Explaining the enormous biodiversity observed in bacterial communities is challenging because ecological theory predicts that competition between species occupying the same niche should lead to the exclusion of less competitive community members. Competitive exclusion should be particularly strong when species compete for a single limiting resource or live in unstructured habitats that offer no refuge for weaker competitors. Here, we describe the 'cheating effect', a form of intra-specific competition that can counterbalance between-species competition, thereby fostering biodiversity in unstructured habitats. Using experimental communities consisting of the strong competitor Pseudomonas aeruginosa (PA) and its weaker counterpart Burkholderia cenocepacia (BC), we show that co-existence is impossible when the two species compete for a single limiting resource, iron. However, when introducing a PA cheating mutant, which specifically exploits the iron-scavenging siderophores produced by the PA wild type, we found that biodiversity was preserved under well-mixed conditions where PA cheats could outcompete the PA wild type. Cheating fosters biodiversity in our system because it creates strong intra-specific competition, which equalizes fitness differences between PA and BC. Our study identifies cheating - typically considered a destructive element - as a constructive force in shaping biodiversity.

Phenotypic plasticity in response to competition is a well-described phenomenon in higher organisms. Here, we show that also bacteria have the ability to sense the presence of competitors and mount fine-tuned responses to match prevailing levels of competition. In our experiments, we studied interspecific competition for iron between the bacterium Pseudomonas aeruginosa (PA) and its competitor Burkholderia cenocepacia (BC). We focused on the ability of PA to phenotypically adjust the production of pyoverdine, an iron-scavenging siderophore. We found that PA upregulates pyoverdine production early on during competition under condition of low iron availability. This plastic upregulation was fine-tuned in response to the level of competition imposed by BC, and seems to confer a relative fitness benefit to PA in the form of an earlier initiation of growth. At later time points, however, PA showed reduced growth in mixed compared to monoculture, suggesting that competitive responses are costly. Altogether, our results demonstrate that phenotypic plasticity in siderophore production plays an important role in interspecific competition for iron. Upregulating siderophore production may be a powerful strategy to lock iron away from competing species, and to reserve this nutrient for strain members possessing the compatible receptor for uptake.

Given the rise of bacterial resistance against antibiotics, we urgently need alternative strategies to fight infections. Some propose we should disarm rather than kill bacteria, through targeted disruption of their virulence factors. It is assumed that this approach (i) induces weak selection for resistance because it should only minimally impact bacterial fitness, and (ii) is specific, only interfering with the virulence factor in question. Given that pathogenicity emerges from complex interactions between pathogens, hosts and their environment, such assumptions may be unrealistic. To address this issue in a test case, we conducted experiments with the opportunistic human pathogen Pseudomonas aeruginosa, where we manipulated the availability of a virulence factor, the iron‐scavenging pyoverdine, within the insect host Galleria mellonella. We observed that pyoverdine availability was not stringently predictive of virulence and affected bacterial fitness in nonlinear ways. We show that this complexity could partly arise because pyoverdine availability affects host responses and alters the expression of regulatorily linked virulence factors. Our results reveal that virulence factor manipulation feeds back on pathogen and host behaviour, which in turn affects virulence. Our findings highlight that realizing effective and evolutionarily robust antivirulence therapies will ultimately require deeper engagement with the intrinsic complexity of host–pathogen systems.

Given the rise of bacterial resistance against antibiotics, we urgently need alternative strategies to fight infections. Some propose we should disarm rather than kill bacteria, through targeted disruption of their virulence factors. It is assumed that this approach (i) induces weak selection for resistance because it should only minimally impact bacterial fitness, and (ii) is specific, only interfering with the virulence factor in question.Given that pathogenicity emerges from complex interactions between pathogens, hosts and their environment, such assumptions may be unrealistic. To address this issue in a test case, we conducted experiments with the opportunistic human pathogen Pseudomonas aeruginosa, where we manipulated the availability of a virulence factor, the iron-scavenging pyoverdine, within the insect host Galleria mellonella. We observed that pyoverdine availability was not stringently predictive of virulence and affected bacterial fitness in nonlinear ways. We show that this complexity could partly arise because pyoverdine availability affects host responses and alters the expression of regulatorily linked virulence factors. Our results reveal that virulence factor manipulation feeds back on pathogen and host behaviour, which in turn affects virulence.Our findings highlight that realizing effective and evolutionarily robust antivirulence therapies will ultimately require deeper engagement with the intrinsic complexity of host-pathogen systems. K E Y W O R D Santivirulence therapy, Galleria mellonella, host effects, opportunistic pathogen, pathogen fitness, pleiotropy, Pseudomonas aeruginosa, siderophore

Most habitats on earth are colonized by diverse bacterial communities, offering ample opportunities for inter-species interactions. While competition for space and nutrients might often dominate such interactions, little is known about whether bacteria can sense competitors and mount specific responses to withstand their attacks. The competition-sensing hypothesis proposes that bacteria can do so through nutrient stress and cell damage cues. Here, we conducted a replicated RNAseq-transcriptomic study to test this hypothesis. We exposed Pseudomonas aeruginosa to either its own spent medium or to the supernatant of its competitor Burkholderia cenocepacia. Compared to controls, we detected significant changes in the transcriptome of P. aeruginosa that entail both general responses to spent medium and specific responses to the competitor. Specifically, genes encoding various competitive traits, including the type-VI secretion system, the siderophore pyoverdine, and the toxins phenazines and hydrogen cyanide, were upregulated when exposed to the competitor supernatant. Similarly, several genes coding for stress response and quorum-sensing regulators were overexpressed. Moreover, we found transcriptional responses to vary as a function of iron availability, whereby metabolically more costly responses were launched under iron rich conditions. Altogether, our results reveal nuanced competitive responses of P. aeruginosa when exposed to B. cenocepacia supernatant, integrating both environmental and social cues.