Tuning the sensitivity of cell-based biosensors for the detection of biocides

The presence of biocides in wastewater can negatively impact the efficiency of wastewater treatment processes, particularly the process of nitrification. In this paper, we describe the development of cell-based biosensors (CBBs) with tunable levels of sensitivity for rapidly detecting the presence and predicting the type and concentration of biocides. The CBB assay developed is performed by first exposing a panel of bacterial strains (E. coli, B. subtilis, B. cereus) to the sample being tested and to the control sample without biocide, and then adding a fluorescent dye (LIVE/DEAD BacLight). We then compare the fluorescence signals generated by the two samples, and the differences in the signals indicate the presence of a biocide, as previously reported in the literature. We found that the sensitivity of the CBB assay can be improved by 'tuning' the type/salinity of the buffer used to suspend the cells, and by changing the number of cells used in the assay. These changes improved the level of detection (LOD) of the biocide Cetyltrimethylammonium bromide (CTAB) from 10 ppm to 0.625 ppm and the biocide Grotan® BK from 500 ppm to 7.8 ppm. With the optimized conditions for each strain, we also establish that the combined response from the panel of bacterial strains can be used to predict the type and concentration of biocide sample tested. Additionally, we provide evidence that the CBB assay can be performed using a compact, commercially available fluorometer. Overall, the significance of this work will improve point-of-use testing and enable the discrimination between biocide-containing samples of similar toxicity and detection of lower toxicity samples, thereby improving the accuracy of the CBB assay.