The resistance levels to carbamate (CB) and organophosphate (OP) insecticides were determined by topical application in 14 field strains of Nilaparvata lugens. The resistance levels to CB and OP were 1.3~47.5-fold and 1.4~14.4-fold higher than a susceptible strain, respectively. A quantitative sequencing (QS) protocol was established to determine the allele frequencies of four acetylcholinesterase point mutations putatively associated with CB and OP resistance. The allele frequencies of the four mutations (G119A, F/Y330S, F331H and I332L) in field strains ranged from ca. 0.0~51.7%, 0.0~88.9%, 2.5~47.7%, 5.1~56.0% and 6.7~57.3%, respectively. The F331H and I332L were tightly linked each other, suggesting these mutations may occur simultaneously. In the correlation analysis, G119A was not well correlated with actual resistance levels (r2= < 0.232), whereas F331H and I332L showed a better correlation with the resistance levels of benzofuranyl methylcarbamates (r2= 0.595). This finding indicates that F331H and I332L mutation frequencies may be used as molecular markers for detecting carbamate resistance in N. lugens. A QS protocol detecting the F331H and I332L mutation frequencies would be employed as a supportive tool for rapid monitoring of CB insecticide resistance levels in N. lugens

The resistance levels against carbamates (CB) and organophosphates (OP) were determined through bioassay and quantitative sequencing (QS) methods in 16 field populations of Nilaparvata lugens. The resistance levels to CB and OP by bioassay were 1.3~47.5-fold and 1.4~14.4-fold higher than a susceptible strain, respectively. The QS protocol was established to determine the allele frequencies of eight point mutations on acetylcholinesterase putatively associated CB and OP resistance. The allele frequencies of four mutations in local populations (G119A, F/S330Y, F331H and I332L) ranged from ca. 0.0~51.7%, 1.0~44.3%, 8.5~57.3% and 7.12~56.6%, respectively. The average prediction limits were –9.6±5.1~7.7±2.9%. The F330Y, F331H and I332L were tightly linked each other, suggesting these mutations may occur simultaneously. In the correlation analysis, G119A was not well correlated with both insecticides (r2= less 0.25), whereas F/S330Y, F331H and I332L showed better correlation with the resistance levels of carbamate (r2=0.590) than organophosphate (r2=0.235). This finding indicates that F/S330Y, F331H and I332L mutation frequencies are suitable for detecting carbamate resistance in N. lugens. QS will be applicable for the rapid monitoring of resistance levels to CB insecticides in N. lugens.

A quantitative sequencing (QS) protocol was established for predicting the frequencies of the A298S and G324A mutations in the diamondback moth (Plutella xylostella) type-1 acetylcholinesterase locus, putatively involved with organophosphate and carbamate insecticideresistance. The nucleotide signal ratio at each mutation site was generated from sequencing chromatograms and plotted against the corresponding resistance allele frequency. Frequency prediction equations were generatedfrom the plots by linear regression, and the signal ratios were shown to highly correlated with resistance allele frequencies (r2>0.987). QS analysis of 15 representative regional field populations of DBM in Korea revealed that the allele frequencies of both A298S and G324A were over 70% in most field populations. As judged by inhibition assay, all populations showed resistance to paraoxon, DDVP, carbaryl, and carbofuran. In addition, different DBM strains exhibited differential sensitivities to both OPs and CBs depending on the structure of inhibitor, implying that the resistance of DBM against OPs and CBs is saturated and widespread in Korea.