Development of halotolerant multi-pesticides degraders for wastewater treatment

The biotreatment performance of high saline wastewater containing multiple pesticides is significantly reduced because the growth and metabolism of activated sludge microbial communities are severely impaired by high osmotic pressure and joint toxicity of multiple pesticides. In this study, Halomonas cupida J9 was metabolically engineered by chromosomal integration of three different pesticide hydrolase genes, to create a halotolerant multi-pesticides degrader. In batch degradation experiments, the finally constructed strain J9U-PVG was able to simultaneously degrade chlorpyrifos (organophosphate), carbaryl (carbamate) and alpha-cypermethrin (pyrethroid) in mineral salt medium supplemented with 4 g/L glucose and 60 g/L NaCl. Total cell lysate of J9U-PVG showed obvious degradation activity against chlorpyrifos, carbaryl and alpha-cypermethrin. Moreover, introduced Vitreoscilla hemoglobin (VHb) can enhance oxygen-sequestering capability of J9U-PVG and introduced green fluorescent protein (GFP) can be used as a biomarker to track the activity and movement of J9U-PVG in natural environments. More importantly, J9U-PVG could simultaneously and completely degrade chlorpyrifos, carbaryl and alpha-cypermethrin in high saline river water. Here, we demonstrate for the first time the successful construction of a halotolerant multi-pesticides degrader. This study underscores the value of H. cupida J9 as a promising chassis for the development of a halotolerant pesticide bioremediation platform. In the future, synthetic biology may serve as a powerful tool for expanding the catalytic repertoire of extremophiles for bioremediation of extreme environments.