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ORIGINAL PAPER
How Abundant are Root-Colonizing Fungi in Southeastern
Louisiana’s Degraded Marshes?
Demetra Kandalepas &Kevin J. Stevens &
Gary P. Shaffer &William J. Platt
Received: 11 January 2009 /Accepted: 15 September 2009 / Published online: 9 March 2010
#Society of Wetland Scientists 2010
Abstract Despite earlier notions that fungi are not impor-
tant in wetlands, it is becoming clear that root endophytes
are abundant in wetlands and potentially can influence plant
community dynamics. Little is known about the effects of
wetland degradation on these fungi. We assessed two
groups of root endophytes in a degrading marsh in
southeast Louisiana that historically was a swamp forest
dominated by Taxodium distichum (baldcypress) and Nyssa
aquatica (water tupelo). We determined percent root
colonization by arbuscular mycorrhizal fungi (AMF) and
dark septate endophytes (DSE) in each of 18 vascular plant
species. Fungi were present in all species that were
assessed. In general, monocots were primarily colonized
by DSE, whereas dicots were primarily colonized by AMF.
Taxodium distichum was heavily colonized by AMF, as was
the non-native, invasive Chinese tallow (Triadica sebifera).
This study is the first to show that wetland plants in a
degraded marsh harbor abundant and diverse root endo-
phytes. These fungi and their interactions with stressed
plants may be important in effective management of
degrading wetlands.
Keywords Arbuscular mycorrhizas .Dark septate
endophytes .Manchac land bridge .Restoration .Wetlands
Introduction
Fungi form symbiotic associations with plants, inhabiting
above- and below-ground tissue. Two types of root-
inhabiting fungi are arbuscular mycorrhizal fungi (AMF)
and dark septate endophytes (DSE). Both types have been
reported in wetland plants (Turner and Friese 1998;
Cornwell et al. 2001; Stevens et al. 2002; Muthukumar et
al. 2004). These root-inhabiting fungi may affect plants by
reducing negative effects of flood and salt stress, enhancing
nutrient uptake, and providing protection from pathogens
and herbivores (Jumpponen 2001).
Until recently, arbuscular mycorrhizal fungi were con-
sidered unimportant in wetlands. Because AMF are aerobic,
anoxic conditions associated with waterlogged soils were
considered limiting (Malloch et al. 1980; Peat and Fitter
1993), and AMF had rarely been found on plant roots in
waterlogged soils. Several studies, however, indicate that
AMF are present and widespread in some wetlands, and
also may influence wetland plant community structure
(Brown and Bledsoe 1996; Stevens and Peterson 1996;
Carvalho et al. 2001; Hildebrandt et al. 2001; Landwehr et
al. 2002; Bauer et al. 2003; Wang et al. 2004). Several
wetland plant species (Cyperaceae, Chenopodiaceae, and
Plumbaginaceae) that were thought to be non-mycorrhizal
(Hirsch and Kapulnik 1998) have been shown to have high
levels of AMF colonization (Hildebrandt et al. 2001;
Muthukumar et al. 2004). The presence of AMF has been
shown to influence plant zonation in salt marshes (Daleo et
al. 2008), and they may play a role in diversifying plant
communities by allowing mycorrhizal plants to compete
D. Kandalepas (*):W. J. Platt
Department of Biological Sciences, Louisiana State University,
Baton Rouge, LA 70803, USA
e-mail: dkanda1@tigers.lsu.edu
K. J. Stevens
Department of Biological Sciences, Institute of Applied Science,
University of North Texas,
Denton, TX 76203, USA
G. P. Shaffer
Department of Biological Sciences,
Southeastern Louisiana University,
Hammond, LA 70402, USA
Wetlands (2010) 30:189–199
DOI 10.1007/s13157-010-0017-y
with sympatric, non-mycorrhizal plants (Weishampel and
Bedford 2006).
Dark septate endophytes in marshes have received less
attention than AMF. Evidence suggests, however, that these
fungi form associations with some wetland plants and may
occur over a wide range of conditions (Addy et al. 2000;
Fuchs and Haselwandter 2004). Dark septate endophyte
colonization has been shown to occur in both bog and fen
species, although bog plant species are typically colonized
mostly by DSE, and fen species are colonized mostly by
AMF. Some studies suggest that community structure may
be influenced as much by DSE as they are by AMF (Fuchs
and Haselwandter 2004; Weishampel and Bedford 2006).
Because root endophytes may have important functional
roles in wetlands, they may be important in restoration of
marshes (Bauer et al. 2003). However, we know little about
plant-fungal interactions and their effects on community
dynamics in degrading wetlands. In this study, we assessed
AMF and DSE colonization of wetland plant species in the
Manchac land bridge, a degrading marsh in southeast
Louisiana. For several decades, there have been unsuccess-
ful efforts to restore this marsh back to a swamp forest
dominated by Taxodium distichum and Nyssa aquatica.
However, only some stunted, remnant T. distichum survived
from these restoration efforts, and Triadica sebifera, a non-
native, invasive tree, became established. We had three
objectives: 1) to determine if root endophytes colonized
plants in this marsh, 2) to determine if a relationship exists
between AMF and DSE colonization, and 3) to compare
patterns of colonization between two broad categories of
vegetation—monocotyledonous (monocots) and dicotyle-
donous (dicots) plants.
We collected roots from a total of 18 species, 14
herbaceous plants, two shrubs (Iva frutescens and Baccha-
ris halimifolia), and two trees (T. distichum and T. sebifera).
We determined the extent of root colonization for each
species and distinguished between AMF and DSE coloni-
zation. This study is an important first step in determining
the role of root endophytes in degrading coastal marshes of
southern Louisiana.
Methods
Study Site
The Turtle Cove Environmental Research Station (N 30°
17′59″, W 90° 20′10″) fronts a 3,200 ha experimental
marsh located on the Manchac land bridge between Lakes
Maurepas and Pontchartrain in southeastern Louisiana
(Fig. 1). Wetlands on this narrow land bridge have been
undergoing rapid degradation over the past century,
primarily from hydrologic alteration by canal and levee
construction. As a result, the Manchac land bridge
Taxodium distichum and Nyssa aquatica swamps have
converted to marsh vegetation, and parts of these marshes
are now converting to open water (Barras et al. 2003;
Shaffer et al. 2009b).
Wetlands of the Manchac land bridge are currently
nutrient limited, with nitrogen (nitrate plus nitrite) rarely
Lake
Maurepas
Fig. 1 Location of Turtle Cove
Environmental Research Station
in southeast Louisiana. Turtle
Cove (arrow) is located in the
Manchac Wildlife Management
Area, on the Manchac Land Bridge
between Lakes Pontchartrain
and Maurepas (Tangipahoa and
St. John the Baptist Parishes)
190 Wetlands (2010) 30:189–199
exceeding 0.05 mg/L (Lane et al. 2003; USGS, National
Water Information System 2007). Phosphorus levels may
reach 0.2 mg/L (Kandalepas 2004), but concentrations
typically are low, averaging 0.055 mg/L (Lane et al. 2003).
In contrast, the adjacent Mississippi River, which histori-
cally was the source of nutrients and fresh water for
wetlands, contains much higher concentrations of nitrogen
and phosphorus (Lane et al. 2003).
Salinity in the Manchac land bridge wetlands is usually
low, ranging from 0.8 ppt to 1.1 ppt (Kandalepas 2004).
However, pulses of salt water from storms frequently
inundate the wetlands, and post-hurricane salt concentra-
tions may exceed 5 ppt (USGS, National Water Information
System 2005; Shaffer et al. 2009b). Following 2 years
(1999, 2000) of drought, Lane et al. (2003) found the
salinity on the Manchac land bridge wetlands reached
12 ppt.
Sampling
We collected randomly selected plants along a 1,160-m
transect from spring 2005 through fall 2006. We chose
plants that broadly represented species of Louisiana
marshes. Additionally, we collected roots from T. distichum
that were planted in an effort to restore the habitat to
cypress swamp, as well as from invasive species typical of
degraded swamp/marsh habitat (e.g., Triadica sebifera and
Alternanthera philoxeroides). With the exception of trees
and shrubs, we collected entire plants to ensure roots
originated from a given species. For large woody plants we
physically followed the roots to their tips with our hands to
find the new growth.
We collected three to five individuals (true reps) of 18 of
the most common wetland plant species in the area. We
assessed these for AMF and DSE colonization (see Table 1).
Plants were uprooted, cleaned of debris, bagged, and
transported to the laboratory. Plants were kept moist and
stored in a refrigerator until further processing. Nomencla-
ture and authorities of plant species were confirmed by the
USDA Natural Resources Conservation Service plants
database (2008). While grasses (e.g., Echinochloa walteri,
Phalaris sp.) were present, we did not sample them because
inflorescences were lacking and thus conclusive identifica-
tion was difficult.
Processing and Assessment
We initiated processing of roots within 24 h of collecting.
Roots waiting to be processed were kept at 4°C. Sediment
and debris were washed from the roots, and a sub-sample of
non-woody roots large enough to fill a 50 ml centrifuge
tube was obtained from each plant and fixed in 50%
ethanol. All healthy roots collected were cleared by
autoclaving in 10% potassium hydroxide (KOH) for 15–
Species
MAGNOLIOPHYTA (ANGIOSPERMS)
Liliopsida (Monocotyledons)
Alismataceae Sagittaria lancifolia L.
Cyperaceae Eleocharis cellulosa Torr.
Eleocharis montevidensis Kunth
Schoenoplectus tabernaemontani (K.C. Gmel.) Palla
Schoenoplectus americanus (Pers.) Volk. Exs Schinz & R. Keller.
Schoenoplectus robustus (Pursh) M.T. Stong
Typhaceae Typha domingensis Pers.
Magnoliopsida (Dicotyledons)
Amaranthaceae Alternantera philoxeroides (Mart.) Griseb.
Amaranthus australis (Gray) Sauer
Asteraceae Symphyotrichum subulatum (Michx.) G.L. Nesom
Baccharis halimifolia L.
Iva frutescens L.
Convolvulaceae Ipomoea sagittata Poir. In Lam.
Euphorbiaceae Triadica sebifera (L.) Small.
Fabaceae Vigna luteola (Jacq.) Benth.
Sesbania herbacea (Mill.) McVaugh
Polygonaceae Polygonum punctatum Ell.
PINOPHYTA (CONIFERS)
Cupressaceae Taxodium distichum (L.) Rich.
Table 1 Vascular plant species
examined for arbuscular mycor-
rhizal fungi and dark septate
endophytes at Turtle Cove
Environmental Research Sta-
tion, Louisiana
Wetlands (2010) 30:189–199 191
20 min, depending on pigmentation, then stained by
autoclaving in 3% Trypan Blue for 15 min (Brundrett et
al. 1996) to make structures associated with AMF and DSE
colonization visible. A subset of the species were processed
as above, but stained with 3% Chlorazol Black E (Brundrett
et al.1984) to achieve higher contrasts between fungal
structures and plant tissues. Tryphan Blue and Chlorazol
Black E were dissolved in a 1:1:1 lactic acid:glycerine:de-
ionized water solution (Brundrett et al. 1996). Roots were
de-stained and stored in a 50% glycerol solution for up to
1 week before mounting on slides in 50% glycerol (Phillips
and Hayman 1970). All stained roots were cut into 1-cm
segments and mounted on multiple slides.
Root colonization was assessed by viewing stained roots
and estimating the proportion of each root that was
colonized by AMF, DSE, or both. We did not identify
specific fungal species, as this was beyond the scope of this
study. We used a Zeiss Axioimage microscope at 200x
magnification and images were obtained with a Zeiss
Axiocam MRC-5 camera. Colonization levels were quan-
tified using a modified grid line intersect procedure
(McGonigle et al. 1990), with 100 fields of view assessed
for each slide. We calculated total colonization as the
percentage of root length in the 100 different fields of view
containing any AMF or DSE fungal structures, including
hyphae, arbuscules, or vesicles. Plants were considered to
form AMF associations, however, only if arbuscules, the
only uniquely distinguishable feature in AMF, were
detected in the roots (McGonigle et al. 1990). If character-
istic AMF hyphae and/or vesicles were found without
arbuscules, the AMF status was deemed unverified, but
colonization by hyphae and vesicles was included in the
total. For assessment of DSE colonization levels, only
hyphae were quantified, because DSE hyphae are distinc-
tive. As a result, total colonization was often greater than
the sum of AMF and DSE colonization. Means and
standard errors for all estimated percentages of roots
colonized by AMF and DSE or both were computed using
SAS software, Version 9.1.3, of the SAS System for
Windows (SAS Institute Inc. 2000–2004). To determine
the relationship between AMF and DSE colonization,
Spearman Rank correlations were conducted using Graph-
Pad InStat (ver 3.06, GraphPad Software, Inc.). To
determine if levels of colonization differed among mono-
cots and dicots, arbuscular, hyphal, vesicular and DSE
colonization levels were compared using Mann-Whitney
tests in GraphPad InStat.
We used nonmetric multidimensional scaling (NMS)
(Kruskal 1964) in Primer v6 (Clarke and Warwick 2006)to
illustrate the pattern of colonization by AMF and DSE in
monocots and dicots. We used ANOSIM (Clarke and
Warwick 2006) to determine if the observed pattern was
different from random. To perform the NMS, we used the
Bray Curtis Similarity Index with 999 permutations. Square
root transformation was used to minimize the influence of
large values.
Results
Both AMF and DSE were visible in stained cells of roots of
wetland plants at the Turtle Cove Biological Research
Station. Figure 2illustrates a cleared, unstained root
(Fig. 2a), stained roots with AMF (Fig. 2b–e), and stained
roots with DSE (Fig. 2f–g). In Fig. 2b–d, arbuscules are
clearly evident. Figure 2(d) and (e) display vesicles. Dark
septate endophytes in Fig. 2(f) and (g) are recognized by
their thickened walls and septate hyphae. In addition,
microsclerotia, segmented structures visible within the plant
cells, also are characteristic of DSE. The physical presence
of these different structures indicated the presence of AMF,
DSE, or both in at least some plants of all species examined
(Table 2).
Most species that we sampled had indications of
colonization by AMF, and every species, moncot or dicot,
had indications of DSE colonization, usually in all
individuals (Table 2). Typically, every perennial dicotyle-
donous plant was colonized by AMF. In contrast, the
presence of AMF was only conclusively confirmed for 1 of
the 7 monocots sampled—Schoenoplectus robustus. The
presence of vesicles and hyphae suggested AMF might be
colonizing many of the monocots, although there were
higher levels of DSE in this group. Three species, Taxodium
distichum, Alternanthera philoxeroides, and Vigna luteola
contained AMF within all individuals examined, but each
contained DSE in only one individual sampled.
Total colonization of roots by AMF and DSE was
substantial for most species. Among monocots, proportions
of roots colonized by both fungal types ranged from 27%
(Sagittaria lancifolia, Typha domingensis) to > 80%
(Schoenoplectus americanus, S. robustus). Among dicots,
root colonization by AMF and DSE ranged from 29–44%
(Amaranthus australis, Sesbania herbacea)to>80%
(Alternanthera philoxeroides, Triadica sebifera).
Overall, there was a negative correlation between AMF
and DSE for, both, monocots (r=−0.65; p=0.0002; n=27)
and dicots (r=−0.44; p=0.0073; n=36). All measures of
AMF colonization were significantly higher in dicots
compared to monocots (mean arbuscular colonization in
dicots was 10.2%±1.8%(SE) vs 0.1% ± 0.1% for monocots,
p<0.0001; mean hyphal colonization was 47.0% ± 3.3% for
dicots vs 28.8%±6.2% for monocots, p<0.05; mean
vesicular colonization was 6.8%±1.5% for dicots vs
0.7%±0.3% for monocots, p< 0.0001). DSE colonization
showed the opposite trend and was significantly higher in
monocots (25.0%±4.5%) compared to the dicots (14.2% ±
192 Wetlands (2010) 30:189–199
Fig. 2 Cleared and stained roots of seven wetland plant species
collected from the Turtle Cove Environmental Research Station in
southeast Louisiana. Figure 2a is a non-colonized root; Figures 2b–e
illustrate colonization by arbuscular mycorrhizal fungi; Figure 2f–g
illustrates colonization by dark septate endophytes. aNon-colonized
area of a root of Schoenoplectus robustus. Cell contents have been
removed by clearing in KOH. Cell walls are visible in epidermal cells
(arrowheads) and cortical cells (*) otherwise cells appear translucent.
Helical secondary cell wall thickenings are evident in the xylem
tracheary elements (arrows). Roots are stained with Chlorazol Black
E. Scale bar=50 µm. bArbuscules (arrowheads) within cortical cells
of Vigna luteola. Non-colonized cells are visible above the colonized
cells. Helical secondary cell wall thickenings (arrows) are visible in
xylem tracheary elements. Roots are stained with Chlorazol Black E.
Scale bar= 100µm. cArbusculate coils of the Paris-type AMF within a
cortical cell of Triadica sebifera. Roots are stained with Chlorazol
Black E. Scale bar=50 µm. dArbusculate coils of the Paris-type AMF
(arrow) and vesicle (arrowhead) within isolated cortical cells of
Taxodium distichum. Roots are stained with Chlorazol Black E. Scale
bar= 100 µm. eVesicles (arrowheads) in the cortex of Aster subulatus.
Roots are stained with Tryphan Blue. Scale bar=100 µm. f: Micro-
sclerotia (arrowheads) and septate hyphae (arrows) in epidermal cells
of Eleocharis cellulosa. Roots are stained with Tryphan Blue. Scale
bar= 50 µm. gSeptate hyphae (arrow) and melanized microsclerotium
(arrowhead) within an epidermal cell of Iva frutescens. Roots are
stained with Tryphan Blue. Scale bar= 20 µm
Wetlands (2010) 30:189–199 193
2.9%; p<0.05). Finally, NMS revealed a significant
difference in colonization between AMF and DSE in the
plant groups examined (ANOSIM, Global R=0.391; p=
0.003, Fig. 3).
Discussion
Our study demonstrated that plants inhabiting degrading
marshes of coastal Louisiana are colonized by AMF and
DSE, and contributes to a growing body of literature
documenting the occurrence of AMF and DSE fungi in
wetland plants. Only four plant species in this study appear
to have been examined previously for mycorrhizal coloni-
zation (see Appendix); to our knowledge this is the first
record of AMF and/or DSE being present in 13 of the
species. All plant species examined contained DSE, and
AMF were indicated in all but one plant species, suggesting
that mycorrhizal fungi can be expected to be ubiquitous
components of southern Louisiana marshes. Moreover,
colonization levels were comparable to those in other U.S.
wetlands (e.g., Cornwell et al. 2001; Bauer et al. 2003;
Weishampel and Bedford 2006), and higher than those
recorded in wetlands of Asia (Kai and Zhiwei 2006)or
Europe (Sraj-Krzic et al. 2006).
Plant species at Turtle Cove Environmental Research
Station are typical of fresh to intermediate brackish marsh,
but often experience salt and flood stress due to natural and
anthropogenic influences. Storm surges from hurricanes
and periodic droughts occasionally concentrate salts in the
soil, thereby bringing about major shifts in vegetation
(Visser et al. 1999,2002). In addition, levees constructed
throughout southeast Louisiana have resulted in loss of
nutrient input that, historically, occurred with the flooding
of the Mississippi River. These hydrologic alterations have
contributed to relative sea-level rise, due to subsidence that
occurs naturally in south Louisiana. As many marshes in
coastal Louisiana, the Manchac land bridge has been
converting to open water (Barras et al. 2003) due to
saltwater intrusion, increased flooding, nutrient starvation,
Table 2 Arbuscular mycorrhizal fungi (AMF), dark septate endo-
phytes (DSE), are indicated as present (+) absent (−) or indeterminable
(?). AMF colonization could not be determined if arbuscules were not
detected, even though characteristic AMF hyphae and/or vesicles were
present. Numbers of plants of each species positively identified as
harboring AMF and/or DSE are indicated in parentheses (number/total
sampled). The proportions of roots containing arbuscules, vesicles,
AMF hyphae, DSE, and total colonization by fungi are presented
mean percentages ± 1 standard error
Taxon Species AMF
(+/−)
DSE
(+/−)
Arbuscules Vesicles AMF
Hyphae
DSE Total
Colonization
(all AMF and
DSE structures)
MAGNOLIOPHYTA (ANGIOSPERMS)
Liliopsida (Monocotyledons)
Alismataceae Sagittaria lancifolia ? (0/5) + (4/5) 0 0.40± 0.40 20.20± 8.62 6.80± 2.48 27.40 ± 10.39
Cyperaceae Eleocharis cellulosa ? (0/3) + (3/3) 0 0.67± 0.67 1.67± 1.67 43.33± 8.11 44.00 ± 8.72
Eleocharis montevidensis ? (0/3) + (3/3) 0 0.33±0.33 7.33 ± 1.45 46.00± 6.81 50.67 ± 5.17
Schoenoplectus tabernaemontani −(0/3) + (3/3) 0 0 0 43.33± 13.74 43.33 ± 13.74
Schoenoplectus americanus ? (0/5) + (4/5) 0 0 62.20± 11.80 19.20± 15.77 83.20 ± 7.06
Schoenoplectus robustus + (3/5) + (5/5) 0.60± 0.25 2.60 ± 1.60 67.80 ± 4.81 9.20 ± 2.82 80.20± 5.67
Typhaceae Typha domingensis ? (0/3) + (3/3) 0 0.33± 0.33 0.33± 0.33 32.33 ± 7.69 33.00 ± 7.57
Magnoliopsida (Dicotyledons)
Amaranthaceae Alternanthera philoxeroides + (5/5) + (1/5) 6.60± 3.26 8.60±2.44 65.40± 4.07 0.80±0.80 81.4±7.94
Amaranthus australis ? (0/3) + (3/3) 0 2.67 ±1.45 18.67±4.26 13.33±3.93 29.00±5.03
Asteraceae Symphyotrichum subulatum + (3/3) + (3/3) 10.00±1.53 26.67± 5.84 49.33 ± 2.33 28.00 ± 6.66 67.00±6.24
Baccharis halimifolia + (3/3) + (3/3) 12.33± 6.44 8.33 ± 3.29 32.67 ±1.45 40.33 ±8.57 61.67±4.48
Iva frutescens + (3/3) + (3/3) 8.67± 1.20 9.67±6.17 28.67± 10.2 42.00±14.93 65.00± 14.52
Convolvulaceae Ipomoea sagittata + (2/3) + (2/3) 2.33± 1.20 0.33±0.33 62.33± 13.28 17.67 ± 8.88 82.33±5.70
Euphorbiaceae Triadica sebifera + (3/3) + (2/3) 21.33± 3.18 2.33 ± 1.86 69.67 ±6.69 2.00±1.15 96.33±1.20
Fabaceae Vigna luteola + (5/5) + (1/5) 27.20± 4.66 4.80 ± 2.40 45.60 ±5.12 0.60±0.60 76.60±8.94
Sesbania herbacea + (2/3) + (3/3) 9.67± 6.49 8.33±6.35 35.33± 14.97 12.00 ± 1.00 44.67±12.73
Polygonaceae Polygonum punctatum + (3/5) + (5/5) 1.00± 0.55 0.80 ± 0.58 49.40 ± 7.81 7.80± 3.20 58.80 ± 10.29
PINOPHYTA (CONIFERS)
Cupressaceae Taxodium distichum + (3/3) + (1/3) 29.33± 1.67 10.67±0.88 55.67±0.33 0.33±0.33 94.33± 0.88
194 Wetlands (2010) 30:189–199
and herbivory by nutria (Myocastor coypus) (Myers et al.
1995; Shaffer et al. 2009b). Together, these factors have
contributed to wetland loss that is greater than anywhere in
the country.
The high density of root-inhabiting fungi found in this
study suggests that fungi may play an important role in this
stressed ecosystem. Mycorrhizae are known to provide
protection against “physiological drought”(Schimper 1898)
caused by salt stress in coastal wetlands. Therefore, plants at
Turtle Cove may rely on their fungal symbionts’ability to
rehydrate them (Hildebrandt et al. 2001). In addition, AMF
can improve tolerance of plants to flooding (Neto et al. 2006)
and influence resource allocation (Stevens and Peterson
2007), which is important when nutrients are limiting.
Benefits provided by AMF and DSE to their individual
plant hosts may extend to the entire community. Wolfe et
al. (2006) showed that presence of AMF influenced wetland
plant community structure. They suggested that wetlands are
affected not only by abiotic factors that change soil properties,
but also by an interaction of abiotic and biotic factors where
AMF provide amelioration for some negative abiotic effects.
In addition, Fuchs and Haselwandter (2004)showedthat
monocots, which dominate fen communities, contain higher
levels of DSE than less abundant, sympatric dicots, which
are heavily colonized by AMF. This suggests that DSE may
confer a competitive advantage for their plant hosts.
One important outcome of this study is that DSE not
only were abundant in this degrading habitat but were
negatively correlatated to AMF in both monocots and
dicots. Dark septate endophytes have not been studied for
their potential role in wetland plants, but there is a growing
body of evidence that suggests these fungi may play an
important ecological role in wetland communities similar to
that of AMF (Jumpponen 2001). The negative correlation
may either suggest competition among fungal types, or that
the two fungal types perform optimally under different
ecological conditions. For instance, it has been shown that
DSE are commonly found in low nutrient environments
(Peterson et al. 2004), suggesting that DSE may interact
with their plant hosts, the environment, and other fungi in
wetlands to influence community structure. The high
density of DSE found in the marsh at the Turtle Cove
Environmental Research Station warrants further study of
these endophytes’roles. Such studies might examine: 1)
effects of changing environmental conditions (e.g.,
increases in flooding and salinity) on DSEs ability to
colonize wetlands plants; 2) the potential effects of DSE on
plant hosts’competitive ability under changing environ-
mental conditions; 3) the role of plant host on DSE
colonization; 4) identification of species of DSE in wetland
plant hosts and under varying ecological conditions; and 5)
possible interactions between DSE and AMF under varying
ecological conditions.
Another important outcome of this study is the docu-
mentation of AMF in T. distichum. Baldcypress is an
important component of southern U.S. swamps and has
been shown to resist wind-throw during intense hurricanes
(Noel et al. 1995; Shaffer and Day 2007; Shaffer et al.
2009a,b). A consequence of the loss of forested wetlands
dominated by T. distichum is, therefore, diminished
hurricane protection (van Heerden et al. 2006; Day et al.
2007; Shaffer et al. 2009a). The Pontchartrain Basin was
once old-growth Taxodium distichum—Nyssa aquatica
swamp, but today it is rapidly converting to marsh and
open water (Barras et al. 2003). Although efforts are
currently being made toward the restoration of this
ecosystem throughout Louisiana, improving this ecosys-
tem’s health and increasing its productivity have yet to be
attained (Shaffer and Day 2007). Mycorrhizal fungi may be
important in attempts to restore degrading wetlands
Fig. 3 Nonmetric multidimensional scaling ordination showing pattern
of colonization among plant groups by AMF and DSE. Graphs A and B
are the same NMS analysis, but each has been overlayed with circles to
indicate either AMF (a)orDSE(b) colonization. The x-axis represents a
gradient from more AMF colonized on the left to more DSE colonized
on the right. 3A illustrates colonization only by AMF and 3B illustrates
colonization only by DSE. The size of the circle indicates the level of
colonization by AMF or DSE within a particular plant group. The larger
the circle, the higher the level of colonization by a particular fungal
type. T=tree, S=shrub, D=dicot, and M=monocot
Wetlands (2010) 30:189–199 195
containing baldcypress. Introducing appropriate AMF and
DSE to the soils may facilitate regeneration of baldcypress.
The importance of AMF in restoration/forest strategies in
terrestrial ecosystems is well documented (Smith and Read
1997), but this has not been examined in coastal systems.
This may be a particularly relevant area to explore given the
relatively high level of AMF colonization in desirable
wetland plant species, especially T. distichum.
Introduction of root endophytes to an ecosystem,
however, should be done with caution. Undesirable
consequences, such as increased likelihood of invasion by
non-native plants, might result from adding root endophytic
fungi to soils in which they have not evolved, or to plants
with which they have not evolved (Collins Johnson et al.
2006). Generalist AMF species often form associations
with exotic plants. These associations, compounded by the
absence of enemies of exotic plants (Richardson et al. 2000;
Keane and Crawley 2002), could facilitate invasion. Some
studies suggest that invasive plants suppress growth of
native plant species by preventing natives from being
colonized by beneficial fungi (Callaway et al. 2008; Nijjer
et al. 2008). Louisiana wetlands are threatened by invasive
species such as Triadica sebifera and Alternanthera
philoxeroides. Both these plant species showed high levels
of AMF colonization in this study. Along with other
stressors, these invasive species might accelerate the current
trajectory toward open water if they reduce or prevent
native plants from forming mycorrhizal associations.
Arbuscular mycorrhizal fungi also exhibit parasitic
characteristics under certain environmental conditions, such
as when nutrients are abundant. For example, Johnson
(1993) found that inoculating plants with AMF isolated
from fertile soils resulted in a net cost in biomass and
fitness for the plant. In contrast, plants grown with AMF
isolated from soils low in nutrients were taller and had more
inflorescences. Therefore, it is crucial that more studies be
conducted to demonstrate the role of root endophytes in
wetlands, as well as environmental effects on plant-fungal
interactions. Determining which fungal species are native to
a particular soil and which species of fungi are associated
with which plant species will allow managers to make
informed decisions when including fungi in conservation/
restoration plans. Studying these fungi and their role in
plant dynamics, however, is subject to the presence of the
plant hosts. Obviously, once a wetland converts to open
water, it will be difficult to determine which fungi were
present, as well as relationships with indigenous plants.
Because wetlands in Louisiana are currently being lost at
rates anticipated to occur soon in other deltaic regions
around the world, research in Louisiana can be used as a
model for studies elsewhere that soon will be affected by
sea-level rise. Thus, identification of the AMF and DSE in
Louisiana coastal wetlands is of some urgency.
Acknowledgments We thank Southeastern Louisiana University for
granting us permission to use their boats and to have access to Turtle
Cove Environmental Research Station, as well as for partially funding
this study. Biograds, LSU’s Biological Sciences graduate student
organization, also provided partial funding for this project. We thank
Jane Gurney and Misty Wellner for assisting with laboratory work,
and Matt Slocum, Rae Crandall, Ellen Leichty, Yalma Vargas-
Rodriguez, Becky Carmichael, Erin Lawrence, Darin Ellair, Mindy
McCallum, Tracy Hmielowski, Heather Passmore, Kyle Harms, and
two anonymous reviewers for their helpful comments.
Appendix
Table 3 Four plant species that were examined in this study were also
examined in previous studies for their mycorrhizal status. We
compared mycorrhizal levels reported in other studies to levels found
in this study. Dark septate endophytes were not examined for any of
these species in any previous studies. Schoenoplectus tabernaemon-
tani was found to be colonized by AMF in three studies (Wetzel and
van der Valk 1996; Cooke and Lefor 1998; Bauer et al. 2003),
whereas we were not able to find any evidence of AMF in our
specimens. We did, however, find that S. tabernaemontani was
heavily colonized by DSE. In addition, many studies do not report
arbuscular colonization separately from total colonization, which
makes it difficult to know what proportion of the root was, indeed,
colonized by AMF. Standard error is reported if it was reported in a
particular study
Plant Species Reference AMF
(arbuscules only)
AMF (Total
colonization)
DSE Notes
Schoenoplectus
tabernaemontani
This study 0% 0% 43.33%± 13.74% -field collections
As Schoenoplectus
maritimus
1. Gouraud
et al. 2008
0% 0% N/A -field collections
-examined roots for all structures
characteristic to AM fungi
as Scirpus validus 2. Bauer et al.
2003
3–85% N/A -field collections
-total colonization includes arbuscules,
as well as vesicles and hyphae
196 Wetlands (2010) 30:189–199
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