Resistance Selection To Malathion And The Mechanisms Of Metabolic Resistance In Bactrocera Dorsalis (Hendel)

The biochemical mechanisms of metabolic resistance of Bactrocera dorsalis (Hendel) to malathion were investigated on the basis of resistance selection in laboratory experiment with a series of techniques such as bioassay, resistance selection, biochemical analysis, and molecular biology. This research would help to completely understand the physiological and biochemical mechanisms of organophosphorus insecticide resistance of the pests in the future, and also possess great practical significance in the formulation of strategies for the delay and management of resistance. The expression profiles of11detoxification enzyme genes in various development al stages and different tissues from the resistant and susceptible strains of B. dorsalis were determined using qPCR method. The results offer the initial evidence to resolve the molecular resistance mechanism of B. dorsalis, and provide strong theoretical basis for the comprehensive management of the oriental fruit fly. The main results are as follows:1Resistance selection of B. dorsalis to malathionThe resistance to malathion of B. dorsalis was selected in the laboratory environment. After discontinuous selections with25generations followed by continuous selections with13generations, B. dorsalis have developed21.1-fold resistance to malathion. Bioassay revealed that the malathion resistant strain obtained15.80-,14.14-,4.14-,2.75-,2.48-fold cross-resistant to phoxim, chlorpyrifos, beta-cypermethrin, abamectin and fipronil, respectively.2The biochemical mechanisms of metabolic resistance of Bactrocera dorsalis (Hendel) to malathion2.1The activities of CarEs, GSTs and P450s in different developmental stages of B. dorsalisCompared to the susceptible strain of B. dorsalis, the activities of CarEs, GSTs and P450s increased significantly in different developmental stages from the resistant strain. It suggested that the development of resistance to malathion might be related to the rise of metabolic enzyme activities. The contribution of the detoxification enzyme in resistance is affiliated with the developmental stage. CarEs play a leading role in the resistance of the larve of B. dorsalis to malathion, GSTs is the biggest contributor to the resistance to malathion in pupa stage, however, P450s play a leading role in the resistance of the adults of B. dorsalis to malathion.2.2The activities of CarEs, GSTs and P450s in different tissues of adults of B. dorsalisThe activities of CarEs, GSTs and P450s in different tissues of adults from the resistant strain were significantly higher than those of the susceptible strain. The degrees of variance among different strains are different in different tissues. The activities of CarEs and GSTs (4.33-and1.63-fold) increased the most in fatbody, the activity of P450s (1.63-fold) increased the most in Malpighian tubules. It suggested that increased activity of CarEs and GSTs in fatbody and the increased activity of P450s in Malpighian tubules induce the resistance.2.3The activities of CarEs, GSTs and P450s in different tissues of the third larva of B. dorsalisExcept for the activity of CarEs in Malpighian tubules, the activities of these enzymes in different tissues of the third larve from the resistant strain were significantly higher than those of the susceptible strain. The activities of CarEs and GSTs (2.65-and1.44-fold) increased the most in Malpighian tubules, the activity of P450s (1.71-fold) increased the most in fatbody. It suggested that increased activity of CarEs and GSTs in Malpighian tubules and the increased activity of P450s in fatbody induce the resistance.3The expression profiles of11detoxification genes from different strains of B. dorsalisIn order to verify whether the11detoxification genes are over-expressed in resistant fruit flies,#PCR with a-Tubulin as the reference gene was employed to determine the relative expression quantity of11genes in different developmental stages from the resistant and susceptible strains of B. dorsalis. As indicated by qPCR,7genes (a-E3、BdGSTd1、BdGSTd2、BdGSTd5、CYP4D46、CYP6A41and CYP6D9) over-expressed in the resistant strain,5genes(α-E5、BdGSTd2、BdGSTd7、CYP4P5and CYP6D9) over-expressed in pupa in the resistant strain,8genes (BdGSTd1、BdGSTd2、 BdGSTd5、CYP4D46、CYP4P5、CYP6A41and CYP6D9) over-expressed in the resistant strain. It suggested that over-expression of these genes may induce the resistance.We also analyzed the expression model of detoxification genes in different tissues of the adults from the resistant and susceptible strains of B. dorsalis. In Malpighian tubules of the resistant strain, BdGSTd2and CYP4D46over-expressed. In fat body of the resistant strain,6genes (CYP4D46、CYP6A41、CYP6D9、BdGSTdl、BdGSTd2and BdGSTd5) over-expressed. In midgut of the resistant strain,4genes(α-E3、 CYP6A41、BdGSTd2and BdGSTd7) over-expressed. It suggested that over-expression of these genes may play an important role in the evolvement of resistance to malathion in B. dorsalis.