Aquatic Macrophyte Detritus - Applied and Environmental Microbiology

APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Jan. 1990, p. 237-244

Vol. 56, No. 1

0099-2240/90/010237-08$02.00/0 Copyright © 1990, American Society for Microbiology

Effects of Acid Stress on Aerobic Decomposition of Algal and Aquatic Macrophyte Detritus: Direct Comparison in a Radiocarbon Assay STEVEN A. SCHOENBERG,t* RONALD BENNER,t ANTHONY ARMSTRONG, PATRICIA SOBECKY, ROBERT E. HODSON Institute of Ecology and Department of Microbiology, University of Georgia, Athens, Georgia 30602

AND

Received 14 August 1989/Accepted 24 October 1989

Radiolabeled phytoplankton and macrophyte lignocelluloses were incubated at pHs 4 and 7 in water from a naturally acidic freshwater wetland (Okefenokee Swamp; ambient pH, 3.8 to 4.2), a freshwater reservoir (L-Lake; pH 6.7 to 7.2), and a marine marsh (Sapelo Island; pH -7.8). The data suggest that acidity is an important factor in explaining the lower decomposition rates of algae in Okefenokee Swamp water relative to L-Lake or Sapelo Island water. The decomposition of algal substrate was less sensitive to low pH (-5 to 35% inhibition) than was the decomposition of lignocellulose (-30 to 70% inhibition). These substrate-dependent differences were greater and more consistent in salt marsh than in L-lake incubations. In both freshwater sites, the extent to which decomposition was suppressed by acidity was greater for green algal substrate than for mixed diatom or blue-green algal (cyanobacteria) substrates. The use of different bases to adjust pH or incubation in a defined saltwater medium had no significant effect on substrate-dependent differences. Although pH differences with lignocellulose were larger in marine incubations, amendment of lakewater with marine bacteria or with calcium, known to stabilize exoenzymes in soils, did not magnify the sensitivity of decomposition to acid stress.

study which examines the effect of acidity on aerobic decomposition of algal material. Based on previous studies in our laboratory (18, 24), Benner et al. (5) proposed that the processing of substrates that rely on extracellular enzymes for their decomposition, such as lignocellulose, will be impaired to a greater extent than will the decomposition of soluble substrates, such as sugars and free amino acids, which can be transported into cells, where the internal pH is metabolically buffered (25). Carbon derived from algae forms a major detrital component in aquatic systems that is intermediate in persistence between simple dissolved substances and complex carbohydrates, such as lignocelluloses from vascular plants. Decomposition rates are about an order of magnitude faster for algae than for leaf litter (10, 11, 20), and the leachable fraction of algae is high (17). If the above hypothesis is true, the pattern for algal decomposition should resemble that of a dissolved substrate and exhibit a reduced sensitivity to pH. In the present study, the effects of acid stress on decomposition of several algal and vascular plant substrates were examined in two wetlands and a lake with a radiotracer procedure that allows us to quantify mineralization rates with the same sampling procedure and time interval. Further experiments are reported which examined whether such effects are modified by intersite differences in the source of the microbiota, water chemistry, or preadaptation of inocula. It is established that low pH is primarily responsible for the reduced mineralization of lignocellulose and subsequent peat formation within the Okefenokee Swamp relative to the salt marsh site (5, 7). The relative effect of pH on the decomposition of algal detritus and the examination of an additional system with low alkalinity described in this paper help to explain how the basis of microbial production can vary across systems which differ naturally in pH regime and how it may be altered in lakes sensitive to acid deposition.

A growing appreciation of the acid precipitation problem has stimulated study of how pH modifies the carbon balance of aquatic ecosystems (16, 27) and microbial processes in particular (13). Stream and lake systems are often classified with respect to the proportion of carbon input derived from outside the system, i.e., allochthonous sources, versus autochthonous production within the system. Although there is some overlap, allochthonous material is primarily of vascular plant origin, and the bulk of autochthonous material is planktonic or periphytic algae. To appraise the impact of pH stress on carbon balance, it is necessary to determine whether acidity affects the decomposition of chemically different substrates in the same way. Some studies suggest that acidity significantly suppresses to varying degrees the decomposition of complex particulate substrates which degrade slowly, such as terrestrial leaves (1, 2, 8, 9, 14), emergent macrophytes (5, 7, 22), or cellophane (19), as well as the decomposition of dissolved substrates which are more rapidly respired, such as glucose (3, 9, 12). On the one hand, the commonly used litter bag method to determine leaf decomposition cannot be directly compared with radiometric assays of simple monomers because it requires much longer incubation times and includes all dissolved organic carbon as a decomposition loss whether it is mineralized or simply leached. On the other hand, the kinetics of a single dissolved substance may not accurately reflect the decomposition of the mixed compounds derived from algae; in fact, we are unaware of a

Corresponding author. t Present address: Institute of Limnology, Uppsala University, Box 557, S-751 22, Uppsala, Sweden. t Present address: Marine Science Institute, University of Texas at Austin, Port Aransas, TX 78373. *

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TABLE 1. Distribution of total carbon and 14C used to evaluate specific activities of warm water extract (3 h, 40°C) and unextracted particulate carbon fractions in labeled phytoplanktona Substrate

Microcystis sp. C. reinhardi Mixed diatoms

%'4C in:

%Total carbon in: Extract

Particulate

Extract

Particulate

29.8 (2.7) 29.8 (1.6) 32.3 (1.0)

70.2 (2.7) 70.2 (1.6) 67.7 (1.0)

66.9 (6.8) 18.0 (1.2) 36.7 (2.0)

33.1 (6.8) 82.0 (1.2) 63.3 (2.0)

Sp act %14C/%total C in: Extract Particulate 2.23 (0.09) 0.61 (0.05) 1.14 (0.10)

0.47 (0.09) 1.17 (0.03) 0.94 (0.04)

a Mean (standard error of the mean) of four replicates. A specific activity of 1.0 indicates completely uniform labeling.

MATERIALS AND METHODS Study sites. L-Lake is a cooling lake on the Savannah River Plant reservation near Aiken, S.C. Although now circumneutral (pH 6.7 to 7.2), the low-alkalinity waters of this region are showing some early effects of acid precipitation (30). Blue-green alga (cyanobacteria) blooms occur during summer or when reactor operation raises the water temperature; no macrophytes were present during our study. Okefenokee Swamp is a large freshwater wetland in southeast Georgia which is naturally acidic (pH 3.8 to 4.2), owing to high concentrations of organic acids. The sedge Carex walteriana is the dominant macrophyte in the area from which inocula were collected for the experiments described below, although mat-forming and planktonic algae are also abundant (28). Located on the coast of south Georgia, Sapelo Island is surrounded by extensive salt marshes of the cordgrass Spartina alterniflora. The water is saline, basic (pH -7.8), and strongly buffered by carbonates. Unlike L-Lake, the pH of the wetlands is stable over the long term and resistant to pH change due to atmospheric deposition. Substrate preparation. Algal substrates from culture or L-Lake were labeled by a modification of the method of Cole et al. (10) and included natural blue-green alga material (Microcystis sp.) collected during a summer bloom, a mixed diatom assemblage obtained after turnover in fall, and a pure culture of the green alga Chlamydomonas reinhardi. Phytoplankton were labeled by the addition of ["4C]bicarbonate given as pulses of 2 to 3 ,uCi/ml initially and after 24 h. Cells were allowed to metabolize for 2 days, after which they were harvested on glass fiber filters, fumed with HCl vapor to remove residual bicarbonate and freeze-dried for storage. To determine the homogeneity of label in the preparations, we extracted each substrate with sterile hot water (3 h, 40°C) and refiltered the material onto a second glass fiber filter. The filtrate and particulate materials were digested by wet oxidation with potassium persulfate, and the total carbon content of each fraction was measured by infrared gas analysis (model 524, Oceanics International Corporation). The radiocarbon content was determined by assay of exhaust gas from the carbon analyzer collected in C02-trapping scintillation cocktail (6). Specific activity is defined as the ratio of % radiocarbon to % total carbon. Uniformly labeled lignocelluloses from the macrophytes C. walteriana and S.

alterniflora were prepared by incubating the plants in

[14C]carbon dioxide gas. A standard extraction scheme was used to separate lignocellulose from other plant components

(6).

Incubation procedure. The inoculum for L-Lake incubations was surface water combined from collections at several locations along the major axis of the lake. Water from the Okefenokee Swamp was collected in the vicinity of Mizell Prairie adjacent to the Suwanee Canal. Salt marsh water was obtained from the Duplin River on Sapelo Island. The

laboratory incubation vessel consisted of a 150-ml stoppered bottle fitted with a long inlet and a short outlet glass stem for purging 14CO2. A minimum of three replicates and one Formalin-killed control were used for treatments, each containing a 25-ml inoculum and a known quantity of radiolabeled substrate (10 to 20 p.g of algae, 5 to 10 mg of lignocellulose). Evolved "4CO2 was collected at 2-day intervals for 14 days by bubbling for 20 min with humidified air into the trapping cocktail. After each gas collection, the pH was measured and readjusted as necessary with dilute HCI or NaOH. A second purging was performed on the last sampling day after injecting 2 ml of 10% phosphoric acid to convert any residual inorganic [14C]bicarbonate into gas. A portion of the incubation medium was then filtered (0.2-p.m pore size; Millipore Corp.) and counted in a gel with Scintiverse I (Fisher Scientific Co.) to determine the amount of residual radiolabeled dissolved organic carbon ([14C]DOC). Decomposition experiments. (i) For one of the algal substrates, we quantified the decomposition rates of the leachable and particulate fractions separately. "Initial leachate" was prepared by soaking Microcystis sp. substrate in sterile water for 1 h at 20°C; this fraction contained about 10% of the total radioactivity. The leachate was isolated from the particulate matter by filtering both the substrate and liquid through a second glass fiber filter. The same procedure was performed for unlabeled substrate, which had been prepared at the same time as the labeled substrate. Treatments consisted of labeled leachate combined with unlabeled particulate and unlabeled leachate combined with labeled particulate. A fresh inoculum of L-Lake water was added to each incubation. (ii) An implicit assumption in the interpretation of the pH experiments is that the sensitivity of the micr bial community to acid stress is not affected by prior exposure. In order to test for short-term adaptation of the microbial community to pH, we compared the decomposition of C. reinhardi substrate in L-Lake water which was adjusted to pH 4 or 7 two days prior to the experiment with decomposition in water preincubated at pH 4 or 7 for 16 days. (iii) A preliminary study was conducted to compare decomposition rates of algae across sites which differed in ambient pH. The design involved incubations of Microcystis sp. and C. reinhardi with inocula from an acidic site (Okefenokee Swamp) and a nonacidic site (L-Lake). (iv) Another series of experiments was designed to isolate the effect of pH from other intersite differences. For both Okefenokee and L-Lake waters, Microcystis sp., C. reinhardi, and the mixed diatom substrates were incubated at a controlled pH of 4 or 7; an additional intermediate treatment of pH 5.5 was included for Microcystis sp. (v) We subsequently determined the effect of acidity on decomposition of algae compared with that of macrophyte lignocellulose by using L-Lake and salt marsh inocula. For

SUBSTRATE-DEPENDENT DECOMPOSITION UNDER pH STRESS

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239

filter sterilized through 0.2-gum pore size Nuclepore membranes, adjusted to pH 7 or 4, then reinoculated with similar concentrations of either L-Lake or salt marsh bacteria which had been concentrated on 0.2-,um-pore-size filters.

c,. 00 a)

a) -o a)

| particulate| |- leachatej

0 0 L.

0--

0

10 DAY

5

15

20

FIG. 1. Decomposition of initial leachate isolated by sterile water extraction and unextracted particulate components of Micro-

cystis

sp.

substrate (mean

±

standard

error

of the mean).

each site, treatment combinations of four substrates (C. walteriana, S. alterniflora, C. reinhardi, and Microcystis sp.) at pHs 4 and 7 were included. Intersite differences were investigated in a series of experiments (vi to ix) using a representative algal substrate, Microcystis sp., and lignocellulose from S. alterniflora. (vi) To control for differences in experimental protocol, inocula from L-Lake were acidified to pH 4 with HCl and then brought up to pH 8.0 with either NaOH, as used in the present study, or with NaHCO3, as was used previously (7). (vii) Dissolved calcium was tested as a factor that may affect sensitivity to acid stress, as it is known to stabilize cellulolytic exoenzymes important in decomposition (26). To test this possibility, decomposition rates were determined at pHs 4 and 7 in L-Lake incubations amended with 400 mg of calcium as CaCl2 per liter and in unamended controls. (viii) Artificial seawater (29) was employed as a defined medium to control for other major constituents of salt as well as the presence of certain trace metals which become toxic when mobilized at low pHs (3). We set up treatments at pHs 4 and 7 using L-Lake water cross-inoculated with salt marsh bacteria, salt marsh water cross-inoculated with L-Lake bacteria, and artificial seawater media with inocula from both sites. (ix) To see if the microbial communities differed in sensitivity to acid stress between sites, L-Lake water was

RESULTS Characterization of the specific activities of warm water extracts and unextracted particulate materials indicated that the homogeneity of label varied among algal substrates (Table 1). In all three substrates, the extract comprised about a third of the total carbon. The specific activity of the extracted material was greater than that of the particulate for Microcystis sp., less than that of the particulate for C. reinhardi, and approximately the same as the particulate for the mixed diatom substrate. Although the strongest bias was observed for Microcystis sp., the decomposition rate for material initially leached in cold water was nearly identical to that of the particulate material (Fig. 1). The similarity in decomposition rates suggests to us that the experimental results are not substantially compromised by nonuniform label distribution. In the preliminary study, decomposition of both algal substrates was significantly reduced by incubation in acidic Okefenokee water relative to L-Lake water (Fig. 2), although the difference varied between the materials. Total mineralization (i.e., the percentage of label recovered over the entire incubation) of Microcystis sp. was inhibited by 39% in Okefenokee water, whereas the inhibition for C. reinhardi was 65%. The second set of experiments indicates that a portion of the difference in decomposition between sites may be attributed to the effects of pH (Fig. 3). In L-Lake, loss rates were reduced at pH 4 relative to pH 7 by 12, 19, and 40%, respectively, for Microcystis sp. mixed diatom, and C. reinhardi substrates. Corresponding differences between treatments for Okefenokee Swamp incubations were 6% for Microcystis sp., 5% for diatom, and 33% for C. reinhardi substrates. The effect of pH on algal decomposition was not as large in the controlled pH experiment as indicated in the preliminary study and was reduced in the Okefenokee Swamp relative to L-Lake water. The sensitivity of decomposition to acid stress was greater for the C. reinhardi than for the Microcystis sp. or mixed diatom substrates at each site. There was no indication that the decomposer sensitivity to pH is affected by prior exposure to acidity. With the C. reinhardi substrate, mineralization in treatments with inoc-

cM

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o a,

a)

l | L-lake |-- Okefenokee|

0 La,

10

DAY

0

5

10

15

20

DAY

FIG. 2. Decomposition (mean + standard error of the mean) of the green alga C. reinhardi and the blue-green alga Microcystis sp. in naturally acidic wetland, Okefenokee Swamp (pH 3.8 to 4.2) and a circumneutral lake, L-Lake (pH -7).

a

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SCHOENBERG ET AL.

L-Lake U 7Ft% M ro 6

Microcystis

,

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DAY

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FIG. 3. Effect of pH manipulation on the decomposition of Microcystis sp., mixed diatom assemblage, and C. reinhardi pure culture in the Okefenokee Swamp and in L-Lake (mean ± standard error of the mean).

ula maintained at pH 4 or 7 for an additional 2 weeks prior to substrate addition showed the same degree of inhibition at a low pH as those with inocula which were adjusted 2 days preceding the experiment (Fig. 4). The influence of pH in parallel incubations using macrophyte lignocelluloses and algal substrates varied between L-Lake and salt marsh sites (Fig. 5). In L-Lake, low pH reduced mineralization rates by similar percentages for both lignocelluloses (33% reduction for C. walteriana and 28% for S. alterniflora) and one algal substrate (34% for C. reinhardi) but by only 15% for Microcystis sp. In parallel salt marsh incubations, decomposition rates of lignocellulose were consistently inhibited to a greater degree (52 and 44% for C. walteriana and S. alterniflora, respectively) than for algal substrate (31 and 22% for C. reinhardi and Microcystis sp., respectively). Within sites, residual [14C]DOC from lignocel-

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pH 4-nonadapted pH 7-nonadapted pH 4-preadapted

pH 7-preadapted

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10

12

14

FIG. 4. Decomposition (mean + standard error of the mean) of C. reinhardi substrate using inocula held for 16 days prior to addition of the substrate at the treatment pH (preadapted) compared with inocula adjusted 2 days prior to addition of the substrate

(nonadapted).

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SUBSTRATE-DEPENDENT DECOMPOSITION UNDER pH STRESS 301-

241

4

Carex

=3

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pH

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60-

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pH

7

4

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FIG. 5. Decomposition (mean + standard error of the mean) of lignocelluloses and whole algal substrates in L-Lake and Sapelo Island salt marsh waters. Percentages represent cumulative mineralization over 14 days and residual dissolved label at the termination of experiments.

lulose

was

1.5 to 3 times greater at pH 4 than pH 7. The

[14C]DOC fraction for incubations of lignocellulose

was

greater in the salt marsh than in L-Lake. For the algal

substrates, this fraction was -fourfold higher and similar between sites. Several additional experiments were done to investigate whether these results were related to water chemistry. Prior results with inocula from another freshwater site, Okefenokee Swamp, showed a tripling of the decomposition rate of lignocellulose over the same pH range adjusted with bicarbonate solution (7). Tests in L-Lake water showed that the decomposition of both lignocellulose and algal substrate is not affected by the use of a different base buffer, NaOH, in the present study (Table 2). At neutral pH, flocculation of cellulase with humic acid in the presence of CaCl2 has been shown to enhance enzyme resistance to proteolysis in soils (26). The presence of abundant Ca2" and some humic acid in salt marsh water raised the possibility that this interaction could augment mineralization of lignocellulose. Addition of CaCl2 to incubations did enhance residual [14C]DOC levels

TABLE 2. Decomposition in L-Lake incubations maintained at pH 8.0 by adjustment with different bases Substrate and end product

S. alterniflora CO2 DOC

% Radioactivity recovered from incubations adjusted witha: NaOH

NaHCO3

7.5 (0.4) 0.6 (0.02)

6.6 (0.01) 0.5 (0.1)

Microcystis sp. 66.0 (1.0) 66.7 (0.8) 6.4 (0.1) 5.7 (0.4) Mean (standard error of the mean) of three replicates. Values represent the percent radioactivity recovered as "4CO2 accumulated over 14-day incubations or as [14C]DOC at the end of the experiment.

CO2

DOC

a

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TABLE 3. Effects of pH on decomposition in L-Lake inoculum amended with calcium % Radioactivity recovered from incubations witha: Substrate and end product

S. alterniflora CO2 DOC

No addition (control)

at:

pH 4

pH 7

4.4 (0.6) 0.9 (0.1)

7.1 (0.3) 0.5 (0.02)

400 mg of calcium liter added at: pH 4 pH 7

6.1 (0.3) 1.5 (0.2)

7.0 (0.9) 0.8 (0.1)

38.7 (1.6) 5.5 (0.1)

61.1 (0.7) 11.9 (1.6)

Microcystis sp. CO2

42.0 (5.9) 63.1 (3.4) 7.7 (0.3) 4.8 (0.1)

DOC

a Mean (standard error of the mean) of three replicates. Values represent the percent radioactivity accumulated over 14-day incubations or as [14C]DOC remaining at the end of the experiment.

by 60 to 67% for S. alterniflora and 147% for Microcystis sp. at pH 7 (Table 3). However, this did not result in significantly greater differences in mineralization of either sub-

strate because of low pH; mineralization of S. alterniflora was actually more similar between pH treatments with calcium, contrary to expectation. In the subsequent experiment (Table 4), decomposition in incubations cross-inoculated with both L-Lake and salt marsh microbiota at pH 4 compared with pH 7 was inhibited to a similar extent in L-Lake, salt marsh, and artificial seawater media, and treatment effects were lower for the alga than for the lignocellulose (58 to 65% suppression for S.

alterniflora and 11 to 21% for Microcystis sp.). The slightly larger pH effect for lignocellulose in the salt marsh and L-Lake compared with artificial seawater could be a result of toxic metals present in natural, but not synthetic, media that are mobilized at low pHs, but the effect of medium source is very weak compared with the large difference due to pH alone. The residual [14C]DOC level in lignocellulose but not algal incubations was up to -twofold greater in both salt marsh media and artificial seawater than in L-Lake media. As a whole, the data provide little evidence that differences in water chemistry affect the sensitivity of decomposition to acid stress. More significantly, results for lignocellulose with crossinoculated L-Lake and salt marsh media were very similar (Table 4), whereas the earlier experiment without crossinoculation (Fig. 5) indicated substantially different sensitivities to acid stress between sites. We used the transplant experiment to examine whether members of the microbial community which decompose lignocellulose in the salt

marsh were more sensitive to acid stress than those of L-Lake (Table 5). As in the experiments discussed above, decomposition of Microcystis substrate was inhibited at low pH to a lesser extent (11 to 29% reduction compared with pH 7) than was decomposition of Spartina lignocellulose (63 to 78% reduction). However, the effect of pH on Spartina mineralization was substantially less, not greater, as expected, with bacteria from the salt marsh than with bacteria from L-Lake. Unlike the earlier experiments with different media, residual [14C]DOC levels were similar between treatments. On this basis, we cannot attribute differences in the response to acid stress between sites to variation in the resident microbial community. DISCUSSION Compounds leached from vascular plant detritus are decomposed much more rapidly than are more stable particulates (4). For algae, leaching is a biphasic, predominantly physical process which solubilizes two-thirds or more of the particulate carbon within a week (17). Continued leaching probably was the main factor which caused decomposition rates to be similar for the initial leachate and particulate matter in our experiment with the blue-green alga Microcystis sp. The slightly greater response of decomposition to pH we observed with C. reinhardi than with other algal substrates may reflect the presence of cellulose-like compounds in its cell wall that are extensively degraded by action of the exoenzyme cellulase (15). In contrast to lignocellulose decomposition, the extensive leaching of algal detritus as a whole is a process which minimizes the role of exoenzymes in favor of dissolved carbon transport as the rate-limiting step of mineralization. In nearly all comparisons, lignocellulose decomposition was more sensitive to pH than was algal decomposition. These results are consistent with the hypothesis that decomposition is more sensitive to pH when exoenzymatic action is required (5). In the present study, pH also had a greater effect on lignocellulose decomposition in the salt marsh relative to L-Lake in one experiment (Fig. 5), but not always (Table 4). The use of a defined salt mixture or the amendment of calcium alone failed to reproduce this effect; these results tend to rule out suspected interactions in salt water, which would enhance the effect of pH, such as improved stability of exoenzymes or the solution of trace metals at low pH. In a subsequent experiment (Table 5), we were also unable to detect larger pH effects in L-Lake media inoculated from the salt marsh site. A comparison between experiments shows that the response of lignocellulose decomposition to the pH in L-Lake is more variable (compare L-Lake data in Fig. 5,

TABLE 4. Effects of pH on decomposition in unfiltered fresh water, unfiltered marine water, and artificial seawater media Substrate and end product

S. alterniflora Co2 DOC

Microcystis sp. CO2 DOC

% Radioactivity recovered from incubation ina: L-Lake water at: Salt marsh water at: Artificial seawater at: L-Lake water_at:_Salt_marsh_water_at:_Artificial_seawater_at: pH 7 pH 4 pH 4 pH 7 pH 4 pH 7

10.0 (0.3) 0.7 (0.1)

28.9 (2.0) 0.6 (0.1)

6.7 (0.8) 1.5 (0.1)

22.3 (1.0) 0.4 (0.1)

9.0 (0.3) 1.4 (0.1)

21.5 (0.8) 1.1 (0.1)

46.6 (1.9) 1.8 (0.1)

57.9 (2.4) 1.9 (0.3)

46.8 (2.1) 2.0 (0.1)

53.2 (1.5) 1.4 (0.1)

37.9 (3.1) 2.5 (0.2)

47.9 (0.7) 1.8 (0.2)

a Mean (standard error of the mean) of three replicates. All treatments were cross-inoculated with bacteria from L-Lake and the salt marsh. Values represent the percent radioactivity recovered as 14CO2 accumulated over 14-day incubations or as dissolved organic [14C]DOC remaining at the end of the experiment.

SUBSTRATE-DEPENDENT DECOMPOSITION UNDER pH STRESS

VOL. 56, 1990

TABLE 5. Effects of pH on decomposition in 0.2-,m-filtered LLake water reinoculated with either L-Lake or salt marsh bacteria Substrate and end product

% Radioactivity recovered from incubations inoculated witha: L-Lake bacteria at: pH 4

S. alterniflora CO2 DOC

pH 7

Salt marsh bacteria at: pH 4

1.8 (0.1) 8.1 (0.4) 1.4 (0.1) 0.12 (0.03) 0.31 (0.02) 0.16 (0.01)

Microcystis sp. CO2 46.9 (1.3) DOC 3.1 (0.2)

52.5 (1.8) 1.9 (0.1)

38.1 (1.1) 3.7 (0.2)

pH 7

3.8 (0.1) 0.14 (0.01) 53.8 (0.7)

1.9 (0.1)

Mean (standard error of the mean) of three replicates. Values represent the percent radioactivity recovered as 14CO2 accumulated over 14-day incubations or as [14C]DOC remaining at the end of the experiment. a

Table 3 [control], Table 4 [L-Lake], and Table 5 [L-Lake]) than it is in salt marsh water (salt marsh data in Fig. 5 and Table 4 [salt marsh]) (7). Our inability in these latter experiments to assign intersite differences such as those in Fig. 5 to chemical or microbial source effects may, in part, be a consequence of the variability in the short term between experiments conducted several months apart. On the other hand, a comparison of pH responses with the same inocula and Microcystis sp. substrate shows very little variation between sites or between experiments. We suggest that functionally distinct decomposer groups are involved in the processing of vascular and nonvascular detritus in these systems. The group involved in decomposition of algae appears to be ubiquitous and stable, whereas the data suggest that the group involved in lignocellulose decomposition may not always be present or active. Because decomposition of lignocellulose was generally more sensitive to acid stress than was decomposition of whole algal substrate, we expect that the impact of acid stress on carbon balance will be minimal where either allochthonous inputs or littoral production of vascular plant carbon is low. This appears to be the case for one lake in which the epilimnetic pH was gradually reduced by acid addition from 6.8 to 5.0 over an 8-year period. Since annual primary production (27) and organic sedimentation rates as inferred from benthic gas flux (21) were both unchanged with progressive acidification, it can be concluded that the decomposition rate in the surface waters was unaffected by pH. Moreover, one would expect the resource base of the microbial food web to shift in favor of nonvascular carbon in acid-stressed systems. Such a transition may explain the enigmatically high microbial production in sedge marshes of the Okefenokee Swamp (24). Based on model calculations (23), only a third of this production can be supported by the dissimilation of lignocellulose. In the present study, the effect of natural acid stress in the Okefenokee on decomposition of all algal substrates was small (5 to 30% inhibition), whereas earlier results (5, 7) with the same experimental procedure showed that decomposition of lignocellulose is substantially inhibited by low pH (-70% inhibition). Assuming a reasonable turnover rate, the standing stock of algae is sufficiently high at this site (28) to propose that a substantial fraction of the carbon entering the microbial food web in this wetland originated as algal detritus rather than vascular plant material.

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ACKNOWLEDGMENTS Technical support was provided in part by laboratory assistants P. Breed and M. Tift. D. Pakulski and E. Sheppard collected salt marsh inocula. J. Wiegel generously permitted use of a carbon analyzer. Research and manuscript preparation were supported by contract DE-AC09-76SRO0-819 between the U.S. Department of Energy and the Savannah River Ecology Laboratory, University of Georgia. LITERATURE CITED 1. Allard, M., and G. Moreau. 1986. Leaf decomposition in an experimentally acidified stream channel. Hydrobiologia 139: 109-117. 2. Andersson, G. 1985. Decomposition of alder leaves in acid lake waters. Ecol. Bull. 37:293-299. 3. Baker, M. D., W. E. Inniss, C. I. Mayfield, and P. T. S. Wong. 1983. Effects of acidification, metals and metalloids on sediment microorganisms. Water Res. 17:925-930. 4. Benner, R., and R. E. Hodson. 1985. Microbial degradation of the leachable and lignocellulosic components of leaves and wood from Rhizophora mangle in a tropical mangrove swamp. Mar. Ecol. Prog. Ser. 23:221-230. 5. Benner, R., D. L. Lewis, and R. E. Hodson. 1989. Biogeochemical cycling of organic matter in acidic environments: are microbial degradative processes adapted to low pH?, p. 33-45. In S. S. Rao (ed.), Acid stress and aquatic microbial interactions. CRC Press, Inc., Boca Raton, Fla. 6. Benner, R., A. E. Maccubbin, and R. E. Hodson. 1984. Preparation, characterization and microbial degradation of specifically radiolabeled [14C]lignocelluloses from marine and freshwater macrophytes. Appl. Environ. Microbiol. 47:381-389. 7. Benner, R., M. A. Moran, and R. E. Hodsos 1985. Effects of pH and plant source on lignocellulose biodeg: idation rates in two wetland ecosystems, the Okefenokee Swamp and a Georgia salt marsh. Limnol. Oceanogr. 30:489-499. 8. Burton, T. M., R. M. Stanford, and J. W. Allan. 1985. Acidification effects on stream biota and organic matter processing. Can. J. Fish. Aquat. Sci. 42:669-675. 9. Carpenter, J., W. E. Odum, and A. Mills. 1983. Leaf litter decomposition in a reservoir affected by acid mine drainage. Oikos 41:165-172. 10. Cole, J. J., G. E. Likens, and J. E. Hobbie. 1984. Decomposition of planktonic algae in an oligotrophic lake. Oikos 42:257-266. 11. Fallon, R. D., and T. D. Brock. 1979. Decomposition of bluegreen algal (cyanobacterial) blooms in Lake Mendota, Wisconsin. Appl. Environ. Microbiol. 37:820-830. 12. Ferroni, G. D., L. G. Leduc, and C. G. Choquet. 1982. Preliminary studies on the use of the heterotrophic activity method to evaluate acid-stress in aquatic environments. Water Res. 17: 1379-1384. 13. Francis, A. J. 1986. Acid rain effects on soil and aquatic microbial processes. Experientia 41:455-465. 14. Francis, A. J., H. L. Quinby, and G. R. Hendry. 1984. Effect of pH on microbial decomposition of allochthonous litter, p. 1-21. In G. R. Hendry (ed.), Early biotic responses to advancing lake acidification. Butterworth Publishers, Stoneham, Mass. 15. Gunnison, D., and M. Alexander. 1975. BEsis for the susceptibility of several algae to microbial decok position. Can. J.

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