Incorporation of nitro-sensitive fluorescent bacteria and nitro-capturing bacteriophages into alginate-cellulose beads as field-deployable biosensor for highly sensitive detection of nitro compounds

Abstract

Bacterial species capable of expressing fluorophores in response toexternal stimuli are actively being utilized as light-activated sensors forvarious applications. These stimuli-responsive bacteria encapsulatedin miniaturized spherical hydrogels (‘microbeads’) are especially useful as a field deployable form of sensors for detecting environmental chemicals, due to the capability of mass production as well as long-range light detection. Herein, genetically engineered bacteria capable of expressing enhanced green fluorescent protein (eGFP+) in response to nitro compounds were encapsulated into alginate-cellulose beads to develop microbead biosensor. Mechanical strength of the conventional alginate microbeads was improved by incorporating anionic cellulose. The encapsulated bacteria proliferated within the microbeads, and eGFP+ expression was proportional to the amount of nitro compounds (e.g. DNT and TNT). Furthermore, the M13 bacteriophage having high binding affinity towards DNT and TNT were also encapsulated into the same microbeads, resulting in increased sensitivity of nitro detection. In addition, the fluorescence emitted from the microbead biosensor deployed on a soil sample was detected at long range (e.g. 20 meters and beyond) using a laser fluorescent scanning system to validate the feasibility as field application and safe long-range detection of explosives.