The enemy of my enemy: A bacterial antagonist rescues Chlamydomonas by inactivating a toxin from a rival

Microbial interactions are shaped by multiple community members, yet the underlying mechanisms remain poorly understood. Here, we investigated a tripartite interaction between the microalga Chlamydomonas reinhardtii and two Pseudomonas bacteria that coexist in the same soil habitat. In pairwise cultures, all three interactions are amensalistic (-like): Pseudomonas protegens Pf-5 antagonizes both C. reinhardtii and Pseudomonas fluorescens HKI0770 without benefit, while P. fluorescens inhibits C. reinhardtii without profiting from the alga. However, in a tripartite culture, P. fluorescens protects C. reinhardtii from P. protegens and promotes algal recovery. This protection is not stopped when the used carbon source, acetate, is supplemented. The P. fluorescens rescue effect is associated with reduced levels of the polyyne toxin protegencin produced by P. protegens . Protegencin levels, detected by Raman microspectroscopy, are decreased by application of spent medium or protein extracts from P. fluorescens , while heat- and protease-treatment abolish this activity. A heat-sensitive, proteinaceous factor, likely an enzyme(s), appears to contribute to protegencin inactivation. Our results show that P. fluorescens rescues C. reinhardtii by inactivating protegencin, despite being inhibited by P. protegens . These findings demonstrate the importance of investigating higher-order effects, since additional community members can transform pairwise antagonistic interactions into conditional protection through toxin inactivation.