C4

State of the art

Mixotrophs combine photosynthesis with phagotrophy to cover their demands in energy and essential nutrients (Rothhaupt 1996a; Flynn et al. 2012). Mixotrophs can be important grazers on bacteria (both heterotrophic and autotrophic), especially in oligotrophic pelagic systems (Berninger et al. 1992; Hartmann et al. 2012; Unrein et al. 2014). Mixotrophs compete with obligately photosynthetic phytoplankton for dissolved nutrients. Yet, they are able to coexist because mixotrophs obtain mineral nutrients by feeding on bacteria (Katechakis and Stibor 2006). Due to their high surface:volume ratio, heterotrophic and autotrophic bacteria dominate the uptake of dissolved nutrients at low concentrations (Rothhaupt and Güde 1992). Given enough light, mixotrophs are able to maintain positive growth rates at low bacterial prey concentrations and are thus able to outcompete purely heterotrophic bacterivores (Rothhaupt 1996b; Tittel et al. 2003). In contrast to heterotrophic bacterivores, mixotrophs do not necessarily remineralize inorganic nutrients, but are able to use them for their own photosynthetic growth (Rothhaupt 1996a, 1997). Remineralization of inorganic nutrients by heterotrophic bacterivores, however, stimulates autotrophic phytoplankton. Due to high C:nutrient ratios under conditions of nutrient limitation, phytoplankton are able to establish high biomass yield if they are stimulated by heterotrophic bacterivores (Rothhaupt 1992). In contrast to autotrophic phytoplankton, mixotrophs are characterized by moderate C:nutrient ratios (Caron et al. 1990). Consequently, total biomasses of protists that form the base of the food web (phytoplankton, mixotrophs, heterotrophic bacterivores) are assumed to be lower under conditions of mixotrophic bacterivory than under conditions of heterotrophic bacterivory.

Re-oligotrophication of Lake Constance went along with a regime shift in the phytoplankton from a high biomass period (HBP) to a low biomass period (LBP) and a concomitant increase (both relative and total) of the biomass of chrysophytes (Jochimsen et al. 2013). Many chrysophytes are known to be mixotrophs (Porter 1988). During the HBP, light limitation was an important factor (Jochimsen et al. 2013).

Preparatory work

KOR and PGK have documented expertises in experimental work with heterotrophic, mixotrophic and phototrophic protists (Rothhaupt 1992, 1996a; b, 1997; Rottberger et al. 2013). DMC studied the trophic upgrading of nutritionally inadequate autotrophic picoplankton by heterotrophic protists (i.e. Paraphysomonas sp., ciliates) for subsequent use by zooplankton (Martin-Creuzburg et al. 2005; Bec et al. 2006). DS will contribute his expertise in respect to bacteria and setting-up, maintaining and sampling of continuous-flow and chemostatic cultures (Buhmann et al. 2012; Kesberg and Schleheck 2013), and the link to the NGS sequencing pipeline (project A2).

Proposed project and role within the RTG

We hypothesize that the regime shift from a HBP to a LBP in Lake Constance can be understood as a shift in the importance of functional groups, from heterotrophic to mixotrophic bacterivores, due to an increased availability of light. Besides the role of vegetation in shallow lakes (Scheffer et al. 1993) and nutrients release from the sediment due to hypolimnic anoxia in eutrophic lakes (Carpenter et al. 1999), this is supposed to be a third and novel mechanism that can cause regime shifts in lake ecosystems.

Thus, our project deals with a central point of the RTG, i.e., the question if the changes at the base of the food web, that were observed and documented during the re-oligotrophication of Lake Constance, can be mechanistically explained by elementary food-web interactions. In our experimental work, we will start with artificial food webs in chemostats that are assembled from laboratory cultures to comprise the relevant functional groups. Treatments will consist of varying light and nutrient levels. In a further step, the chemostat experiments will be performed with an inoculum of seston from Lake Constance that in principle should contain representatives of all functional groups. The different protist communities that are expected to develop, depending on the light and nutrient levels, should differ in the severity and nature of bacterial mortality, which is expected to impact on the bacterial community composition and diversity. The impact on the bacterial community composition and abundance will be determined through the 16S rRNA amplicon-sequencing pipeline (with help by the doctoral researcher of project A2), which will allow for a very fine-scaled correlation analysis regarding negative interactions (grazing) between bacteriovorous protist and bacterial taxa. In addition to the correlation of abundance data obtained by the NGS pipeline, the doctoral researchers of projects A2 and C4 are planned to conjointly explore possibilities to feed BrdU/¹³C -labelled bacteria to protists, in order to identify important taxa of the heterotrophic/mixotropic protist contingent directy (through sequencing of BrdU/¹³C-labelled protist DNA; see therefore also project description A2). Additionally, the organisms that constitute the base of the food web in dependence on the light and nutrient levels in the different treatments should differ in their resource quality for subsequent consumers. The food quality of mixotrophs, which may produce harmful secondary metabolites (chlorosulfolipids), will be explored in collaboration with project B2. The project is strongly linked to projects A2 and B3 investigating the role of mixotrophs in the oligotrophic food web.