A Function-Driven Single-Cell Approach to Unveiling and Cultivating Hidden Microplastic-Degrading Microbes from Insect Gut Microbiota

<title>Abstract</title> Biodegradation is a sustainable strategy to address global microplastics (MPs) pollution but is constrained by the lack of efficient degrading microbes and effective tools to harness them. Here, we developed a function-driven single-cell approach to precisely identify and recover MPs-degrading microorganisms from complex microbiota by integrating isotope-labeled single-cell Raman spectroscopy with targeted cell sorting, sequencing and culturing. Using heavy water and MPs as the sole carbon source, Raman spectroscopy effectively identified active microbes capable of degrading multiple types of MPs from insect gut microbiota. Raman-guided single-cell sorting and sequencing revealed seven previously unrecognized polystyrene degraders and mapped key enzymes involved in each degradation stage. Furthermore, live-cell Raman-guided sorting enabled the cultivation of rare but highly active polystyrene degraders often missed by conventional methods. This “screen-first, culture-second” single-cell approach offers a powerful and scalable platform to accelerate MPs biodegradation and supports the development of microbial solutions to mitigate global plastic pollution.