In nature, microbes interact antagonistically, neutrally, or beneficially. To shed light on the effects of positive interactions in microbial consortia, we introduced metabolic dependencies and metabolite overproduction into four bacterial species. While antagonistic interactions govern the wild-type consortium behavior, the genetic modifications alleviated antagonistic interactions and resulted in beneficial interactions. Engineered cross-feeding increased population evenness, a component of ecological diversity, in different environments, including in a more complex gnotobiotic mouse gut environment. Our findings suggest that metabolite cross-feeding could be used as a tool for intentionally shaping microbial consortia in complex environments. IMPORTANCE Microbial communities are ubiquitous in nature. Bacterial consortia live in and on our body and in our environment, and more recently, biotechnology is applying microbial consortia for bioproduction. As part of our body, bacterial consortia influence us in health and disease. Microbial consortium function is determined by its composition, which in turn is driven by the interactions between species. Further understanding of microbial interactions will help us in deciphering how consortia function in complex environments and may enable us to modify microbial consortia for health and environmental benefits.

Bacteroidales are the most abundant Gram-negative bacteria of the healthy human colonic microbiota, comprising nearly 50% of the colonic bacteria in many individuals. Numerous species and strains of gut Bacteroidales are present simultaneously at high concentrations in this ecosystem. Studies are revealing that gut Bacteroides has numerous antibacterial weapons to antagonize closely related members. In this study, we identify a new diffusible antibacterial toxin produced by Bacteroides fragilis 638R, designated BSAP-4. This is the fifth antibacterial toxin produced by this strain and the second toxin of this strain with a membrane attack complex/perforin domain (MACPF). We identify the target molecule of sensitive cells as a ␤-barrel outer membrane protein (OMP) with calycin-like domains. As with other MACPF toxins, the gene encoding the target in sensitive strains is in the same genetic region as bsap-4 in producing strains. A comparison of B. fragilis strains showed there are two sensitive variants of this OMP that are 87% similar to each other and 50% similar to the resistant OMP. Unlike other MACPF toxins, there are numerous B. fragilis strains that harbor the resistant OMP without bsap-4. Several OMP variants from strains that are BSAP-4 resistant under the conditions of our assay confer BSAP-4 sensitivity to Bacteroides thetaiotaomicron when constitutively expressed. Using a reporter assay, we show that the BSAP-4 receptor gene is differentially expressed in sensitive and resistant strains leading to apparent BSAP-4 resistance under the conditions of our assay, despite harboring the BSAP-4 target gene. IMPORTANCE The intestinal microbiota is a diverse microbial ecosystem that provides numerous benefits to humans. The factors that govern its establishment and stability are just beginning to be elucidated. Identification and characterization of antimicrobial toxins produced by its members and their killing range are essential to understanding the role of antagonism in community composition and stability. Here, we identify a fifth antimicrobial toxin produced by a single Bacteroides fragilis strain and identify its target. The finding of such a large number of toxins that antagonize competing members suggests that this feature substantially contributes to the fitness of these bacteria. In addition, these toxins may have applications in genetically engineered gut bacteria to allow engraftment or to antagonize a potentially pathogenic member.

In nature, microbes interact antagonistically, neutrally or beneficially. To shed light on the effects of positive interactions in microbial consortia we introduced metabolic dependencies and metabolite overproduction into four bacterial species. While antagonistic interactions govern the wildtype consortium behavior, the genetic modifications alleviated antagonistic interactions and resulted in beneficial interactions. Engineered cross-feeding increased population evenness, a component of ecological diversity, in different environments including in a more complex gnotobiotic mouse gut environment. Our findings suggest that metabolite cross-feeding could be used as a tool for intentionally shaping microbial consortia in complex environments.ImportanceMicrobial communities are ubiquitous in nature. Bacterial consortia live in and on our body and in our environment and more recently, biotechnology is applying microbial consortia for bioproduction. As part of our body, bacterial consortia influence us in health and disease. Microbial consortia function is determined by its composition, which in turn is driven by the interactions between species. Further understanding of microbial interactions will help us deciphering how consortia function in complex environments and may enable us to modify microbial consortia for health and environmental benefits.