A dip-stick type biosensor using bioluminescent bacteria encapsulated in color-coded alginate microbeads for detection of water toxicity

Abstract

The use of genetically engineered bioluminescent bacteria, in which bioluminescence is induced by different modes of toxic action, represents an alternative to acute toxicity tests using living aquatic organisms (plants, vertebrates, or invertebrates) in an aqueous environment. A number of these bacterial strains have been developed, but there have been no attempts to develop a hand-held type of biosensor for monitoring or identification of toxicity. We report a facile dip-stick type biosensor using genetically engineered bioluminescent bacteria as a new platform for classification and identification of toxicity in water environments. This dip-stick type biosensor is composed of eight different optically color-coded functional alginate beads that each encapsulates a different bioluminescent bacterial strain and its corresponding fluorescent microbead. These color-coded microbeads exhibit easy identification of encapsulated microbeads, since each microbead has a different color code depending on the bioluminescent bacterial strain contained and improved cell-stability compared to liquid culture. This dip-stick type biosensor can discriminate different modes of toxic actions (i.e. DNA damage, oxidative damage, cell-membrane damage, or protein damage) of sample water tested by simply dipping the stick into the water samples. It was found that each color-coded microbead emitted distinct bioluminescence, and each dip-stick type biosensor showed different bioluminescence patterns within 2 hours, depending on the toxic chemicals contained in LB medium, tap water, or river water samples. This dip-stick type biosensor can, therefore, be widely and practically used in checking toxicity of water in the environment primarily in situ, possibly indicating the status of biodiversity.

Stimuli-responsive engineered living materials

“…Taking advantage of encapsulation methods to create a device with multiple responses to various toxic compounds, a dip-stick type biosensor that produces bioluminescence in response to a pool of contaminants has been developed. 147 The dip-stick biosensor consisted of eight bioluminescent bacterial strains encapsulated in color-coded alginate microbeads that were entrapped in a laser cut transparent glass (Fig. 4A).…”

“…4B). 147 This approach shows a significant advancement for the development of stimuli-responsive ELM devices that detect multiple chemical contaminants with specificity and produce a readable and easy to measure response. An overview of the applications described in this section is given in Table 1.…”

Engineered Living Hydrogels

“…Second, when multiple types of cells are incorporated in a single hydrogel matrix and each cell type is exclusively sensitive to a specific signal, various signals can be sensed in an efficient manner (Figure 11b, middle). [22,46,192,193] For example, a dip-stick type living sensor that incorporates eight bacterial strains can detect eight different toxins simultaneously (Figure 11b, middle). [193] Third, the distribution of different microbial cells and their connectivity allow engineered living hydrogels to perform advanced computing functions, such as signal digitization, signal amplification, Boolean logic gates, and data storage (Figure 11b, bottom).…”

Creation of a new portable biosensor for water toxicity determination

“…Increasing alginate concentration (from 0.5 to 2% (w/v)) decreased their light response. The immobilization of cells in calcium alginate has been widely studied and used in many applications [32][33][34][35][36]. Diffusion of ethanol decreased with increasing alginate concentration [37].…”