Alginate-εPLL core-shell hydrogel beads as a tool for an effective, stable and scalable microbial encapsulation

Synthetic microbial consortia offer significant potential for biotechnological applications. However, achieving stable and reproducible co-cultures is challenging due to competitive dynamics and unbalanced growth rates among species. Here, we present an effective method for microbial encapsulation based on alginate-based core-shell hydrogel beads coated with ε-poly-L-lysine (εPLL-HB). This method ensures complete microbial containment while allowing for consistent growth inside the beads of several microbial species with a wide variability in their features. Compared to chitosan and α-poly-L-lysine, two of the most used coating agents for this kind of encapsulation, εPLL demonstrated superior performance in avoiding cellular escape across diverse culture conditions and for all the microbial strains tested, while allowing their proliferation inside the capsule. εPLL-HB enabled the construction of spatially organized co-cultures, effectively balancing populations between microorganisms with different growth rates. Furthermore, εPLL-HB provided protection against toxic compounds in lignocellulosic-derived media and maintained their encapsulation efficacy and viability after long-term storage at -80°C. The superior microbial containment, structural integrity and chemical resistance of εPLL-HB, combined with their low price and ease of preparation, makes them a versatile tool for engineering synthetic microbial consortia, with broad applicability in biotechnological processes.