The mixotrophic chrysophyte Ochromonas grazing on the toxic and bloom-forming cyanobacterium Microcystis aeruginosa (Alleen Engelse tekst)

Cyanobacterial blooms regularly occur in many eutrophic lakes throughout the world and can cause considerable harm to animals including humans due to the production of toxins. One of the most common freshwater cyanobacteria that forms dense surface scums during late summer is the colony forming species Microcystis aeruginosa. Microcystis species produce the hepatotoxic microcystins that are toxic to many potential grazers of Microcystis like cladocerans, copepods, and rotifers. The toxicity of many cyanobacteria is one reason why their food quality and edibility is comparatively low.
However, the mixotrophic flagellate Ochromonas has been shown to graze efficiently on Microcystis and has even been suggested as a possible biological control agent against blooms of Microcystis.
As a mixotroph Ochromonas has the capability to perform both photosynthesis and heterotrophic nutrition (uptake of particulate or dissolved organic matter). This way of nutrition makes the interaction between Ochromonas and Microcystis particularly interesting, because Ochromonas can act both as a predator and as a competitor (for dissolved nutrients or light) of Microcystis.

Objectives
Different aspects of the interaction between the toxic bloom-forming cyanobacterium Microcystis aeruginosa and the mixotrophic flagellate Ochromonas shall be investigated in this project, giving opportunities for several student projects:

1. Effect of grazing by Ochromonas on the toxin production in microcystins
Whether Microcystis is able to up regulate the production of microcystins in response to grazing (inducible defense against grazing) shall be investigated in grazing experiments with Ochromonas grazing on different strains of M. aeruginosa. To do so, Microcystis will be sorted from a mixed culture containing also Ochromonas using a flow cytometer to be able to distinguish the microcystins in Microcystis from those already taken up and possibly accumulated in Ochromonas. Both microcystins and the mRNA of the genes encoding for enzymes of the microcystin production shall be quantified.

Techniques: batch cultures, flow cytometry, light microscopy, ELISA (enzyme-linked immunosorbent assay) and/ or HPLC (high performance liquid chromatography) for microcystin analysis and RT-qPCR.

2. Effects of Microcystis on Ochromonas growth and grazing rates
In grazing experiments using the toxic M. aeruginosa strain PCC 7806 and its microcystin-deficient mutant the effect of microcystins on the growth and grazing rates of Ochromonas shall be investigated.

Techniques: batch cultures, flow cytometry, functional and numerical response.

3. Factors determining the relative importance of photosynthesis and phagotrophy to the nutrition of Ochromonas
Since both photosynthesis and phagotrophy (uptake of particulate prey) require the investment of energy into the enzymatic machinery needed for each pathway there is a trade off between investment into one or the other of these pathways in mixotrophs. Whether photosynthesis or phagotrophy contribute more to the overall energy-budget depends on prey availability and light intensity. But also other environmental factors like temperature and pH might have an impact. Because growth of purely heterotrophic organisms depends more strongly on temperature than the growth of photoautotrophs does, a similar trend for the two nutritional pathways within mixotrophs could also be expected. This would mean that the relative importance of phagotrophy in mixotrophs increases with increasing temperature.
The effect of temperature on the relative (and absolute) contribution of photosynthesis and phagotrophy to the growth of Ochromonas shall be determined for different combinations of prey availability and light intensity in culture experiments.

Techniques: batch cultures, measurement of primary production by incorporation of C13 or C14, Phyto-PAM measurements of electron transfer efficiency in Photosystem II, determination of grazing rates by microscopical techniques, analysis of particulate (and dissolved) C, N and P, flow cytometry.

4. Population dynamics of Microcystis and Ochromonas under different levels of nutrient or light limitation
A system where two species compete for a shared resource (in this case dissolved nutrients) and at the same time one of them (Ochromonas) feeds on the other (Microcystis) is called a system with intraguild predation. For such systems it has been shown theoretically that the possibility of coexistence for these two species depends on the availability of the resource. For low resource availability only the prey species that usually has the higher resource affinity can attain positive growth rates. For intermediate resource availabilities both species can coexist and for high resource availabilities the predator might reach high enough population densities to exclude the prey species from the system and grow on the resource alone.
These theoretical predictions shall be tested experimentally in P-limited chemostats over a gradient of phosphorus concentrations using Microcystis and Ochromonas as model organisms. Depending on the interests of the student these experiments could be extended for the effect of, for example, light limitation or inorganic carbon availability.

Techniques: Chemostat experiments, analysis of dissolved and particulate C, N and P, flow cytometry.

For more information
Susanne Wilken, tel: +31 (0)294 239 394, e-mail: s.wilken@nioo.knaw.nl