Microencapsulation of engineered bacteria towards whole-cell-based environmental biosensing.

Introduction: Legitimate concerns about the quality of environmental waters are on the rise. As a consequence, efforts are being made by both the scientific community and policymakers to develop proficient methods for the monitoring and detection of pollutants of emerging concerns (PECs). In line with the European Union's "Zero Pollution" action plan, there is a growing need to develop real-time, multiplexed, on-site monitoring systems. The study explores the encapsulation of engineered Methods: Genetically modified Results: Encapsulation strategy using alginate-PLL capsules revealed a good mechanical stability and resistance to saline water, with minimal cell leakage. About 75% of encapsulated cells remained viable after 45 days, and fluorescence increase upon induction was still observed after 2 months of storage at room temperature without nutrient supply. These findings suggest that a subpopulation of cells entered a dormant or low-metabolic state, enabling long-term sensing under non-ideal conditions. Higher cell loads correlated with stronger responses, likely due to an increased fraction of metabolically reactivable cells. Encapsulated cells were also reusable, showing measurable sensing response for five sensing cycles within 2 weeks. Discussion: Results demonstrate that response dynamics are primarily governed by reduced metabolic activity and limited oxygen access, rather than by analytes diffusion through the hydrogel matrix.