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Stuttgart, January 2005
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Verh. Internat. Verein. Limnol.
Landscape-scale patterns of nitrogen fixation by cyanobacteria
A. Patoine, M. Graham, P.R. Leavitt, and R. Hesslein
Introduction
Cyanobacteria blooms are characteristic of eutrophic
lakes and are often a threat to water consumption
(MILLIE et al. 1999). Lakes of the prairie ecozone of
Canada are particularly prone to cyanobacteria
blooms harbouring nitrogen-fixers (Aphanizomenon,
Anabaena, Gloeotrichia spp.), in part because of
phosphorus (P)-rich soils and high irradiance
regimes (BARICA 1987). However, despite the prevalence of cyanobacteria in phytoplankton of some
lakes, there is little agreement on whether nitrogen
fixed from the atmosphere (N2) is quantitatively incorporated into the aquatic food web. On one hand,
their colonial nature and potential toxicity make heterocystous cyanobacteria poor food for zooplankton
(GHADOUANI et al. 2004). On the other hand, historical correlations between cyanobacterial abundance
and stable isotope signals in plankton and lake sediments (TALBOT 2001) suggest that atmospheric N2
fixed by cyanobacteria may be an important source
of nitrogen (N) to lake ecosystems.
Little is known of how the importance of N sources
may vary as a function of lake position within the
greater landscape. Many chemical properties of lakes
are strongly influenced by the position of a lake within the hydrologic or topographic landscape, such that
headwater lakes tend to exhibit lower nutrient concentrations and algal densities than do downstream
sites (SORANNO et al. 1999). Based on these patterns,
we hypothesized that the intensity of N2 fixation by
cyanobacteria should increase with distance from
headwaters, and that fixed N2 should be more completely incorporated into the primary and secondary
consumers of downstream lakes.
To test these hypotheses, we analyzed seasonal
variability in N isotope content of algae and zooplankton communities in five chained prairie lakes
during 1995. Nitrogen fixation and assimilation by
zooplankton should be reflected by concomitant declines in the 15N contents of algae and zooplankton,
as light atmospheric N2 (δ15N = 0‰) is incorporated
into relatively 15N-rich consumers (5–10‰).
Umbruch
Key words: landscape, cyanobacteria, N fixation, N
concentration
Material and methods
Site description – The Qu’Appelle River drains
53,000 km2 of predominantly (76%) farmland as it
flows eastward ~340 km from Lake Diefenbaker
reservoir through a series of eight fluvio-glacial
lakes, including Buffalo Pound, Katepwa and
Crooked lakes (Fig. 1). The effective drainage area
increases regularly with lake order (Fig. 2A). At the
same time, soluble reactive phosphorus concentrations increase, while N:P ratios of dissolved nutrients
decline as a function of lake position (Fig. 2B-C).
Cyanobacteria are a common feature of Qu’Appelle
Valley lakes (BARICA 1987) and were present before
European settlement (HALL et al. 1999).
Sampling and analyses – Diefenbaker, Buffalo
Pound, Last Mountain, Katepwa and Crooked lakes
were sampled bi-weekly May-August 1995. Integrated water samples from the surface to one meter above
the sediments were used for phytoplankton taxonomic analyses and heterocyst counts. Water samples for
N-isotopic analyses of particulate organic matter
(δ15NPOM) were screened through 153-µm mesh net
and filtered onto glass fiber filters. The screening
procedure was effective at removing crustaceans and
rotifers, while maintaining a representative algal assemblage characteristic of each lake (GRAHAM 1997).
Zooplankton were collected with a 243-µm mesh
conical plankton net. Adult zooplankton were sorted
to species (Daphnia galeata mendotae, Leptodiaptomus sp., Diacyclops thomasi, Leptodora kindtii),
dried, and frozen until isotope analyses. Mass spectrometry analyses of dried POM and zooplankton
followed HESSLEIN et al. (1989). Isotope content
(δ15N) was defined as permille (‰) deviation from
the atmospheric standard and was calculated as:
[(Rsample-Rstandard)/Rstandard] × 1000, where R is
the isotope ratio (15N/14N).
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©2005 E. Schweizerbart’sche Verlagsbuchhandlung, D-70176 Stuttgart
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Fig. 1. Qu’Appelle Valley drainage basin and lakes included in this study.
Results
The 15N content of algae (as POM) showed significant variations among lakes (Kruskal-Wallis Test Statistic = 9.9, P = 0.043), with lower
values and greater seasonal variability in smaller, more productive lakes (Buffalo Pound,
Katepwa, Crooked) than in larger Diefenbaker
and Last Mountain Lakes (Fig. 2D). Most of
this variability was inversely related to the absolute density of N2-fixing cyanobacteria (r2 =
0.76, P<0.001, Fig. 3A), suggesting that
cyanobacteria influenced the overall isotopic
signature of POM. Further, most of this trend
was associated with small productive lakes that
exhibited a wide range of cyanobacteria densities (103 to 105 cells mL–1) relative to larger
sites (<104 cells mL–1).
Fixed atmospheric N2 was assimilated into
zooplankton only in downstream Katepwa and
Crooked lakes (Fig. 4). At these sites, invertebrate d15N declined significantly as a function
of the density of N2-fixing cyanobacteria. In
contrast, there was no evidence of any negative
relationship between N2-fixers and zooplank-
ton isotopic signature in upstream Buffalo
Pound Lake, suggesting zooplankton were not
using cyanobacteria as a principal food source.
Over the entire range of N2-fixing cyanobacteria relative density, the N-isotopic signature of
zooplankton in upstream Buffalo Pound Lake
tended to be lower relative to downstream
lakes, probably because of inter-lake differences in baseline isotopic composition. No pattern was recorded in either of the large lakes
(Diefenbaker and Last Mountain) due to very
low cyanobacterial densities (Fig. 3A).
Patterns of seasonal development of N2-fixing cyanobacteria differed substantially among
lakes (Fig. 3B). In upstream Buffalo Pound
Lake, blooms were smaller (<20 000 cells
mL–1), and shorter-lived (<20 d) relative to
downstream Katepwa and Crooked lakes.
Estimates of the total amount of N2 fixed into each lake showed heterocystous cyanobacteria can contribute up to 8% of the average N
pool (mean N concentration x lake volume)
recorded in upstream Buffalo Pound lake, and
more than 100% in downstream lakes (Table 1).
Although these proportions were estimated
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A. Patoine, Landscape – scale patterns
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Fig. 2. Landscape patterns in A) drainage area, B) soluble reactive phosphorus (SRP), C) nitrogen to
phosphorus mass ratio (as the sum of nitrate and ammonium relative to SRP) and D) nitrogen isotopic
content of particulate organic matter (POM). Letters represent Diefenbaker Reservoir (D), Buffalo
Pound Lake (B), Last Mountain Lake (L), Katepwa Lake (K), and Crooked Lake (C). Box and whiskers
represent the 1995 seasonal variability. Centre vertical lines represent the median. Hinges represent the
first and third quartiles and span the interquartile range (IQR). Upper/lower whiskers show the range of
values that fall within the upper/lower inner fences i.e. upper/lower hinges +/– 1.5(IQR). Asterisks represent values between the inner fences and outer fences (upper/lower hinges +/– 3(IQR)). Empty circles
represent values that fall outside the outer fences.
from the growth season only and were derived
from different methodologies (physiological
estimates of N2 fixation rate, total N concentrations), they still suggest N2 fixation by heterocystous cyanobacteria is a more important N
source in downstream lakes than in upstream
lakes.
Discussion
If chemical properties of lake water are determined in part by a lake’s position in the land-
scape, then so should some of its biological
properties (SORANNO et al. 1999). Here, we
showed that filamentous N2-fixing cyanobacteria, despite their apparently low palatability,
were assimilated by zooplankton in the most
downstream of a set of chained prairie lakes.
This strong landscape pattern is consistent with
observed declines in N:P ratios in downstream
lakes (Fig. 2C), a factor known to favour development of heterocystous cyanobacterial
blooms. In the Qu’Appelle lakes, this tendency
to N limitation downstream arises because
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In contrast to lowland sites, atmospheric N2
did not appear to contribute substantially to food
webs of upstream Buffalo Pound Lake, despite a
high proportion of heterocystous cyanobacteria
in phytoplankton (Fig. 4). This pattern likely
arises because absolute densities of N2-fixers
were low at all upstream sites (Fig. 3A, Table 1)
and because inorganic N contents of these lakes
were already relatively high (Fig. 2C). Thus, in
this case, fixed N2 represented only a small proportion of total N standing stock (~8%) relative
to downstream lakes and was less likely to influence the overall elemental composition of the
consumer community. On-going analyses will
reveal whether temporal patterns in N2 fixation
are also observed in other years. In addition,
mass-balance studies are being used to evaluate
the relative importance of alternate N sources,
including urban wastewater (δ15N ~17 ‰) and
agricultural fertilizers (~0‰) (P.R. LEAVITT et al.
unpubl. data).
Conclusion
Fig. 3. Nitrogen isotopic content of particulate organic matter as a function of density of nitrogenfixing cyanobacteria in Qu’Appelle Valley lakes
(A) and seasonal development of nitrogen-fixing
cyanobacteria (B). Numbers represent lake order
on a West to East transect (1: Diefenbaker Reservoir; 2: Buffalo Pound Lake; 3: Last Mountain
Lake; 4: Katepwa Lake; 5: Crooked Lake). Regression line follows the equation Y = 19.4 – 1.5
(log X), r2 = 0.76, P<0.001.
lakes may act as net exporters of dissolved P
(KENNEY 1990), leading to an “amplification”
process whereby P accumulates in downstream
lakes. The importance of N2 fixation in downstream lakes suggests internal sources of N
such as sediment return were insufficient to alleviate N-limited phytoplankton growth
(LEVINE & SCHINDLER 1992).
Despite the filamentous and unpalatable nature of
cyanobacteria, N from these phytoplankton is apparently incorporated into zooplankton communities.
However, the degree of assimilation of fixed N2 by
invertebrates was dependent on lake position in the
landscape and was greatest in downstream lakes
where low N:P ratios promoted dense and long-lived
populations of N2-fixing cyanobacteria. At present, it
is unclear whether fixed N2 is directly incorporated
from cyanobacteria into invertebrates or whether it is
first recycled from cyanobacteria into more palatable
algae or bacteria.
References
BARICA, J., 1987: Water quality problems associated
with high productivity of prairie lakes in Canada: a
review. – Water Qual. Bull. 12: 107–115.
GHADOUANI, A., PINEL-ALLOUL, B., PLATH, K., CODD,
G.A. & LAMPERT, W., 2004: Effects of Microcystis
aeruginosa and purified microcystin-LR on the feeding behavior of Daphnia pulicaria. – Limnol.
Oceanogr. 49: 666–679.
GRAHAM, M.D., 1997: Omnivory and selective feeding
by zooplankton along a lake production gradient:
complementary 15N isotope and gut pigment analyses. – M.Sc. Thesis, Univ. of Regina, Regina, Canada.
HALL, R.I., LEAVITT, P.R., QUINLAN, R., DIXIT, A.S. &
SMOL, J.P., 1999: Effects of agriculture, urbanization,
and climate on water quality in the northern Great
Plains. – Limnol. Oceanogr. 44: 739–756.
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A. Patoine, Landscape – scale patterns
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Fig. 4. Nitrogen isotopic content of Daphnia (bottom row), Copepoda (middle row) and Leptodora (top
row) as a function of the relative density of cyanobacteria in upstream Buffalo Pound Lake (left colum)
and downstream lakes (middle and right column). Numbers represent sampling events, from 1 (May 4)
to 9 (August 31). Missing sampling events indicate that lake zooplankton densities were too low for sufficient material to be collected for mass-spectrometry analyses. Coefficients of determination and associated probability values are indicated.
HESSLEIN, R.H., FOX, D.E. & CAPEL, M.J., 1989: Sulfur,
carbon, and nitrogen isotopic composition of fish
from the Mackenzie River delta region and other Arctic drainages. Canadian Data Report of Fisheries and
Aquatic Sciences 728, iv + 11 pp.
KENNEY, B.C., 1990: Dynamics of phosphorus in a
chain of lakes: the Fishing Lakes. National Hydrology Institute, Paper no. 44, 18 pp., Saskatoon,
Saskatchewan, Canada.
LEVINE, S.N. & SCHINDLER, D.W., 1992: Modification of
the N:P ratio in lakes by in situ processes. – Limnol.
Oceanogr. 37: 917–935.
MILLIE, D.F., DIONIGI, C.P., SCHOFIELD, O., KIRKPATRICK,
G.J. & TESTER, P.A., 1999: The importance of understanding the molecular, cellular, and ecophysiological
bases of harmful algal blooms. – J. of Phycol. 35:
1353–1355.
SORANNO, P.A., WEBSTER, K.E., RIERA, J.L., KRATZ,
T.K., BARON, J.S., BUKAVECKAS, P.A., KLING, G.W.,
WHITE, D.S., CAINE, N., LATHROP, R.C. & LEAVITT,
P.R., 1999: Spatial variation among lakes within landscapes: ecological organization along lake chains. –
Ecosystems 2: 395–410.
TALBOT, M.R. 2001: Nitrogen Isotopes in Palaeolimnology. – In: LAST, W.M. & SMOL, J.P. (Eds.): Tracking
Environmental Change Using Lake Sediments, Vol. 2,
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Table 1. Estimates of the amount of nitrogen fixed by cyanobacteria during the 1995 growing season,
May-August. Computations are based on an average nitrogen fixation rate of 5.91 × 10–18 mol N2 s–1 heterocyst –1, and an average cell length of 7 µm (WALSBY 1985). Nitrogen fixation was assumed to occur
uniformly during six hours per day.
Buffalo Pound
Katepwa
Crooked
Lake volume (millions of cubic meters)
87.5
233.2
120.9
Nitrogen-fixing cyanobacteria density (millions
5,132
33,757
44,993
per cubic meter of water, 1995 seasonal average)
Number of heterocysts per 100 µm length of
2.1
6.9
7.1
filament (1995 seasonal average)
Time span during which heterocysts were present 93
76
71
in collected samples (days)
Estimated amount of nitrogen fixed by cyano22
1033
686
bacteria (tonnes)
Estimated 1995 total nitrogen concentration
3,246
2,654
5,120
(mg m–3)*
Estimated amount of nitrogen (tonnes)
284
619
619
Proportion of nitrogen atmospherically derived
8%
169%
111%
* TN concentrations for 1995 were estimated from total inorganic nitrogen (TIN, nitrogen and ammonium) and average TIN/TN ratios of 0.14, 0.18 and 0.10 for Buffalo Pound, Katepwa, and Crooked
Lakes respectively in subsequent years (1998, 1999, 2002).
Physical and Geochemical Methods: 401–439. Kluwer Academic Press, The Netherlands.
WALSBY, A.E., 1985: The permeability of heterocysts to
the gases nitrogen and oxygen. – Proc. of the Royal
Soc. of London B 226: 345–366.
Authors’ addresses:
A. PATOINE, M. GRAHAM, P.R. LEAVITT, Department of
Biology,
University
of
Regina,
Regina,
Saskatchewan, S4S 0A2, Canada. E-mail: [email protected]
R. HESSLEIN, Fisheries and Oceans Canada, Freshwater Institute, Winnipeg, Manitoba, R3T 2N6, Canada.