Response Mechanisms Of Tribolium Castaneum And Callosobruchus Maculatus Exposure To Physical Agents

Ultraviolet(UV), hypoxia, electron beam(e Beam) are important physical stress agents which could reduce insect fitness. These agents are usually used to control pests. Insects can respond to these physical stress agents, which refers to gene expressions, protein translations, and morphological changes. At last, insects will tolerate physical stresses. Study on strategies of insect response to physical agents could help us understand the mechanism of insects adapt the pest physical control methods. In this study, we used Tribolium castaneum(Herbst) and Callosobruchus maculatus(Fabricius) as targets. We studied on the response mechanisms of T. castaneum exposure to UV and the response mechanisms of C. maculatus exposure to hypoxia and e Beam, respectively. The results are below:1. UVA stress induces expressions of heat shock protein genes(Hsps) and cytochrome P450 enzyme genes(P450s) in T. castaneum adults.In this study, it was found that an intensity of 1000 μw/cm2 UVA exposure could induce expressions of heat shock protein genes(Hsp27、Hsp68、Hsp83) and cytochrome P450 enzyme genes(CYP6BQ4、CYP6BQ8). The three of Hsps were up-regulated by 4.1-8.9 folds(P < 0.05). The two of P450 s were up-regulated by 2.2 and 2.9 folds(P < 0.05). The expressions of these genes were induced at first 2 h of UVA exposure and then reduced, which showed similar pattern. To analyze the regulatory regions of these induced genes, it was found that they all have common transcription factor binding sites, such as HSF、KROX, NF-kappa B, AFB2, and dl etc. These transcription factors may regulate transcriptions of Hsps and P450 s to respond to UVA stress in T. castaneum.2. UVB stress inhibits ecdysteroid biosynthesis and delays metamorphosis of final instar larvae in T. castaneumIn this study, it was found that a dose of 20 k J/m2 UVB exposure could cause 100 % death of final instar larvae in T. castaneum. A dose of 10 k J/m2 UVB exposure could significantly reduce emergence rate to 61.1%(P < 0.001), pupal length by 0.36 mm and pupal weight by 0.48 mg(P < 0.001), respectively. UVB irradiation could delay larval-pupal metamorphosis of larvae. Insect metamorphosis is critically managed by ecdysteroid signaling. Therefore, in this study, expressions of key genes in ecdysteroid pathway were analyzed. It was found that the expression profiles of ecdysteroidogenesis genes(Trcptth, sad, and shd) and response genes(Ec R and Br) were down-regulated by 5.4、3.6、2.7、2.9, and 5.8 folds after UVB exposure. In addition, UVB disturbed pulse of ecdysteroid signaling, which triggers metamorphosis of larvae.3. Stress response, nervous regulation, energy metabolism, and damage repair etc. are involved in response to UVB stress in final instar larvae in T. castaneumDigital gene expression profiling method was used to analyze the differentially expressed genes of T. castaneum at 4 h and 24 h post a dose of 10 k J/m2 UVB irradiation, respectively. Hundreds of differentially expressed genes were identified under UVB stress. Four hours after UVB exposure, there were 222 genes up-regulated and 88 genes down-regulated. Twenty-four hours after UVB exposure, there were 544 genes up-regulated and 452 genes down-regulated. After GO enrichment analysis, the results showed that four hours after UVB exposure, peptidoglycan metabolic process genes, glycosaminoglycan catabolic process genes, peptidoglycan catabolic process genes, response to stress genes, aminoglycan catabolic process genes were significantly enriched(P < 0.001). Twenty-four hours after UVB exposure, organic substance catabolic process genes, proteolysis genes, catabolic process genes, carbohydrate derivative catabolic process genes, and macromolecule catabolic process genes were very significantly enriched(P < 0.0001). To analyze the pathways of differentially expressed genes, it was found that four hours after UVB exposure, tyrosine metabolism and Wnt signaling pathway were activated; Twenty-four hours after UVB exposure, protein processing in endoplasmic reticulum, glycolysis/gluconeogenesis, ECM-receptor interaction, and proteasome pathways were activated. These results indicated with the time post UVB exposure increasing, the much more genes participated in UVB stress response in larvae of T. castaneum. At early stage post UVB exposure(4 h), stress response and nervous regulation were involved in response to UVB stress. At later stage(24 h), the larvae could manage energy metabolism, control organ formation, repair DNA lesions, and induce proteolysis to respond to UVB stress.4. Hypoxia stress influences transferring from parental generation to offspring induces hypoxia resistance of offspring in C. maculatusIn this study, it was found that hypoxia treatment(2% O2+18% CO2+80% N2 mixture gases) had impacts on reproduction 4th instar larvae, pupae and adults of parental generation of C. maculatus. Hypoxia treated parental adults reduced fecundity per female by 21 eggs(P < 0.001). The hatch rate was reduced the most significantly by 9.9%(P <0.001) when pupae were treated by hypoxia. Hypoxia treated parents, which could induce hypoxia resistance of offspring in C. maculatus. When the 4th instar larvae of F1 generation were exposed to hypoxia treatment, which their parents suffered from, the hatch rates of eggs laid by survivor were not impacted(P > 0.05). Compared with F1 generation from hypoxia treated parents, the basal gene expressions of hypoxia response genes Cat L was significantly down-regulated by 2.8 folds(P < 0.05), and Hsp27 was significantly up-regulated by 1.6 folds(P < 0.05) in F1 generation from hypoxia untreated parents. These results indicated that hypoxia stress influences could transfer from parental generation to offspring, and induce hypoxia resistance of offspring in C. maculatus, which helps offspring overcome hypoxia stress.5. e Beam stress seriously harms to development and reproduction in C. maculatusIn this study, it was found that di ff erential radiosensitivity was detected during egg development in C. maculatus. Early and intermediate stages of eggs never hatched after exposure to a dose of 0.03 k Gy, whereas a substantial portion of black-headed(i.e. late) eggs survived irradiation even at 0.12 k Gy. However, further development of the hatched larvae was inhibited. Although midgut protein digestion remained intact, irradiated larvae(0.06 – 0.12 k Gy) failed to develop into normal living adults; rather, they died as pupae or abnormally eclosed adults, suggesting a detrimental e ff ect of e Beam on metamorphosis. Emerged irradiated pupae had shorter longevity and were unable to produce any eggs at 0.06 k Gy or 0.9 k Gy. At the dose range from 0.06 k Gy to 2 k Gy, eggs laid by irradiated adults were not viable. e Beam treatment shortened adult longevity in a dose-dependent manner. Reciprocal crosses indicated that females were more sensitive to e Beam exposure than their male counterparts. Dissection of the female reproductive system revealed that e Beam treatment prevented formation of oocytes.