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.