When treating the 3rd instar larvae of the diamondback moth (DBM), Plutella xylostella, with sublethal doses (LC10) of chlorantraniliprole, indoxacarb and spinosad via leaf dipping, their tolerance to insecticides was significantly enhanced. By analyzing the differentially expressed genes (DEGs), we found a number of genes that respond commonly or specifically to the test insecticides. With the criteria of p value < 0.05 and Log2FC > 1/ < -1, a total of 476, 367 and 410 genes were determined to respond specifically to chlorantraniliprole, indoxacarb and spinosad, respectively. Gene Ontology (GO) analysis revealed that the cuticle reorganization is commonly associated in all treatments and the oxidative stress-related process is also shown in all insecticides except spinosad. Finally, the DEGs seemingly related with enhanced tolerance were chosen for further characterization, and reliability of the transcriptome data were confirmed by quantitative PCR. The functional categories of these DEGs included mostly detoxification related genes, cuticle proteins, energy metabolism and transcriptional regulation. While the commonly responding DEGs suggest that they are likely involved in defense against common intoxication process, the DEGs specifically responded to each insecticide suggests the presence of unique tolerance mechanisms to each insecticide depending on their different structure and mode of action. Their possible roles in the tolerance/resistance development were discussed.

RNA interference (RNAi)-based strategy has been developed to control various phytophagous chewing pests. However, only a few cases of RNAi-based control success have been reported for sucking pests, suggesting that sucking pests likely ingest less amount of transgenic subcellular hairpin RNA (or dsRNA). In this study, as the basic information for the establishment of ingestion RNAi against sucking pests, feeding amount and time course of plant subcellular fractions of the four sucking pest species (Frankliniella occidentalis, Frankliniella intonsa, Tetranychus urticae and Nilaparvata lugans) were determined by quantitative PCR (qPCR). Adults of the four species were starved for 24 h and then fed with kidney bean leaf (F. occidentalis, F. intonsa, T. urticae) or rice leaf (N. lugens) for 48 h. The leaf-fed adults were collected every 6-h interval and their genomic DNA was extracted. The ingested fractions of chloroplast and nuclear were quantified using rubisco and 50s rRNA as marker genes, respectively. The ingested amount of rubisco and 50s rRNA genes in F. occidentalis, F. intonsa and T. urticae showed rapid increasing pattern after feeding and then slightly reduced over time. In contrast, N. lugens neither ingest nuclear nor showed any distinct feeding pattern of chloroplast. These results demonstrate that F. occidentalis, F. intonsa and T. urticae ingest both chloroplasts and nucleus along with cytosol as cell-feeders but N. lugens, a phloem sap feeder, does not ingest nucleus during sucking. Our findings further suggest that ingestion RNAi-based control strategy would work better for cell-feeding sucking pests compared to phloem sap-feeding sucking pests.

To identify genes that commonly respond to the treatment of different insecticides and are responsible for the toleranceenhancement, transcriptomic profiles of larvae treated with sublethal doses of the five insecticides were compared withthat of untreated control. A total of 117,181 transcripts with a mean length of 662 bp were generated by de novo assembly,of which 35,329 transcripts were annotated. Among them, 125, 143, 182, 215 and 149 transcripts were determined tobe up-regulated whereas 67, 45, 60, 60 and 38 genes were down-regulated following treatments with these five insecticides.The most notable examples of commonly responding over-transcribed genes were two cytochrome P450 genes and ninecuticular protein genes. In contrast, several genes composing the mitochondrial energy generation system were significantlydown-regulated in all treated larvae. Considering the distinct structure and mode of action of the five insecticides tested,the differentially expressed genes identified in this study appear to be involved in general chemical defense at the initialstage of intoxication. Their possible roles in the tolerance/resistance development were discussed.

To identify genes that commonly respond to the treatment of different insecticides, 3rd instar larvae of the diamondback moth, Plutella xylostella, were treated with sublethal doses (<LC10) of chlorantraniliprole, cypermethrin, dinotefuran, indoxacarb and spinosad via leaf dipping. Then, transcriptomic profiles of treated larvae were compared with that of untreated control. A total of 117,181 transcripts in average with a mean length of 662 bp were generated by de novo assembly, of which 35,329 transcripts were annotated. Among them, 207, 153, 336, 360, and 262 transcripts were determined to be up-regulated whereas 117, 47, 92, 115, and 81 genes were down-regulated following treatments with chlorantraniliprole, cypermethrin, dinotefuran, indoxacarb and spinosad, respectively. Finally, with the criteria of >10 X fold change (FC) and p < 0.05 or >4 X FC, p < 0.05 and q < 0.2, the genes commonly over-transcribed in all treated insects were selected and their over-transcription levels were confirmed by quantitative PCR. These commonly responding genes included three cytochrome P450 genes (Cyp303a1, Cyp6a20 and CYP9E2), three cuticle protein genes (LM-8, LM-19 and TM-A3A), lavesin-1, acyl-CoA D11 desaturase, glucose dehydrogenase, nose resistant to fluxetine protein 6, chorion peroxidase and protein yellow. As the five test insecticides have distinct structure and mode of action, the genes identified in this study were suggested to be involved in general chemical defense at the initial stage of intoxication. Their possible roles in tolerance and resistance development were further discussed.

Acetylcholinesterase (AChE) is an enzyme for hydrolyzing neurotransmitter acetylcholine. Soluble form of AChE is generated via alternative splicing and functions as a bioscavenger in Dropsophila melanogaster. In this study, effects of ethanol and acetic acid on the soluble AChE expression were investigated. Treatment of ethanol and acetic acid results in over-expression of soluble AChE in the abdomen in a dose-dependent manner. However, no apparent change in AChE expression was observed in the head. Our finding suggests that the soluble AChE is involved in chemical defense against high concentration of ethanol, which is a common by-product from fermented food，and acetic acid, the main metabolite of ethanol. Thus, high level of ethanol and acetic acid resistance in D. melanogaster appears to be evolved via the induction mechanism of soluble AChE expression.

Acetylcholinesterase (AChE) is a hydrolase that hydrolyzes the neurotransmitter acetylcholine. Soluble form of AChE is generated via alternative splicing and functions as a bioscavenger in Dropsophila melanogaster. In this study, effects of acetic acid on the soluble AChE expression were investigated. Treatment of acetic acid resulted in over-expression of soluble AChE in the abdomen in a dose-dependent manner. The soluble AChE was determined to be expressed in the fat body. However, no apparent change in AChE expression was observed in the head. Our finding suggests that the soluble AChE is involved in chemical defense against high concentration of acetic acid, which is a common by-product in fermenting foods. The high level of acetic acid resistance in D. melanogaster, thus, appears to have been evolved via the induction mechanism of soluble AChE expression.