，Medium-N
and High-N addition significantly altered soil chemical
properties ( Tab． 1 ) ． High-N treatment significantly
decreased soil pH by 12. 5%，SOC concentration in
High-N addition plots was significantly lower
(18. 1% ) than that in the Control plots (P ＜ 0. 05) ．
Moreover，concentrations of soil total N，NH4
+ -N and
NO3
－ -N significantly increased in Medium-N and
High-N treatment plots in comparison with the Control
plots ( P ＜ 0. 05 ) ． Soil available P and total P
concentrations were not significantly influenced by N
additions (P ＞ 0. 05) (Tab． 1) ．
Tab． 1 Soil chemical properties after one-year N addition①
T pH(H2 O)
SOC /
( g·kg － 1 )
Avail． P /
(mg·kg － 1 )
Total P /
( g·kg － 1 )
Total N /
( g·kg － 1 )
NH4
+ -N /
(mg·kg － 1 )
NO3
－ -N /
(mg·kg － 1 )
C 4. 55 ± 0. 04a 15. 70 ± 0. 78a 2. 04 ± 0. 24a 0. 19 ± 0. 01a 0. 98 ± 0. 09c 13. 59 ± 1. 59c 8. 72 ± 0. 81c
L 4. 44 ± 0. 12a 14. 59 ± 0. 49ab 1. 98 ± 0. 32a 0. 18 ± 0. 02a 1. 01 ± 0. 09bc 16. 76 ± 2. 28c 11. 68 ± 1. 13c
M 4. 33 ± 0. 43ab 13. 76 ± 2. 47ab 1. 70 ± 0. 37a 0. 28 ± 0. 06a 1. 21 ± 0. 12ab 23. 97 ± 2. 29b 16. 76 ± 0. 94b
H 3. 98 ± 0. 08b 12. 86 ± 0. 92b 1. 63 ± 0. 16a 0. 21 ± 0. 08a 1. 32 ± 0. 11a 29. 04 ± 1. 76a 21. 68 ± 3. 44a
①Values are mean ± standard errors for the sampling plots ( n = 3 ) ． T: Treatments; C: Control; L: Low-N; M: Medium-N; H: High-N．
Significant differences (P ＜ 0. 05，Duncan’s test) among treatments are indicated by different letters．
SOC of all treatments included four carbon
functional groups: Alkyl-C ［( 0 － 47 ) × 10 － 6］，O-
alkyl C［(47 － 112 ) × 10 － 6］，Aromatic-C［( 112 －
165) × 10 － 6］ and Carbonyl-C［(165 － 215) × 10 － 6］．
High-N treatment led a significant decline in the
proportion of O-alkyl C by 6. 1% and increase in the
ratio of Alkyl-C and O-alkyl C by 6. 1% (P ＜ 0. 05)
( Tab． 2 ) ． Other variables including Alkyl-C，
Aromatic-C， Hydrophobic-C， Hydrophilic-C and
Hydrophobic-C /Hydrophilic-C were not significantly
influenced by N addition (P ＞ 0. 05) (Tab． 2) ．
Tab． 2 Comparisons of chemical composition of SOC among treatments①
Variables Control Low-N Medium-N High-N
Alkyl-C (% ) 22. 2 ± 1. 6a 23. 1 ± 1. 0a 22. 5 ± 1. 8a 22. 4 ± 0. 8a
O-alkyl C (% ) 45. 6 ± 1. 3a 45. 9 ± 1. 4a 43. 8 ± 0. 9ab 42. 8 ± 0. 6b
Aromatic-C (% ) 20. 3 ± 0. 9a 18. 9 ± 1. 1a 21. 5 ± 1. 3a 20. 5 ± 0. 9a
Carbonyl-C (% ) 11. 9 ± 1. 6a 12. 1 ± 0. 9a 12. 2 ± 0. 8a 14. 3 ± 1. 9a
Alkyl-C /O-alkyl C 0. 49 ± 0. 01b 0. 50 ± 0. 01b 0. 51 ± 0. 01ab 0. 52 ± 0. 01a
Hydrophobic-C(% ) 42. 6 ± 2. 6a 42. 1 ± 1. 8a 43. 6 ± 2. 1a 42. 9 ± 1. 5a
Hydrophilic-C(% ) 57. 4 ± 2. 9a 58. 1 ± 3. 2a 56. 1 ± 2. 1a 57. 1 ± 3. 2a
Hydrophobic-C /Hydrophilic-C 0. 74 ± 0. 02a 0. 71 ± 0. 05a 0. 77 ± 0. 01a 0. 76 ± 0. 02a
①Hydrophobic-C = Alkyl-C + Aromatic-C，Hydrophilic-C = O-alkyl C + Carbonyl-C; Values are mean ± standard errors． Significant differences
(P ＜ 0. 05，Duncan’s test) among treatments are indicated by different letters．
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第 6 期 张晓文等: 氮添加对银杏幼林土壤有机碳化学组成及土壤微生物群落的影响
2. 2 Soil microbial biomass and activity
Medium-N and high-N treatments significantly
decreased MBC concentration and increased MBN
concentration relative to the Control treatment ( P ＜
0. 05 ) ( Fig． 1a， b ) ． Soil respiration in High-N
treatment plot ［(43. 2 ± 1. 2) mg·m － 2 h － 1 CO2-C］
was 10. 3% lower compared with the Control plots
［(48. 1 ± 2. 7 ) mg·m － 2 h － 1 CO2-C］( Fig． 2 ) ． In
addtion total microbial biomass，bacterial biomass and
fungal biomass in Medium-N and High-N treatment
plots decreased compared to the Control plot ( P ＜
0. 05) (Fig． 3a，b，c) ．
Fig． 1 Microbial biomass C ( a) and Microbial biomass N ( b)
in soil samples after one-year N additions
Significant differences ( P ＜ 0. 05， Duncan’s test ) among
treatments are indicated by different letters． Error bars show
standard errors ( n = 3) ．
2. 3 Soil microbial community structure
The ratio of fungi to bacterial PLFAs was
significantly increased by high-N treatment ( 9. 5% )
compared to the control plots (P ＜ 0. 05) ( Fig． 3d) ．
Futhermore， Medium-N and High-N treatments
significantly decreased the relative abundance of gram-
positive bacterial PLFAs by an average of 4. 7% and
8. 5%， but significantly increased the relative
abundance of actinomycetes PLFAs by an average of
8. 0% and 7. 0% relative to the control plots ( P ＜
0. 05) (Fig． 4) ．
Principal Components Analysis ( PCA ) of the
Fig． 2 Comparisons of soil respiration among treatments
Values are means for three months (March 2013 to May 2013) ．
Significant differences ( P ＜ 0. 05，Duncan’s test ) among
treatments are indicated by different letters． Error bars show
standard errors ( n = 9) ．
microbial community composition， defined by the
PLFA profile using 23 individual PLFAs，demonstrated
that the first two axes explained 36. 6% and 19. 6% of
the total variation in microbial communities ( Fig． 5a，
5b)， respectively． According to the patterns in the
PCA plot，the Control treatment samples were so close
to the Low-N treatment samples，showing that Low-N
treatment did not significantly affect soil microbial
community compositions relative to the Control
treatment ( Fig． 5a ) ． In addition， Medium-N and
High-N treatment samples were located in different
quadrants and clearly separated from the Control and
Low-N treatment samples by PC2， indicating that
Medium-N and High-N treatments significantly altered
the microbial community structure， the main reason
might be their lower relative abundance of gram-
positive bacterial PLFAs and higher relative abundance
of actinomycetes PLFAs under N addition compared to
the Control samples (Fig． 5a，Fig． 4) ．
For individual PLFAs， Gram-positive bacterial
PLFAs such as a15: 0，i15: 0 and i16: 0 got higher
scores on PC1，other PLFAs such as 18: 0 2OH also
got higher scores on PC1． While on PC2， Gram-
negative bacterial PLFAs such as cy17: 0 and 16: 1
2OH got higher scores． Additionally， other PLFAs
such as Gram-positive bacterial PLFAs ( i14: 0，i17:
0 and a17: 0) and AMF PLFAs biomarker 16: 1ω5c
got higher scores on PC2 as well (Fig． 5b) ．
2. 4 Soil microbial community structure and soil
chemical properties
The correlations between soil microbial community
and soil chemical properties were analyzed by ＲDA and
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林 业 科 学 50 卷
Fig． 3 Comparisons of soil total PLFAs ( a)，bacterial PLFAs ( b)，fungal PLFAs ( c)，
and F∶ B ( d) among treatments after one-year N addition
F∶ B: the ratio of fungal to bacterial PLFAs． Significant differences ( P ＜ 0. 05，Duncan’s test) among treatments are
indicated by different letters． Error bars show standard errors ( n = 3) ．
Fig． 4 Ｒelative abundances of the individual
PLFAs (mol% ) in soil samples
G + : The proportion of gram-positive bacterial PLFAs． G － : The
proportion of gram-negative bacterial PLFAs; Fungal: The
proportion of fungal PLFAs． Actino． : The proportion of
actinomycetes PLFAs． AM: The proportion of AM fungal PLFAs．
Significant differences ( P ＜ 0. 05， Duncan’s test ) among
treatments are indicated by different letters． Error bars show
standard errors ( n = 3) ．
the significance of environmental variables present in
the ordination was determined by Monte Carlo
permutation tests (P ＜ 0. 05) ． The results showed that
seven soil variables，including soil pH，SOC，NH4
+，
NO3
－，TN，available P and total P，explained 48. 5%
of the variation in soil microbial community composition
(Fig． 6 ) ． Soil microbial community composition was
Fig． 5 Phospholipid fatty acid (PLFA) pattern in soil
samples from different N addition plots ( a) and PC
loadings of the individual PLFAs ( b)
Values are means ( n = 3) with bidirectional error bars of axis 1 and 2．
For both the PCA plots，values on the PC1 and PC2 axes represent the
percent variation explained by PC1 and PC2，respectively．
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第 6 期 张晓文等: 氮添加对银杏幼林土壤有机碳化学组成及土壤微生物群落的影响
significantly related to NO3
－ (F = 4. 09，P = 0. 002)
and total P (F = 2. 03，P = 0. 028)，with ＲDA axis1
explaining 33. 6% of the variance and ＲDA axis 2
explaining 14. 9%，respectively (Fig． 6) ． The Control
plot samples were almost clearly separated from
Medium-N and High-N treatment samples by ＲDA axis
2． In addition，NO3
－ and total P showed a negative
association with ＲDA axis 1 (Fig． 6) ．
The proportional abundance of gram-positive
bacterial PLFAs was positively correlated with pH，SOC
and C /N， and negatively correlated with Total N，
NH4
+ -N and NO3
－ -N (P ＜ 0. 01) (Tab． 3) ． For the
proportional abundance of gram-negative bacterial
PLFAs，it was negatively correlated with pH，SOC，
Avail． P and C /N (P ＜ 0. 05) (Tab． 3) ． In addition，
the proportional abundance of fungal PLFAs was
positively correlated with C /N ( P ＜ 0. 05 ) and
negatively correlated with Total P，Total N，NH4
+ -N
and NO3
－ -N ( P ＜ 0. 05) ( Tab． 3 ) ． Moreover，the
proportional abundance of actinomycetes PLFAs was
positively correlated with Total N，NH4
+ -N and NO3
－ -N
(P ＜ 0. 01)，but negatively correlated with pH，SOC
and C /N ( Tab． 3 ) ． Furthermore， the proportional
abundance of AMF PLFAs was not significantly
correlated with soil chemical properties ( P ＞ 0. 05 )
(Tab． 3) ．
Fig． 6 Ｒedundancy Analysis (ＲDA) results of soil microbial
community composition and soil chemical properties
In ＲDA plots，values on the x and y axes represent the percent
variation explained by ＲDA axis 1 and ＲDA axis 2，respectively
(P ＜ 0. 05) ． SOC and TN refer to soil organic carbon and soil total
nitrogen，respectively．
Tab． 3 Pearson’s correlation coefficients between the proportional abundance of
main microbial communities and soil chemical properties①
Community pH SOC Avail． P Total P Total N NH4
+ -N NO3
－ -N C /N
G + (mol% ) 0. 682 * 0. 690 * 0. 279 － 0. 193 － 0. 807＊＊ － 0. 893＊＊ － 0. 882＊＊ 0. 815＊＊
G － (mol% ) － 0. 628 * － 0. 629 * － 0. 658 * 0. 319 0. 445 0. 344 0. 406 － 0. 582 *
Fungal (mol% ) 0. 573 0. 414 0. 400 － 0. 638 * － 0. 650 * － 0. 643 * － 0. 642 * 0. 599 *
Actino． (mol% ) － 0. 721＊＊ － 0. 671 * － 0. 480 0. 260 0. 859＊＊ 0. 815＊＊ 0. 839＊＊ － 0. 857＊＊
AMF (mol% ) － 0. 391 － 0. 244 － 0. 252 0. 482 0. 035 0. 046 － 0. 034 － 0. 094
①G + : The proportion of gram-positive bacterial PLFAs; G － : The proportion of gram-negative bacterial PLFAs; Fungal: The proportion of fungal
PLFAs; Actino． : The proportion of actinomycetes PLFAs; AMF: The proportion of AM fungal PLFAs． * and ＊＊ denote significant correlations at P ＜
0. 05 and P ＜ 0. 01，respectively．
3 Discussions and conclusions
3. 1 The effects of N addition on soil organic
carbon and soil pH
In this study， High-N addition significantly
decreased soil organic carbon content ( Tab． 1 ) ． N
fertilization has been observed to stimulate ecosystem C
loss ( Liu et al．， 2009 ) and such a decline was
consistent with a recent report by Wei et al． (2011) ．
A possible mechanism is that N addition stimulated
SOC decomposition more than plant production，
leading to a net loss of ecosystem C (Mack et al．，
2004)，and another one is that higher atmospheric N
deposition resulted in higher C loss by increasing
heterotrophic respiration and dissolved organic carbon
leaching ( Bragazza et al．，2006 ) ． In addition，N
addition have significant effects on C inputs processes
from plants， such as amount of litterfall and litter
decomposition (Knorr et al．，2005)，fine root biomass
( Wallenstein et al．， 2006 ) and root exudation
(Bowden et al．，2004)，which were closely related to
soil organic carbon content dynamics．
High-N treatment significantly decreased soil pH
(Tab． 1)，such a finding was consistent with the result
from a N addition study on larch plantation (Hu et al．，
2010) ． A decline in soil pH may be resulted from
NH4
+ uptake by plants，nitrification of NH4
+ in soils，
and NO3
－ leaching ( Matthew et al．，2006 ) ． In a
121
林 业 科 学 50 卷
mature subtropical forest of China，Lu et al． (2009)
reported that the ecosystem was sensitive to high N
addition， and soil acidification was significantly
enhanced after continuous two-year N additions (50 －
150 kg·hm － 2 a － 1N) ． Furthermore，Fan et al． (2007)
found that exchangeable base cations ( e． g． Ca2 + and
Mg2 + ) reduced with increasing N addition in a
subtropical Chinese fir plantation after three years of N
addition ( 60 － 240 kg·hm － 2 a － 1 N )， the nitrate
leached out was accompanied by positively charged
“counter-ions”，the base cations K +，Ca2 + and Mg2 +，
resulting in the further acidification of the leached soil，
or hydrogen and aluminum ions，which may cause the
acidification of receiving ecosystems．
3. 2 Effects of N addition on soil microbial
biomass and community composition
Medium-N and High-N additions significantly
decreased MBC concentration in this study (Fig． 1a) ．
Decreased soil organic carbon and pH may be the main
reason of reduced MBC， according to the related
studies by Sarathchandra et al． ( 2001 ) and
Wallensteina et al． (2006) ． Additionally，Medium-N
and High-N application significantly increased
microbial biomass N in the present study ( Fig． 1b)，
mainly because soil microbial biomass serves as both an
important source and sink for plant available nutrients，
and N availability was increased after N addition and
consequently immobilized by microbes，which led to an
increase in soil microbial biomass N and a decline in
nutrient loss ( Garcia et al．， 1994; Wang et al．，
2008) ．
Medium-N and High-N additions significantly
decreased total microbial biomass，bacterial biomass
and fungal biomass，these findings are corresponding
with the results obtained in field studies ( Fig． 3 )，
where N fertilization decreased soil microbial biomass
by an average of 11% to 35% (Ｒamirez et al．，2012) ．
One explanation is the decrease in soil acidity，soil pH
is a major factor influencing the structure of the soil
microbial community and individual microbial PLFAs，
the strongest influence of soil pH was on the fungal and
bacterial growth ( Bth et al．，2003; Ｒousk et al．，
2010)，so the declines of bacterial biomass and fungal
biomass in this study may be due to the higher growth
inhibition of Medium-N and High-N additions to
bacterial and fungal communities，consequently leading
a decline in soil total biomass． In addition， soil C
availability and plant belowground C allocation play
dominant roles in regulating soil microbial population
size and community composition． The influences of N
addition on soil microbial biomass and community
structure are strongly linked to changes of plant
production and belowground C supply ( Hu et al．，
2010)，so another possible mechanism to explain the
reduction of soil microbial biomass is the changes of
belowground C under N additions． In the present
study，High-N treatment significantly increased the
ratio of fungal to bacterial，such a result was consistent
with a recent report from a comparable pH gradient in
an arable soil，where the fungal∶ bacterial growth ratio
also increased 50-fold between pH 8. 3 and 4. 0
(Ｒousk et al．，2009) ． In addition，Liu et al． (2013)
reported that F ∶ B ratios were significant higher in the
N-addition plots comparing to the control plots after
four years N addition in a tropical forest of subtropical
China．
In the present study，Medium-N and High-N
additions significantly altered the soil microbial
community structure ( Fig． 5a ) ． Allison et al．
(2008) summarized the results of studies previously
reported on the soil microbial community composition
response to the mineral fertilization ( N /P /K )，and
the majority of these studies demonstrate that
composition is sensitive to disturbance． More than
80% of the mineral fertilization ( N /P /K ) studies
found significant effects of disturbance on microbial
composition． And this study proved that soil microbial
community composition in a young plantation of
Ginkgo biloba was significantly sensitive to Medium-N
and High-N additions．
3. 3 Effect of N addition on soil respiration
High-N treatment led a significant reduction in
soil respiration after 12 months N addition in this study
( Fig． 2 )， which was also found in other forest
ecosystems ( Mo et al．，2007; Burton et al．，2004;
Chen et al．，2002) ． Ｒamirez et al． (2012) found that
decreases in soil respiration by N addition ranged
between 8% and 15% ． The reductions in soil
respiration after nitrogen additions may be possibly
related to the following mechanisms: 1 ) N additions
decreased autotrophic respiration from plant roots，
decrease in soil respiration was related to the reduction
in standing root biomass and root exudation in N-
fertilized plots ( Bowden et al．， 2004 ); 2 )
221
第 6 期 张晓文等: 氮添加对银杏幼林土壤有机碳化学组成及土壤微生物群落的影响
Heterotrophic respiration from the microbial community
may be reduced by N addition，which was indicated by
that the decomposition rates of litter and soil organic
matter were suppressed (Mo et al．，2007; Jiang et al．，
2010) ． In the present study， soil respiration as an
indices of soil microbial activity was measured for three
months only to determine the difference of soil
microbial activity among treatments rather than the
dynamics of CO2 emission，such a experiment design
was also reported in other N addition studies ( Liu et
al．，2013 )， so the mean soil respiration for three
months as an indicator of soil microbial activity was
appropriate． In contrast，several neutral effects of N
addition on soil respiration have also been reported
across different ecosystems． For example，Jiang et al．
(2010) found that nitrogen deposition (during growing
season) tended to decrease CO2 emission， but the
differences caused by nitrogen deposition were not
significant in a short-term simulated nitrogen deposition
experiment in an alpine meadow on the Qinghai-
Tibetan Plateau． Similarly，Liu et al． (2013) reported
that soil respiration did not significantly change after
four years N addition in a tropical forest． So it is，
therefore，that effects of N addition on soil respiration
were different across ecosystems and need a better
understanding．
3. 4 Conclusion
The effects of N additions on soil microbial
biomass and community structure in a young plantation
of G． biloba were concluded that: 1) Low-N treatment
did not significantly affect soil microbial biomass; 2)
Medium-N and High-N treatments significantly
decreased MBC ( C ) concentration but increased
microbial biomass nitrogen ( N ) concentration; 3 )
Medium-N and High-N treatments significantly
decreased soil total microbial biomass，fungal biomass，
and bacterial biomass; 4 ) Medium-N and High-N
treatments significantly decreased the relative
abundance of gram-positive bacterial PLFAs， and
increased fungal ∶ bacterial ratio and the relative
abundance of actinomycetes PLFAs; 5 ) High-N
treatment significantly decreased soil pH，respiration，
SOC concentration and the proportion of O-alkyl but
increased Alkyl∶ O-alkyl． Our conclusion indicated that
the responses of soil microbial biomass and community
structure in a young plantation of G． biloba varied due
to the amount of N loaded．
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