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

Area of Science:

• Environmental Science
• Biotechnology
• Materials Science

Background:

• Rising concerns about environmental water quality necessitate advanced monitoring methods for pollutants of emerging concern (PECs).
• There is a need for real-time, multiplexed, and on-site monitoring systems aligned with the European Union's "Zero Pollution" action plan.
• Engineered microorganisms offer potential for sensitive and specific pollutant detection.

Purpose of the Study:

• To develop and characterize encapsulated engineered *Pseudomonas putida* cells for environmental water monitoring.
• To create a miniaturized, biocompatible, and sustainable sensing element for integration with optical-electrochemical systems.
• To evaluate the stability, reusability, and response dynamics of the encapsulated biosensors.

Main Methods:

• Genetically modified *P. putida* cells were encapsulated in sub-200 µm alginate-poly-L-lysine (PLL) microcapsules.
• Capsule morphology and structure were analyzed using Laser Scanning Confocal and cryo-Scanning Electron microscopy.
• Cell viability, fluorescent response, storage stability, and response-time were evaluated under various conditions.

Main Results:

• Alginate-PLL capsules demonstrated good mechanical stability and resistance to saline water with minimal cell leakage.
• Encapsulated cells remained viable for over 45 days, with detectable fluorescence after 2 months of storage without nutrients.
• Higher cell loads improved responses, and encapsulated cells were reusable for at least five sensing cycles over two weeks.

Conclusions:

• Encapsulation in alginate-PLL microcapsules provides a robust platform for long-term bacterial biosensing.
• The sensing response is primarily limited by reduced metabolic activity and oxygen diffusion, not analyte diffusion.
• This technology offers a promising foundation for developing on-site, real-time monitoring systems for water pollutants.