Data for: Top-down control and species composition non-linearly influence the short-term response of experimental food webs to a nutrient pulse perturbation

Extreme weather events intensify with global change and frequently disturb aquatic ecosystems via e.g., nutrient pulses through surface runoff. While there is accumulating evidence that the associated short-term responses of plankton communities depend on their top-down control (TC), the actual shape of TC-community response relationships remains unclear. We therefore conducted 24 flow-through (chemostat) experiments to investigate algal and rotifer biomass responses to a nutrient pulse along a gradient of TC values. To generate this gradient, we used a trait-based method instead of varying the inoculation species densities. We complemented our analysis by estimating non-measured nutrient dynamics and feeding interactions via Bayesian modelling. We found support for hump-shaped relationships. At low TC, algal species were strongly nutrient-depleted, limiting their immediate growth potential after the pulse, and thereby one of the rotifer species. At high TC, algae were not nutrient-limited, thus algae and rotifers hardly responded. At intermediate TC, responses were larger because algae immediately exploited the pulse, and rotifers exploited the increased algal biomass production without preventing an algal bloom. This study showed the importance of investigating community responses to resource pulses along a gradient of TC values, thereby improving the understanding of the ecological stability of food webs under global change.

Methods
The aim of this study is to investigate the response in biovolume of phytoplankton (algae) and zooplankton (rotifers) after a nutrient pulse perturbation. We notably test two types of relationships:

the hump-shaped relationship between biovolume responses of algae and the top-down control at the pulse perturbation
the hump-shaped relationship between biovolume responses of rotifers and the top-down control at the pulse perturbation

In the following, you can find a summary of the methods. Please see the published study for more details.

We cultivated the four algal species Chlamydomonas reinhardtii (strain No. SAG 11-32b), Monoraphidium minutum (SAG 243-1), Chlorella vulgaris (SAG 211-11b), and Cryptomonas sp. (SAG 26-80), and the three herbivorous rotifer species Brachionus calyciflorus sensu strictu (isolated from the Milwaukee Harbor, Wisconsin, USA), Cephalodella sp. (obtained from Florian Altermatt, Campus pond), and Lecane sp. (isolated by Christina Schirmer, Löschteich Golm, Potsdam, Germany). Stock cultures were reared in 300-mL flasks containing 200 mL of Woods Hole Culture medium (WC medium) at pH 7, 19.5°C, and a 16:8 hour day:night cycle. B. calyciflorus and Cephalodella sp. were fed once per week with C. reinhardtii, and Lecane sp. with C. vulgaris. Three days before inoculation, we diluted algal stock cultures with fresh medium to guarantee exponential growth.

We ran chemostats (i.e., flow-through systems) in 1-L bottles filled with 800 mL of nitrogen-reduced WC medium (inorganic nitrogen N0 = 80 μmol N ⋅ L −1 rather than 1 mmol N ⋅ L −1) at 19.5° C, under constant light conditions, and with a dilution rate δ = 0.2 d −1. After one to two weeks, we spiked the nitrogen concentration to a total of 400 μmol N ⋅ L −1 and the phosphorus concentration in the same ratio to avoid a shift in the limiting macronutrient. Then, we measured the short-term responses of algae and rotifers over 12 days.

We conducted a total of 24 chemostats comprising the four algal species, differing in the identity and number of rotifer species, and timing of the pulse. Varying these factors generated a gradient of top-down control (TC). With this information in mind, we designed two treatments: (i) rotifer monocultures pulsed 17 days after inoculation (12 chemostats, i.e., 4 per rotifer species); and (ii) rotifer polycultures comprising all the rotifer species and pulsed 9 or 13 days after inoculation (12 chemostats, i.e., 6 per date).

To obtain time series of species biovolumes, we sampled 25 mL of each chemostat three times a week, and daily during the first five days after the perturbation. A 10 mL subsample fixed with Lugol’s solution was used to determine algal density and size spectra with a particle counter (CASY; Schärfe, Germany), and rotifer density with a microscope (Zeiss Axioskop 2, Germany). Then, we calculated algal and rotifer biomass as their respective total biovolume (in μm3 ⋅ mL −1) using densities and size spectra for algae, and densities and fixed species-specific volumes for rotifers (1.85 ⋅ 106 μm3 for B. calyciflorus, 7.37 ⋅ 104 μm3 for Cephalodella sp., and 3.45 ⋅ 104 μm3 for Lecane sp.). Algal species composition was analysed via flow cytometry (BD Accuri C6 Flow Cytometer, BD Biosciences), accounting for differences in algal morphology and pigmentation. Distinguishing M. minutum from C. vulgaris was not possible, hence they were considered together as one morphological group in the analyses.

Following the framework of Hillebrand et al (2018) and Urrutia-Cordero et al (2022), biomass responses were quantified using the metric of overall ecological vulnerability (OEV), which integrates multiple dimensions of ecological stability such as resistance, resilience, and recovery. A large OEV value means that a trophic level or a community strongly reacts to a perturbation because of pronounced biomass changes in comparison to the pre-perturbation conditions. The trophic level or community is thus vulnerable to a perturbation, indicating low ecological stability. We quantified the short-term responses over a maximum of 12 days after the pulse and standardised these responses by the sampling period of each chemostat and obtained thereby the average biomass responses per day.

We expect biomass responses of algae and rotifers to be influenced by the top-down control (TC). TC is the log2 value of the ratio between the total algal biomass per chemostat with rotifers at the pulse and the average total algal biomass in the absence of rotifers at the pulse (i.e., carrying capacity of 4 ⋅ 107 μm3 ⋅ mL −1). We also determined the relative biomasses, i.e., the species biomasses divided by the trophic level biomass.

Data analysis was performed in R 4.5.0 and RStudio environment using notably the libraries tidyverse, PKNCA, cmdstanr, vegan, brms, gridExtra, scales, readxl, stringr, and deSolve. For each trophic level and each treatment separately (12 rotifer monocultures or 12 rotifer polycultures) or both treatments together, we fitted quadratic regressions (y ∼ a + bx + cx2 ) to log2 avgOEV and TC data. Because of the small number of observations, we used Bayesian inference, which produces accurate estimates of parameter uncertainty even in small datasets. We considered a relationship as hump-shaped if (i) the quadratic term c of the regression was negative, and (ii) if the predicted maximum $-b/2c was located within the observed range of TC. We calculated posterior probabilities as

P(shape)=P(c<0 AND TCmin < -b/2c < TCmax).

Furthermore, we obtained insights into non-measurable nutrient dynamics and species interactions, and, thus, into top-down and bottom-up control, via Bayesian inference. For this, we fitted the predictions of a differential equation model to the experimental timeseries of trophic level biomasses, and estimated posterior probability distributions of all the model parameters.