Influence of Calcium depletion on susceptibility of Daphnia magna to disease Roisin Donohoe, Clayton Cressler, William Nelson 3 Treatments: 1) 2.5 mg Ca/L 2) 5 mg Ca/L 3) 10 mg Ca/L 20 controls 40 exposed 20 controls 20 controls 40 exposed 40 exposed Measuring daphnia body size Department of Biology, Queen’s University Ca <1.5 mg/L Help me! Pasteuria spores • hostparasite system: Daphnia magna (daphnia) and Pasteuria ramosa (Pasteuria) • Pasteuria infections, passed from host to host via transmission spores, cause gigantism and castration in daphnia • prior research: looked at the effects of food and temperature on daphnia growth/ reproduction in conjunction with a Pasteuria infection • daphnia are dependent on calcium (used to rebuild carapace following molting) • in nature, daphnia are increasingly exposed to calciumdepleted environments (concentrations < 1.5 mg/L) due to previous displacement of calcium from soils via acidic deposition (acid rain) 3 • for this reason, there is interest in looking at the effect calcium deficiency has on the susceptibility of daphnia to infections like that caused by Pasteuria Introduction Methods • Day 0: all daphnia placed into assigned Ca treatment 4 • Day 10: daphnia in “exposed” conditions exposed to Pasteuria spores • Day 18: Pasteuria spores removed • Day 36: experiment terminated, all remaining exposed daphnia frozen pending further analysis (spore counting) • Daphnia were measured/neonates were counted every other day Hypotheses Inducing calcium deficiencies will have an effect on Pasteuriaexposed daphnia by: 1) increasing the overall growth and decreasing the number of neonates produced 2) increasing the infection success of Pasteuria (more infected daphnia, more spores) Further Reading 1) Cressler, C., Nelson, W.A., Day, T. and E. McCauley. Starvation reveals the cause of infectioninduced castration and gigantism, Proceedings of the Royal Society B 2014. 2) Ebert, D., Rainey, P., Embley, T.M. and D. Scholz. Development, life cycle, ultrastructure and phylogenetic position of Pasteuria ramosa Metchnikoff 1888: rediscovery of an obligate endoparasite of Daphnia magna straus, Proceedings of the Royal Society B 1996. 3) Smol, J.P. The power of the past: using sediments to track the effects of multiple stressors on lake ecosystems. Freshwater Biology 2010. 4) Tan, Q. and W. Wang. The regulation of calcium in Daphnia magna reared in different calcium environments, Limnology and Oceanography 2009. Total Growth (mg) Calcium Concentration (mg/L) Combined Effect of Calcium Concentration and Pasteuria Infection on Average Growth Figure 1: • Infected daphnia did reach a statistically larger size (0.42 ± 0.007 mg) than uninfected daphnia (0.36 ± 0.003 mg), without taking calcium into account (F(2,131)= 53.207, p=0.000) • addition of calcium concentration as a second independent variable showed that the combination of calcium concentration and infection did not significantly affect the total growth of the daphnia Results Calcium Concentration (mg/L) Figure 2: • infected daphnia did not have statistically fewer neonates over their lifetime (23.40± 5.418 neonates) compared to uninfected daphnia (26.65± 7.278), without the effects of calcium • addition of calcium concentration as a second independent variable saw no significant interactions between the calcium concentration/Infection success and total reproduction Total Reproduction Combined Effect of Calcium Concentration and Pasteuria Infection on Average Reproduction Figure 3: • significant difference in the number of spores found in daphnia living at different calcium concentrations (p=0.004); difference lies between daphnia found at 2.5 vs. 10 mg Ca/L • high degree of correlation between spore density and calcium concentration (R=0.676, R square=0.457) 250000 450000 650000 850000 1050000 1250000 1450000 1650000 1850000 0 2.5 5 7.5 10 12.5 Spore Density (per daphnia) Spore Densities at Various Calcium Concentrations Calcium Concentration (mg/L) Average Spore Density Conclusions inducing calcium deficiencies does affect the infection success of Pasteuria increases vulnerability of daphnia at low calcium, increases spore density at high calcium fewer daphnia were infected than expected based on previous experiments 1 likely due to later exposure to spores Pasteuria also appears to be calcium dependent lower numbers of spores at low calcium concentrations