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Agro-Ecosystems, 2 (1975) 127—132
127
© Elsevier Scientific Publishing Company, Amsterdam — Printed in The Netherlands
THE RELATIONSHIP BETWEEN NITRATE CONCENTRATION IN THE
SOUTHERN APPALACHIAN MOUNTAIN STREAMS AND TERRESTRIAL
NITR1FIERS
R.L. TODD, W.T. SWANK,* J.B. DOUGLASS,* P.O. KERR,** D.L. BROCKWAY^" and
C.D. MONKtt
Department of Agronomy and Institute of Ecology, University of Georgia, Athens, Ga. 30102
(U.S.A.)
*Coweeta Hydrologic Laboratory, U.S. Forest Service, Franklinf, N.C. 28734 (U.S.A.)
**Department of Microbiology, University of Georgia, Athens, Ga. 30602 (U.S. A)
'U.S. Environmental Protection Agency, Southeast Environmental Research Laboratory,
Athens, Ga. 30601 (U.S.A.)
ft Department of Botany, University of Georgia, Athens, Ga. 30602 (U.S.A.)
Contribution No. 243 from the Deciduous Forest Biome, US-IBP.
(Received April 8th, 1975)
ABSTRACT
Todd, R.L., Swank, W.T., Douglass, J.E., Kerr, P.O., Brockway, D.L. and Monk, C.D., 1975.
The relationship between nitrate concentration in the Southern Appalachian mountain
streams and terrestrial nitrifiers. Agro-Ecosystems, 2: 127—132.
The nitrate content of stream water and the nitrifying bacterial population of the terrestrial horizon were measured in three southern Appalachian watersheds over a 22-month
period. The watersheds studied were a fescue grass catchment, a 15-year old white pine plantation, and a mature undisturbed hardwood forest. Monthly averages of nitrate-nitrogen in
stream water from the three watersheds were 730, 190, and 3 ppb respectively; the respective
nitrifying populations averaged 16,000, 175 and 22 per gram of dry weight for each 40 cm
soil profile. These populations were concentrated in the upper 10 cm of the profile (grass =
98%, white pine = 90%, and hardwood = 88%). A correlation is evident between the number
of nitrifying bacteria in the soil from gaged watersheds and the NO3 content of the streams.
Nitrifying activity appears to be dependent on vegetation type and suceessional stage.
INTRODUCTION
Nitrogen transformations occurring within the biosphere are regulated almost
completely by terrestrial and aquatic microorganisms. The biological formation
of nitrate and/or nitrite from compounds containing reduced nitrogen (ammonia)
is a process termed nitrification.
Nitrification in forest ecosystems was reviewed by Chase et al. (1968). Proposals for the importance of the nitrifying process on the functioning of forested
ecosystems have included an increase of nitrifying populations (Smith et al.,
128
1968) and nitrate losses (Likens et al., 1969) from a cutover New Hampshire,
watershed. Rice and Pancholy (1972) suggest such increases are not due to
elimination of the competition for nitrate by the vegetation but result from a
de-repression of the nitrification process. They propose that the suppression of
nitrification is a prime factor in the establishment of a climax forest vegetation.
Recent evidence (Rice and Pancholy, 1973; 1974) has been presented for the
role of tannins and other polyaromatic compounds as natural occurring inhibitors of nitrification.
The collation of several investigators' efforts at the US- IBP Eastern Deciduous
Forest Biome study site, located at Coweeta Hydrologic Laboratory in western
North Carolina, allows an in-depth assessment of the role nitrification plays in
nutrient cycling within southern Appalachian forested watersheds. In this communication the terrestrial nitrifying populations are correlated with the nitrate
content in the streams for a 22-month period from an area manifesting a grassto-forest serai stage after 4 years, a 15-year-old white pine plantation, and a
mature undisturbed hardwood forest. The treatment histories and physical
properties of these adjacent catchments with similar soil types under differing
vegetative regimes have been described in detail by Johnson and Swank (1973).
MATERIALS AND METHODS
Composite samples were collected at biweekly intervals over a 22-month
period from the soil solum from at least two locations on each catchment. The
samples were transported back to the laboratory at 5°C and processed within
24 h. The number of chemoautotrophic nitrifying microorganisms was determined using a modification of the procedure described by Alexander and Clark
(1965). The chemoautotrophic nitrifying population is defined in this investigation as one forming nitrate (NO3~) in a chemically defined medium in which
ammonium (NH 4 + ) is the sole nitrogen source and calcium carbonate the only
added energy source. Nitrate formation in the incubation medium was detected
following 3 weeks incubation at 25°C (Alexander and Clark, 1965). Population
density was estimated by the most probable number (MPN) method outlined by
Alexander (1965). Results are expressed as numbers per gram of dry weight soil,
and reported values reflect the mean of each month's set of determinations.
Nitrate was quantified in the stream water from each watershed on a twiceweekly basis. Samples, collected just above the weir settling basin, were filtered
within 2 h after their collection (0.45 p.m, Millipore) and stored at low (5°C)
temperature in sterile polyethylene or polypropylene bottles. Nitrate was analyzed by the automated cadmium reduction method (Environmental Protection
Agency, 1971). Reproducibility of the nitrate determination at 100 ppb was ±
1 ppb (McSwain, 1973). Average concentration values were weighted for streamflow for each drainage basin.
129
RESULTS
The monthly mean of nitrifying bacteria (number/g of soil) and the average
NO3-N concentration of stream water are illustrated in Figs 1—3. Data in Fig. 1
are taken from a watershed in the fourth and fifth year of a grass-to-forest succession. Similar activities for a 15-year white pine plantation and a mature mixed
hardwood catchment are represented in Figs 2 and 3, respectively. The monthly
plottings generally show low levels of NO3-N and nitrifying bacteria throughout
the year for the mature hardwood cover. In contrast, the pine and grass covered
watersheds show large seasonal fluctuations in bacteria and NO3-N levels; the
annual maximum and minimum levels for the two variables tend to coincide.
In Table I the distribution of the nitrifying populations in the soil profiles is
compared to the mean annual NO3-N content of the stream water. For a 40- cm
profile 98% of the microbial nitrifying population is located within the upper
10 cm of the grass watershed profile; for the pine the value is 90% and for the
hardwood 88%.
DISCUSSION
A correlation of nitrate content in first order mountain streams and terrestrial
nitrifying bacterial populations from their respective drainage watersheds is observed. The distribution of these populations occurs in the upper division of the
e
?5
O
CD
O 4
1.4
Gross
• Nitrifying Bacteria
1.2
1.0
0.8
<
K.
UJ
CD
CD
Z! I
u_
E0
I—
0.6
0.4
0.2
6
8
10
12
2
4
10
0
12
1972
1971
TIME (MONTHS)
Fig. 1. Comparison of the terrestrial nitrifying bacteria and stream nitrate (NO3) content for
the fourth and fifth year of a grass-to-forest successional vegetation.
130
1.4
Pine
• Nitrifying Bacteria
• N0 3 -N
o
o
O 4
1.2
1.0
0.8
0.6
QQ
0.4
0.2
U.
4
6
8
10
12
6
1971
8
10
0
12
1972
TIME (MONTHS)
Fig.2. Comparison of the terrestrial nitrifying bacteria and stream nitrate (NO 3 ) content for
the fifteenth and sixteenth year of a white pine successional vegetation.
O
CO
o
o
O 4
1.4
Hardwood
• Nitrifying Bacteria
• N0 3 -N
1.2
1.0
0.8
1'
UJ
0.6
§2
CD
O
0.4
0.2
10
12
2
4
1971
10
12
1972
TIME (MONTHS)
Fig. 3. Comparison of the terrestrial nitrifying bacteria and stream nitrate (NO 3 ) content for
a mature hardwood vegetation.
131
TABLE I
Comparison of NO3-N content (weighted annual mean) in stream run-off with mean number
of nitrifying bacteria in the soil
Forest type
NO3-N
(ppm)
Nitrifying bacteria (number/g oven-dry soil)
by soil depth
Total
0—10 cm
20—40 cm
10—20 cm
Grass- to-forest
succession
White pine
0.792
0.190
15,750
160
90
4
250
13
16,000
175
Mature hardwood
0.003
15
2
5
22
profile as would be expected for these organisms. The observation that nitrate
loss and nitrifying populations decrease as vegetation approaches climax supports
the hypothesis proposed by Rice and Pancholy (1972). They propose that the
nitrifiers are inhibited in the climax so that ammonium nitrogen is not oxidized
to nitrate as readily as in the successional stages. Climax vegetation can either inhibit the nitrification process (Rice and Pancholy, 1973) or can better utilize
ammonium-nitrogen directly than can the preceding serai stages.
In spite of the positive correlation between lower nitrate losses with decline
in nitrifying populations, it may be premature to speculate that these activities
are responsible for nitrate loss from the terrestrial ecosystems. However, it certainly appears that the nitrate content of these streams could be largely due to
biological activity in the terrestrial environment and not to biological activity
within the stream.
ACKNOWLEDGEMENT
Research supported by the Eastern Deciduous Forest Biome, US-IBP, funded
by the National Science Foundation under Interagency Agreement AG-199,
BMS69-01147 A09 with the Energy Research and Development Administration —
Oak Ridge National Laboratory.
REFERENCES
Alexander, M., 1965. Most-probable-number method for microbial populations. In: C.A.
Black et al. (Editors), Methods of Soil Analysis, Part 2. Am. Soc. Agron.,Madison, Wise.,
pp. 1467—1472.
Alexander, M. and Clark, F.E., 1965. Nitrifying bacteria. In: C.A. Black et al. (Editors),
Methods of Soil Analysis, Part 2. Am. Soc. Agron., Madison, Wise., pp. 1477—1483.
Chase, F.E., Corke, C.T. and Robinson, J.B., 1968. Nitrifying bacteria in soil. In: T.R.G.
Gray and D. Parkinson (Editors), The Ecology of Soil and Bacteria. Univ. of Toronto Press,
Toronto, Ont, pp. 593—611.
Environmental Protection Agency, 1971. Methods for chemical analysis of water and wastes.
Environm. Prot. Agency, 16020, pp. 175—183.
132
Johnson, P.L. and Swank, W.T., 1973. Studies on cation budgets in the southern Appalachians
on four experimental watersheds with contrasting vegetation. Ecology, 54: 70—80.
Likens, G.E., Bormann, F.H. and Johnson, N.M. 1969. Nitrification: Importance to nutrient
losses from a cutover forested ecosystem. Science, 163: 1205—1206.
McSwain, M.R., 1973. Procedures for chemical analysis of streamflow and precipitation at the
Coweeta Hydrologic Laboratory. Eastern Deciduous Forest Biome Rep. No. 73, p.12.
Rice, E.L. and Pancholy, S.K., 1972. Inhibition of nitrification by climax ecosystems. Am. J.
Bot., 59: 1033—1040.
Rice, E.L. and Pancholy, S.K., 1973. Inhibition of nitrification by climax ecosystems. II. Additional evidence and possible role of tannins. Am. J. Bot., 60: 691—702.
Rice, E.L. and Pancholy, S.K., 1974. Inhibition of nitrification by climax ecosystems. III.
Inhibitors other than tannins. Am. J. Bot, 81: 1095—1103.
• Smith, W., Bormann, F.H., and Likens, G.E., 1968. Response of chemoautotrophic nitrifiers
to forest cutting. Soil Sci., 106: 471—473.