Phytoplankton essential fatty acid and phosphorus content constraints on Daphnia somatic growth and reproduction

We performed 12 experiments where the herbivorous zooplankter Daphnia pulex was fed three different phytoplankton food types (cyanophytes, chlorophytes, or cryptophytes) at two phosphorus (P) deficiency levels (C : P < 400 or 600) and with two different fatty acid (FA) supplements. Phosphatidylcholine liposome amendments were used to manipulate P availability and/or deliver FA in order to simultaneously test the relative importance of phytoplankton P and FA limitation as constraints on Daphnia somatic growth rate and reproduction. We used multiple regression analyses to test the data set for six main effects and three interaction terms. The six main effects tested were phytoplankton taxa, FA supplementation, FA type (EPA or FA-mix), direct (P-D) and indirect (P-ID) phosphorus limitation, and P deficiency level. Food quality was most strongly affected by phytoplankton taxa followed by P-ID, FA supplementation, the interaction between phytoplankton taxa and P-ID, and P-D. Daphnia fed cryptophytes grew 0.18 day21 faster and had an additional 2.6 eggs individual21 than Daphnia fed cyanophytes. Indirect P limitation reduced Daphnia somatic growth rates by 0.16 day21 and egg production by 2.6 eggs individual21. Direct P limitation reduced Daphnia growth rates by 0.04 day21 and egg production by 1.2 eggs individual21. FA supplementation improved growth by 0.09 day21 and egg production by 2.2 eggs individual21. These results suggest that FA supplementation exerted stronger effects on Daphnia somatic growth rate and reproduction than did direct P limitation. Furthermore, these results suggest that phytoplankton taxa and indirect P limitation had the greatest effects on Daphnia growth and reproduction. Phytoplankton phosphorus content and fatty acid composition have been previously identified as factors that may play important roles in herbivorous zooplankton nutrition (Urabe and Watanabe 1992; Müller-Navarra 1995b; Sterner and Schulz 1998). The phosphorus limitation hypothesis is based on the observation that the ratio of carbon (C) to phosphorus (P) in herbivorous zooplankton is nearly constant (Sterner and Hessen 1994), while this ratio in lake seston can vary widely across and within systems. Empirical evidence suggests that for phosphorus rich zooplankton such as Daphnia (C : P < 93), the dietary phosphorus limitation threshold occurs in the range of seston C : P . 150–375 (Urabe et al. 1997; DeMott 1998; Brett et al. 2000). In Daphnia, a large fraction of the P pool is found in nucleic acids and phospholipids (Vrede et al. 1999). Vrede et al. (2002) demonstrated that the RNA : DNA ratio in Daphnia galeata was highly correlated with somatic growth rate and increased as the dietary C : P ratio was decreased down to a threshold of approximately 200. To maintain constant C : P ratios, daphnids are known to make physiological adjustments such as increasing their carbon excretion and respiration rates when food becomes P deficient (Darchambeau et al. 2003). The low food quality of P-limited algae may also be due to morphological changes that cause some phytoplankton to become less digestible when nutrient stressed (Lürling and Van Donk 1997; Van Donk et al. 1997). The fatty acid limitation hypothesis is based on the observation that phytoplankton food quality is sometimes strongly correlated with the availability of certain polyunsaturated fatty acids (PUFA) (Müller-Navarra 1995b; Müller-Navarra et al. 2000; Wacker and Von Elert 2001). Essential fatty acids (EFA) are necessary in cell membranes and also as precursors for molecules involved in immune responses (Vance and Vance 1985). Fatty acids with double bonds in the n3 and n6 positions of the fatty-acid molecule are considered to be essential because most animals lack the specific desaturases required to synthesize them de novo (Vance and Vance 1985). Controlled laboratory growth experiments also indicate that dietary v3 PUFA enrichment can have significant effects on Daphnia somatic growth rates and egg production (Von Elert 2002; Ravet et al. 2003; Becker and Boersma 2005). Food quality experiments that have used natural lake seston to compare phytoplankton P and PUFA limitation have reported mixed results. In one study it was found that Daphnia growth rates increased when supplementing lake seston from a mesotrophic system with both P and PUFA (Boersma et al. 2001). In another study, DeMott and Tessier (2002) observed only weak responses to both P and PUFA supplementation in six lakes intended to represent a natural gradient in resource quality. 1 Corresponding author ([email protected]). Acknowledgments We thank Thomas Lumley at the University of Washington’s Department of Biostatistics for suggesting the statistical design for the regression analyses used in this study. We also thank Dick Lee of the Skidaway Institute of Oceanography for his insights on the metabolic pathways and bioavailability of phospholipids. Stephanie Cartier, Dag Olaf Hessen, and two anonymous reviewers provided editorial comments on this manuscript. Anne Liston at the University of California in Davis assisted with the fatty acid analyses. This project was supported by the National Science Foundation (0075591) grant to M.T.B. Limnol. Oceanogr., 51(5), 2006, 2438–2452 E 2006, by the American Society of Limnology and Oceanography, Inc.