| Summary: | <p>The evolution of multicellular organisms from single cells has puzzled scientists for centuries. Separating this problem into smaller steps has been illuminating. The first step is multicellular group formation and the second is group transformation into an obligate multicellular entity where cells are fixed in a multicellular lifestyle and cannot reverse back to being single cells. Relatedness among cells is known to be fundamental in this major transition to an obligate lifestyle, but the role of ecology has not been addressed to such an extent. For instance, in the ecological literature, predation is known to induce defensive group formation in freshwater algae, and this has been mainly investigated as a means of harvesting algae. However, the role of algal predation in the context of multicellularity and social evolution has not been investigated as much. I use the freshwater algae <em>Chlorella sorokiniana, Chlorella vulgaris</em>, and <em>Scenedesmus obliquus</em> exposed to the predators <em>Ochromonas, Tetrahymena thermophila</em>, and <em>Daphnia magna</em>, to address why and how algae form multicellular groups. Answering why algae form groups helps us illuminate the costs and benefits involved in this cooperative behaviour. Answering how these algae form groups helps us understand whether these algae are likely to make a transition to obligate multicellularity, since we know that all obligate multicellular organisms have evolved via cells remaining stuck to their parent cell after division, not by cell aggregation. We found that these three algal species form multicellular groups in response to the three different predators, and in three predator-prey combinations this multicellular group formation was induced just by predator cues. These multicellular groups formed via aggregation of single cells and by cells remaining attached to the parent cell after division. Also, multicellular groups could form between different algal species. This group formation is costly under conditions of limited resources in <em>C. sorokiniana</em>, and provides a benefit in terms of avoiding ingestion by small predators, such as <em>Ochromonos danica</em>, but not by the larger predator <em>D. magna</em>. The benefit of multicellular group formation in the latter case may be that cells in multicellular groups are better able than single cells to kill their predator once they are ingested. Finally, this group formation is reversible. These results indicate that our system has not made a major transition to obligate multicellularity, but has made the first step of facultative multicellular group formation. This facultative feature of our system has therefore been ideal in allowing us to reveal the costs and benefits of multicellular group formation.</p>
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