Grazing rates and functional diversity of uncultured heterotrophic flagellates.

Aquatic assemblages of heterotrophic protists are very diverse and formed primarily by organisms that remain uncultured. Thus, a critical issue is assigning a functional role to this unknown biota. Here we measured grazing rates of uncultured protists in natural assemblages (detected by fluorescent in situ hybridization (FISH)), and investigated their prey preference over several bacterial tracers in short-term ingestion experiments. These included fluorescently labeled bacteria (FLB) and two strains of the Roseobacter lineage and the family Flavobacteriaceae, of various cell sizes, which were offered alive and detected by catalyzed reporter deposition-FISH after the ingestion. We obtained grazing rates of the globally distributed and uncultured marine stramenopiles groups 4 and 1 (MAST-4 and MAST-1C) flagellates. Using FLB, the grazing rate of MAST-4 was somewhat lower than whole community rates, consistent with its small size. MAST-4 preferred live bacteria, and clearance rates with these tracers were up to 2 nl per predator per h. On the other hand, grazing rates of MAST-1C differed strongly depending on the tracer prey used, and these differences could not be explained by cell viability. Highest rates were obtained using FLB whereas the flavobacteria strain was hardly ingested. Possible explanations would be that the small flavobacteria cells were outside the effective size range of edible prey, or that MAST-1C selects against this particular strain. Our original dual FISH protocol applied to grazing experiments reveals important functional differences between distinct uncultured protists and offers the possibility to disentangle the complexity of microbial food webs.