Effects Of Diallyl Trisulfide On Chitin Structural Chemistry And Chitin Synthase A Of Sitotroga Cerealella Olivier(Lepidoptera:Gelechiidae)

The Angoumois grain moth,Sitotroga cerealella Olivier?Lepidoptera:Gelechiidae?,is the most severe and destructive pest of stored commodities around the globe,particularly on wheat,rice and maize.Although broad spectrum insecticides are currently effective in controlling this devastating pest,however the indiscriminate heavy use leads to development of pesticide resistance,environmental pollution,and public health hazards,and also contributes to climate change.Therefore,it is very urgent to find an alternative to conventional chemical insecticides in order to control the severe moths.Previously,we confirmed that diallyl trisulfide?DAT?,an active substance from garlic essential oil,reduced oviposition of females after a fumigation at the sublethal concentration(LC₂₀=0.015?L/L);meanwhile,through transcriptome sequencing,we found that DAT significantly downregulates the expression of chitin synthase A?CHSA?gene.To gain a broader insight into the relationship between the moth reproduction and CHSA gene,we assessed the effects of DAT and ds RNA of CHSA gene of S.cerealella?ds Scer CHSA?on the morphology,physiology,and oviposition of adults S.cerealella.Meanwhile,we characterized and evaluated the effect of DAT on the whole content and structural chemistry of chitin of adults S.cerealella,studied by Fourier-transform infrared spectroscopy?FTIR?,thermogravimetric analysis?TGA?,X-ray diffraction?XRD?,differential scanning calorimetry?DSC?,and scanning electron microscopy analysis?SEM?in order to identify the type of chitin,analyze the chemical elements,thermal stability,crystallinity,and morphology of moth chitin.The study revealed that:1)S.cerealella chitin synthase A?Scer CHSA?was expressed in the adult stage and observed higher in the cuticle compared with head and fat bodies.The results indicated that DAT fumigation significantly downregulated the m RNA expression of Scer CHSA at 12h,18h and 24h,and inhibited moth oviposition at the sublethal concentration;Moreover,silencing of Scer CHSA through RNAi resulted in the oviposition reduction?ds Scer CHSA=59,ds GFP=98;P<0.001?and mortality of the moth?ds Scer CHSA=60%,ds GFP=16.6%;P<0.001?.All results mean that Scer CHSA could possess the characteristics to regulate the reproduction of moths.2)DAT-treated and ds Scer CHSA-injected groups showed morphological alteration compared with control.The hardness and arrangement of the integuments and cuticle structure were thinner and damaged compared with control groups.Moreover,all samples from the DAT and ds Scer CHSA groups showed a discontinuous,shrink and translucent cuticle that was thinner and apart from the epidermis,altered and disrupted inner fat bodies tissues.Furthermore,the morphological and histological alterations in the cuticle were similar to the group treated with DAT.3)The chitin content of male and female moths in controls was found to contain 18- 20%chitin per body weight.There was no significant difference observed between the amount of chitin extracted from control males and females.In contrast, significantly less chitin?8-10%?was extracted from the DAT-treated groups compared with their respective controls?P<0.001?.4)Thea-form of chitin was confirmed and the effects of DAT on the structure of pest insects were performed that the thermal stability and crystallinity of chitin were significantly reduced compared with controls by FTIR,TGA,XRD,and DSC. (LC₂₀=0.015?L/L)could downregulate the expression of Scer CHSA at the transcriptional level,decrease the cuticular chitin content,and cause histological and morphological alteration of the chitin,and finally inhibit the moth oviposition.All results prove that DAT possesses the great potential for being used in pest control as a friendly bio-pesticide and establishes the basis for its use as an alternative to chitin synthesis inhibitors.