Construction of spores for portable bacterial whole-cell biosensing systems

Whole-cell sensing systems based on living genetically engineered bacteria are known to have high sensitivity, selectivity, and rapid response times. Although these systems have found applications in biomedical and environmental analyses, their limited shelf life and transportability still restrict their use for on-site monitoring of analytes. To that end, we have developed a new method for the long-term preservation, storage, and transport of whole-cell biosensing systems that is based on bacterial spores, a dormant form of life. Specifically, we have empl oyed spore-forming bacteria such as Bacillus subtilis and Bacillus megaterium for development of luminescent sensing systems for two model analytes, namely, arsenic and zinc. These sensing cells were converted to spores, which can then be revived (germinated) at a later time to generate viable and metabolically active cells. Herein, we demonstrate that these spore-based sensing systems retained their analytical performance, in terms of detection limit, dynamic range, and reproducibility, after storage at room temperature for at least 6 and 8 months, respectively, as well as after three cycles where the cells alternated between being dormant or active, i.e., sporulation-germination cycles. The ability to cycle the sensing cells between active and dormant states prolongs the cell's lifetimes and increases their robustness and ruggedness, thus making them more amenable for field applications. In addition, the small size of spores allows for their easy transport and incorporation in miniaturized portable devices. Finally, we envision that this novel strategy could expand the use of whole-cell biosensors for on-site sensing not only in mild environments but also in harsh environments and locations where there is no easy access to a laboratory, e.g., in developing countries.