J. Great Lakes Res. 31:144–154 Internat. Assoc. Great Lakes Res., 2005

Behavioral response of Lake Michigan mendotae to Mysis relicta

Scott D. Peacor1,2,*, Kevin L. Pangle1,2, and Henry A. Vanderploeg2 1Michigan State University Department of Fisheries and Wildlife 13 Natural Resources Building East Lansing, Michigan 48824-1222 2Great Lakes Environmental Research Laboratory (NOAA) 2205 Commonwealth Boulevard Ann Arbor, Michigan 48105-2945

ABSTRACT. We performed laboratory experiments to determine if Mysis relicta induce changes in the behavior of Daphnia mendotae collected from Lake Michigan. Laboratory results indicate that Daphnia perceived Mysis kairomones and responded by changing their vertical position in cylinders. Experiments using different resource levels, and two procedures to examine the potential effects of the chemical cues from Mysis or from particulate matter or associated with capture and defecation of prey, sug- gest that Daphnia detect Mysis via a chemical cue. This is the first laboratory study that we are aware of that indicates that a species from the Great Lakes responds behaviorally to an predator. Our findings support the hypothesis that changes in vertical distribution of zooplankton associ- ated with changes in invertebrate predator density, observed in previous Great Lakes studies, is due to behavioral responses to reduce risk. It is important to understand and quantify such responses, because predator-induced changes in prey behavior represent trait-mediated interactions that can poten- tially strongly affect prey growth rates, and indirectly affect resources, competitors, and predators of the prey. INDEX WORDS: Daphnia mendotae, Mysis relicta, Lake Michigan, vertical migration, , kairomone.

INTRODUCTION Ramcharan and Sprules 1991, Loose and Dawidow- In aquatic systems, predation risk is believed to icz 1994,Van Gool and Ringelberg 1998, reviewed be an important factor in determining the vertical in Tollrian and Harvell 1999). Thus, not only has position of many species in the water column (Zaret behavior (and other traits) been selected to reduce and Suffern 1976, Bowers and Vanderploeg 1982, predation risk, but behavior can be modified de- Ohman et al. 1983, Hayes 2003). Predation pro- pending on short-term changes in predation risk. vides a strong selective force on behavior, and Invertebrate predators are recognized as playing many species respond to environmental cues, mi- a critical role in the structure of aquatic food webs, grate according to diel cycles, or respond to including those of the Great Lakes food webs changes in light level, in a manner that reduces spa- (Bowers and Vanderploeg 1982, Branstrator and tial overlap with visual predators. More recently, re- Lehman 1991, Johannsson et al. 1994, Schulz and searchers have found that species can perceive the Yurista 1998). To fully understand the effects of in- presence of predators, often via chemical cues, and vertebrate predators it may be important to deter- change the degree to which they migrate in an mine if they affect zooplankton prey phenotype adaptive manner to increase fitness (Dodson 1988, such as vertical position. If zooplankton prey re- spond to invertebrate predators by modifying their behavior and other phenotypic traits to reduce pre- *Corresponding author. E-mail: [email protected] dation risk, this could affect prey consumption rates

144 Behavioral response of Daphnia to Mysis 145

of resources and vulnerability to predators (i.e., to Arbor, Michigan, and cultured separately in 4-L the predator inducing the change and to other vessels in aged Lake Michigan water under experi- predators). By inducing change in zooplankton prey mental temperatures (20°C). Food conditions con- phenotype, a predator can therefore affect the fit- sisted of 6 mg/L of Nanochloropsis limnetica (SAG ness of both the responding prey, and species with 18.99, University of Gottingen, Sag, Germany), and which the prey interacts. Such interactions are lighting was maintained at a 14 h light:10 h dark termed trait-mediated interactions (Abrams et al. regime. Mysis were also collected in Lake Michigan 1996) in order to differentiate them from density- near Muskegon, at sites approximately 75–100-m mediated interactions that occur through density ef- total depth, using a 1-m diameter net with 1,000-µm fects (i.e., predation). Trait-mediated interactions mesh made after sunset. Mysis were maintained in can contribute strongly to the net effect of a preda- the laboratory in a 30-L aquarium at 4°C. tor (Turner and Mittlebach 1990, Huang and Sih In general, the experiments were designed to 1991, Peacor and Werner 2001, Peacor and Werner evaluate the vertical movement (i.e., upward or 2004, Romare and Hansson 2003, reviewed in Dill downward) of D. mendotae in response to Mysis et al. 2003, and Werner and Peacor 2003). Further, kairomones in clear acrylic cylinders (e.g., Loose theory predicts that trait-mediated effects will affect and Dawidowicz 1994, Pijanowska 1997, and Van population dynamics and the stability of communi- Gool and Ringelberg 1998). D. mendotae (standard ties (reviewed in Bolker et al. 2003). body length range, 1.5–2.2 mm) were transferred In Lake Michigan, the invertebrate predator with a wide bore pipette into 60-cm tall, 26-mm di- Mysis relicta is a voracious predator of cladocerans, ameter, clear acrylic cylinders that were filled with including Daphnia spp. (Bowers and Vanderploeg experimental water (e.g., water with Mysis 1982). It has been hypothesized that this vulnerabil- kairomone and control water, see below) and sub- ity to Mysis contributes to a vertical diel migration merged vertically in a transparent, 40-L aquarium. by Daphnia spp. that minimizes spatial overlap, and The aquarium acted as a water bath, and uniform therefore vulnerability (Bowers and Vanderploeg water temperature was regulated at 20°C using an 1982, Spencer et al. 1999, Clarke and Bennet external chiller unit. The cylinders were illuminated 2003). Based on these patterns, we hypothesized by diffused light from directly above using three, that Mysis induce a change in Daphnia mendotae 50W halogen bulbs. Light intensity was kept con- behavior, in particular vertical migration patterns, stant over the duration of the experiment, with a and we tested this hypothesis with laboratory exper- photon flux density of 16.0 µEinst ˙ m–2 at the top iments. Our results indicate that D. mendotae of the cylinder, and 4.0 µEinst ˙ m–2 at the bottom. changed vertical position in experimental columns No resources were added to the cylinders, except in in response to Mysis kairomones (i.e., chemical Exp. 2 in which the effect of resource level was ex- cues produced by Mysis). Our results suggest that amined. D. mendotae potentially modify their vertical mi- To quantify the vertical position of D. mendotae, gration patterns in the field in response to Mysis the cylinders were demarcated in 10-cm intervals, presence and absence. Such induced changes in be- creating six vertical sections that were assigned the havior could contribute to the net effect of Mysis on values 1 to 6, with the lowest being closest to the D. mendotae density and also indirectly affect other bottom. D. mendotae were allowed to acclimate to species in Lake Michigan food web through trait- experimental conditions for 1 h before observations mediated interactions. were initiated. In Experiment 1, the vertical posi- tion of all D. mendotae in each cylinder was METHODS recorded every 20 min over a 4-h period (12 obser- General Experimental Approach vations total). Experiments 1–3 were similarly con- ducted, however over a 4.5-h period, with We conducted four laboratory experiments to in- observations recorded every 30 minutes (10 obser- vestigate the behavioral response of D. mendotae to vations total). Mysis using two isofemale lines of D. mendotae, la- beled clone-A and clone-B, originating from two females collected in Lake Michigan approximately Specific Experimental Treatments 8 km offshore of Muskegon, Michigan. D. mendo- Experiment 1 was conducted on 23 Jul 03 to tae clones were transported to the Great Lakes En- evaluate the behavioral response of clone-A and vironmental Research Laboratory (NOAA) in Ann clone-B D. mendotae to Mysis kairomones. The ex- 146 Peacor et al.

periment was carried out as a 2 × 2 factorial design Three additional experiments were conducted on with two clones and treatments with and without 5 Aug 03, 29 Aug 03, and 7 Nov 03, to examine if Mysis cue, thus four treatments total. In the treat- the presence of bacteria or debris associated with ment with Mysis cue, which we denote the “Mysis Mysis consumption of D. mendotae served as a re- water” treatment, Mysis were transferred into re- source that could contribute to the behavioral re- frigerated well water in 1-L containers (density, 10 sponses of D. mendotae to Mysis. Resource Mysis ˙ L–1) 5 days prior to the experiment, and fed on conditions can affect the magnitude of predator-in- the first and last day of this period with both clone-A duced changes in prey behavior (Werner and Anholt and clone-B D. mendotae (approximately three D. 1993, Grand and Dill 1999, Weber 2001, Weetman mendotae ˙ Mysis–1). In the treatment without Mysis and Atkinson 2002). These experiments also served cue, the control treatment was well water main- to test the robustness of the results of Experiment 1 tained under similar condition with no Mysis or D. and to shed insight into the origin of the kairomone mendotae added. Mysis were held at a lower tem- to which D. mendotae respond. The general meth- perature (4°C) than used in the experiment (20°C) ods were similar to that of Experiment 1, and some and would die if quickly transferred to this warm minor deviations are listed in Table 1. temperature. In order to allow the temperature of Experiment 2 consisted of a 2 × 2 × 3 factorial the experimental water to warm to experimental design with two clones (A and B), two predator cue conditions, we therefore removed Mysis from the levels (control water and Mysis water), and three re- water, and transferred the remaining water, denoted source levels. We used 0, 0.5, and 3 mg/L of N. lim- “Mysis water,” and the control water to a 20°C in- netica to represent low, intermediate, and high cubator the evening before the experiment. Two resource level treatments. Two replicate cylinders hours prior to the experiment, the Mysis water and were designated to each treatment (24 cylinders control water were passed through a 64-µm filter to total), and each cylinder received either eight clone- remove large debris and then transferred into re- A or eight clone-B D. mendotae. spective cylinders. We used a randomized factorial Experiment 3 consisted of three treatments: con- block design; each of the four treatments was repre- trol, Mysis water, and a third treatment denoted sented in each of five spatial blocks, with each con- “ultra-filtered Mysis water.” As in other experi- secutive four cylinders representing a block (20 ments, all experimental water was passed through a cylinders aligned side by side in total). Each cylin- 64-µm filter prior to the experiment; however, the der received either five clone-A or six clone-B D. ultra-filtered Mysis water was passed through an mendotae (we used different numbers depending on additional 0.46-µm filter. Past research has found availability). that D. mendotae can selectively filter particles

TABLE 1. Summary of methods that varied in Experiments 1, 2, 3, and 4. Procedures Experiment 1 Experiment 2 Experiment 3 Experiment 4 Treatments Mysis water Mysis water and three Mysis water and ultra- Mysis water and particulate- (in addition to resource levels filtered Mysis water free Mysis water control)

Clones A and B A and B B B

Mysis water 5 4 4 2 production period (d)

Mysis diet 3 Daphnia · 4 Daphnia · 4 Daphnia · 4 Daphnia · during Mysis–1 on Mysis–1 on Mysis–1 on Mysis–1 on incubation days 1 and 5 days 1 and 4 days 1, 2, 3, and 4 day 1

Experimental 17-h period in 14-h period in 1-h period using 13.5-h period water thermal incubator at 20°C incubator at 20°C warm-water bath in incubator at 20°C acclimation* * Time period and method used to warm water originally at 4°C to 20°C used in experiment. Behavioral response of Daphnia to Mysis 147

down to approximately 0.6 microns (Gophen and behavioral response of each treatment was evalu- Geller 1984); therefore, ultra-filtration should elim- ated based on two parameters: 1) the average inter- inate all potential food resources for D. mendotae. val occupied by D. mendotae in a cylinder (this Four replicate cylinders were designated to each metric is nearly equivalent to the average height of treatment, and each cylinder received six clone-B the D. mendotae), and 2) the fraction of the D. men- D. mendotae. Note that in Experiment 3, Mysis and dotae in the bottom half of the cylinder. control water were warmed with a warm water bath In each experiment, a fraction of the D. mendo- (Table 1), and therefore warmed more quickly than tae were not included in the observations because in the other experiments in which the water was they either died or adhered to air bubbles on the placed in the incubator to slowly warm over night. side of the cylinder during the observation period. This difference in protocol was due to logistical We quantified this loss using the metric, percent constraints, but we believe it unlikely to affect the loss (l), which was calculated as l = 100 ˙ (1 – Nf qualitative nature of the results. /Ni), where Ni was the initial number of individuals Experiment 4 consisted of three treatments: con- counted in each cylinder, and Nf was the final num- trol water, Mysis water, and a third treatment de- ber of individuals counted in each cylinder. The noted “particulate-free Mysis water” treatment. This percent loss was 1.71 ± 5.26 (mean ± stdev), 4.6 ± latter treatment was designed to include kairomones 8.2, 17 ± 13, and 9.8 ± 11.5, in Experiments 1, 2, 3, but no particulate matter. The Mysis water treatment and 4, respectively. Percent loss was analyzed in was created using protocols similar to those de- two ways to determine if it created confounding ef- scribed in Experiment 1. To create the particulate- fects on experimental parameters. First, we ana- free Mysis water treatment, 80 D. mendotae (4 D. lyzed if there were treatment effects on percent loss mendotae ˙ Mysis–1 ) were added to two 1-L using analysis of variance (ANOVA). Second, the beakers 3 days prior to the experiment, and Mysis two behavioral responses (above) were analyzed were added. After 2 hours, the Mysis from both using analysis of covariance (ANCOVA), with per- beakers were transferred into new beakers with well cent loss designated as a covariate to experimental water in order to separate the Mysis from large par- parameters. Both tests revealed no significant ef- ticulate matter associated with consuming Daphnia. fects caused by percent loss, and therefore, factors After an additional 24 hours (sufficient time for any that contributed to loss of D. mendotae did not con- consumed daphnia to have passed through the found the results on behavior. Mysis), the Mysis were again transferred into final beakers with well water which served as the partic- Statistical Analysis ulate-free Mysis water treatment. This procedure ensured that the particulate-free Mysis water con- We analyzed untransformed data using a full fac- tained Mysis, but contained much less (almost no) torial ANOVA following the generalized linear particulate matter or bacteria associated with cap- model (GLM) procedures (SAS Institute 1990). ture and defecation of D. mendotae relative to the Data from Experiments 1 and 2 were blocked by regular Mysis water treatment. Indeed, fecal mater- clonal group and simple main effects of Mysis were ial was visually apparent at the bottom of the Mysis evaluated within each block. Our null hypothesis water treatment beakers, but not in the particulate- for all experiments was that there would be no sig- free Mysis water beakers. Mysis were removed from nificant difference (α = 0.05) among treatments. both the regular Mysis water treatment and the par- When significant differences among treatments ticulate-free Mysis water 13.5 h prior to the experi- were detected, a Tukey pair-wise comparison was ment (approximately 2 days after Mysis were used to separate the means. Based on Lilliefors isolated from particulate matter in the manner de- tests, the data did not deviate from normality. scribe above). Seven cylinders were designated for each treatment (21 cylinders total) and six clone-B RESULTS D. mendotae were transferred into each cylinder. The experiments indicated that D. mendotae col- lected from Lake Michigan changed their position Quantification of Behavioral Response in the experimental cylinders in response to Mysis The number of D. mendotae counted in each in- water, a result consistent with D. mendotae re- terval in each cylinder was averaged over the exper- sponse to Mysis kairomones. In Experiment 1, both imental observations. In each experiment, the clones placed in Mysis water occupied lower posi- 148 Peacor et al.

TABLE 2. Factorial ANOVA on treatment effects on average interval position and fraction of Daphnia in the bottom half of the cylinder in Experiment 1. Simple main effects in Mysis treat- ments were also determined within each clone. Asterisks denote significant effects (P < 0.05). Interval Fraction in position bottom half Sources df FPFP Mysis 1, 16 33.06 < 0.01* 19.43 < 0.01* Clone 1, 16 30.81 < 0.01* 22.47 < 0.01* Mysis × Clone 1, 16 5.56 0.031* 4.5 0.05* Mysis (clone A) 1, 16 6.46 0.02* 2.44 0.11 Mysis (clone B) 1, 16 24.04 < 0.01* 22.96 <0.01*

tions in the cylinders (Table 2, Fig. 1), on average, than in control water. Figure 1a illustrates the per- centage of D. mendotae observed at each of the six demarcated intervals. In the Mysis water, D. mendo- tae were much more unevenly distributed than in the control treatment, with a much higher percent- age (77% higher) of D. mendotae occupying the lowest 10-cm section. The difference in distribution is further shown in Figures 1b and 1c, which illus- trate the effect of treatment on the two metrics we used to quantify the vertical distribution. In Mysis water, the average interval occupied by D. mendo- tae, and the fraction of D. mendotae in the bottom FIG. 1. Vertical distribution of clone-A and half of the cylinder, were significantly lower and clone-B D. mendotae in control water (C) and higher than in the control, respectively. Mysis water (M) in Experiment 1. (a) Fraction of Experiment 2 did not reveal effects of resource clone-B Daphnia that occupied each of the 6 10 level on D. mendotae behavior (Fig. 2a, Table 3). cm sections, where 1 was the lowest section (re- Unfortunately, the difference in D. mendotae posi- sults of clone-A similar). (b) Average section occu- tion in the cylinders within a treatment showed pied by Daphnia in a cylinder, where lower higher variation in this experiment for unknown numbers represent Daphnia lower in the cylinder. reasons, and therefore, any potential effects were Error bars are the standard error (SE) between less likely to be evident. However, there clearly was cylinders within a treatment. (c) Fraction of the not a very strong effect of the resource. Because Clone A (A) and Clone B (B) Daphnia in the bot- there was no effect of resource, we collapsed the tom half (mean ± SE) of the cylinder (i.e., Daph- design and combined all different resource levels as nia that occupied one of the bottom 3 of 6 one treatment. Examined in this way, the main ef- segments). fect of the Mysis on both measures of D. mendotae distribution was not significant (Table 3). However, there was a significant clone by Mysis interaction. mendotae to move higher (Fig. 3) than in the con- This interaction arose because clone-A was signifi- trol, however, this response was much weaker or cantly higher in the Mysis water treatment (Figs. 2b non-existent in the ultra-filtered Mysis water treat- and 2c, Table 3) than in the control, but there was a ment. The average interval occupied by D. mendo- non-significant effect of Mysis in the opposite di- tae in Mysis water was significantly greater (F2,3 = rection on clone-B (Figs. 2b and 2c, Table 3). 9.75; P = 0.01) than in control and ultra-filtered In Experiment 3, Mysis water caused clone-B D. Mysis water. In addition, the fraction of D. mendo- Behavioral response of Daphnia to Mysis 149

TABLE 3. Factorial ANOVA on treatment effects on average interval position and fraction of Daphnia in the bottom half of the cylinder in Experiment 2. Resource effects on both parame- ters were not significant (F = 1.08, P = 0.37; F = 0.72, P = 0.51, respectively), so they were removed from the analysis. Simple main effects in Mysis treatments were also determined within each clone. Asterisks denote significant effects (P < 0.05). Interval Fraction in position bottom half Sources df FPFP Mysis 1, 20 2.50 0.13 2.31 0.14 Clone 1, 20 14.58 < 0.01* 16.38 < 0.01* Mysis × Clone 1, 20 8.30 0.01* 7.28 0.01* Mysis (clone A) 1, 20 9.96 < 0.01* 8.90 < 0.01* Mysis (clone B) 1, 20 0.84 0.37 0.69 0.42 FIG. 2. Vertical distribution of clone-A and clone-B Daphnia in Experiment 2. (a) Vertical dis- tribution of Daphnia (response of clone-A and (Fig. 4a). As a result, D. mendotae in both Mysis clone-B averaged) in Mysis water (M) and control water treatments occupied a significantly higher av- water (C) at the low (Low R), intermediate (In- erage interval (F2,6 = 11.62; P < 0.01) than those in term. R) and high (High R) resource level. (b) Ef- control water. D. mendotae in both Mysis water fect of Mysis water on the average interval treatments also had a significantly lower (F2,6 = position of clone-A and clone-B. Squares and cir- 9.88; P < 0.01) fraction of individuals in the bottom cles for clone-A and clone-B, respectively, and half of the cylinder than those in the control water. empty and filled symbols for control (C) and Importantly, the particulate-free Mysis water treat- Mysis (M) water, respectively. (c) The effect of ment, which was designed to greatly reduce particu- Mysis water on the fraction of clone-A and clone- late matter, had a strong effect on behavior and was B Daphnia in the bottom half of the cylinder. not significantly different from the regular Mysis Error bars represent SE. water treatment (Fig. 4).

tae in the bottom half of the cylinder in Mysis water was significantly lower than in control and ultra-fil- tered Mysis water (F2,3 = 7.59; P = 0.01). In con- trast, while there was a small trend for the ultra-filtered Mysis water in the same direction as the Mysis water, neither the average interval occu- pied, nor the fraction in the bottom half, were sig- nificantly different than the control (P > 0.4 for both responses). In Experiment 4, Mysis water also caused clone- B D. mendotae to move higher into the water cylin- der (Fig. 4). There was a high percentage of D. FIG. 3. Vertical position of clone-B Daphnia in mendotae in the bottom 10-cm interval in the con- control water (C), Mysis water (M), and Mysis trol treatment, but this pattern was reversed in the water filtered with a fine filter (F-M). (a) Average regular Mysis water and particulate-free Mysis interval position and (b) fraction of Daphnia in treatments, in which the proportion of D. mendotae the bottom half of the cylinder. Bars are means ± was much higher in the top two 10-cm sections SE’s of four replicates per treatment. 150 Peacor et al.

man and Atkinson 2002). However, in Experiment 2, large changes in resource level had no effect on the behavior of the D. mendotae, nor on the re- sponse of the D. mendotae to the Mysis water. Fur- ther, in Experiment 4, the response of the D. mendotae without this particulate matter was strong, and response variables were not signifi- cantly different between the treatment with, and without, the particulate matter. This suggests that rather then responding to particulate matter, D. mendotae more likely responded to chemical cues (i.e., kairomones) emitted by Mysis. Experiment 3 suggests that these chemicals are large enough to be removed by a 0.46-µm filter, as Mysis water that had been filtered through such a fine filter had little or no effect on Mysis vertical position. This may offer a clue into the identity and properties of the kairomones, which are not well-known or under- stood in invertebrate predator-prey systems (Lass and Spaak 2003). Note that Experiment 4 suggests that the cue is not associated with alarm signals from conspecifics (Pijanowska 1997) because D. mendotae responded to water that never contained conspecifics, but rather only contained Mysis previ- ously fed D. mendotae. While this is the first laboratory study we are aware of that demonstrates a behavioral response of a Great Lakes zooplankton species to a predator, several field studies have indicated shifts in zoo- plankton vertical position associated with changes FIG. 4. Vertical position of clone-B Daphnia in in invertebrate predator density. For example, control water (C), Mysis water (M), and “particu- Benoit et al. (2002) used seasonal time series to ex- late-free Mysis water” (PF-M) in which particu- amine the vertical distribution of zooplankton be- late matter from consumption and defecation of fore and after the invasion of Cercopagis pengoi in the Daphnia they were fed before the experiment Lake Ontario. They found that juvenile copepods was greatly reduced (see text). Bars are means ± occupied deeper waters after Cercopagis invaded in SE’s of seven replicates per treatment. Figures (a), 1999. They hypothesized that this was due to either (b) and (c) as in Figure 1. predator avoidance behavior, or localized depletion. Further, Lehman and Caceres (1993) showed that the cladocerans Bosmina longirostris, D. mendotae, DISCUSSION and D. pulicaria shifted their vertical distributions The principal finding of this study is that Mysis following the invasion of Bythotrephes longimanus relicta from Lake Michigan induced a change in the in Lake Michigan offshore from Grand Haven, behavior of a prey species, Daphnia mendotae. Michigan. Similarly, Johannsson (O. Johannsson, Clearly this response must be olfactory, because vi- Department of Fisheries and Oceans Canada, 3 Mar sual or tactual cues were absent in the experimental 04) has found that the average depth of Bosmina cylinders. Experiments 2, 3, and 4 lend insight into and Dapnia in Lake Erie is inversely related to the origin of the Mysis cue. It is possible that partic- Bythotrephes densities. These studies are suggestive ulate matter and bacteria associated in the Mysis of zooplankton response to invertebrate kairomones water served as a resource that could confound the in the Great Lakes. Note, however, that other mech- response of the D. mendotae to kairomones, given anisms could lead to these patterns. For example, that induced traits of Daphnia have been shown to zooplankton may be responding to other factors be affected by resource level (Weber 2001, Weet- correlated with increases in predator density, or the Behavioral response of Daphnia to Mysis 151

changes in distribution in predator presence could tive and reduces predation risk. In the case of the be due to density effects (i.e., differential predation Mysis relicta-Daphnia mendotae interaction in on clones with higher spatial overlap with the Lake Michigan, we would predict that it would be predator). adaptive for D. mendotae to migrate up in response Surprisingly, the response of D. mendotae from to cues of Mysis presence. This is because, at night, the same clone was qualitatively different in differ- Mysis migrate up from the hypolimnion and lower ent experiments. For example, in Mysis water (i.e., metalimnion, into the upper metalimnion and lower water that previously contained Mysis), clone-B oc- epilimnion. D. mendotae have been shown to si- cupied a lower position in Experiment 1 than in the multaneously migrate away from the regions inhab- control, did not respond significantly in Experiment ited by Mysis to higher regions in the epilimnion 2, and occupied a higher position in Experiments 3 (Bowers and Vanderploeg 1982). Bowers and Van- and 4. While we are aware of studies that show dif- derploeg argue that this coordinated movement up- ferent predators can induce migration in different ward by Mysis and D. mendotae represents an directions (Ohman et al. 1983, Hayes 2003) we are evolutionary refugium of Daphnia from Mysis that unaware of experiments that show the same preda- minimize contact between the two species. Similar tor species (and size class) can induce such a differ- simultaneously migrations between Daphnia and ent response in the same prey. This ambiguity in the Mysis have also been observed in Lake Pend response is indicative of the complexity of the re- Orielle, Idaho (Clarke and Bennet 2003) and Flat- sponse of Daphnia to different environmental cues head Lake, Montana (Spencer et al. 1999). and/or internal states such as hunger level (Weet- Our study suggests that D. mendotae not only man and Atkinson 2002). While we attempted to may have evolved to move upward to avoid Mysis hold all conditions constant (e.g., Daphnia were at certain times of the day, but that they can also cultured in a similar manner and fed the day before perceive Mysis presence, and thus modify the de- each experiment), differences in the resources and gree to which they migrate in an adaptive manner in light levels in the culturing environment of Daph- response to variation in risk from Mysis. This is an nia, in their handling during transfer to the experi- important distinction, because if D. mendotae mod- ment, or in the preparation of Mysis cue, may have ify the degree to which they migrate as a function caused the Daphnia to respond differently (Weber of Mysis density, then the immediate effects of 2001). While more research is required to under- Mysis on D. mendotae extend beyond the effect stand the complexity of the response, the ambiguity they have on D. mendotae density. Induced changes does not change our principal result that Lake in behavior could affect D. mendotae population Michigan D. mendotae can perceive and respond to growth rates through changes in temperature the presence of Mysis kairomones. (Lueke et al. 1990, Loose and Dawidowicz 1994), Note that the data (Figs. 1 and 2) suggest a dif- resource availability (Gielbelhausen and Lampert ference in clonal response to a predator cue, which 2001) and risk to other predators (Liljendahl-Nur- has been reported by other investigators for Daph- minen et al. 2003). For example, Loose and Dawid- nia (Dodson 1988, Weber and Declerck 1997, owicz (1994) have shown that in the presence of Weber and Van Noordwijk, 2002, Michels and De fish kairomone, Daphnia magna will migrate to Meester 2004). We hesitate to make this claim here colder temperatures that cause a 62% reduction in as the experiment was not designed to account for intrinsic growth rate, a value that most likely will subtle differences in their culturing environment. A contribute strongly to the net effect of the predator design which raises multiple groups of each clone (Peacor and Werner 2004). Similarly, Winder et al. using a factorial design would be needed to avoid (2003) found that the vertical migration exhibited pseudoreplication and thereby isolate clonal differ- by Daphnia galeata was associated with a large ences from those that arise from subtle differences thermal cost to growth and development rates in a in the culturing environment (Hurlbert 1984). high mountain lake. Further, such behavioral modi- Predator-induced changes in vertical position and fications can lead to trait-mediated indirect interac- diel migration timing of Daphnia and other zoo- tions, in which predators indirectly affect species plankton in both freshwater and marine systems through their effect on prey traits (such as behav- have been well documented (Frost and Bollens ior). Such indirect interactions are becoming in- 1992, Fortier et al. 2001, Irigoien et al. 2004, re- creasingly recognized as playing a significant role viewed in Hayes 2003, and Lass and Spaak 2003). in the net effect of predators (Turner and Mittlebach Typically, movement is in a direction that is adap- 1990, Huang and Sih 1991, Peacor and Werner 152 Peacor et al.

2001, Romare and Hansson 2003, reviewed in Dill that occur on the scale they do in the Lake Michi- et al. 2003, and Werner and Peacor 2003), and are gan. However, such laboratory experiments can predicted to affect population and community struc- contribute to our knowledge of the system and be ture (Abrams 1984, reviewed in Bolker et al. 2003). an integral part of a larger comprehensive study One factor that could influence an adaptive re- (Werner 1998). But this is only a first step, and sponse of Lake Michigan D. mendotae to Mysis is more studies are needed to gain further understand- the recent invasion of Bythotrephes (Lehman 1987). ing and predictive capabilities. First, we must estab- Bythotrephes is a visual predator, and therefore it lish if the signals of Mysis are strong enough in the may be adaptive for D. mendotae to respond to field to elicit strong responses. In our laboratory ex- Bythotrephes by swimming lower in the water col- periment, the concentration of the kairomone was umn. Indeed, Lehman and Cáceres (1993) found likely much higher than that found in the field. One that D. mendotae was lower in the water column potential procedure is to use water from areas in- after Bythotrephes invasion. Whether the presence habited and not inhabited by Mysis in similar exper- of this new species has affected the response to iments as those performed here. It would also be Mysis would depend on several factors, including helpful to examine the response to Mysis in more how fast zooplankton evolve to a changing predator realistic conditions, across a range of light levels assemblage, and whether zooplankton such as D. and temperature levels (Mysis are typically found at mendotae can differentiate cues of different preda- much lower temperatures and light levels than used tors. There are several studies that indicate that in our experiment for convenience). Further, it will Daphnia can differentiate between fish and inverte- be important to combine survey and experimental brate predators (e.g., Dodson 1988, Brett 1992, Weber and Van Noordwijk 2002, Dzialowski et al. studies to establish (1) how potential changes in 2003), and different invertebrate predators (e.g., vertical distribution will affect Daphnia growth rate Chaoborus and Notonecta, Dodson 1988, see also in the field, and (2) how factors such as resource Nesbitt et al. 1996). We have recently conducted level and temperature affect the degree of migra- pilot studies (Pangle, unpublished data) that also tion, and in turn affect the consequences to growth suggest that D. mendotae can differentiate between rate. We believe that a combined field, laboratory, Mysis and Bythotrephes cues, however more work and modeling effort is required to understand the is needed. Laboratory experiments could further consequences of predator-induced behavioral re- shed insights into the interactive effects of cues sponses of zooplankton prey in the Great Lakes. from multiple predators, including fish (e.g., will cues fish overshadow those from Mysis?). In addi- CONCLUSION tion, laboratory experiments could be used to eluci- To gain an increased understanding of the poten- date potential interactive effects of zooplankton tial effects of invertebrate predators such as Mysis responses to predator density and conspecific den- on their prey in the Great Lakes, it is important to sity, or even behavioral responses to the cues aris- ing by other conspecific individuals responding to understand the potential effects they exert by induc- predation risk (Relyea 2002, Ralph Tollrian, per- ing changes in prey phenotype, including behavior. sonal communication, reviewed in Elgar 1989, Ecologists are becoming increasingly aware that Lima 1990, Roberts 1996, Peacor 2003). How such responses can contribute to the net conse- Great Lakes zooplankton will respond to different quences of species interaction in many systems, in- predators, competitors, resources, and interactive cluding freshwater. That is, not only are such effects between these factors, are complex and in- responses interesting from an evolutionary and be- teresting questions that deserve further attention. havioral standpoint, but they may play a critical This study indicates that Mysis induce changes in role in the structure and dynamics of food webs. behavior of Lake Michigan Daphnia. In the field, However, phenotypic responses to predators and such a response could affect Daphnia population potential consequent trait-mediated interactions growth rate (e.g., through changes in temperature, have not received attention in the study of Great resource availability, or predation from other preda- Lakes ecosystems. This may be particularly impor- tors), and indirectly affect resources, competitors, tant in describing the effects of recent invasions of and predators of Daphnia. Clearly it is not possible predatory cladocerans (i.e., Bythotrephes longi- to make direct inferences from laboratory experi- manus and Cercopagis pengoi) and of future ments performed in small cylinders to interactions invaders. Behavioral response of Daphnia to Mysis 153

ACKNOWLEDGMENTS Smith, V.H. 2003. Predator-induced phenotypic plas- We thank J. Cavoletto, R. Pichlova, and K. ticity in the exotic cladoceran Daphnia lumholtzi. Freshw. Biol. 48:1593–1602. Rosenkrands for assistance in collecting zooplank- Elgar, M.A. 1989. Predator vigilance and group size in ton and performing experiments, and D. Hondorp mammals and birds: a critical review of the empirical for providing Mysis. We thank C. Darnell, O. Jo- evidence. Biological Reviews of the Cambridge Philo- hannsson, J. Makarewicz, J. Williams, and two sophical Society 64:13–33. anonymous reviewers for their comments on this Fortier, M., Fortier, L., Hattori, H., Saito, H., and manuscript. We wish to acknowledge support for Legendre, L. 2001. Visual predators and the diel verti- this research from the Fishery Research Program of cal migration of copepods under Artic sea ice during the Great Lakes Fishery Commission, the Michigan the midnight sun. J. Plankton Res. 23:1263–1278. Agricultural Experimental Station, and the Cooper- Frost, B.W., and Bollens, S.M. 1992. Variability of diel ative Institute for Limnology and Ecosystems Re- vertical migration in the marine planktonic copepod search. This is GLERL contribution 1353. Pseudocalanus newmani in relation to its predators. Can. J. Fish. Aquat. Sci. 49:1137–1141. Gielbelhausen, B., and Lampert, W. 2001. Temperature REFERENCES reaction norms of Daphnia magna: the effect of food Abrams, P.A. 1984. Foraging interactions and interac- concentration. Freshwater Biol. 46:281–289. tions in food webs. Am. Nat. 124:80–96. Gophen, M., and Geller, W. 1984. Filter mesh size and ——— , Menge, B.A., Mittelbach, G.G., Spiller, D., and food particle uptake by Daphnia. Oecologia Yodzis, P. 1996. The role of indirect effects in food 64:408–412. webs. In Food webs: dynamics and structure, eds. G. Grand, T.C., and Dill, L.M. 1999. The effect of group Polis and K. Winemiller, pp. 371–395. New York, size on the foraging behaviour of juvenile coho NY: Chapman and Hall. salmon: reduction of predation risk or increased com- Benoit, H.P., Johannsson, O.E., Warner, D.M., Sprules, petition? Anim. Behav. 58:443–451. W.G., and Rudstram, L.G. 2002. Assessing the impact Hayes, G.C. 2003. A review of the adaptive significance of a recent predator invader: The population dynam- and ecosystem consequences of zooplankton diel ver- ics, vertical distribution, and potential prey of Cerco- tical migration. Hydrobiol. 503:163–170. pagis pengoi in Lake Ontario. Limnol. Oceanogr. Huang, C.F., and Sih, A. 1991. Experimental studies on 47:626–635. direct and indirect interactions in a three trophic-level Bolker, B., Holyoak, M., Krivan, V., Rowe, L., and stream system. Oecologia 85:530–536. Schmitz, O. 2003. Connecting theoretical and empiri- Hurlbert, S.H. 1984. Pseudoreplication and the design of cal studies of trait-mediated interactions. Ecology ecological field experiments. Ecol. Monogr. 84:1101–1114. 54:187–211. Bowers, J.A., and Vanderploeg, H.A. 1982. In situ Irigoien, X., Conway, D.V.P., and Harris, R.P. 2004. predatory behavior of Mysis relicta in Lake Michigan. Flexible behaviour of zoo- Hydrobiol. 93:121–131. plankton in the Irish Sea. Mar. Ecol. Prog. Ser. Branstrator, D.K., and Lehman, J.T. 1991. Invertebrate 267:85–97. predation in Lake Michigan—regulation of Bosmina longirostris by Leptodora kindtii. Limnol. Oceanogr. Johannsson, O.E., Rudstam, L.G., and Lasenby, D.C. 36:483–495. 1994. Mysis relicta—assessment of metalimnetic Brett, M.T. 1992. Chaoborus and fish-mediated influ- feeding and implications for competition with fish in ences on Daphnia longispina population structure, Lakes Ontario and Michigan. Can. J. Fish. Aquat. Sci. dynamics, and life-history strategies. Oecologia 89: 51:2591–2602. 69–77. Lass, S., and Spaak, P. 2003. Chemically induced anti- Clarke, L.R., and Bennet, D.H. 2003. Seasonal zooplank- predator defenses in plankton: a review. Hydrobiol. ton abundance and size flucuations across spatial 491:221–239. scales in Lake Pend Orielle, Idaho. J. Freshwater Lehman, J.T. 1987. Palearctic predator invades North Ecol. 18:277–290. American Great Lakes. Oecologia 74:478–480. Dill, L.M., Heithaus, M.R., and Walters, C.J. 2003. ———, and Caceres, C.E. 1993. Food-web responses to Behaviorally mediated indirect interactions in marine species invasion by a predatory invertebrate: communities and their conservation implications. Bythotrephes in Lake Michigan. Limnol. Oceanogr. Ecology 84:1151–1157. 38:879–891. Dodson, S. 1988. The ecological role of chemical stimuli Liljendahl-Nurminen, A., Horppila, J., Malinen, T., Elo- for the zooplankton-predator-avoidance behavior in ranta, P., Vinni, M., Alajarvi, E., and Valtonen, S. Daphnia. Limnol. Oceanogr. 33:1431–1439. 2003. The supremacy of invertebrate predators over Dzialowski, A.R., Lennon, J.T., O’Brien, W.J., and fish—factors behind the unconventional seasonal 154 Peacor et al.

dynamics of cladocerans in Lake Hiidenvesi. Arch. Schulz, K.L., and Yurista, P.M. 1998. Implications of an Hydrobiol. 158:75–96. invertebrate predator’s (Bythotrephes cederstroemi) Lima, S.L. 1990. The influence of models on the inter- atypical effects on a pelagic zooplankton community. pretation of vigilance. In Interpretation and Explana- Hydrobiol. 380:179–193. tion in the Study of Behaviour: Vol. 2. Expla- Spencer, C.N., Potter, D.S., Bukantis, R.T., and Stan- nation, Evolution and Adaptation, ed. M. Bekoff and ford, J.A. 1999. Impact of predation by Mysis relicta D. Jamieson, pp. 246–267. Westview Press. on zooplankton in Flathead Lake, Montana, USA. J. Loose, C.J., and Dawidowicz, P. 1994. Trade-offs in diel Plankton Res. 21:51–64. vertical migration by zooplankton: the costs of preda- Tollrian, R., and Harvell, C.D. 1999. The ecology and tor avoidance. Ecology 75:2255–2263. evolution of inducible defenses. Princeton, N. J: Lueke, C., Vanni, M.J., Magnuson, J.J., Kitchell, J.K., Princeton University Press. and Jacobson, P.T. 1990. Seasonal regulation of Turner, A.M., and Mittelbach, G.G. 1990. Predator Daphnia populations by planktivorous fish: implica- avoidance and community structure: interactions tions for the spring clear-water phase. Limnol. among piscivores, planktivores, and plankton. Ecol- Oceanogr. 35:1718–1733. ogy 71:2241–2254. Michels, E., and De Meester, L. 2004. Inter-clonal varia- Van Gool, E., and Ringelberg, J. 1998. Light-induced tion in phototactic behavior and key life-history traits migration behaviour of Daphnia modified by food in a metapopulation of the cyclical parthenogen and predator kairomones. Anim. Behav. 56:741–747. Daphnia ambigua: the effect of fish kairomones. Weber, A. 2001. Interactions between predator Hydrobiol. 522:221–233. kairomone and food level complicate the ecological Nesbitt, L.M., Riessen, H.P., and Ramcharan, C.W. interpretation of Daphnia laboratory results. J. Plank- 1996. Opposing predation pressures and induced ver- ton Res. 23:41–46. tical migration responses in Daphnia. Limnol. ——— , and Declerck, S. 1997. Phenotypic plasticity of Oceanogr. 41:1306–1311. Daphnia life history traits in response to predator Ohman, M.D., Frost, B.W., and Cohen, E.B. 1983. kairomones: genetic variability and evolutionary Reverse diel vertical migration: an escape from inver- potential. Hydrobiol. 360:89–99. tebrate predators. Science 220:1404–1407. , and Van Noordwijk, A. 2002. Swimming behav- Peacor, S.D. 2003. Phenotypic modifications to conspe- ——— ior of Daphnia clones: differentiation through preda- cific density: a new mechanism arising from predation tor infochemicals. J. Plankton Res. 24:1335–1348. risk assessment. Oikos 100:409–415. Weetman, D., and Atkinson, D. 2002. Antipredator reac- , and Werner, E.E. 2001. Effects of predators on ——— ecological communities: the role of adaptive prey tion norms for life history traits in : behavior. Proc. Natl. Acad. Sci. U. S. A. 98: dependence on temperature and food. Oikos 98: 3904–3908. 299–307. Werner, E.E. 1998. Ecological experiments and a ———, and Werner, E.E. 2004. How dependent are species-pair interaction strengths on other species in research program in community ecology. In Experi- the food web? Ecology 85:2754–2763. mental ecology: issues and perspectives, eds. W.J. Pijanowska, J. 1997. Alarm signals in Daphnia? Oecolo- Resetarits and J. Bernardo, pp. 3–26. Oxford: Oxford gia 112:12–16. University Press. Ramcharan, C.W., and Sprules, W.G. 1991. Predator- ———, and Anholt, B.R. 1993. Ecological consequences induced behavioral defense and its ecological conse- of the trade-off between growth and mortality-rates quences for two calanoid copepods. Oecologia mediated by foraging activity. Am. Nat. 142:242–272. 86:276–286. ———, and Peacor, S.D. 2003. A review of trait-medi- Relyea, R.A. 2002. Competitor-induced plasticity in tad- ated indirect interactions in ecological communities. poles: consequences, cues, and connections to preda- Ecology 84:1083–1100. tor-induced plasticity. Ecological Monographs Winder, M., Boersma, M., and Spaak, P. 2003. On the 72:523–540. cost of vertical migration: are feeding conditions Roberts, G. 1996. Why individual vigilance declines as really worse at greater depths? Freshwater Biol. group size increases. Animal Behaviour 51: 48:383–393. 1077–1086. Zaret, T.M., and Suffern, J.S. 1976. Vertical migration in Romare, P., and Hansson, L.A. 2003. A behavioral cas- zooplankton as a predator avoidance mechanism. Lim- cade: Top-predator induced behavioral shifts in plank- nol. Oceanogr. 21:804–813. tivorous fish and zooplankton. Limnol. Oceanogr. 48:1956–1964. Submitted: 5 August 2004 SAS Institute. 1990. SAS/STAT user’s guide, version 6, Accepted: 10 February 2005 4th edition. Cary, North Carolina: SAS Institute. Editorial handling: Joseph P. Makarewicz