Ecological Archives E095-235-A2

Jeffrey T. Wright, James E. Byers, Jayna L. DeVore, Erik E. Sotka. 2014. Engineering or food? mechanisms of facilitation by a habitat-forming invasive seaweed. Ecology 95:2699–2706. http://dx.doi.org/10.1890/14-0127.1

Appendix B. Detailed methods for the use of Gracilaria as a food resource by Gammarus.

Gammarus used in feeding experiments were collected from nearby sites and held in a shaded outdoor flow-through tank (~ 1000 L) for ~ 5–10 days prior to experiments. During this time Gammarus had access to unlimited live Gracilaria and Spartina detritus as food. Experiments were done in the lab at ~ 25°C in plastic containers with 200 ml of seawater. No-choice experiments to determine the feeding rate of Gammarus were done by offering live Gracilaria or Spartina detritus to five Gammarus adultsper container. Pieces of each diet that were visually free of epiphytes were collected from the mudflat on the day of the experiments, cleaned of sediment, blotted to a constant wet mass, and weighed. Pieces ranged from 65 – 90 mg wet weight. There were 11 replicates with amphipods for each diet and the same number of replicates without amphipods to control for autogenic changes in seaweed mass. Adjacent segments of the same thallus or piece of detritus were used for each pair of amphipod and control replicates. Amphipod and control treatments were randomly assigned to containers. The experiment ran for 72 h after which all pieces were blotted to a constant wet mass, reweighed, and the differences in feeding rates analyzed with a two-way analysis of variance (ANOVA, diet × herbivore presence/absence). Differential feeding rates between diets are indicated by an interaction between these factors (Peterson and Renaud 1989). Tukey's post hoc test within the herbivore treatment was then used to determine differences among the means of each diet after a correction for autogenic change, using the mean square error from the two-way ANOVA in the Tukey's post hoc tests (Peterson and Renaud 1989).

Feeding rates in no-choice trials do not always reflect feeding preferences. To directly assess feeding preference, we performed feeding choice experiments with five Gammarus per container under similar experimental conditions. Preweighed live Gracilaria and Spartina detritus (65 – 100 mg wet weight, N = 20 replicates with and without amphipods) were both presented to Gammarus, which were allowed to feed for 72 hours before pieces were reweighed. Feeding choice by Gammarus was determined by calculating the difference between the mass loss of live Gracilaria and Spartina detritus for each replicate, and performing a t test on these differences between the herbivore plus and herbivore minus (autogenic control) treatments (Peterson and Renaud 1989).

Samples for stable isotope analysis were oven dried at 55°C and ball milled to a fine powder or, in the case of Gammarus, ground with a glass rod prior to analysis. All individuals were analyzed separately. Analyses were conducted by the University of Georgia Stable Isotope Laboratory using a NA1500 CHN Analyzer (Carlo Erba Strumentazione, Milan, Italy) coupled to a continuous-flow isotope-ratio mass spectrometer (Thermo, San Jose, CA, USA).

For the survivorship experiment, Gammarus were collected from nearby sites and held under the same conditions prior to experiments as amphipods used in feeding experiments. For live Gracilaria and Spartina detritus treatments, amphipods were given ~ 80 mg pieces of one food type, while for the sediment treatment, the bottom of the petri dish was covered to a depth of ~ 2 mm with sediment scraped from the top ~2 mm of the mudflat. The Gracilaria, Spartina and sediment was changed every ~4 days. Faeces were removed and water changed every 3–4 days for 28 days.

Literature cited

Peterson, C. H. and P. E. Renaud. 1989. Analysis of feeding preference experiments. Oecologia 80:82–86.


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