Optimization of A Bioassay Method for Specific Activity of Acetylcholinesterase of B Biotype Bemisia tabaci (Hemiptera: Aleyrodidae)
Abstract
The whitefly Bemisia tabaci (Gennadius), a major pest of protected and outdoor crops worldwide, has developed resistance to a wide range of insecticides including organophosphates (OPs) and carbamates (CBs). The biochemical target of OPs and CBs is acetylcholinesterase (AChE). The objective of this study was to optimize AChE bioassay conditions including enzyme concentration, substrate concentration, pH, temperature and reaction time in order to quickly and accurately monitor specific activity of AChE and monitor resistance of B. tabaci to insecticides. The results of an orthogonal array test showed that the optimum combination of levels was A3B3C4D5E1 in which the enzyme concentration was 20 adults/mL, substrate concentration was 7.0 × 10-4 mol/L, pH was 7.5, temperature was 45 °C, and reaction time was 5 min. The specific activity of AChE from B Biotype B. tabaci was 41.23 nmol · min-1 · mg-1 under this set of conditions. This optimum combination proved to be reliable and accurate for testing the specific activity of AChE.La mosca blanca Bemisia tabaci (Gennadius), una plaga importante en cultivos sembrados en lugares protegidos y en el campo en todo el mundo, ha desarrollado resistencia a una amplia gama de insecticidas, incluyendo organofosfatos (OF) y carbamates (CBs). El enfoque bioquimico de los OFs y carbamates es acetilcolinesterasa (AChE). El objetivo de este estudio fue el optimizar las condiciones del bioensayo de AChE incluyendo la concentración de enzima, la concentración de sustrato, pH, temperatura y tiempo de reacción. Los resultados de una prueba de matriz ortogonal mostró que la combinación óptima de los niveles fue A3B3C4D5E1 en el que la concentración de enzima fue 20 adultos /ml, concentracién de sustrato de 7.0 × 10-4 mol/L, pH de 7.5, temperatura de 45 °C y la reacción tiempo de 5 min. La actividad específica de AChE en el biotipo B de B. tabaci fue 41.23 nmol · min-1 · mg-1 bajo estas condiciones. Esta combinación óptima demostró ser fiable y precisa para probar la actividad especifica de AChE.
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