Genomic and proteomic studies on the effects of the insect growth regulator diflubenzuron in the model beetle species Tribolium castaneum

Abstract

SUMMARY Several benzoylphenyl urea-derived insecticides such as diflubenzuron (DFB, Dimilin®) are in wide use to control various insect pests. Although this class of compounds is known to disrupt molting and to affect chitin content, their precise mode of action is still not understood. To gain a broader insight into the mechanism underlying the insecticidal effects of benzoylphenyl urea compounds, we conducted a comprehensive study with the model beetle species and stored product pest Tribolium castaneum (red flour beetle) utilizing genomic and proteomic approaches. DFB was added to a wheat flour-based diet at various concentrations and fed to larvae and adults. We observed abortive molting, hatching defects and reduced chitin amounts in the larval cuticle, the peritrophic matrix and eggs. Electron microscopic examination of the larval cuticle revealed major structural changes and a loss of lamellate structure of the procuticle. We used a genomic tiling array for determining relative expression levels of about 11,000 genes predicted by the GLEAN algorithm. About 6% of all predicted genes were more than 2-fold up-or down-regulated in response to DFB treatment. Genes encoding enzymes involved in chitin metabolism were unexpectedly unaffected, but many genes encoding cuticle proteins were affected. In addition, several genes presumably involved in detoxification pathways were up-regulated. Comparative 2D gel electrophoresis of proteins extracted from the midgut revealed 388 protein spots, of which 7% were significantly affected in their levels by DFB treatment as determined by laser densitometry. Mass spectrometric identification revealed that UDP-N-acetylglucosamine pyrophosphorylase and glutathione synthetase were up-regulated. In summary, the red flour beetle turned out to be a good model organism for investigating the global effects of bioactive materials such as insect growth regulators and other insecticides. The results of this study recapitulate all of the different DFB-induced symptoms in a single model insect, which have been previously found in several different insect species, and further illustrate that DFB-treatment causes a wide range of effects at the molecular level.

Population bulk segregant mapping uncovers resistance mutations and the mode of action of a chitin synthesis inhibitor in arthropods

“…S1 B and C). These specific symptoms are nearly identical to those observed with benzoylureas in insects (12). To establish the effect of etoxazole on chitin deposition in mites, we treated synchronized female deutonymphs with etoxazole and collected the teleiochrysalis stage just before ecdysis.…”

Characterization of a Chitin Synthase Encoding Gene and Effect of Diflubenzuron in Soybean Aphid, Aphis Glycines

“…Earlier, it has been reported that CHS expression changes of up to two-fold can occur even among control insect samples 17. Further, results from previous studies do not reveal a consistent pattern on the effect of DFB and other Benzoylphenylureas on insect CHS expression 14, 17, 18, 43. DFB has been found to enhance the CHS1 expression in A.quadrimaculatus 14 but not in D. melanogaster 17 and T. casteneum 18.…”

Cloning, Expression and Enzymatic Characterization of Chitin Deacetylase 4 from Hyphantria Cunea (Drury)

“…The three main constituents of the cuticle are proteins, lipids and chitin, the latter chitin, a homopolymer comprising β-(1-4)-linked N-acetyl-D-glucosamine residues, is one of the most abundant, easily obtained and renewable natural polymers, second only to cellulose and it is commonly found in many invertebrates and in the cell walls of most fungi and algae [1][2][3]. In the last few years, research shows that the enzymes in chitin metabolism are the critical target for insecticides [4][5][6].…”