98
C.E. Goulden et al.
Grazing rates were calculated from the individual clearance rates (Peters, 1984)
of 16 food items, representing >95% of the algal biomass present. We used
analysis of covariance (ANOCOVA; Snedecor and Cochran, 1967) to calculate
clearance rates and significance levels from differences in In (cell concentration)
between treatments with and without Daphnia. The ANOCOVA simultaneously
provided estimates of nutrient effects (for the N plus P coaddition treatment) and
their significance levels. ANOCOVA was an appropriate analytical model because the variances were homogeneous across all treatments and, with the exception of Chrysochromulina, slopes were homogeneous (all P >.1) for treatments
with and without added Nand P. For Chrysochromulina, the clearance rate was
estimated from the NP treatments alone. Average length and biovolume (e.g., cell
contents excluding walls, colonial mucilage) were estimated for each food item.
Total biovolume is the product of average cell volume and cell concentration.
Ingestion rate is the product of total biovolume of a food item (10 3 ,.....m 3 • ml- I)
and clearance rate (mI· Daphnia -I . d - I).
Parallel samples of the original lake water used for the microparticle supplements were screened (40 ,.....m) to remove the largest, mostly inedible algae, collected on precombusted glass fiber filters (Whatman GFIF), and frozen. These
were analyzed for organic carbon and nitrogen (Carlo-Erba Elemental Analyzer).
Total phosphorus was determined as phosphate (John, 1970) following combustion and extraction in boiling dilute HCI.
5.2.4. Algal Counting and Autoradiography
Preserved algal samples (5 ml) were filtered onto 1-,.....m Nucleopore polycarbonate membranes. These were immediately inverted onto gelatin-coated slides and
quick-frozen (cryogenic aerosol), and the filters were stripped away (Hewes and
Holm-Hansen, 1983). After drying (~35°C), slides were mounted in 30%
glycerol (for cell counting) or dipped in Kodak NTB3 emulsion for 14C track
autoradiography (Knoechel and DeNoyelles, 1980; Knoechel and Kalff, 1979,
1976). Following exposure (14 d at 4°C) and processing (7 minutes Kodak D-19,
10 minutes Kodak Fixer, 15 minutes I 0% [mass . vol- I] sodium thiosulfate pentahydrate, with intermediate rinses in tap water), slides were dried and then
mounted with 30% glycerol. Track data were compiled per cell (Synura petersenii, Cryptomonas marssonii) or per unit filament length (Aphanizomenon flosaquae) and are rescaled for plotting as a proportion of the mean rate for the freshly
collected lake treatment.
5.3. Results
5.3.1. Supplement Experiments with Natural Populations
Responses to dietary supplementations were judged from the magnitude and
statistical significance of any increase in clutch size, relative to the control.
C.E. Goulden et al.
Grazing rates were calculated from the individual clearance rates (Peters, 1984)
of 16 food items, representing >95% of the algal biomass present. We used
analysis of covariance (ANOCOVA; Snedecor and Cochran, 1967) to calculate
clearance rates and significance levels from differences in In (cell concentration)
between treatments with and without Daphnia. The ANOCOVA simultaneously
provided estimates of nutrient effects (for the N plus P coaddition treatment) and
their significance levels. ANOCOVA was an appropriate analytical model because the variances were homogeneous across all treatments and, with the exception of Chrysochromulina, slopes were homogeneous (all P >.1) for treatments
with and without added Nand P. For Chrysochromulina, the clearance rate was
estimated from the NP treatments alone. Average length and biovolume (e.g., cell
contents excluding walls, colonial mucilage) were estimated for each food item.
Total biovolume is the product of average cell volume and cell concentration.
Ingestion rate is the product of total biovolume of a food item (10 3 ,.....m 3 • ml- I)
and clearance rate (mI· Daphnia -I . d - I).
Parallel samples of the original lake water used for the microparticle supplements were screened (40 ,.....m) to remove the largest, mostly inedible algae, collected on precombusted glass fiber filters (Whatman GFIF), and frozen. These
were analyzed for organic carbon and nitrogen (Carlo-Erba Elemental Analyzer).
Total phosphorus was determined as phosphate (John, 1970) following combustion and extraction in boiling dilute HCI.
5.2.4. Algal Counting and Autoradiography
Preserved algal samples (5 ml) were filtered onto 1-,.....m Nucleopore polycarbonate membranes. These were immediately inverted onto gelatin-coated slides and
quick-frozen (cryogenic aerosol), and the filters were stripped away (Hewes and
Holm-Hansen, 1983). After drying (~35°C), slides were mounted in 30%
glycerol (for cell counting) or dipped in Kodak NTB3 emulsion for 14C track
autoradiography (Knoechel and DeNoyelles, 1980; Knoechel and Kalff, 1979,
1976). Following exposure (14 d at 4°C) and processing (7 minutes Kodak D-19,
10 minutes Kodak Fixer, 15 minutes I 0% [mass . vol- I] sodium thiosulfate pentahydrate, with intermediate rinses in tap water), slides were dried and then
mounted with 30% glycerol. Track data were compiled per cell (Synura petersenii, Cryptomonas marssonii) or per unit filament length (Aphanizomenon flosaquae) and are rescaled for plotting as a proportion of the mean rate for the freshly
collected lake treatment.
5.3. Results
5.3.1. Supplement Experiments with Natural Populations
Responses to dietary supplementations were judged from the magnitude and
statistical significance of any increase in clutch size, relative to the control.