Development of a handheld PEST-pen device for the rapid detection of organophosphate insecticides on food and clothing

ABSTRACT The neurotoxicity of OP pesticides in humans may result from dermal or inhalation exposure on clothing or machinery or orally from the residue on foodIn the latter contextthe health consequences associated with the use of organophosphateOPpesticidesespecially in childrencould be reduced by monitoring produce and eliminating the consumption of OP contaminated foodPlantVax currently employs the Nicotiana benthamiana plant expression system to produce high levels of recombinant human acetylcholinesteraserHuAChEwhich is used in devices eg the FLIR CADpenfor the detection of trace amounts of organophosphateOPnerve agents for military purposesIn this proposalPlantVax aims to develop a simplifiedhand held inexpensive PEST pen to rapidly detect phosphorothionate insecticides on foodclothing and machineryWhile the new device will utilize the same esterase based embedded chemistry as the CADpen with a yellow to red color as a read outimportant features need to be altered to increase the rate of color change efficacy of the new deviceFirstlythe inhibition constantskiobtained for carboxylasesCESare higher for insecticides than HuAChE and will initially be incorporated into the pen spongeSecondly since phosphorothionate insecticides are inactive and need to be metabolically converted by oxidative desulfuration into the active oxon form by cytochrome Pin the liver for their insecticidal actionthe PEST pen may need to integrate features of both thion oxidation to an oxon as well as CES AChE inhibition into a single deviceThe innovation in this proposal will be the identification of novel indicator molecules with high inhibition rates for incorporation into the spongeThese includeithe wildtype or GD mutant form of the carboxylesterase enzyme from the Australian blow fly Lucilia cuprinaLc Ewhich exhibits high kinetic parameters for thion forms oriimutant AChE enzyme with increased rates of inhibition by oxon embedded together with a Pin the presence of a cytochrome c reductaseNADPHto expedite conversion to the oxonformsBased on the design of the successful CADpenseveral PEST pen prototypes with different indicator molecules will be produced and tested within a year Project NarrativeWhile organophosphateOPinsecticides greatly increase yields of agricultural cropsthey can cause neurotoxicity and even death in humans as a result from dermal or inhalation exposure on clothing or orally from residue on foodIn order to eliminate consumption of OP contaminated foodPlantVax plans to develop a simplehand heldinexpensive device called a PEST pen to monitor and rapidly detect OP insecticides on foodclothing and machinery