Tunable lysis of functionalized nanocultures for efficient microbial and nucleic acid recovery

The cultivation of low-abundance and fastidious microorganisms remains a critical challenge in microbiology. To overcome this, we developed nanocultures —a microencapsulation platform that enables high-throughput cultivation of microbes under near-native conditions. These nanoliter-sized polymeric microcapsules are generated via droplet microfluidics and provide a mechanically robust yet tunable environment for microbial growth, isolation, and interrogation. By engineering the capsule shell chemistry, we control the mechanical properties to allow on-demand lysis and efficient retrieval of intracellular contents. We assessed three physical disruption methods—osmotic swelling, sonication, and bead beating—to evaluate their performance in releasing viable cells and nucleic acids from nanocultures. Osmotic swelling yielded the highest recovery of both viable microbes and high-integrity nucleic acids, outperforming the other methods. This approach preserves the biological relevance of the cultivated communities while enabling downstream processing for species identification and functional analysis. Nanocultures offer a scalable, high-throughput strategy to culture diverse microbial species in environments that mimic natural habitats. Their mechanical tunability provides precise control over microcapsule lysis and content recovery, positioning them as a powerful tool for microbial discovery, diagnostics, and synthetic ecology. • We developed nanocultures for microbial encapsulation, cultivation, and scalable downstream processing. • Engineered polymeric microcapsules exhibit tunable elasticity for controlled burst release. • Osmotic swelling, sonication, and bead beating were tested for efficient cell and nucleic acid recovery. • Osmotic swelling is the most effective method for high yield and integrity of cells, DNA, and RNA from nanocultures.