Associations of Methanotrophs with the Roots and Rhizomes of Aquatic Vegetationt
←
→
Page content transcription
If your browser does not render page correctly, please read the page content below
APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Sept. 1994, p. 3220-3227 Vol. 60, No. 9
0099-2240/94/$04.00+0
Copyright © 1994, American Society for Microbiology
Associations of Methanotrophs with the Roots and
Rhizomes of Aquatic Vegetationt
GARY M. KING*
Darling Marine Center, University of Maine, Walpole, Maine 04573
Received 25 April 1994/Accepted 8 July 1994
Results of an in vitro assay revealed that root-associated methane consumption was a common attribute of
diverse emergent wetland macrophytes from a variety of habitats. Maximum potential uptake rates (Vmaxp)
varied between about 1 and 10 ,umol g (dry weight)-' h-1, with no obvious correlation between rate and gross
Downloaded from http://aem.asm.org/ on March 16, 2020 by guest
morphological characteristics of the plants. The Vmaxp corresponded to about 2 x 108 to 2 x 109 methano-
trophs g (dry weight) -, assuming that root-associated methanotrophs have cell-specific activities comparable
to those of known isolates. Vm.p varied seasonally for an aquatic grass, Calamogrostis canadensis, and for the
cattail, Typha latifolia, with highest rates in late summer. Vm,, p was well correlated with ambient temperature
for C. canadensis but weakly correlated for T. latifolia. The seasonal changes in Vmaxp, as well as inferences from
apparent half-saturation constants for methane uptake (Kapp; generally 3 to 6 ,uM), indicated that oxygen
availability might be more important than methane as a rate determinant. In addition, roots incubated under
anoxic conditions showed little or no postanoxia aerobic methane consumption, indicating that root-associated
methanotrophic populations might not tolerate variable oxygen availability. Hybridization of oligodeoxynucle-
otide probes specific for group I or group II methylotrophs also varied seasonally. The group 11-specific probe
consistently hybridized to a greater extent than the group I probe, and the relative amount of group II probe
hybridization to C. canadensis root extracts was positively correlated with Vmaxp.
Methane transport through aboveground plant tissues often Holtzapfel-Pschorn et al. (20, 21) have reported that methane
exceeds diffusive and ebullitive fluxes across the sediment- oxidation (presumably in the rhizosphere) consumed up to
water (or air) interface in vegetated wetlands (e.g., see refer- 80% of total methane production in vegetated rice paddy soils.
ences 11, 12, 16, 37, and 43). In rice paddies and natural Stimulation of methane efflux from rice plants incubated with
wetlands, up to 90% of the total methane emission occurs via nitrogen headspaces or acetylene, a potent inhibitor of meth-
plants (e.g., see references 7, 8, 27, 28, 36, and 38), with the ane oxidation (1), has provided additional evidence for rhizo-
relative extent of emission varying as a function of shoot spheric methane consumption.
development. At present, the significance of plant emissions The rhizospheres (or roots) of other aquatic plants also
appears independent of physiognomy, even though absolute oxidize methane. Epp and Chanton (14) have reported meth-
emission rates vary substantially among wetland systems. For ane consumption by roots of Pontederia cordata and Sagittaria
example, plant emissions dominate methane flux in a Canadian lancifolia. Increases in methane efflux from the leaves of
fen (primarily Carex spp.; low total methane flux [42]), the "weed" species incubated in the presence of acetylene or
Florida Everglades (primarily Cladium jamaicense; intermedi- nitrogen provide other evidence (21) for oxidation. King et al.
ate total flux [43]), and rice paddies (high total flux [8, 36, 38]). (26) have observed rapid methane oxidation by live, sediment-
The uniformly large contribution of plants to total wetland free roots of several plant species from the Florida Everglades.
emissions suggests that belowground plant surface area and These observations suggest that root tissues and their meth-
gas phase transport within plant tissues are more important anotrophic associations are an important locus controlling
determinants of methane dynamics than is diffusional transport methane flux to the atmosphere.
across the sediment-water interface. The net result is that In this study, basic characteristics of root-associated meth-
wetland plants provide not only the reduced carbon fuel that anotrophy have been assessed by using an in vitro approach
drives methanogenesis but also the exhaust system that vents it with multiple plant species from a number of different systems.
to the atmosphere. Activities have been measured on a seasonal basis and corre-
Although the mechanisms and dynamics of methane emis- lated with temporal variations in the hybridization of oligode-
sions from aboveground plant tissues have been well docu- oxynucleotide probes specific for sequences from the 16S
mented, the controls of methane flux into belowground plant rRNA of group I and group II methylotrophic bacteria. Results
tissues are poorly known. In addition to physical processes of these analyses indicate that associations of methanotrophs
(e.g., diffusion), biological controls likely determine the char- with aquatic plants are very common, that activities are
acteristics of methane transport into roots and rhizomes. In probably limited by oxygen availability, and that group II
particular, root-associated, bacterial methanotrophy probably methylotrophs dominate the associations.
functions as a filter that removes much of the methane influx.
DeBont et al. (13) have observed enhanced populations of
methane-oxidizing bacteria in the rice rhizosphere, and MATERIALS AND METHODS
Field sites. Plants for methane uptake and production assays
were collected primarily from two wetlands. Vilhelmsborg S0,
Phone: (207) 563-3146. Fax: (207) 563-3119. Electronic mail ad- near Aarhus, Denmark (described previously by King [22, 23]),
dress: GKing@Maine.Maine.edu. contained a diversity of herbaceous emergent macrophytes in a
t Contribution 275 from the Darling Marine Center. silty sand of the riparian zone of a stream feeding a small lake.
3220VOL. 60, 1994 ROOT-ASSOCIATED METHANOTROPHY 3221 The degree of submergence of this zone fluctuated with stream 1 to 3%; methane depletion was monitored at intervals until level. A second site located near Newcastle, Maine, was final concentrations were
3222 KING APPL. ENVIRON. MICROBIOL.
TABLE 1. Methane consumption by live, sediment-free roots
_
140.0
and rhizomesa
E
120.0 Species, habitat Type Rate ± SE Locale Dateb
E
c 100.0 Veronica beccabunga H 6.4 ± 1.9 DK Ju-Jl
Mentha aquatica L. H 1.7-3.9 DK Ju-Jl
s80.0 Lycopus europeus L. H 2.8 ± 0.5 DK Ju-Jl
.C Ranunculus sceleratus L. H 1.7 ± 0.3 DK Ju-Jl
E 60.0 Ronpia amphibia (L.) Bess H 1.0-1.6 DK Ju-Jl
Alisma plantago aquatica L. WP 4.5 ± 1.2 DK Ju-Jl
c-) 40.0 Typha latifolia L. C 5.4 ± 1.1 DK Ju-Jl
0.
co Sparganium erectum B 4.5 ± 0.2 DK Ju-Jl
cu 20.0 Glyceria maxima G/S 5.1 ± 1.6 DK Ju-Jl
Carex spp. GIS 0.1-4 ME YR
0 10 20 30 40 50 Calamagrostis canadensis G/S 0.1-5.3 ME YR
Downloaded from http://aem.asm.org/ on March 16, 2020 by guest
Time (hr) Typha latifolia B 0.1-3.5 ME YR
FIG. 1. Methane consumption by washed, sediment-free excised Cladium jamaicense, peat G/S 0.7 ± 0.4 FL Oc
roots and rhizomes of M. aquatica from Vilhelmsborg S0; the symbols Cladium jamaicense, marl G/S NDC FL Oc
0, M, and M represent time course data from 1-g (fresh weight) Eleocharis interstincta G/S ND FL Oc
samples from three individual plants. Sagittaia lancifolia, peat WP 0.5 ± 0.04 FL Oc
Sagittaria lancifolia, marl WP ND FL Oc
Cninum americanum Am 0.3 ± 0.1 FL Oc
duced by incubating the membranes at -80°C for 24 to 48 h a
Vmaxp are micromoles gdw-1 hour-'. Location codes are as follows: DK,
with Kodak X-Omat-AR X-ray film and an intensifying screen. Vilhelmsborg S0, Denmark; ME, Newcastle, Maine; FL, Florida Everglades (see
reference 26). Broad taxonomic groupings are as follows: H, herbs; G/S, grasses
After the autoradiograms had been developed, the membranes and sedges; C, cattails; B, bur reeds; WP, water plantain; Am, amyrillis.
were sectioned and the bound radioactivity corresponding to b Ju, June; Jl, July; Oc, October; YR, periodically through a seasonal cycle.
each sample slot was determined by liquid scintillation count- C ND, not determined.
ing. Crude nucleic acid extracts derived from the cell pellets of
Methylobacter albus BG8 (group I; formerly Methylomonas
albus BG8 [3]) were used as a positive control for the 10-y
probe and a negative control for 9-ao; extracts of Methylosinus Vma,p (Table 1) were typically estimated from the linear
depletion of methane at saturating concentrations. However,
tnichosporium OB3b (group II) served as a positive control for for two species, S. erectum and M aquatica, Vmp were also
9-ao and a negative control for 10-,y; a commercial preparation calculated from nonlinear parameter estimation based on
(Sigma Chemical Co.) of 16S rRNA from Escherichia coli was progress curves (Fig. 1). These rates compared favorably with
used as a negative control for both probes. Group I and II estimates based on a linear regression of uptake at the
methanotrophs have been previously defined as coherent beginning of the progress curve incubations (Table 2) and with
clades on the basis of a variety of physiological and biochem- initial rates of depletion by separate samples at saturating
ical traits; for example, group II but not group I meth- methane concentrations (Table 1). In addition, progress curve
anotrophs fix nitrogen, possess an incomplete tricarboxylic acid analyses provided estimates of Kapp; these values were similar
cycle, and assimilate methane via serine (see references 3 and to estimates calculated by assuming that Kapp = Vmap k-1
24 and references therein). (Table 2). The first-order rate constant, k, was determined
from regression analysis of the region of the progress curves
RESULTS where methane depletion was exponential. Rate constants
determined from incubations of C. canadensis with 100 ppm of
Methane consumption. The washed, excised roots and rhi- methane, and the corresponding Vm,p values determined
zomes of all of the macrophytes sampled at Vilhelmsborg S0 from incubations with 1- to 3% methane, were also used to
consumed methane actively and without a lag (Fig. 1). Sub- estimate Kapp (Table 2). All of the Kapp values ranged between
stantial differences in Vm,p were observed among the taxa 3 and 6 ,uM, irrespective of the plant species examined or
(Table 1), but these differences were not associated with any calculation procedure.
apparent taxonomic characteristics (e.g., aboveground mor- Vmaxp varied seasonally over a 30-fold range (Fig. 3A) for
phology). Variability within pooled root samples was generally roots and rhizomes of T. latifolia and C. canadensis incubated
relatively low, with a few exceptions. Variation for replicate at ambient field temperatures. Through winter, spring, and
samples from individual plants was more substantial for three early summer, changes in VmS,p basically paralleled changes in
taxa (Sparganium erectum, Ronpia amphibia, and M. aquatica field temperatures, although anomalously low Vmnp were
[Table 2]), but uptake estimates based on the means from observed for T. latifolia in April 1992. In addition, Vm,p for
three individuals for each of these species were comparable in both T. latifolia and C. canadensis remained high while tem-
magnitude and variability to estimates from a separate assay peratures were relatively low in September 1992. Vm,p for C.
based on pooled samples from multiple individuals (Table 1). canadensis were typically greater than Vm,p for T. latifolia, but
Methane consumption also occurred without a lag for roots the differences were statistically significant (t test; P c 0.05)
and rhizomes of plants collected near Newcastle, Maine (Fig. only during the late spring-summer months. Seasonality in
2; Tables 1 and 2). Levels of variability for these pooled methane consumption was also evident for incubations of C.
samples were comparable to those for samples from Vilhelms- canadensis at ambient laboratory temperatures (22 to 25°C;
borg So. Vmap for T. latifolia from Denmark were also Fig. 3B). In particular, Vm,p for methane consumption in-
comparable to rates for the same species collected in Maine at creased through spring and summer, as did rates for samples
a similar time. incubated at ambient field temperatures. Vmaxp declinedVOL. 60, 1994 ROOT-ASSOCIATED METHANOTROPHY 3223
TABLE 2. Summary of kinetic data for washed, sediment-free roots A
and rhizomesa
Species and sample type (n) Vmap + 1 SE Kapp ± 1 SE 4.0 25
R amphibia _= 3.5
A (3)
B (3)
0.6
1.0
± 0.2
± 0.2
.c.3O 3.0
20 >
cm
C (3) 1.4 0.1 2.5
Mean A, B, C 1.0 ± 0.3 E 15
Pooled (3) 1.3 ± 0.2 5.7 ± 0.8 2.0 3
a) c0
_
S. erectum am 1.5 CD
(D
A (3) 3.4 ± 0.9 CL
C
B (3) 2.0 ± 0.5 4) 1.0 ,i
C (3) 2.1 ± 0.6 co$
Mean A, B, C 2.5 ± 0.7 ; 0.5
Downloaded from http://aem.asm.org/ on March 16, 2020 by guest
CD
Pooled (3) 3.6 ± 0.4 4.1 ± 0.8
NPE (6) 1.7 ± 0.3 1.3 ± 0.6 0.0
50 100 150 200 250 300 350
M. aquatica Julian day
A (3) 1.5 ± 0.2 B
B (3) 1.5 ± 0.3
C (3) 2.2 ± 0.3 6.0
Mean A, B, C 1.7 ± 0.5
Pooled (3) 2.0 ± 0.3 3.8 ± 0.9 ~c 5.0
0
NPE (3) 3.9 ± 2.2 4.2 ± 2.7
C canadensis (pooled) (3) 1.6 ± 0.4 5.8 ± 0.5 4.0
a All rates are in micromoles gdw-1 hour- 1, and Kapp is in micromolar. NPE, 'a 0
CD 3.0
nonlinear parameter estimation for kinetic data. For R amphibia, S. erectum, and I.-
75
M. aquatica, samples either are from individual plants (A, B, or C) or are 0)
replicates from the pooled roots of multiple plants; all other data are based on CD 2.0
replicates from pooled material only. Numbers of replicates are given in 0
parentheses. 0
1.0
0 0
0 c
0.0
sharply during late fall and remained low but measurable 50 100 150 200 250 300 350
through the winter. Julian day
On the basis of an analysis of the natural logarithm of FIG. 3. (A) Seasonality of methane consumption by triplicate
methane uptake data, the activity of C. canadensis was strongly samples of washed sediment-free roots and rhizomes of C. canadensis
correlated with ambient field temperature during an annual (0) and T. latifolia (O), incubated at ambient field temperatures (a).
cycle, with the exception of a point in late fall (Fig. 4). A much The standard error bars (± 1 standard error) for C. canadensis are
weaker correlation was observed for T. latifolia (unpublished representative of errors (coefficients of variation, about 15%) for all
samples. The arrow indicates anomalously low activity for T. latifolia.
(B) Methane consumption by washed sediment-free roots and rhi-
zomes of C. canadensis incubated at 22 to 25°C. Uptake rates are
means of triplicates from pooled root samples with coefficients of
E variation typically 10 to 15%.
I-I 10.0
0a
co
.0.
8.0 data). The apparent activation energy for methane uptake by
C
CU
0
co
C. canadensis derived from an analysis of the data according to
0
6.0 the Arrhenius relationship was 97.3 kJ °K-' mol' Because of .
C.)
a) the weak correlation between temperature and activity, activa-
co tion energies were not calculated for T. latifolia.
4.0 Oligonucleotide hybridization with RNA extracts. Seasonal-
{L
E
a
ity was also evident in the binding of radiolabeled oligonucle-
a) otide signature probes to total root and rhizome nucleic acid
2.0 extracts. Liquid scintillation assays provided a relative indica-
0.
m
tion of the extent of hybridization by the probes, since the
~0'a w > probes were used at equivalent specific activities and compa-
cDs
a) 0.0 rable stringencies. Extracts from both C. canadensis and T.
C: 0 10 20 30 40 50 latifolia hybridized with both the group I and group II meth-
Time (hr) ylotroph probes (Fig. 5), with the strongest signals in spring
FIG. 2. Methane consumption by washed, sedimei nt-free excised and summer. The group I and II probes hybridized with the
Newcastle,
roots of T. latifolia, C. canadensis, and a Carex sp. fri om appropriate positive controls, but not with the negative con-
Maine. Initial headspace methane concentrations we re 5,000 ppm trols. For both plant extracts and all sampling dates, the group
(about 200 nmol cm-3); the arrow indicates atmospherric methane. II probe hybridized to a greater extent than did the group I3224 KING APPL. ENVIRON. MICROBIOL.
-
A
1 .5
c-
1.0
8000
1-
7000
Ecm 0.50
E0L 6000
a 0.0 -0
r 5000
-0.50 a)
.0
O 4000
0.
CZ -1.0 'D
CL 3000
E
0
0-
(i) -1.5 0
m 2000
CD -2.0 1000
a)
Downloaded from http://aem.asm.org/ on March 16, 2020 by guest
-c -2.5
co co) CD co cr) cII cII
0
CZ o o oo o o o 0 F Ap M Ju S
LC)
Co t
Lt LO
LO) 0 LO) 0
(D
LO
(D B
c) Co Cf) C) C') Co C)
TV1 (Kelvin)
5000
FIG. 4. C. canadensis methane consumption rates versus incuba-
tion temperature (T). The regression line excludes anomalously low
consumption rates from September (O). The relationship between 4000
uptake and temperature is described by ln uptake = 40.2 + EQ
11,700 * T-', r2 = 0.97 (PVOL. 60, 1994 ROOT-ASSOCIATED METHANOTROPHY 3225
8000 and the more general differences in uptake rates among taxa
(Table 1) indicated that species-specific characteristics, e.g.,
E 7000 aerenchyma architecture, root morphology, and root physiol-
0.
a) 6000 ogy, among others, constrain the extent and activities of
CD methanotrophic associations.
.0
0 5000 The differential association of methanotrophs with aquatic
a plants and differences in responses to seasonal changes in
0. 4000 temperature and plant physiology have important implications
0
for predicting the behavior of wetland systems in the context of
3000 climate change. Current climate change models treat wetland
CD
2000 gas exchange as a parameter that changes mostly as a function
of wetland area or hydrology (e.g., see references 33 and 34).
0 Wetland area and hydrology are certainly key determinants,
1000
. but future emissions may not be accurately predictable without
Downloaded from http://aem.asm.org/ on March 16, 2020 by guest
0 considering the impact of changes in wetland species compo-
0.00 1.00 2.00 3.00 4.00 5.00 6.00 sition and variabilities in plant-methanotroph interactions.
Methane oxidation Seasonality was also evident in the hybridization signals
FIG. 6. Correlation (r = 0.95) between bound type II oligode observed with oligonucleotide probes for 16S rRNA (Fig. 6
oxynucleotide probes and Vmp for C. canadensis (C)). Data for T and 7). Probe binding to extracts from the roots and rhizomes
latifolia (a) are shown without a correlation (r = 0.2'0). Anomalous of C. canadensis correlated well with maximum potential
points (U and O; elevated rate and low-level bindinl g, respectively) methane uptake rates, if data from late September were
were excluded from the correlation analysis. excluded from the analysis (Fig. 6). In late September, meth-
ane uptake rates for C. canadensis were high, even though
ambient temperatures had declined sharply, the plants had
temperature (22 to 25°C) (Fig. 3B) provided e *vidence
ence that
begun to senesce, and the level of probe hybridization was
changes in methanotrophic populations contril
3A).
that
In
ied
(Fgd
relatively low. This pattern may have been the result of a
smaller but more active methanotrophic population. Lower
dynamics of subsurface methane consumption I
general, methane uptake was greatest in the lal sprlng and (Fsring
me 3a)nd
rates of root respiration accompanying senescence and de-
creased temperatures might temporarily have reduced compe-
summer when the plant hosts were most active r
This suggested that changes in methanotrophic populations petabolically. tition between methanotrophs and plant tissues for oxygen,
thus enhancing uptake.
were coupled to plant processes, with root oxyg(enation likely The weak correlation between probe hybridization and
of greatest significance. However, the extent of seasonal uptake rates for T. latifolia suggests that several factors may
change was not equally apparent in the two taxa examined, confound interpretation of the results. Perhaps the most
Changes in activities associated with T. latifolfia roots and significant limitation is the fact that the 9-a and 10--y probes
rhizomes were much less pronounced than in C7. canadensis. are specific for methylotrophs, but not methanotrophs (40). As
The differences between taxa in seasonal responses (Fig. 3) a consequence, an unknown fraction of the observed signal
may arise from bacteria that utilize nonmethane C1 substrates
(e.g., methanol). Methylotrophs (e.g., the pink-pigmented fac-
Anoxic Oxic ultative methylotrophs) have been previously documented as
common components of the phyllosphere (e.g., see reference
18 200 9) and likely occur in the rhizosphere as well. The relative
'3 K: abundances of methylotrophs versus methanotrophs may vary
cm 15
1 CD both temporally and spatially for a given plant and vary among
_p, plant taxa. Other confounding factors include the variability in
E
^ 12 ECD ribosome (i.e., oligonucleotide target) content as a function of
c 120 x physiological status and variations among the methanotrophic
4nz and methylotrophic taxa in ribosome content for a given
0 9
ig physiological state.
:0 80 m In spite of these caveats, the probe data indicate that
0 0 bacteria from the group II methanotroph-methylotroph cluster
C% s_ 40Qco (17) dominate root and rhizome associations throughout the
_ > year for both C. canadensis and T. latifolia. A similar pattern
CD
has been observed for limited analyses of RNA extracts from
0 o 0 the methane-consuming roots of a Carex sp. obtained from a
0.00 20.0 40.0 60.0 80.0 100 different site (25a), and for extracts of the floating, stemless
Time (hr) Lemna minor (17). Naphthalene oxidation by C. canadensis
plant roots (25a) is consistent with domination by group II
FIG. 7. Methane production and postanoxia methanie consumption methanotrophs, since group I methanotrophs typically cannot
by roots and rhizomes of C. canadensis (open symbols) and T latifolia facilitate this process (5).
(closed symbols). Roots were incubated initially undei rsanoxic
condi-
tions for the time period indicated. Subsequently, root swere aerated
Although it is not possible by using existing data to assign
and consumption of added methane under oxic conditi definitively the methanotrophic activity of aquatic plants to
was compared with uptake by fresh roots incubated in parallel under either group I or group II taxa, it appears that the phyllosphere
oxic conditions only (A and A). Error bars indicate ± 1 standard error generally favors or promotes associations of group II species.
for triplicates; error bars are within the symbols fcwr all methane Group II methanotrophs also appear to dominate in methane-
consumption assays. enriched soils (4, 30). In contrast, group I methanotrophs3226 KING APPL. ENVIRON. MICROBIOL.
appear to dominate the metalimnia of two lakes (17) and the ing methane transport to the atmosphere (e.g., see reference
symbionts of mytilid mussels (e.g., see reference 6). It is 18).
tempting to speculate that the diazotrophic character of group The results of incubations with both C. canadensis and T.
II methanotrophs provides a selective advantage in systems latifolia also indicated that methanogenesis is strongly sensitive
where nitrogen is potentially limiting (e.g., methane-enriched to oxygen, as expected, and that methanotrophic activity does
soils and plant roots), while greater growth yields provide an not appear to recover from relatively brief exposures to anoxia
advantage for group I methanotrophs in systems where nitro- (Fig. 7). The sensitivity of root-associated methanotrophy to
gen is relatively abundant or where methane is limiting (e.g., anoxia differs distinctly from the sensitivities observed for
the water column and animal tissues). This pattern conforms to peats, sediments, and cultures (22, 25, 26, 31, 32). In each of
the outcome of competition studies using the group I (M. albus the latter two cases, it is evident that methanotrophs possess a
BG8) and the group II (M. trichosporium OB3b) meth- notable ability to survive anoxia, and to rapidly consume
anotrophs in continuous-flow reactors (15). A more detailed methane after reintroduction of oxygen. The anoxia intoler-
analysis of root-associated methanotroph phylogeny and the ance of root-associated methanotrophy observed in this study
determinants of methanotroph distribution should provide may arise from toxic effects of fermentation end products
important insights into the controls and dynamics of root- resulting from both plant cell and microbial activity. Whether
Downloaded from http://aem.asm.org/ on March 16, 2020 by guest
associated activities in situ. Recently developed signature a similar intolerance of anoxia occurs in situ is unclear.
oligonucleotide probes specific for various phylogenetic and However, it is certain that the roots of at least some aquatic
physiological subgroups of methanotrophs, as well as probes plants are variably oxygenated because of diurnal and seasonal
specific for methylotrophs, will allow much greater resolution shifts in plant metabolism. As a consequence, root-associated
of these issues (4). methanotrophy may be restricted to tissues that are continu-
Kinetic data (Table 1) provide insights into other controls of ously oxygenated. Any such limitation could prove an impor-
root-associated methane uptake. In spite of differences in root tant determinant of the relative significance of root-associated
and rhizome morphology, the Kapp values for all plants used in methanotrophy for the magnitude of wetland methane fluxes.
this study were about 3 to 6 ,uM. These values are comparable In conclusion, the rhizoplane, and likely the interior, of the
to results from sediments and some cultures (see review in roots and rhizomes of aquatic vegetation promotes the growth
reference 24) but are generally lower than dissolved methane of methanotrophs, thereby substantially expanding the volume
concentrations in the root zones of typical wetlands (e.g., see of the methanotrophic zone below the sediment surface. The
references 10, 35, and 44). Pore water methane concentrations activity of root-associated methane-oxidizing bacteria varies
on the order of 10 to 100 ,uM are often observed, with values seasonally and among plant species, may be dominated by
ofVOL. 60, 1994 ROOT-ASSOCIATED METHANOTROPHY 3227
the marine environment: ultrastructural, biochemical and molec- 27. Nouchi, I., and S. Mariko. 1993. Mechanism of methane transport
ular studies, p. 315-328. In J. C. Murrell and D. P. Kelly (ed.), by rice plants, p. 336-352. In R. S. Oremland (ed.), Biogeochem-
Microbial growth on C, compounds. Intercept Ltd., Andover, istry of global change: radiatively active trace gases. Chapman &
United Kingdom. Hall, New York.
7. Chanton, J. D., and J. W. H. Dacey. 1991. Effects of vegetation on 28. Nouchi, I., S. Mariko, and K. Aoki. 1990. Mechanism of methane
methane flux, reservoirs, and carbon isotopic composition, p. transport from the rhizosphere to the atmosphere through rice
65-92. In T. D. Sharkey, E. A. Holland, and H. A. Mooney (ed.), plants. Plant Physiol. 94:59-66.
Trace gas emissions by plants. Academic Press, New York. 29. Remsen, C. C., E. C. Minnich, R. S. Stevens, L. Bucholz, and M. E.
8. Cicerone, R. J., and J. D. Shetter. 1981. Sources of atmospheric Lidstrom. 1989. Methane oxidation in Lake Superior sediments.
methane: measurements in rice paddies and a discussion. J. J. G. Lakes Res. 15:141-146.
Geophys. Res. 86:7203-7209. 30. Ringelberg, D. B., J. D. Davis, G. A. Smith, S. M. Pfiffner, P. D.
9. Corpe, W. A., and S. Rheem. 1989. Ecology of methylotrophic Nichols, J. S. Nickels, J. M. Henson, J. T. Wilson, M. Yates, D. H.
bacteria on living leaf surfaces. FEMS Microbiol. Ecol. 62:243- Kampbell, H. W. Read, T. T. Stocksdale, and D. C. White. 1989.
250. Validation of signature polarlipid fatty acid biomarkers for alkane-
10. Crill, P. M., K. B. Bartlett, R. C. Harriss, E. Gorham, E. S. Verry, utilizing bacteria in soils and subsurface aquifer materials. FEMS
D. I. Sebacher, L. Madzar, and W. Sanner. 1988. Methane flux Microbiol. Ecol. 62:39-50.
Downloaded from http://aem.asm.org/ on March 16, 2020 by guest
from Minnesota peatlands. Global Biogeochem. Cycles 2:371-384. 31. Roslev, P., and G. M. King. 1993. TTC, a new water-soluble
11. Dacey, J. W. H. 1981. Pressurized ventilation in the yellow water formazan dye, and applications in starvation responses of bacteria.
lily. Ecology 62:1137-1147. Appl. Environ. Microbiol. 59:2891-2896.
12. Dacey, J. W. H., and M. J. Klug. 1979. Methane efflux from lake 32. Roslev, P., and G. M. King. 1994. Survival and recovery of
sediments through water lilies. Science 203:1253-1254. methanotrophic bacteria starved under oxic and anoxic conditions.
13. DeBont, J. A. M., K. K. Lee, and D. F. Bouldin. 1978. Bacterial Appl. Environ. Microbiol. 60:2602-2608.
methane oxidation in a rice paddy. Ecol. Bull. 26:91-96. 33. Roulet, N. T., R. Ash, W. Quinton, and T. Moore. 1993. Methane
14. Epp, M. A., and J. P. Chanton. 1993. Rhizospheric methane flux from drained northern peatlands: effect of a persistent water
oxidation determined via the methyl fluoride inhibition technique. table lowering on flux. Global Biogeochem. Cycles 7:749-769.
J. Geophys. Res. 98(D):18413-18422. 34. Roulet, N. T., T. R. Moore, J. Bubier, and P. Lafleur. 1992.
15. Graham, D. W., J. A. Chaudhary, R. S. Hanson, and R. G. Arnold. Northern fens: methane flux and climate change. Tellus 44(B):
1993. Factors affecting competition between type I and type II 100-105.
methanotrophs in two-organism, continuous flow reactors. Mi- 35. Sass, R. L., F. M. Fisher, and P. A. Harcombe. 1990. Methane
crob. Ecol. 25:1-17. production and emission in a Texas rice field. Global Biogeochem.
16. Grosse, W., H. B. Buchel, and H. Tiebel. 1991. Pressurized Cycles 4:47-68.
ventilation in wetland plants. Aquat. Bot. 39:89-98. 36. Schutz, H., A. Holzapfel-Pschorn, R. Conrad, H. Rennenberg, and
17. Hanson, R. S., B. J. Bratina, and G. A. Brusseau. 1993. Phylogeny W. Seiler. 1989. A 3-year continuous record on the influence of
and ecology of methylotrophic bacteria, p. 285-302. In J. C. daytime, season and fertilizer treatment on methane emission
Murrell and D. P. Kelly (ed.), Microbial growth on C1 compounds. rates from an Italian rice paddy. J. Geophys. Res. 94(D):16405-
Intercept Ltd., Andover, United Kingdom. 16416.
18. Happell, J. D., J. P. Chanton, G. J. Whiting, and W. J. Showers. 37. Schutz, H., P. Schroder, and H. Rennenberg. 1991. Role of plants
1993. Stable isotopes as tracers of methane dynamics in Ever- in regulating the methane flux to the atmosphere, p. 29-63. In
glades marshes with and without active populations of methane T. D. Sharkey, E. A. Holland, and H. A. Mooney (ed.), Trace gas
oxidizing bacteria. J. Geophys. Res. 98:14771-14782. emissions by plants. Academic Press, New York.
19. Heyer, J., Y. Malashenko, U. Berger, and E. Budkova. 1984. 38. Schultz, H., W. Seiler, and R Conrad. 1989. Processes involved in
Verbreitung methanotropher Bakerien. Z. Allg. Mikrobiol. 24: formation and emission of methane in rice paddies. Biogeochem-
725-744. istry 7:33-53.
20. Holtzapfel-Pschorn, A., R. Conrad, and W. Seiler. 1985. Produc- 39. Sebacher, D. I., R. C. Harriss, and K. B. Bartlett. 1985. Methane
tion, oxidation and emission of methane in rice paddies. FEMS emissions to the atmosphere through aquatic plants. J. Environ.
Microbiol. Ecol. 31:343-351. Qual. 14:40-46.
21. Holtzapfel-Pschorn, A., R. Conrad, and W. Seiler. 1986. Effects of 40. Tsien, H. C., B. J. Bratina, K. Tsuji, and R S. Hanson. 1990. Use
vegetation on the emission of methane from submerged paddy of oligodeoxynucleotide signature probes for identification of
soil. Plant Soil 92:223-233. physiological groups of methylotrophic bacteria. Appl. Environ.
22. King, G. M. 1990. Dynamics and controls of methane oxidation in Microbiol. 56:2858-2865.
a Danish wetland sediment. FEMS Microbiol. Ecol. 74:309-323. 41. Tsuji, K., H.-C. Tsien, R S. Hanson, S. R DePalma, R. Scholtz,
23. King, G. M. 1990. Regulation by light of methane emission from a and S. LaRoche. 1990. 16S ribosomal RNA sequence analysis for
Danish wetland. Nature (London) 345:513-515. determination of phylogenetic relationship among methylotrophs.
24. King, G. M. 1992. Ecological aspects of methane oxidation, a key J. Gen. Microbiol. 136:1-10.
determinant of global methane dynamics. Adv. Microbial Ecol. 42. Whiting, G. J., and J. P. Chanton. 1992. Plant-dependent CH4
12:431-468. emission in a subarctic Canadian fen. Global Biogeochem. Cycles
25. King, G. M. 1993. Ecophysiological characteristics of obligate 6:225-232.
methanotrophic bacteria and methane oxidation in situ, p. 303- 43. Whiting, G. J., J. P. Chanton, D. S. Bartlett, and J. D. Happell.
313. In J. C. Murrell and D. P. Kelly (ed.), Microbial growth on C, 1991. Relationships between CH4 emission, biomass, and CO2
compounds. Intercept Ltd., Andover, United Kingdom. exchange in a subtropical grassland. J. Geophys. Res. 96(D):
25a.King, G. M. Unpublished data. 13067-13071.
26. King, G. M., H. Skovgaard, and P. Roslev. 1990. Methane 44. Wilson, J. O., P. M. Crill, K. B. Bartlett, D. L. Sebacher, R. C.
oxidation in sediments and peats of a subtropical wetland, the Harriss, and R L. Sass. 1989. Seasonal variation of methane
Florida Everglades. Appl. Environ. Microbiol. 56:2902-2911. emissions from a temperate swamp. Biogeochemistry 8:55-71.You can also read
