Utilizing Incoherent Feedforward Loops for Enhanced Response Stability and Coordination of Microbial Consortia-Based Biosensors

Abstract Synthetic multicellular systems can accomplish complex tasks. However, their asynchronous design requires precise adjustments, often resulting in undesired output variations. Here, we created bacterial consortia designed for multi-marker detection, achieving member coordination by activating an array of AND logic gates controlled by a shared quorum sensing signal. Our experimental results demonstrate that successful coordination was achieved when the shared signal was generated by an incoherent feedforward loop (IFFL), leading to a low-level signal that maintains a consistent response across 20 cell generations. These characteristics provide a stable relationship in molecule synthesis and prevent amplitude amplification during signal processing, enabling the IFFL to coordinate the asynchronous activities among consortium members effectively. We examined the consortia-based biosensor by monitoring Lactate and Heme in humanized fecal samples for 40 hours. Our findings provide insights into the IFFL system's dynamic response and long-term stability, making them valuable for biotechnological applications e.g., scaling up synthetic multicellular systems.