Two acetylcholinesterase genes, Acel and Ace2, have been fully cloned and sequenced from both oranophoshate-resistant and susceptible clones of cotton aphid. Comparison of both nucleic acid and deduced amino acid sequences revealed considerable nucleotide polymorphisms. Further study found that two mutations occurred consistently in all resistant aphids. The mutation F139L in Ace2 corresponding to F115S in Drosophila acetylcholinesterase might reduce the enzyme sensitivity and result in insecticide resistance. The other mutation A320S in Acel abutting the conserved catalytic triad might affect the activity and insecticide sensitivity of the enzyme. Phylogenetic analysis showed that insect acetylcholinesterases fall into two subgroups, of which Acel is the paralogous gene whereas Ace2 is the orthologous gene of Drosophila AChE. Both subgroups contain resistance-associated AChE genes. To avoid confusion in the future work, a nomenclature of insect AChE is also suggested in the paper.

gossypii Glover. The procedure involved filtration on a sephadex G-25 column, separation with sephadex G-200 and procainamide offinity column. AChE from both susceptible and resistant strains were purified to a single band as resolved on denaturing polyacrylamide gel electrophoresis (SDS-PAGE). The specific activity increased by 35, 100- and 33, 680-fold with a yield of 30.3 and 29.8%, respectively. The molecular mass of the purified AChE was about 63, 500 Dalton as etermined by SDS-PAGE. However, three bands resolved on PAGE gel electrophoresis, leading to the inference that native AChE exists in three forms. The optimum conditions for measuring the activity of purified AChE with kinetic method were 0.02M phosphate buffer, pH7.2, 0.02mM 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB), AND 25℃. Investigation also revealed that crude extract and purified AChE had different kinetic characteristics and inhibiroty properties. They responded differently to varied DTNB, ATChI, and phosphate buffer ion concentrations, as well as pH, temperature, and inhibitors. The purified AChE was more sensitive to eserine, methamidophos, and pirimicarb. Especially for resistant aphids, the sensitivity of purified AChE to methamidophos and pirimicarb was enhanced 6.43 and 11.73 times, respectively. We infer that one or more factors in the crude extract from the resistance stain have more influence on AChE sensitivity. Further study is needed to investigate the basis of these observations.

Insensitive acetylcholinesterase was identified as a esistance mechanism by comparing biochemical analysis with a laboratory selected monocrotophos resistant cotton bloodworm (RR: 200) and the susceptible strain The cDNA enCODING AChE was cloned by the method of RACE (rapid amplification of cDNA ends). The complete AChE gene deduced from the cDNA consisted of a putative signal peptide of 32 amino acid residues, a mature protein of 615 residues, 5' untranslated regions (UTR) of 315bp and 3' UTR of 324bp. The Coding sequence had a high degree of Homology to the AChE from other insect species reported in the GenBank. After comparing analysis of the entire AChE gene sequence from 5 resistant and 6 susceptible cotton bollworm individuals, nine mutations were identified. One of them, the Ala/Thr mutation, is likely to be responsible for the AChE insensitivity to monocrotophos.

The activity and electrophoretic mobility of nonspecific esterases, and the sensitivity of acetylcholinesterase (AChE) to organophosphate and carbamate insecticides, was determined for 21 strains or clones of Aphis gossypii of worldwide origin and related to patterns of insecticide resistance disclosed by diagnostic dose bioassays with demeton-S-methyl, pirimicarb, and permethrin. Results confirmed previous work showing the occurrence of two forms of mutant AChE, both conferring insensitivity to pirimicarb and representing the primary mechanism of resistance to this insecticide. One of these mutant AChE enzymes also caused resistance to demeton-S-methyl in its own right, whereas the other only conferred resistance in combination with moderate or high levels of nonspecific esterase activity. Lack of any correlation between permethrin resistance and quantitative or qualitative changes in esterases implied a distinct, as yet uncharacterised, mechanism of resistance to pyrethroids, possibly based on insensitivity in the target protein for these pesticides.

Dynamics of pyrethroid resistance in a field population of cotton bollworm (Helicoverpa armigera) was demonstrated by continuous monitoring with twin discriminating dosages, and the influencing factors were also experimentally analysed. Resistance in a field population in China increased rapidly in the 3rd and 4th generations when population density became higher and insecticides were applied repeatedly, then decreased suddenly during over-wlntering and slowly in the 1st and 2nd generations when insecticide spraying was suspended. Resistance increase could be countered by dilution as a result of immigration of susceptible moths from corn fields, which were found to be a natural refuge for this pest in China, The reduction of resistance during over-wintering and the 1st and 2nd generations was affected by the lower fitness of resistant cotton bollworms to low temperature and disadvantages in reproduction. The possibilities of managing the resistance in field populations on the basis of these observations are discussed.