Study On The Relationship Between P-glycoprotein And Abamectin Resistance In Tetranychus Cinnabarinus (Boisduval)

The Tetranychus cinnabarinus is the most harmful crop mite pests. Owing to its strong reproductive ability, short life cycle, limited movement area, high inbreeding rate and exposure rate to acaricides, it has quickly developed resistance to acaricides. Abamectin (ABM) is a 16-membered macrocyclic lactone agent and shows high insecticidal, nematicidal, and acaricidal activities.With the application of abamectin, different insect species has developed resistance to this agent. P-glycoprotein (Pgp), which belongs to the B subfamily of the ABC transporters, works as an ATP-dependent efflux pump, and plays a critical role in drug efflux and lipid transport in normal cells. Recently several reports showed that Pgp was involved in ABM resistance.Resistance caused by Pgp is also called multidrug resistance (MDR). This study foucuses on the relationship between P-glycoprotein and abamectin resistance in T. cinnabarinus. The main results obtained from this research are as follows:1. An abamectin resistant strain (AbR) of T. cinnabarinus was selected, and the resistance ratio was over 20-fold at LC50. After 12 h pretreatment with the Pgp specific inhibitor verapamil, the toxicity of abamectin to susceptible strain (SS) and AbR increased by 19.91% and 74.51%, respectively. The preliminary results showed that Pgp influenced more obviously on AbR than SS and was associated with resistance to abamectin.2. The affinity of Pgp ATPase was detected in SS and AbR. The Km and Vmax were 1.835 ± 0.142 mM,0.097 ± 0.036μmol/mg/min in SS, and 1.595 ± 0.353 mM,0.108 ± 0.054 μmol/mg/min in AbR. It showed that there was no significant difference between SS and AbR.3. The Pgp ATPase activity assay showed that the activity of Pgp ATPase in AbR was significantly higher than in SS. After exposure to LC30, activity of Pgp ATPase did not change much in SS, butsignicantly increased in AbR.4. Two putative Pgp genes, TcPgpl and TcPgp2, were cloned and characterized.The complete ORF of TcPgpl and TcPgp2 consisted of 3885 and 3879 nucleotides, which encoded 1295- and 1293- amino acid peptide, respectively. The deduced TcPgpl and TcPgp2 sequences all possessed conservative sites such as the Walker A and B motifs, the Signature C motif, and the D and H loops.5. Comparison of the amino acid sequences of these two genes between SS and AbR showed no difference. The mRNA levels of TcPgpl and TcPgp2 were detected by qPCR in eggs, larvae, nymphs and adults of SS. The results showed that TcPgpl expressed at a same level in all stages, however, TcPgp2 expressed significantly higher in eggs and larvae than nymphs and adults. Although the mRNA levels of TcPgpl and TcPgp2 between SS and AbR adult females showed no difference, after exposure to LC30, mRNA levels of TcPgpl and TcPgp2 did not change much in SS, but signicantly increased in AbR.