Grazing of heterotrophic flagellates on viruses is driven by feeding behaviour

The trophic interactions between viruses, bacteria and protists play a crucial role in structuring microbial communities and regulating nutrient and organic matter flux. Here, we show that the impact on viral density by heterotrophic flagellates is related to their feeding behaviour (feeding on sedimented particles Thaumatomonas coloniensis, filter feeding of suspended particles - Salpingoeca sp., and actively searching raptorial feeding - Goniomonas truncata). Phage MS2 was co-incubated with flagellates and the natural bacterial and viral community originating from the same groundwater habitats where the flagellates were isolated. Three complementary assays, i.e. flow cytometry, qPCR and plaque assay, were used for enumeration of total viruses, total MS2 phages, and free and infectious MS2, respectively, to provide insights into the grazing mechanisms of the flagellates on viruses. Phage MS2 was actively removed by the suspension feeders T. coloniensis and Salpingoeca sp. in contrast with the actively raptoriale grazer G. truncata. The decline of viral titre was demonstrated to be caused by ingestion rather than random absorption by both qPCR and locating protein fluorescently labelled MS2 inside the flagellates. Further, we indicate that phages can be used as a minor carbon source for flagellates. Collectively, these data demonstrate that eliminating viruses can be an important function of protists in microbial food webs, carbon cycling and potentially water quality control.