AHLs-driven quorum sensing modulates competitive interspecies interactions in synthetic biofilms community among fish spoilage bacteria

Abstract

Food spoilage microorganisms often reside in dense mixed-species biofilms mediated by quorum sensing (QS). However, the regulatory mechanisms underlying interspecific interactions within spoilage microecosystems remain poorly understood. In this study, an experimental mixed-species biofilms model consisting of Pseudomonas fluorescens, Shewanella baltica, and Acinetobacter johnsonii was established to examine interspecies interactions at different temperatures and to explore regulatory role of N-acyl-L-homoserine lactones (AHLs). Genome sequencing revealed that compared to only LuxR receptor in S. baltica and A. johnsonii, P. fluorescens possesses a complete LuxI/R-type QS system and numerous genes related to biofilms formation and iron acquisition. Phenotypic results confirmed that P. fluorescens, as the dominant strain in the mixed-species biofilms, enhanced its competitiveness at low temperatures by repressing S. baltica metabolism. Meanwhile, S. baltica and A. johnsonii could eavesdrop on AHLs signaling secreted by P. fluorescens, particularly C₄-HSL. C₄-HSL-mediated QS modulated interspecies competition within the biofilms community and contribute to increased disinfectant resistance, likely through enhanced antioxidant capacity. Transcriptomic analysis revealed that C₄-HSL activated multiple genes expression involved in oxidative stress defense to mitigate intracellular ROS accumulation, including antioxidant enzyme synthesis, pyoverdine production for iron chelation and competitive advantage, and the upregulating ABC transporters and two-component systems to facilitate substance transport. Thus, these findings demonstrate that AHL-mediated interspecies communication plays a critical role in maintaining biofilms stability and provide mechanistic insights into QS-driven bacterial adaptation, offering potential strategies for targeted biofilms control.