Plant Soil (2010) 331:181–191

DOI 10.1007/s11104-009-0244-2

REGULAR ARTICLE

Arbuscular mycorrhizal fungi in two mangroves

in South China
Yutao Wang & Qiu Qiu & Zhongyi Yang &
Zhijian Hu & Nora Fung-Yee Tam & Guorong Xin

Received: 30 August 2009 / Accepted: 25 November 2009 / Published online: 15 December 2009
# Springer Science+Business Media B.V. 2009

Abstract The symbiosis between arbuscular mycor-
rhizal fungi (AMF) and mangrove plant species was
investigated in two mangrove swamps in south China.
AMF were mostly found in the form of hyphae and
were commonly associated with all the mangrove
species we investigated. Six AMF species belonging
to the genera Glomus or Acaulospora were identified.
Multiple step-wise linear regression analyses showed
that hydrological conditions and phosphorus levels in
the rhizosphere were the main abiotic factors affecting
the colonization of mangrove species by AMF. A
greenhouse experiment was conducted to evaluate the
effects of AMF inoculation on the growth and nutrient
Responsible Editor: Peter Christie.
Y. Wang : Q. Qiu : Z. Yang (*) : Z. Hu : G. Xin (*)
State Key Laboratory of Biocontrol,
School of Life Sciences, Sun Yat-sen University,
Guangzhou 510275, China
e-mail: lssxgr@mail.sysu.edu.cn
e-mail: adsyzy@mail.sysu.edu.cn
N. F.-Y. Tam
Department of Biology and Chemistry,
City University of Hong Kong,
Hong Kong, China
N. F.-Y. Tam
Futian-CityU Mangrove RandD Centre,
Futian,
Shenzhen, China
uptake of a true mangrove plant species, Sonneratia
apetala B. Ham. The inoculated AMF significantly
improved growth, resulting in greater plant height,
diameter at ground level and plant biomass, as well as
increased absorption of N, P and K. These findings
suggest that AMF play important roles in mangrove
ecosystems.
Keywords Arbuscular mycorrhizal fungi (AMF) .
Mangrove . Hydrological conditions . Soil properties .
Sonneratia apetala
Abbreviations
AMF arbuscular mycorrhizal fungi
FT Futian Mangrove Area locating in Shenzhen,
Guangdong, China
ZH Zhuhai Mangrove Area locating in Zhuhai,
Guangdong, China
Introduction
Arbuscular mycorrhizal fungi (AMF) are found in most
terrestrial ecosystems, spanning tropical (Janos 1980;
Shi et al. 2006), temperate (Muthukumar and Udaiyan
2000) and arctic-alpine ecosystems (Haselwandter and
Read 1980). The importance and functional signifi-
cance of AMF in terrestrial ecosystems have been well
documented. As AMF require oxygen to survive,
182
early studies postulated that hypoxic conditions
could prevent their survival. Over the last two and
a half decades, the presence of AMF in wetland
plants has received increased attention, and there is
strong evidence that AMF commonly occur in
wetland ecosystems (Miller and Bever 1999; Miller
2000). Despite this attention, the functional roles of
the AM symbiosis in wetland ecosystems are poorly
understood.
Mangrove forests are generally characterized by
high levels of productivity, standing biomass and litter
production when compared to other terrestrial plant
communities (Lugo and Snedaker 1974). They are
one of the most important wetland ecosystems,
fulfilling essential ecological functions and harboring
precious natural resources (Rönnbäck 1999). The
saline and anaerobic conditions in the rhizosphere of
mangrove plants make it difficult to estimate the
abundance of AMF. Previous studies have shown that
these fungi are absent (Mohankumar and Mahadevan
1986), rare (Kothamasi et al. 2006) or ubiquitous
(Sengupta and Chaudhuri 2002; Kumar and Ghose
2008) in mangrove ecosystems. To date there has
been no research on the effects of AMF on mangrove
plants.
The aims of this study were to determine: (i) whether
AMF occur in the investigated mangrove ecosystems;
(ii) the main abiotic factors that influence the formation
of AM symbioses in mangrove ecosystems; and (iii)
how the growth and nutrient uptake of mangrove plants
is affected by inoculation with AMF.
Materials and methods
Field experiment
Study sites and sample collection
The Pearl River is the largest and most important river
in south China. The Futian Mangrove Area (FT) (22°
30′–22°39′N, 113°53′–114°05′E) and the Zhuhai
Mangrove Area (ZH) (22°23′–22°27′N, 113°36′–
113°39′E) are located on the east and west banks of
the Pearl River estuary, respectively. The FT site is
situated within the Futian Nature Reserve of the
Shenzhen Special Economic Zone, which covers an
area of 111 hectares of mangrove forest. The ZH site
is situated in the Qi Ao Mangrove Nature Reserve in
Plant Soil (2010) 331:181–191
Zhuhai and the mangrove forest covers an area of 58
hectares. Geographically, both sites are located close
to the northern limit of mangrove development, where
there is an irregular semi-diurnal tide, i.e. two
irregular high tides per 24 h period, with a mean tidal
range of 1.9 m. The average annual precipitation,
daily temperature and water salinity in ZH and FT are
1,964 mm, 22.4°C, 15‰ and 1,957 mm, 22.5°C,
18‰, respectively.
We investigated seven true mangrove species at FT
that are clearly distributed at different tidal heights.
The horizontal zonation pattern, landward to seaward
is: Bruguiera gymnorhiza Lam, Acanthus ilicifolius
Linn, Kandelia candel Druce, Avicennia marina
Vierh, Aegiceras corniculatum Blanco, Sonneratia
apetala B. Ham and Sonneratia caseolaris Engler. At
ZH, six true mangrove species (including A. ilicifo-
lius, S. apetala, A. corniculatum, K. candel, B.
gymnorhizal and S. caseolaris) and two semi-
mangrove species (Heritiera littoralis Ait. and Acros-
tichum aureum L.) were sampled. For A. ilicifolius,
samples were collected at both high and low tide
levels. At both sites, the average time the site was
flooded by tidal water (referred to, hereafter, as
flooding hours) for each mangrove plant species was
estimated by field observations (Table 1). Based on
the flooding hours and the moisture content in the
rhizospheric soil, both sites were divided into three
different intertidal zones: high, middle and low
(Table 1). In September and November 2007, root
and rhizosphere soil samples were collected from five
representative adult individuals of each plant species
1–2 h before high tide. For the root sample, only the
juvenile nutritive roots attached to the plant were
collected. The soil that remained adhered to the root
after gentle shaking (i.e. the rhizosphere soil) was also
collected. Each soil sample was divided into two
parts: one part was used for the analysis of physical
and chemical properties, and the other for AMF
identification.
Analysis of mycorrhizal colonization
Rinsed root samples were cleared with 10% KOH
at 90°C for approximately 40 min, and then
stained with trypan blue (Phillips and Hayman
1970). Percentage root colonization was determined
using the line intersect method (see McGonigle et al
1990) and we scored 200 intersects on 40 root
Plant Soil (2010) 331:181–191 183

Table 1 Hydrological con-

ditions of the area where the Site Species Intertidal zone FHa Mb
investigated species occurred
in the Futian Mangrove area FT B. gymnorhiza High (approximately 1.5 m from the lowest tide) 1.0 0.34±0.04
(FT) and the Zhuhai A. ilicifolius 2.5 0.40±0.04
Mangrove area (ZH) K. candel Middle (approximately 1.0 m from the lowest tide) 3.5 0.48±0.05
A. marina Low (approximately 0.4 m from the lowest tide) 10.0 0.63±0.02
A. corniculatum 10.5 0.64±0.08
S. caseolaris 11.0 0.60±0.02
S. apetala 11.5 0.59±0.02
a ZH H. littoralisc High (approximately 1.5–2.0 m from the lowest tide) 0 0.34±0.02
flooding hours: estimated
hours when the surface layer A. aureurmc 1.0 0.34±0.02
was underwater for each 24 h A. ilicifolius (H) 1.0 0.33±0.03
period S. apetala 1.5 0.36±0.02
b
moisture content of the rhi- A. corniculatum Middle (approximately 1.0 m from the lowest tide) 3.0 0.46±0.03
zosphere soil (Mean ± SD,
n=5); H and L represent the K. candel 3.5 0.48±0.02
high and low tide level B. gymnorhiza 3.5 0.49±0.02
c
semi-mangrove species; S. caseolaris Low (approximately 0.5 m from the lowest tide) 10.0 0.59±0.01
the same legend applies to A. ilicifolius (L) 11.0 0.59±0.01
other tables

segments per root sample using a compound micro-
scope (Carl Zeiss, Axiostar plus, Germany) at 100×
magnification. Special care was taken to discriminate
between the hyphae of AM and other fungi in roots,
and the pictures available from INVAM (http://
invam.caf.wvu.edu/fungi/taxonomy/speciesID.htm)
were used as references. Any septate hyphae or
hyphae that could not be stained were excluded.
Photographs were taken of the typical structures of
the hyphae.
Soil analysis
The soil samples were passed through a 0.25-mm
sieve before determining the organic matter content,
total N, P and K; samples used to determine pH,
electrical conductivity, available N, P and K were
passed through a 1-mm sieve. These analyses were
based on the methods described by Page et al. (1982).
The pH was measured in a 1:2.5 soil:water paste,
using a digital pH meter (Basic PB-20, Sartorius AG,
Goettingen, Germany). Electrical conductivity was
measured in the centrifuged supernatant of a 1:5 soil:
water extract. Organic matter content was determined
by the Walkley-Black acid digestion method. Total N
(digested with sulfuric acid) and available N
(extracted by 2 M KCl) were measured by a titration
of the distillates after Kjeldahl sample preparation and
analysis. Total P (digested with HNO3) and available
P (extracted by 0.05 M HCl-1/2H2SO4) were mea-
sured by molybdenum blue colorimetry. Total K
(digested with HNO3) and exchangeable K (extracted
by 0.1 M HCl) were measured using atomic absorp-
tion spectrophotometry (Analyst AA 100, Perkin-
Elmer, USA).
Trap culture and AMF identification
To prepare the AMF inocula for the pot experiment
and propagate more healthy spores for identifica-
tion, trap culture experiments were conducted by
taking the rhizosphere soil of S. apetala at ZH and
soil samples from ZH or FT as the inocula; we
followed the INVAM method (http://invam.caf.wvu.
edu/), except that 2 g of NaCl was added to each
1 kg of soil to ensure the AMF were propagated in
saline conditions. Zea mays (L.) was used as the
catch plant.
The AMF spores used for identification were
obtained from both the rhizosphere soil samples
(100 g) and the trap cultures (25 g) following the
wet sieving and decanting method (An et al. 1990).
Intact and healthy spores were then stored in tap
water. Some spores were mounted on slides in
polyvinyl-lactic acid-glycerine (PVLG; Koske and
Tessier 1983) and a mixture of PVLG with Melzer’s
184
(1:1; v/v; Brundrett et al. 1994). Gentle pressure was
applied to the cover slip to break the spore wall. The
spores were identified according to Schenck and
Peréz (1990) and INVAM.
Pot experiment
Plant and fungal materials
A pot experiment was conducted to determine the
effects of AMF on the growth and nutrient uptake
of mangrove plants. S. apetala was chosen as the
experimental species. It is a true mangrove species
from the Sonneratiaceae family, and it commonly
grows in newly formed sand or mudflats on the outer
fringes of mangrove forests. In China, S. apetala has
been widely used as a pioneer species for the
rehabilitation of bare mudflat areas and to facilitate
the re-colonization of other mangrove species be-
cause of its fast growth rate and high adaptability
(Ren et al. 2008).
Trap cultures from rhizosphere soil of S. apetala at
ZH were used as the fungal inoculum. After the
successful trapping process, AMF species were
identified and the mixtures containing AMF spores,
hyphae and mycorrhizal root fragments were stored in
zip lock plastic bags for more than 30 days before use
as the inoculum.
Experimental design
The pot experiment was conducted in a greenhouse at
Sun Yat-sen University. It consisted of a randomized
complete block design with two treatments: mycor-
rhizal (AM) and non-mycorrhizal (CK) treatments.
Each treatment was applied to five pots, each pot
(24 cm height, 18 cm diam. top, 13 cm diam. bottom)
was sterilized and contained 2.5 kg (dry weight) of
soil collected from ZH. The soil was air-dried and
autoclaved (121°C, 2 h) before being added to the
pot. The electrical conductivity, organic matter con-
tent and pH value of the soil were: 2.8±0.1 dS m−1,
3.1±0.3% and 7.7±0.0, respectively; and the concen-
trations of total N, P and K and available N, P and K
were 1.1±0.1, 0.8±0.0 and 53.8±1.7 gkg−1 and 130.7±
14.6, 14.1±0.2 and 536.9±18.4 mg kg−1, respectively.
For the AM pots, 100 g of prepared inoculum (trap
cultures of S. apetala from ZH) was thoroughly mixed
with the prepared soil. The same amount of sterilized
Plant Soil (2010) 331:181–191
inoculum was mixed with the soil used for the CK
pots. The CK pots also received 50 ml each of a
bacterial filtrate obtained by passing the suspensions
of inoculum through a paper filter (Whatman No.1,
particle retention: 11µm) (Jayachandran and Shetty
2003). On 5th May 2008, five surface sterilized S.
apetala seeds were sown in each pot. After the
appearance of euphylls (on 25th May), plants were
thinned to two seedlings per pot. Plant height and
diameter at ground level were measured every
15 days from 10th June. Plants were regularly
watered with de-ionized water, and supplemented
with Hoagland’s solution (but without P) biweekly at
a rate of 50 ml per pot.
Harvest and measurements
All plants were harvested after 110 days of growth
and divided into stem, leaf and root samples. A
small subsample of fine branch roots was random-
ly sampled to identify whether AM colonization
had occurred. The plant tissues were oven dried at
70°C for 72 h, weighed, and ground. The concen-
trations of N, P and K in the plant tissues were
analyzed.
Statistical analysis
A parametric One-Way Analysis of Variance (ANOVA),
followed by the Least Significant Difference (LSD) at
the 5% confidence level, was used to determine any
differences in colonization rates as well as rhizosphere
soil variables among different mangrove species at each
site. The same test was used for comparisons between
the plant height, diameter at ground level and biomass
(root, stem and leaf) measurements for the mycorrhizal
and non-mycorrhizal treatments in the pot experiment.
To determine the relationships between the soil variables
and the AM colonization rate, a multiple linear regres-
sion analysis was employed to develop predictive
models of AM colonization. The step-wise method was
used for variable selection, and the stepping criteria
employed for entry and removal were based on the
significance level of the F-value and set at 0.05 and
0.10, respectively. Correlation analysis was also con-
ducted to evaluate relationships between selected soil
variables and AMF colonization rates. All statistical
analyses were performed using SPSS Base 16.0 (SPSS
Inc., USA).
Plant Soil (2010) 331:181–191
Results
AM colonization
At both mangrove sites, all investigated plant species
showed associations with AMF (Figs. 1, 2 and 3).
Intracellular hyphae and hyphal coils were the
dominant structures. Vesicles were also found in all
species, while arbuscular structures were rare or
absent. At both sites, total AM colonization rates of
the species located at the low tide level were
significantly lower than those at the middle and high
tide levels (p<0.01).
AMF species
A number of different AMF species, including
Glomus intraradices Schenck and Smith, G. mosseae
(Nicol and Gerd) Gerd and Trappe, Acaulospora
scrobiculata Trappe at ZH and G. intraradices at


TCR ¼ 1:21 MC þ 0:19 TP þ 1:20 R2 ¼ 0:71; F ¼ 39:33; P ¼ 0:001 ;
TCR ¼  1:07 MC þ 12:66 OM þ 0:22 TNþ0:01APÞ þ 0:82 R2 ¼ 0:71; F ¼ 23:06; P ¼ 0:001 :
where MC, TP, OM, TN and AP represent soil
moisture, total P, organic matter, total N and
available P, respectively. The standardized coeffi-
cients of moisture and total P content at FT were
100% TC HC
80%
60%
PRC
40%
20%
0%
B.c A.i K.c
Mangrove species (FT)
Fig. 1 Percentage root colonization (PRC) of AMF in
different mangrove species at FT. The error bars represent
the standard deviations (SD) of five replicates. TC: total
colonization rate (calculated by summing the number of
occurrences of vesicles, arbuscules or hyphae, including times
when the structures occurred together as a single record), HC:
185
FT, were identified from the soil samples. A total of
six different Glomalean spore types were identified
from the trap cultures at ZH and FT. In addition to the
species directly identified from the soil samples,
Glomus aggregatum Schenck and Smith, G. geo-
sporum (Nicol and Gerd) Walker, G. rubiforme (Gerd.
& Trappe) Almeida & Schenck at ZH, and G.
mosseae at FT, were also identified. Besides, one
AMF spore type in soil samples from FT and two
AMF spore types in trap cultures from ZH which
could not be identified based on their morphological
characteristics were observed.
Properties of the rhizosphere soil
The properties of the rhizosphere soil from the different
mangrove species are shown in Table 2. The linear
models used to estimate the correlation between the
total AM colonization rate (TCR) and important soil
properties for FT and ZH (Table 3) were:


−0.53 and −0.38, indicating that an increase in
moisture content or total P content significantly
inhibited AM colonization (P<0.05). At ZH, AM
colonization rates were significantly inhibited by
VC AC
A.m A.c S.c S.a
colonization rate of hyphae, VC: colonization rate of vesicles,
AC: colonization rate of arbuscules; B.c: B. gymnorhizal, A.i:
A. ilicifolius, K.c: K. candel, A.m: A. marina, A.c: A.
corniculatum, S.c: S. caseolaris, S.a: S. apetala; The same
legend applies to the other figures
186 Plant Soil (2010) 331:181–191

100% TC HC VC AC

80%

60%

RPC
40%

20%

0%
H.l* A.a* A.i (H) S.a A.c K.c B.c S.c A.i (L)
Mangrove species (ZH)

Fig. 2 Percentage root colonization (PRC) of AMF in levels), S.a: S. apetala, A.c: A. corniculatum, K.c: K. candel,
different mangrove or semi-mangrove species at ZH. H.l: B.c: B. gymnorhizal, S.c: S. caseolaris; *: semi-mangrove
Heritiera littoralis, A.a: Acrostichum aureurm, A.i: A. species
ilicifolius (followed by H and L represent high and low tide

increased soil moisture or total N content (P<0.05),
while colonization rates were significantly improved
by increased soil organic matter or available P
content (P<0.05).
The results of the correlation analysis between AM
colonization and the selected single soil variables are
shown in Fig. 4. At FT, the total P content was
significantly negatively correlated with the AMF
colonization rate (P<0.05). At ZH, organic matter
and available P content were both significantly
positively correlated with the AMF colonization rate,
while total N content showed a negative correlation
(P<0.05).
Effects of inoculated AMF on the growth and nutrient
uptake of S. apetala
Four AMF species were identified from the AM
inocula. They were G. intraradices, G. mosseae, G.
aggregatum and G. geosporum, of which the two
former species were dominant. In the mycorrhizal
treatment, the percentage root colonization of the
inoculated AMF ranged from 8.5–15.5%, with an
average of 11.5%. In the CK treatment, no AMF
structures were observed.
The plant height and diameter at ground level are
shown in Fig. 5. One and a half months after

Fig. 3 Typical AMF struc

tures observed in mangrove
plant species
Plant Soil (2010) 331:181–191 187

Table 2 Electrical conductivity (EC), organic matter content (OM), pH, total P (TP), available P (AP), total N (TN) and available N
(AN) of rhizosphere soil in different true mangrove and semi-mangrove species

Site Species Intertidal zone EC (dS m−1) OM (%) pH TP AP TN AN

(g kg−1) (mg kg−1) (g kg−1) (mg kg−1)

FT B. gymnorhiza High 5.4±0.4 2.4±0.3 6.13±0.15 0.46±0.05 63.0±7.6 0.77±0.07 85.8±8.9

A. ilicifolius 8.6±0.3 2.7±0.4 5.86±0.14 0.44±0.03 113.6±11.6 1.21±0.50 165.9±17.5
K. candel Middle 9.8±0.3 6.0±1.2 6.11±0.52 0.92±0.09 120.4±9.8 1.61±0.20 165.2±18.9
A. marina Low 14. 9±0.8 12.0±0.9 6.82±0.41 1.12±0.07 134.6±7.9 2.70±0.24 272.1±12.1
A. corniculatum 11.7±0.5 12.7±0.8 5.35±0.54 1.28±0.10 172.5±17.0 3.53±0.26 235.1±18.0
S. caseolaris 11.9±0. 6 7.1±1.2 6.60±0.65 1.45±0.19 99.8±9.3 2.22±0.25 171.9±9.9
S. apetala 13.3±0.7 5.4±0.9 6.64±0.10 2.08±0.18 203.9±27.4 2.43±0.16 204.2±20.7
ZH H. littoralis* High 1.7±0.2 4.0±0.5 7.54±0.12 0.90±0.20 12.1±0.7 1.66±0.04 118.4±18.9
A. aureurm* 2.5±0.1 3.4±0.9 7.34±0.18 0.81±0.06 13.3±0.7 1.97±0.19 167.0±8.6
A. ilicifolius (H) 2.5±0.1 2.5±0.6 7.46±0.29 0.67±0.10 11.5±2.2 1.25±0.14 165.9±24.8
S. apetala 2.7±0.3 3.0±0.4 7.27±0.22 0.84±0.11 18.1±1.8 1.58±0.08 144.4±11.3
A. corniculatum Middle 2.9±0.3 3.5±0.4 6.87±0.16 0.90±0.07 19.4±0.8 1.99±0.13 208.1±17.1
K. candel 3.3±0.3 4.0±0.4 6.98±0.39 0.92±0.08 17.6±2.3 1.92±0.19 135.4±6.2
B. gymnorhiza 2.9±0.7 3.7±0.6 6.87±0.30 0.98±0.05 19.8±1.2 1.91±0.47 137.6±11.2
S. caseolaris Low 2.9±0.1 3.4±0.5 6.77±0.19 0.88±0.11 13.7±2.0 1.99±0.05 150.3±13.1
A. ilicifolius (L) 2.1±0.1 2.6±0.2 7.29±0.09 0.77±0.02 4.6±1.5 2.06±0.16 168.2±24.2

Means and standard deviations are for five replicates

inoculation, the plant height and diameter at ground
level for AM plants were significantly higher than
those for CK plants (p<0.05). The biomass (DW, per
plant) of leaves, stems and roots of the AM plants
were all significantly greater than those of the CK
plants (p<0.01) (Fig. 6).
The concentrations of root P and leaf K in the AM
plants were significantly higher than those in the CK
plants (p<0.05). The concentrations of root N, stem
N, leaf N, leaf P and stem K were significantly lower
in the AM plants compared to the CK plants (p<0.05)
(Table 4). However, because the biomass of the AM
plants was much higher than those of the CK plants,
total amounts (concentration × biomass, per plant) of
root and stem N, P and K, and leaf P and K
accumulated by the AM plants were significantly
greater than those of the CK plants (p<0.01).
Discussion
AMF occurrence in the investigated mangrove
ecosystems
The presence of AMF in wetland ecosystems has
been recently reported a lot (Ipsilantis and Sylvia
2007). However, few studies have been conducted in
mangrove ecosystems. In our investigation, AMF
structures were present in all investigated species,
but few arbuscules were found at either site. Accord-

Table 3 Coefficients of the

multiple step-wise linear Site Variable Unstandardized coefficients Standardized coefficients ta Sig.a
regression analysis between
AM colonization rate and FT Moisture −1.21 −0.53 −3.92 0.000
the selected soil variables Total P −0.19 −0.38 −2.85 0.008
(excluded variables are not ZH Moisture −1.07 −0.43 −3.67 0.001
shown)
Available P 0.01 0.24 4.49 0.000
Organic matter 12.66 0.45 −2.38 0.023
a
t-test of regression Total N −0.22 −0.29 2.38 0.023
coefficients
188 Plant Soil (2010) 331:181–191

Fig. 4 Correlations (a) (b)

between AMF percentage 100 100
FT ZH
root colonization (PRC) and
a soil total P content (FT), b 80 80
r = 0.37, P =0.015
soil available P content
60 60

PRC (%)

PRC (%)
(ZH), c soil organic matter
r =-0.76, P =0.000
content (ZH) and d soil total
40 40
N content (ZH)
20 20

0 0
0.0 1.0 2.0 3.0 0 10 20 30
Total P (ppt) Available P (ppm)

(c) (d)
100 FT 100 ZH
r =0.36, P =0.013
80 80

PRC (%)

PRC (%)
60 60

40 40
r =-0.50, P =0.000
20 20

0 0
15 25 35 45 55 1.0 1.5 2.0 2.5
Organic matter (ppt) Total N (ppt)

ing to Gallaud’s descriptions to ‘Paris-type’ and structure; this was also reported by Sengupta and
‘Arum-type’ arbuscules (see Smith and Smith 1997), Chaudhuri (2002). It might possibly be explained by
both arbuscule types were present in mangrove the high moisture and salt levels in mangrove
species, and ‘Paris-type’ arbuscules were observed ecosystems, but further research is needed.
more frequently. Sengupta and Chaudhuri (2002) and Miller and Bever (1999) identified two possible
Kumar and Ghose (2008), who investigated the mechanisms by which AMF could survive in hypoxic
Sundarban mangrove swamps in the Ganges river conditions. Since AMF are aerobic microorganisms,
estuary in India, report similar results. Estuarine we propose that the common occurrence of AMF
mangrove habitats, as recent alluvial landforms, differ symbioses in mangrove ecosystems is related to the
from marine salt marshes in their origin (Sengupta well developed aerenchyma that has been described in
and Chaudhuri 2002). This may explain why AMF adult mangrove species (Allaway et al. 2001). When
were not reported in mangrove species of a marine flooded, AMF may survive by relying on the oxygen
salt marsh by Mohankumar and Mahadevan (1986). provided by the aerenchyma. This is supported by our
In our study, hyphae were the dominant AMF observation of little or no AMF colonization of

Fig. 5 a Plant height (H) 45 0.8

and b diameter at ground (a) AM (b)
40 0.7
level (DGL) for S. apetala CK
35 0.6
at different times during the
pot experiment. AM: 30 0.5
DGL (cm)
H (cm)

mycorrhizal treatment; CK: 25

0.4
non-mycorrhizal treatment; 20
0.3
means and standard devia- 15
tions are from 10 replicates 0.2
10
0.1
5
0
0
9-Aug

24-Aug
10-Jul

25-Jul
10-Jun

25-Jun
9-Aug

24-Aug
10-Jul

25-Jul
10-Jun

25-Jun

Date Date
Plant Soil (2010) 331:181–191 189

2.5 2002; Carvalho et al. 2004). The composition of the

AM CK a
AMF species in mangrove ecosystems studied indicates
2.0
possible fungal adaptation to high salinity and flood
Biomass (g)

1.5 a
a conditions.
b
1.0 b b Abiotic factors influencing the formation of AM
symbiosis in mangrove ecosystems
0.5

0.0
Root Stem Leaf
Fig. 6 Average biomass per plant (DW) of roots, stems and
leaves in mycorrhizal (AM) and non-mycorrhizal (CK) treat-
ments; means and standard deviations are from 10 replicates;
values with different letters are significantly different at p<0.05
level
seedlings of the investigated mangrove species, in
which aerenchyma had not yet become well devel-
oped (data not shown).
Catch plants are useful for propagating some AMF
species for identification. However, biases are often
introduced by plant preference for certain AMF
species and other environmental factors. In the
present study, we used the catch plant strategy with
the above limitations in mind.
In total, six AMF species belonging to the genera
Glomus or Acaulospora were identified from both sites.
Among them, G. mosseae is known for its worldwide
distribution in many ecosystems, including some wet-
lands and salt marshes. G. intraradices has been
reported to be a salt-tolerant species (Sannazzaro et al.
2004; Sannazzaro et al. 2006). These two species were
observed at both ZH and FT. G. geosporum is often
dominant in European salt marshes (Landwehr et al.
Because each mangrove species is usually located in a
particular intertidal zone, both the results of multiple
regression and linear correlation analysis could not take
the effects of different species on AMF colonization into
account. From our study, the hydrological conditions
(rhizosphere soil moisture content and flooding hours)
and soil P level are the main abiotic factors affecting
AMF colonization of mangrove species.
Hydrological conditions
Several field studies have shown that AMF coloniza-
tion of wetlands is generally higher in drier areas
compared to the wetter, more anaerobic areas (Miller
2000), because of the aerobic requirements of AMF.
In our investigation, AMF symbiosis was found in
mangrove plants that experienced flooded conditions
for more than 11 h each day, indicating that some
AMF species had the ability to survive in highly
hypoxic conditions. In contrast, obvious suppression
of AMF colonization by long flooding time was also
observed at both sites. The multiple linear regression
analyses showed that moisture content was the most
important factor that inhibited AMF colonization in
both mangrove swamps; similar findings were also
reported by Miller (2000).

Table 4 Concentrations
and total amount (concen- Variables Concentration (g kg−1) Total amount (mg)
tration × biomass, per plant)
of N, P and K in roots, AM CK AM CK
stems and leaves of AM and
CK plants at the end of the Root N 4.63±0.64 b 6.30±1.81 a 5.53±0.88 A 4.21±1.22 B
pot experiment Stem N 4.05±0.30 b 7.14±1.73 a 4.82±0.64 A 4.01±0.61 B
Leaf N 12.16±0.97 b 17.49±3.08 a 18.68±4.74 A 14.90±3.39 B
Root P 1.32±0.24 a 1.13±0.07 b 1.65±0.56 A 0.76±0.09 B
Stem P 0.91±0.17 a 1.04±0.10 a 1.12±0.31 A 0.60±0.12 B
Leaf P 1.56±0.10 a 1.35±0.19 b 2.39±0.65 A 1.14±0.22 B
Means and standard devia-
tions are from 10 replicates; Root K 9.79±0.50 a 10.41±1.13 a 11.99±2.98 A 6.94±0.50 B
values with different letters Stem K 7.47±0.72 b 9.68±1.77 a 8.84±1.12 A 5.51±0.70 B
are significantly different at Leaf K 9.31±0.38 a 8.55±1.04 b 14.50±4.27 A 7.38±1.77 B
p<0.05 level
190
Soil P level
Concentrations of available P in ZH and FT were very
different; levels were low at ZH (3.0–21.4 mg kg−1)
and high at FT (52.1–228.0 mg kg−1). In terrestrial
ecosystems, a negative correlation between AMF
colonization and soil P concentration has been
commonly reported. Some studies have also sug-
gested that decreased colonization occurs in wetlands
as a result of high phosphorus levels (Wetzel and van
der Valk 1996). However, for many reasons, inhibi-
tion of AMF colonization by increased soil P seldom
occurs in wetland field studies (Kelly et al. 2004). We
found that at FT an increase in the total P in soil
decreased the AMF colonization rates; however, at
ZH, the opposite trend was observed. It has been
previously reported that low soil P levels could also
inhibit AMF colonization in terrestrial ecosystems
(Brundrett et al. 1999; Azcón et al. 2003; Chen et al.
2008). From our results, we suggest that an inverted
‘U’ relationship between AMF colonization and soil
P, i.e. soil P at high or low enough levels inhibits
AMF colonization (Koide and Li 1990; Koide 1991),
could also occur in wetland ecosystems,.
Effect of inoculated AMF on growth and nutrient
uptake of mangrove plants
Previous studies have shown that AMF can have positive
(Miller and Sharitz 2000; Fougnies et al. 2007) or
neutral (Stevens et al. 2002; Ipsilantis and Sylvia 2007)
effects on the performance of wetland species. Dunham
et al (2003) also reported that AMF can reduce the
growth of its host plant (Typha latifolia, a ubiquitous
wetland plant) despite increased mineral nutrition and
photosynthetic activity. However, to our knowledge, no
research has been conducted on the effects of AMF on
mangrove species, owing mainly to the difficulties in
artificial culture of mangrove species (Sengupta and
Chaudhuri 2002).
In a preliminary study, we found that inoculated
AMF had positive impacts on the growth and
nutrient uptake of S. apetala. The lower concen-
trations of root N, stem N, leaf N, leaf P and stem K
in AM plant tissues can be attributed to the dilution
resulting from increased biomass and the compara-
tively low increase in plant N, leaf P and stem K
absorption. Although the soil that we used contained
relatively high levels of N and K, the inoculated
Plant Soil (2010) 331:181–191
AMF significantly improved the total absorption of
N, P and K. It is therefore reasonable for us to infer
that AMF play important roles in mangrove ecosys-
tems. Furthermore, AMF can influence the compo-
sition and diversity of the plant community (van der
Heijden 2002; Urcelay and Diaz 2003; Wolfe et al.
2006). Considering the large effects on mangrove
species, AMF could be important drivers of plant
community composition in mangrove ecosystems.
However, further research is needed to consider the
combined effect of saline and periodic flooded
conditions.
Acknowledgements This research was financially supported
by the National Natural Science Foundation of China (NSFC,
U0633002; 30871475) and a research grant from the Research
Grants Council of the HKSAR (Project No. CityU 1406/06M),
Zhang Hong-da Science Research Fund of Sun Yat-Sen University
and the Project of Science and Technology of Zhuhai City
(200901022). We thank Wang You-shan (Beijing Academy of
Agriculture and Forestry Sciences) for her help with the AMF
identification, and Luan Tian-gang, Xia Fang-fang, Yang Qiong,
Zhong Yin and Wang Yuan-yuan (Futian-CityU Mangrove R and
D Centre) for their assistance in sampling. We also thank Dr.
Janice Martin (University of Wales, Aberystwyth, UK), Dr. John
Blackwell (University of Sheffield, Sheffield, UK), Roger T.
Koide (Penn State University, University Park, USA) and Chen
Bao-ming (Sun Yat-sen University, Guangzhou, China) for
assistance with English grammar.
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