Abstract
As the result of the extensive use and high contamination of organophosphate (OP) insecticides in agricultural products and the environment, especially in developing countries, a simple and convenient means for OP detection and screening was investigated. Whole cells from the recombinant clone BpGP carrying methyl parathion (MP) degrading, mpdB, gene originating from MP-degrading Burkholderia cepacia and possessing MP-degrading activity were immobilized onto the polystyrene surface of the wells of microtiter plates using adsorption method, followed by cross-linking with glutaraldehyde. The immobilized cells were used as a bio-component for spectrophotometric detection of OP compounds, with MP as a representative insecticide. Scanning electron microscope and cell count analyses revealed highly dense, rod shaped bacterial cells stably attached to the well surface. The system was most stable at 25°C and could be used to detect MP repetitively up to 46 rounds, although it was still active at about 45% until round 100. For MP analysis, the linear range of the system was 2-200 µg/mL and the detection limit was 2 µg/mL. The tests on spiked and real samples were also performed and the results revealed high detection efficiency. This immobilized cell microtiter plate system has a great potential to be utilized as an alternative for OP detection and screening with the advantages of simplicity, stability, ease and convenience for assembly and use, and low cost.
Highlights
Organophosphate (OP) compounds, a group of neurotoxic substances, have been extensively used as pesticides and insecticides in agriculture worldwide, especially in many developing countries
The mpdB gene was expressed in E. coli as methyl parathion hydrolase (MPH) that fused with GST (MPH-GST)
The cells used for immobilization were from the clone BpGP containing mpdB gene cloned into plasmid pGEX 6P-1 and transformed into E. coli BL21
Summary
Organophosphate (OP) compounds, a group of neurotoxic substances, have been extensively used as pesticides and insecticides in agriculture worldwide, especially in many developing countries. These substances have been used as chemical warfare agents as well. OP insecticides exert their toxicity by inhibiting acetylcholinesterase (AChE) which hydrolyses a neurotransmitter acetylcholine, resulting in its accumulation in the synapses of the nervous system (Abou-Donia, 2003) This causes a wide range of OP toxicity symptoms involving nervous system disorder, teratogenicity, carcinogenicity, impairment of the reproductive system and detrimental impact on biological macromolecules (Ahmadizad Firozjaei et al, 2015; Kumar et al, 2016). Due to high threats to human health and the ecosystem resulting from OP contamination in food products and the environment, sensing devices that are simple, rapid and sensitive are required for effective control and monitoring of these substances
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