Interspecies interactions reduce selection for a biofilm‐optimized variant in a four‐species biofilm model

Summary Multispecies biofilms are structured and spatially defined communities, where interspecies interactions impact assembly and functionality. Here, we compared the spatial organization and growth of bacterial cells in differently composed biofilm communities over time to determine links between interspecies interactions and selection for biofilm phenotypes of individual species. An established model community consisting of Stenotrophomonas rhizophila , Xanthomonas retroflexus , Microbacterium oxydans and Paenibacillus amylolyticus was used. It was found that interspecies interactions led to varying levels of selection for a new colony phenotype of X . retroflexus , depending on the presence/absence of other species. When M . oxydans was absent, X . retroflexus was not able to establish in the top layers of the biofilm, which led to selection for a hyper‐matrix forming phenotype of X . retroflexus that successfully established in the biofilm top layers. No such phenotypic X . retroflexus variants were identified in the presence of M . oxydans . These findings indicate that interspecies interactions may lead to favourable localization of individual species in a multispecies biofilm and thereby reduce selection for competitive phenotypes.