Effects of colonial morphology on zooplankton utilization of algal resources during blue-green algal (Microcystis aeruginosa) blooms

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1 Limnol. Oceanogr., 32(3), 1987, , by the American Society of Limnology and Oceanography, Inc. Effects of colonial morphology on zooplankton utilization of algal resources during blue-green algal (Microcystis aeruginosa) blooms Rolland S. Fulton 1112 and Hans W. Paerl Institute of Marine Sciences, University of North Carolina, 3407 Arendell Street, Morehead City Abstract Experiments were conducted to test the following proposed mechanisms whereby blooms of colonial blue-green algae (Microcystis aeruginosa) may more negatively affect large-bodied cladocerans than other zooplankton taxa: that colonial morphology most strongly interferes with the feeding of larger cladocerans, and that larger cladocerans more readily ingest toxic or nonnutritious colonial blue-greens. Experiments were conducted with both colonial and unicellular M. aeruginosa, in order to distinguish the effects of coloniality from chemical inhibitory factors associated with bluegreen algae. Experiments with colonial M. aeruginosa provided some support for both mechanisms, particularly with the cladoceran Diaphanosoma brachyurum. However, some small-bodied species associated with blue-green algal blooms did not conform to the proposed mechanisms, showing relatively strong reductions in clearance rates in the presence of colonial M. aeruginosa (Bosmina longirostris, Ceriodaphnia quadrangula) or relatively high consumption of colonial M. aeruginosa (Brachionus cafyciflorus, B. Iongirostris). Our results indicate that, contrary to earlier proposals, neither body size, taxonomic position, nor association with blue-green algal blooms are good predictors of the response to colonial M. aeruginosa. It has often been noted that blooms of colonial or filamentous blue-green algae are associated with changes in zooplankton composition, including losses of large cladocerans and increases in importance of smaller cladocerans, rotifers, and copepods (Gliwicz 1977; Edmondson and Litt 1982; Richman and Dodson 1983; Orcutt and Pace 1984). Among the explanations for such changes in zooplankton composition is that the colonial morphology of the blue-green algae most strongly inhibits large-bodied cladocerans. Two major, but not necessarily mutually exclusive, mechanisms have been proposed for the greater susceptibility of large cladocerans to colonial or filamentous blue-green algae (Gliwicz 1977; Webster and Peters 1978). First, it has been suggested that colonial or filamentous algae clog the filtering appendages of larger cladocerans, reducing their feeding rates on co-occurring nutritious food sources or increasing their respiration rates or both. Several studies have supported this mechanism, showing that feeding or respiration rates of larger I Supported in part by North Carolina Sea Grant Project R/MER-5 and by NSF BSR * Present address: Department of Biology, George Mason University, 4400 University Drive, Fairfax, Virginia cladocerans are most strongly affected by the presence of filamentous blue-greens (Webster and Peters 1978; Richman and Dodson 1983; Porter and McDonough 1984). However, this mechanism has not been examined with more loosely organized colonial blue-greens, such as Microcystis aeruginosa. Moreover, these studies do not clearly demonstrate that the effects on feeding and respiration are due solely to the filamentous morphology; toxic or inhibitory chemicals produced by the blue-green algae could also contribute to the effect. Lampert ( 198 1, 1982) has shown that unicellular M. aeruginosa can strongly inhibit cladoceran feeding rates, presumably due to a chemical effect. Webster and Peters (1978) found that cellulose fibers had less effect on cladoceran reproduction than did blue-green filaments, again suggesting that chemical factors contribute to the inhibitory effect of blue-green filaments. Porter and McDonough (1984) reported that unicells, toxic filaments, and nontoxic filaments of Anabaena all stimulated similar rejection rates by Daphnia, implicating taste as another factor. The second proposed mechanism is that larger cladocerans more readily ingest colonial or filamentous blue-greens and are thereby more strongly affected by any toxic chemicals that these algae may possess. Most 634

2 Efects of coloniality on herbivory 635 blue-green algae studied have been found to be toxic or of poor nutritional value (Arnold 197 1; Porter and Orcutt 1980; Lampert 198 1; Holm and Shapiro 1984; Nizan et al. 1986), although there are a few reports of successful population growth on a diet of blue-green algae in cladocerans (DeBernardi et al ) and rotifers (Starkweathcr and Kellar 1983). There is little evidence for this mechanism, beyond the observations that the maximum size of inert particles that is eaten increases with cladoceran size (Burns 1968; Gliwicz 1969). More recent studies indicate that this relationship of herbivore size to food size is not so strong (Bogdan and Gilbert 1984). Some copepods and small cladocerans have been found to feed more selectively than large daphnid cladocerans (Poulet and Marsot 1978; Alcaraz et al. 1980; Koehl and Strickler 198 1; DeMott 1982; DeMott and Kerfoot 1982), but there is little evidence that this selectivity is used to avoid feeding on colonial blue-greens. The Neuse River, North Carolina, has developed recurring blue-green algal blooms, dominated by the colonial species M. aeruginosa (Tedder et al. unpubl. rep.; Paerl 1983). Associated with the summer 1983 bloom were high densities of the rotifer Brachionus calycij lorus and several smallbodied cladocerans, including Bosmina longirostris, Bosminopsis dietersi, Diaphanosoma brachyurum, and Moina micrura, while large-bodied cladocerans were almost absent. Growth experiments with strains of M. aeruginosa isolated from the river showed that they were either toxic or of insufficient nutritional value to support growth and reproduction for several of the zooplankton species in the river (Fulton and Paerl in press). In this study we tested the two proposed mechanisms whereby colonial M. aeruginosa may have a size-selective effect on the herbivore community. We did feeding experiments with M. aeruginosa of both unicellular and colonial morphology, in order to distinguish the effects of coloniality from other factors, such as chemical inhibitory effects. Experiments were carried out with a wide range of sizes and taxa of zooplankton in order to determine if there are any size- or taxon-specific responses to colonial M. aeruginosa. We thank D. Bowles for field assistance, M. Haibach and J. Tucker for aid with algal cultures, V. Page and H. Page for assistance with figure preparation, and W. Lampert, K. Porter, and G. Wyngaard for comments on the manuscript. Methods Most species of zooplankton used in the experiments were taken from established cultures of animals originally isolated from the Neuse River. Animals were cultured in filtered (Whatman 934-AH) Neuse River water at 20 C under a variable photoperiod, and fed a mixture of Chlamydomonas reinhardi and Ankistrodesmus falcatus. The experiments with Eurytemora afinis, B. dietersi, and some of the experiments with Ceriodaphnia quadrangula were done with animals collected from the river l-2 d previously. Algae used in the experiments were C. reinhardi obtained from Carolina Biological Supply and two strains of M. aeruginosa isolated from the Neuse River. Chlamydomonas reinhardi is a flagellated green alga, spherical in shape, with average diameter of 7.03 pm and average dry biomass of 9.94 X 10-5 hg cell- I. Cells of M. aeruginosa are also spherical in shape, with average diameter of 3.86 pm and average dry biomass of 7.74 x 1O-6 pg cell-l. Strain 1 of M. aeruginosa, isolated in summer 1983, was largely unicellular in growth form during the study period (average colony size 1.40 cells colony-l). Strain 2, isolated in October 1984, was maintained in a colonial morphology, average size ranging from 11.1 to cells colony- * in different experiments (colony sizes based on counts of at least 100 colonies). Chlamydomonas reinhardi and M. aeruginosa Strain 1 were cultured in ASM-J medium (Parker 1982). Microcystis aeruginosa Strain 2 was initially grown in ASM-J, but we altered the medium when it began to grow unicellularly. The new medium was water from cultures of Daphnia ambigua to which we added ASM-J nutrients before filter sterilization. These cultures were filtered through a 20-pm filter two to three times weekly and large colonies retained on the mesh were resuspended in fresh medium. Controlled experiments have

3 636 Fulton and Paerl indicated that it was the filtration procedure that selected for large colonies in these cultures and not some dissolved product associated with the D. ambigua medium. Algal cultures were not axenic, but we used algae from exponentially growing cultures, in which bacterial biomass has been observed to be relatively low (J. Tucker pers. comm.). There were three treatments for each experiment. In treatment 1, we measured filtering rates when the animals fed on C. reinhardi at a concentration of 10,000 cells ml- I (cell volume concn, 1.82 mm3 liter- ). In treatment 2, we measured filtering rates on C. reinhardi in a mixture consisting of C. reinhardi at 10,000 cells ml- I and M. aeruginosa at a concentration of 100,000 cells ml-l (total cell volume concn, 4.83 mm3 liter-l). This concentration of M. aeruginosa is within the range observed in the Neuse River during the summer 1983 bloom (Paerl unpubl. data). In treatment 3, we measured filtering rates on M. aeruginosa at a concentration of 100,000 cells ml- I (cell volume concn, mm3 liter- ). We tested two major hypotheses with this procedure. The first, that the presence of M. aeruginosa will reduce feeding rates on C. reinhardi, was tested by comparing filtering rates on C. reinhardi in the algal mixture (treatment 2) with those when feeding on C. reinhardi alone (treatment 1). The second was that filtering rates will be lower on M. aeruginosa (treatment 3) than on C. reinhardi in the algal mixture (treatment 2). Experiments were carried out with the copepod E. a&is, the rotifers B. calyciflorus and Platyias patulus, and the cladocerans B. longirostris, B. dietersi, C. quadrangula, D. brachyurum, D. ambigua, and Simocephalus serratulus. For six species the experiment was carried out with both the colonial Strain 2 and the unicellular Strain 1, to distinguish the effects of the colonial morphology from other potential inhibitory factors associated with M. aeruginosa, such as toxic or unpalatable substances. Finally, we repeated the experiment with Strain 2 in a unicellular form to make sure that any differences in responses to the two strains were due solely to differences in morphology and not to chemical differences between the strains. WC obtained Strain 2 in a largely unicellular form (avg size, 2.02 cells colony- I) by growing it in ASM-J medium and using only cells which passed through a 20- pm mesh net. This experiment was done with two herbivores, D. ambigua and D. brachyurum. We used high cell densities in the herbiv- or-y experiments because we were interested in the effects of M. aeruginosa at densities typically occurring in algal blooms and because other experiments indicated that these strains of M. aeruginosa had negligible effects on zooplankton feeding at lower cell densities (Fulton and Paerl in press, and unpubl. data). A complication of the interpretation of these experiments is that filter- ing rates would be expected to be reduced in treatments with higher algal cell volume, if the food concentration exceeds the incipient limiting level (e.g. Porter et al. 1982). This factor is particularly important in the testing of our first hypothesis, since the higher cell volume in the algal mixture treat- ment may reduce filtering rates, regardless of any inhibitory effect of M. aeruginosa. To examine the magnitude of this effect of increased cell volume, we conducted a preliminary experiment in which we measured filtering rates on C. reinhardi at cell volume concentrations equal to those in the treatments used in testing the first hypothesis (1.82 and 4.83 mm3 liter- ). This experiment was conducted with E. afinis, D. ambigua, D. brachyurum, and S. serratulus. The differences in cell volume concentration among treatments are less important in the testing of our second hypothesis, because on the basis of cell volume concentrations alone, filtering rates should be higher on M. aeruginosa than in the algal mixture, which is the opposite of our prediction. Filtering rates were measured by uptake of radiolabeled algal cells. Algae were labeled with 8.1 &i NaH14C ml- l of culture medium 2 d before each experiment. Experimental suspensions of C. reinhardi and unicellular M. aeruginosa were prepared by centrifugation (2,800 rpm for 4 min) and resuspension in Whatman 934- AH filtered, autoclaved Neuse River water. The colonial M. aeruginosa was filtered through a 20-pm mesh net, then resuspend-

4 Efects of coloniality on herbivory 637 ed in filtered, autoclaved river water. Algal concentrations were initially determined by cell counts; later we measured absorbance at 720 nm, deriving cell concentrations from a previously established calibration curve (r2 > 0.99 for both species). Experiments were carried out in darkness, at 20 C on a slow-speed (30 rpm) orbital shaker. The number of animals per replicate varied from 3-5 for the large cladoceran S. serratulus to for the rotifers. Animals were selected with pipettes and allowed to acclimate for 1 h before each experiment in 100 ml of filtered, autoclaved Ncuse River water having the appropriate concentration of unlabeled algae. The experiment was begun by adding 100 ml of radioactively labeled food suspension. Feeding was allowed for 10 min, after which the animals were rinsed and anesthetized in chilled, carbonated water, filtered onto Whatman 934-AH filters, then quickly frozen with liquid N2 and subsequently freezedried (see Paerl 1984). This preservation method avoids loss of radiotracer in liquid media (Holtby and Knoechel 198 1) and also preserves most individuals in good condition for accurate measurements of body size. One or two aliquots of the feeding suspen- sion were taken both at the beginning and end of the experiment, filtered through Whatman 934-AH filters, air-dried, fumed with HCl for 15 min, allowed to dry again, placed in scintillation vials with 5 ml of Fisher Scintillene cocktail, and counted in a Beckman LS-7000 liquid scintillation counter after 2 h storage in darkness at room temperature. The lengths of a subsample of the frcezedried zooplankton (usually five per replicate) were measured with an ocular micrometer ; the zooplankton were then transferred to scintillation vials, fumed with HCl for 15 min, dried, maintained in darkness for 24 h at room temperature in 5 ml of Fisher Scintiverse, and counted in the scintillation counter. In preliminary experiments carried out with D. ambigua and E. afinis, we compared this method of pro- cessing the zooplankton with a more common method of solubilizing the zooplankton in 0.5 ml of Protosol for 12 h at 40 C, and then adding 5 ml of Scintiversc and 150 ~1 of concentrated HCl and maintaining samples in darkness for 12 h at room temperature bcforc counting. We could find no significant difference in counts per minute per replicate recorded by these two methods. Counting efficiency, determined by an.extcrnal standard, exceeded 90% for both phytoplankton and zooplankton samples. On a single date we carried out three rep- licates and one control with dead animals (heat- or Formalin-killed) for each treatment. They were repeated on 2-5 dates for a total of 6-15 replicates for each expcriment. The only exception was the experimcnt with B. dietersi; it could not be cultured and was found in sufficient numbers in the river on only one date during the study period. Accordingly, we have only three replicates for this species. WC calculated filtering rates according to the formula in Porter et al. (1982), corrected for uptake of radiotracer by dead animals. We analyzed the results of each experiment by a randomized-block analysis of variance (ANOVA), considering each date as a block. We used contrasts with single degrees of freedom to test our two major hypotheses. These contrasts are not orthogonal so we adjusted the significance level for them by the Dunn-Sidak method (Sokal and Rohlf 198 1). The adjusted significance level was P < The analysis was performed on log-transformed filtering rates if variances of untransformed filtering rates were heterogeneous (Cochran s test, Winer 197 1). In the one case in which the transformation did not make the variances homogeneous (the experiment with E. afinis feeding on Strain 2 M. aeruginosa) we used a Kruskal- Wallis test, followed by a nonparametric multiple comparisons test (Sokal and Rohlf 1981). For comparisons among hcrbivorc spccics, or among experiments utilizing different strains of M. aeruginosa, we carried out analyses of relative clearance rates. Relative clearance rates were arcsine-transformed and subjected to a t-test, or a oncway ANOVA followed by Tukey s HSD multiple comparisons test. If the variances of the arcsinc-transformed proportions were not homogeneous, we used an approximate t-test assuming unequal variances, or a

5 638 Fulton and Paerl RELATIVE SPECIES CLEARANCE RATES 20 I 40 I 60 Daphnia ambigua Diaphanosoma brachyurum Simocephalus serratulus -- Eurytemora affinis - P Fig. 1. Clearance rates on Chlamydomonas reinhardi at a cell volume concentration equivalent to that in the algal mixture treatment (4.83 mm3 liter- ), expressed as a percentage of those when feeding on C. reinhardi at 10,000 cells ml- (1.82 mm3 liter-*). Mean + 1 SE. Kruskal-Wallis test followed by Dunn s nonparametric multiple comparisons test for unequal sample sizes (Hollander and Wolfe 1973). Results Increasing the total cell volume of C. reinhardi to an amount equivalent to that in the algal mixture treatment significantly reduced the clearance rates of all species tested (P < ) to about 50-60% of the rates at the lower concentration of C. reinhardi (Fig. 1). For all three types of M. aeruginosa, the relative clearance rates of D. ambigua in the algal mixtures (Fig. 2) were significantly lower (P < 0.05) than those at the same total volume of C. reinhardi (Fig. l), demonstrating that M. aeruginosa had an inhibitory effect beyond that expected by the increased cell volume. The relative clearance rate of D. brachyurum in the algal mixture including Strain 1 M. aeruginosa (Fig. 2) was also significantly lower (P < ) than at the same total volume of C. reinhardi, but there were no significant differences for either morphology of Strain 2 M. aeruginosa. Like D. ambigua, clearance rates of E. afinis (Fig. 3C) and S. serratulus (Fig. 4A) were inhibited significantly more by colonial M. aeruginosa than by an equivalent cell volume of C. reinhardi (P < ). However, for unicellular M. aeruginosa, the relative clearance rates of E. afinis in the algal mixture did not differ significantly from those at the same total volume of C. reinhardi. For D. ambigua and D. brachyurum, both strains of M. aeruginosa caused significant reductions in clearance rates on C. reinhardi, compared to the rates when feeding on C. reinhardi alone (P < ) (Fig. 2). Strain 2 had a smaller inhibitory effect than Strain 1: for D. ambigua the relative clearance rate in the algal mixture was significantly lower for Strain 1 than for either colonial or unicellular Strain 2 (P < 0.05, Tukey s HSD test); for D. brachyurum, the relative clearance rate in the algal mixture was significantly lower for Strain 1 than for colonial Strain 2 (P < 0.05, Dunn s test) but did not differ significantly from unicellular Strain 2. Clearance rates on unicellular M. aeruginosa were either significantly higher than those on C. reinhardi in the algal mixture (D. ambigua feeding on Strain 2 and D. brachyurum feeding on Strain 1, both P < ) or did not differ significantly (D. ambigua feeding on Strain 1 and D. brachyurum feeding on Strain 2) (Fig. 2). In sharp contrast were the results with Strain 2 colonial M. aeruginosa; clearance rates of D. ambigua on Strain 2 colonies were 7 1.6% of those on C. reinhardi in the algal mixture (P < ), while those of D. brachyurum were only 24.5% (P < ). Results with B. longirostris and C. qua- drangula were similar to those with D. ambigua and D. brachyurum, respectively. Clearance rates of both herbivores on C. reinhardi were significantly reduced by the presence of either unicellular or colonial M. aeruginosa (P < O.OOOl), although the unicellular strain had a greater effect than the colonial strain (Fig. 3A, B). Clearance rates of B. longirostris on unicellular M. aeruginosa were significantly higher than those on C. reinhardi in the algal mixture (P < ) while those on colonial M. aeruginosa were slightly but significantly lower (77.7%, P < ) (note that the signifi-

6 Efects of coloniality on herbivory 639 Ia A. Daphnia ambigua C.reinhardi lo,ooo/ml q C.reinhar& lo,ooo/ml + Moeruginosa ,000/ml q M.aeruginosa l00,000/ml Strain 1 Unicellular Strain 2 Unicellular Strain 2 Colonio I Fig. 2. Relative clearance rates in the three treatments for cxpcriments with Strain 1 unicellular Microcystis aeruginosa and Strain 2 unicellular and colonial M. aeruginosa. Clearance rates expressed as a percentage of those exhibited in treatment 1 -feeding on Chlamydomonas reinhardi at 10,000 cells ml-. Mean f 1 SE. cance of this result is not apparent in Fig. 3A, because the randomized block ANOVA partitions among-date variability from residual variability, but the error bars in the figure incorporate both among-date and residual variability). Clearance rates of C. quadrangula on unicellular M. aeruginosa were also significantly higher than those on C. reinhardi in the algal mixture (P < ), while those on colonial M. aeruginosa were much lower (20.2%, P < ). Clearance rates of the copepod E. afinis on C. reinhardi were also significantly reduced in the presence of either morphology of M. aeruginosa (P < 0.01) (Fig. 3C). Unlike the cladocerans, E. afinis consumed very little of either unicellular or colonial M. aeruginosa; clearance rates on both strains were < 16% of those on C. reinhardi in the algal mixture (P < 0.01). Unlike the crustaceans, clearance rates of the rotifer B. calyczjlorus were little affected by high densities of M. aeruginosa. There were no significant differences among treat- ments in the experiment using unicellular M. aeruginosa (Fig. 3D). Clearance rates of B. calycijlorus on C. reinhardi were slightly, but significantly, reduced in the presence of colonial M. aeruginosa (74.2%, P < ). Unlike all other species, clearance rates of B. calyctflorus on colonial M. aeruginosa were significantly higher than those on C. reinhardi in the algal mixture (P < ). Results from different dates in which the colony size of M aeruginosa varied indicated that B. calyczjlorus feeds most intensively on medium-sized colonies. On two dates when the average colony size was cells colony- I, clearance rates of B. calyctjlorus on M. aeruginosa were almost double (19 1%) those on C. reinhardi in the algal mixture (P < ), but on two other dates when average colony size was larger (I cells colony- ) the clearance rates did not differ significantly. Thus the relative filtering rates were highest on medium-sized colonies and were similar on unicells and large colonies. We have directly

7 640 F&on and Paerl Bosmina A. - longirostris Ceriodaphnia quadrangula strain I. Unicellular strain 2 Colonial Strain 1 Unicellular Strain 2 Colon ia I Fig. 3. Relative clearance rates in the three treatments for experiments with Strain 1 unicellular Microcystis aeruginosa and Strain 2 colonial M. aeruginosa. Symbols as in Fig. 2. Mean + 1 SE. observed B. calyc$!orus consume colonies in a size range of pm with no apparent difficulty. Clearance rates on C. reinhardi in the algal mixture, relative to those fed C. reinhardi alone, differed significantly among species (P < ) (Fig. 4A). There are evidently two major groups of species (although this assertion cannot be statistically verified by the extremely conservative Dunn s nonparametric comparisons test): the two rotifers and D. brachyurum, whose clearance rates were relatively little affected by the presence of colonial M aeruginosa ( %), and the other crustaceans, whose clearance rates were severely reduced by colonial M. aeruginosa ( %). Clearance rates on colonial AL aeruginosa, relative to those on C. reinhardi in the algal mixture, also differed significantly among species (P < ) (Fig. 4B). The herbivores occur in three general groups: a group which consumes very little ( %) colonial M. aeruginosa (P. patulus, C. quadrangula, D. brachyurum, and E. af- finis), the other crustaceans, which consume substantial amounts of colonial M. aeruginosa ( %), and B. calyciflorus, which, as indicated above, revealed significantly higher clearance rates on colonial M. aeruginosa than on C. reinhardi in the algal mixture. Discussion Clearance rates of all the cladocerans tested were as high or higher on unicellular M. aeruginosa than on C. reinhardi in the algal mixture (Figs. 2, 3). The contrast between the results with both strains of unicellular M. aeruginosa and the results with colonial M. aeruginosa clearly demonstrate that the reduced clearance rates on colonial M. aeruginosa are due solely to the colonial morphology and not to other chemical inhibitory factors. Porter and Orcutt (1980) also reported that Daphnia magna had higher feeding rates on unicellular than on filamentous Anabaena, but our study is the first to demonstrate that some cladocerans commonly associated with blue-green algal

8 Eflects of coloniality on herbivory 641 SPECIES RELATIVE CLEARANCE RATES LENGTH (mm) 0.25 I I I -- Platyias patulus 0.13 Brachionus -- calyciflorus 0.23 Bosminopsis dietersi 0.31 I I ( I I I -- Bosmino longirostris 0.4 I I I, -~ Ceriodaphnia quadrangula 0.67 Diophanosoma brochyurum 0.84 Daphnia -- ambigua 1.08 Eurytemora -- affinis I.37 Simocephalus serratulus I.49 El BI Ss Da Db Pp Bc 4 Pp Ea Cq Db Ss Da Bd BI 4 Bc Fig. 4. Relative clearance rates for a range of sizes and taxa of herbivore in experiments with Strain 2 colonial Microcystis aeruginosa. Mean -t 1 SE. Shown below figures arc significant differences (P < 0.05) among species as determined by Dunn s nonparametric multiple comparisons test (two-letter codes are abbreviations for genus and species name). A. Clearance rates on Chlamydomonas reinhardi in the algal mixture (C. reinhardi 10,000 cells ml I and M. aeruginosa 100,000 cells ml I), expressed as a proportion of those when feeding on C. reinhardi alone. B. Clearance rates when feeding on M. aeruginosa at 100,000 cells ml- relative to those when feeding on C. reinhardi in the algal mixture. blooms (0. brachyurum, C. quadrangula) do consume less of the potentially toxic or nonnutritious blue-greens than do some larger bodied species (e.g. D. ambigua, S. serratulus), solely because of the colonial morphology (assuming our procedure for producing strains of differing sizes did not also change chemical characteristics). It may be noted that there were some colonies of a readily filterable size during the colonial ikf. aeruginosa experiments. Even during the experiments with the largest average colony size, the modal size class was always l-3 cells colony- 9 presumably due to colony breakage during or after filtering. The low clearance rates on colonial M. aeruginosa exhibited by D. brachyurum, C. quadrangula, and P. patulus may be exclusively on these small colonies. This study also supports the hypothesis that clearance rates on nutritious food of some species associated with blue-green algal blooms (D. brachyurum, B. calyciflorus) are less affected by the presence of colonial M. aeruginosa than are clearance rates of some relatively large cladocerans that do poorly during blooms (D. ambigua, S. serrat&us). In the case of D. brac hyurum this result is again due to the colonial morphology, as its clearance rates on C. reinhardi are severely affected by unicellular M. aeruginosa (Fig. 1). However, clearance rates of B. cazycz@ Zorus are little affected by either morphology of M. aeruginosa. Three factors may contribute to the reduced clearance rates on C. reinhardi in the presence of M. aeruginosa: interference with feeding because of the colonial morphology, chemical inhibitory effects (Lampcrt 198 1, 1982), and increased food concentrations if the food level exceeds the incipient limiting level (e.g. Porter et al. 1982). The experiments presented here allow a separation of the effects of these three factors. For cladocerans, the primary factors inhibiting clearance rates on C. reinhardi in the presence of M. aeruginosa were the increased food concentration and inhibitory chemical effects. Clearance rates of all cladocerans on C. reinhardi were inhibited as much or more by unicellular M. aeruginosa as by the colonial form, indicating that effects of inter-

9 642 Fulton and Paerl ference with feeding by the colonial morphology were minor relative to the joint effects of inhibitory chemicals and increased food concentration associated with M. aeruginosa blooms. The more important effect of the colonial morphology is that it makes M. aeruginosa unavailable to cladocerans restricted to feeding on small particles (e.g. D. brachyurum). The greater inhibitory effect of unicellular Strain 1 M. aeruginosa was apparently due to a greater toxicity of this strain (Fulton and Paerl in press). The colonial morphology had a greater inhibitory effect on the feeding of E. afinis and B. calyczjlorus; clearance rates of both species were inhibited more by the colonial form than by the unicellular form, although the effect was small in both cases. The absence of a strong inhibitory effect of either morphology of M. aeruginosa on clearance rates of B. calycijlorus (Fig. 4D) indicates that the incipient limiting level was higher for B. calyczjlorus than for the crustaceans, and that its feeding was not inhibited by toxic or noxious chemicals produced by M. aeruginosa. The higher incipient limiting level of B. calyczflorus is also evident in previous studies of the functional response of B. calyczjlorus and cladocerans (Starkweather 1980; Porter et al. 1982). Strain 1 of M. aeruginosa was toxic to cladocerans but not to B. calyciforus (Fulton and Paerl in press), which probably accounts for the absence of an inhibitory effect of chemicals produced by M. aeruginosa on clearance rates of B. calyciflorus. It is possible that M. aeruginosa has a greater inhibitory effect on zooplankton clearance rates than does an equivalent cell volume concentration of C. reinhardi because the gelatinous sheath surrounding M. aeruginosa disproportionately increases total food volume. We have observed, by toluidine blue staining, that the sheath is much more strongly developed around colonies than around unicells. Therefore, if the increased volume due to the sheath reduces zooplankton filtering rates, then the colonial morphology should have a much stronger inhibitory effect than the unicellular form. This difference in inhibition did not occur; in most cases filtering rates were inhibited as much, or more by the unicellular mor- phology. We conclude that the sheath does not have a significant inhibitory effect on clearance rates. The strong avoidance of both unicellular and colonial M. aeruginosa by E. afinis indicates that this copepod avoids or rejects M. aeruginosa on some basis other than particle size, most likely due to toxic or unpalatable chemicals associated with M. aeruginosa cells. This inference is consistent with the known chemosensory feeding abilities of copepods (Poulet and Marsot 1978; Alcaraz et al. 1980; Koehl and Strickler 198 1). Gut content analyses by Clarke (1978) also indicated little or no feeding by copepods on M. aeruginosa of either colonial or unicellular morphology. However, there are reports of consumption of M. aeruginosa by some copepods (Schindler 197 1; Infante 1978). Comparisons of the response to M. aeruginosa among all species tested show few consistent patterns with body size, among taxa, or even between species associated with blue-green algal blooms and those that do poorly under such conditions (Fig. 4). Only the results with D. brachyurum and P. pat&us support both of the hypothetical mechanisms: their filtering rates were low on colonial M. aeruginosa and were relatively little affected by the presence of colonial M. aeruginosa. Ceriodaphnia quadrangula and E. afinis consumed little colonial M. aeruginosa, but their clearance rates on C. reinhardi were strongly affected by the presence of colonial M. aeruginosa. O Brien and DeNoyelles (1974) reported that filtering rates of Ceriodaphnia reticulata were unaffected by the presence of colonial M. aeruginosa, but they did not measure either the concentration of M. aeruginosa or the colony sizes. Clearance rates of the two closely related rotifers (both of subfamily Brachioninae) on C. reinhardi were both relatively unaffected by the presence of colonial M. aeruginosa, but their clearance rates on M. aeruginosa were at the opposite extremes of the species tested. Although B. longirostris and B. dietersi have been associated with blue-green algal blooms (Gliwicz 1977; Fulton and Paerl in press), their responses to colonial M. aeruginosa departed little from those of D. ambigua

10 Efects of coloniality on herbivory 643 and S. serratulus. The ability of B. longirostris to consume substantial amounts of colonial M. aeruginosa is consistent with previous studies indicating that it can consume relatively large particles for its size (Bogdan and Gilbert 1982; Bleiwas and Stokes 1985). Although B. longirostris has been reported to be capable of complex selective feeding behavior (Bogdan and Gilbert 1982; DeMott 1982; DeMott and Kerfoot 1982), in our experiments it did not exhibit the strong avoidance of M. aeruginosa shown by E. afinis. The results presented here indicate that herbivores associated with blue-green algal blooms exhibit two distinctly different behaviors. One group, exemplified by D. brachyurum, shows a combination of the two hypothetical mechanisms tested in this paper: it consumes little M. aeruginosa in the colonial morphology, and thereby it consumes little of the toxic or inhibitory chemicals possessed by M. aeruginosa, while its clearance rates on co-occurring more nutritious algae are relatively unaffected by the prcscnce of colonial M. aeruginosa. The other group, represented by B. Zongirostris and especially B. calyciflorus, consumes substantial amounts of colonial M. aeruginosa. These species must exhibit a greater ability to utilize M. aeruginosa as a nutritional source or at least a greater tolerance to the toxins to succeed during blooms of M. aeruginosa. References ALCARAZ, M., G.-A. PAFFENH~FER, AND J. R. STRICK- LER Catching the algae: A first account of visual observations on filter-feeding calanoids. Am. Sot. Limnol. Oceanogr. Spec. Symp. 3: New England. ARNOLD, D. E Ingestion, assimilation, survival, and reproduction by Daphnia pulex fed seven spccics of blue-green algae. Limnol. Oceanogr. 16: BLEIWAS, A. H., AND P. M. STOKES Collection of large and small food particles by Bosmina. Limnol. Oceanogr. 30: BOGDAN, K. G., AND J. J. GILBERT Seasonal patterns of feeding by natural populations of Keratella, Polyarthra, and Bosmina: Clearance rates, sclcctivities, and contributions to community grazing. Limnol. Oceanogr. 27: , AND Body size and food size in freshwater zooplankton. Proc. Natl. Acad. Sci. 81: BURNS, C. W The relationship between body size of filter-feeding Cladocera and the maximum size of particle ingested. Limnol. Oceanogr. 13: CLARKE, N. V The food of adult copepods from Lake Kainji, Nigeria. Freshwater Biol. 8: 32 l DEBERNARDI, R., G. GIUSSANI, AND E. L. PEDRETTI The significance of blue-green algae as food for filter-feeding zooplankton: Expcrimcntal studies on Daphnia spp. fed by Mycrocystis aeruginosa. Int. Ver. Theor. Angew. Limnol. Verh. 21: DEMOTT, W. R Feeding selectivities and relative ingestion rates of Daphnia and Bo$mina. Limnol. Oceanogr. 27: , AND W. C. KERFOOT Competition among cladocerans: Nature of the interaction bctween Bosmina and Daphnia. Ecology 63: EDMONDSON, W. T., AND A. H. LITT Daphnia in Lake Washington. Limnol. Oceanogr. 27: FULTON, R. S., III, AND H. W. PAERL. In press. Toxic and inhibitory effects of the blue-green alga Microcystis aeruginosa to herbivorous zooplankton. J. Plankton Res. GLIWICZ, Z. M Studies on the feeding ofpelagic zooplankton in lakes with varying trophy. Ekol. Pol. 17: Food size selection and seasonal succession of filter feeding zooplankton in a eutrophic lake. Ekol. Pol. 25: HOLLANDER, M., AND D. A. WOLFE Nonparametric statistical methods. Wiley. HOLM, N. P., AND J. S. SHAPIRO An examination of lipid rescrvcs and the nutritional status of Daphnia pulex fed Aphanizomenon flos-aquae. Limnol. Oceanogr. 29: 1137-l 140. HOLTBY, L. B., AND R. KNOECHEL Zooplankton filtering rates: Error due to loss of radioisotopic label in chemically preserved samples. Limnol. Oceanogr. 26: INFANTE, A. DE Natural food of herbivorous zooplankton of Lake Valencia (Venezuela). Arch. Hydrobiol. 82: KOEHL, M. A. R., AND J. R. STRICKLER Copepod feeding currents: Food capture at low Reynolds number. Limnol. Oceanogr. 26: LAMPERT, W Inhibitory and toxic effects of blue-green algae on Daphnia. Int. Rev. Gesamten Hydrobiol. 66: Further studies on the inhibitory effect of the toxic blue-green Microcystis aeruginosa on the filtering rate of zooplankton. Arch. Hydrobiol. 95: NIZAN, S., C. DIMENTMAN, AND M. SHILO Acute toxic effects of the cyanobacterium Microcystis aeruginosa on Daphnia magna. Limnol. Oceanogr. 31: O BRIEN, W. J., AND F. DENOYELLES, JR Filtering rate of Ceriodaphnia reticulata in pond waters of varying phytoplankton concentrations. Am. Midl. Nat. 91:

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