Enhanced Biodegradation of the Organophosphate Pesticide Diazinon Using phoU Knockout Mutant of Synechocystis sp. PCC 6803
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Diazinon is a widely used organophosphate insecticide that persists in agricultural runoff and poses risks to aquatic ecosystems and public health. In this study, we investigated the potential of a genetically engi neered cyanobacterium, Synechocystis sp. PCC 6803, for efficient diazinon removal under varying environ mental conditions. A ΔphoU mutant lacking the negative regulator of the Pho regulon was employed to maintain constitutive phosphate uptake and metabolism independent of external phosphate availability. The ΔphoU strain exhibited sustained expression of alkaline phosphatase (ALP), resulting in ALP activity 2.5-fold higher than that of the wild-type (WT). Under different phosphate concentrations (0-1.6 mM), ΔphoU completely degraded diazinon (20 mg/l) within 11 days, with a degradation rate constant (k) up to 2.1-fold higher than WT under phosphate-replete conditions. Concurrently, ΔphoU accumulated intracellular polyphosphate (poly-P) at levels up to four times higher than WT. Light intensity further modulated diazi non degradation and phosphate metabolism. Under high light intensity (2000 μmol photons/m2/s), the ΔphoU strain showed a 2.5-fold increase in diazinon degradation rate compared to WT, accompanied by a maximum poly-P accumulation of 10.95 mg/g FCW. Correlation analysis revealed a strong positive relation ship between degradation rate and poly-P accumulation, indicating that organophosphate pesticide degra dation is functionally linked to intracellular phosphate metabolism. These findings demonstrate that genetic deregulation of the Pho regulon enhances both diazinon degradation and phosphate assimilation, highlighting the ΔphoU mutant as an effective cyanobacterial platform for organophosphate pesticide bioremediation under environmentally relevant conditions.
Abstract
Diazinon is a widely used organophosphate insecticide that persists in agricultural runoff and poses risks to aquatic ecosystems and public health. In this study, we investigated the potential of a genetically engi neered cyanobacterium, Synechocystis sp. PCC 6803, for efficient diazinon removal under varying environ mental conditions. A ΔphoU mutant lacking the negative regulator of the Pho regulon was employed to maintain constitutive phosphate uptake and metabolism independent of external phosphate availability. The ΔphoU strain exhibited sustained expression of alkaline phosphatase (ALP), resulting in ALP activity 2.5-fold higher than that of the wild-type (WT). Under different phosphate concentrations (0-1.6 mM), ΔphoU completely degraded diazinon (20 mg/l) within 11 days, with a degradation rate constant (k) up to 2.1-fold higher than WT under phosphate-replete conditions. Concurrently, ΔphoU accumulated intracellular polyphosphate (poly-P) at levels up to four times higher than WT. Light intensity further modulated diazi non degradation and phosphate metabolism. Under high light intensity (2000 μmol photons/m2/s), the ΔphoU strain showed a 2.5-fold increase in diazinon degradation rate compared to WT, accompanied by a maximum poly-P accumulation of 10.95 mg/g FCW. Correlation analysis revealed a strong positive relation ship between degradation rate and poly-P accumulation, indicating that organophosphate pesticide degra dation is functionally linked to intracellular phosphate metabolism. These findings demonstrate that genetic deregulation of the Pho regulon enhances both diazinon degradation and phosphate assimilation, highlighting the ΔphoU mutant as an effective cyanobacterial platform for organophosphate pesticide bioremediation under environmentally relevant conditions.