RESUMO
The acetylcholinesterase, carboxylesterase, and cytochrome P450 monooxygenase activities of three strains of Oryzaephilus srinamensis (L.) were examined to better understand biochemical mechanisms of resistance. The three strains were VOS49 and VOSCM, selected for resistance to malathion and chlorpyrifos-methyl, respectively, and VOS48, a standard susceptible strain. Cross-resistance to malathion and chlorpyrifos-methyl was confirmed in VOS49 and VOSCM. Acetylcholinesterase activity was not correlated to resistance among these strains. VOS49 and VOSCM showed elevated levels of carboxylesterase activity based on p-nitrophenylacetate, alpha-naphthyl acetate, or beta-naphthyl acetate substrates. PAGE zymograms showed major differences in caboxylesterase isozyme banding among strains. VOSCM had one strongly staining isozyme band. A band having the same Rf-value was very faint in VOS48. The VOS49 carboxylesterase banding pattern was different from both VOSCM and VOS48. Cytochrome P450 monooxygenase activity was based on cytochrome P450 content, aldrin epoxidase activity, and oxidation of organophosphate insecticides, all elevated in resistant strains. The monooxygenase activity varied with insecticide substrate and resistant strain, suggesting specific cytochromes P450 may exist for different insecticides. The monooxygenase activity of the VOS49 strain was much higher with malathion than chlorpyrifos-methyl as substrates, whereas VOSCM monooxygenase activity was higher with malathion than chlorpyrifos-methyl as substrates. Results are discussed in the context of resistance mechanisms to organophosphate insecticides in O. surinamensis.
Assuntos
Clorpirifos/análogos & derivados , Clorpirifos/metabolismo , Inibidores da Colinesterase/metabolismo , Besouros/enzimologia , Inseticidas/metabolismo , Malation/metabolismo , Acetilcolinesterase/metabolismo , Animais , Carboxilesterase , Hidrolases de Éster Carboxílico/metabolismo , Clorpirifos/farmacologia , Inibidores da Colinesterase/farmacologia , Besouros/efeitos dos fármacos , Sistema Enzimático do Citocromo P-450/metabolismo , Fenitrotion/metabolismo , Fenitrotion/farmacologia , Resistência a Inseticidas , Inseticidas/farmacologia , Dose Letal Mediana , Malation/farmacologiaRESUMO
Esterases from a fenitrothion-resistant strain (VOSF) of the saw-toothed grain beetle, Oryzaephilus surinamensis (L.), are presumed to play a role in conferring resistance to malathion, fenitrothion, and chlorpyrifos-methyl. Colorimetric assays showed a significant positive correlation between increased resistance to fenitrothion in strains of O. surinamensis examined and elevated esterase hydrolytic activity to substrates of p-nitrophenyl acetate, alpha-naphthyl acetate, and beta-naphthyl acetate. Esterase zymograms showed different banding patterns between VOSF and an insecticide-susceptible strain, VOS48. A major esterase in the VOSF strain, not detected in VOS48, was purified and characterized by chromatographic and electrophoretic techniques. On the basis of SDS-polyacrylamide gel eletrophoresis, the molecular mass of the purified esterase from VOSF was 130 kDa and consisted of two 65 kDa subunits. Additional properties of this enzyme are discussed.