| The soybean aphid, Aphis glycines Matsumura(Hemiptera:Ahpididae), is an economically damaging pest in Asia and North Americawhere it is invasive and causes yield losses of the soybean Glycine max(L.). In addition to direct damage caused by feeding, soybean aphids can transmit plant viruses, reduce pod set, support the growth of sooty mold through excreted honeydew, indirectly distort growth and further reduce seed yield in soybean.Lambda-cyhalothrin, as a kind of effective pesticide long-term application of prevention and treatment to soybean aphid, which has produced resistance to it. The mechanisms of insect resistance include the reduction of cuticular penetration, the enhancement of detoxification metabolism and reduction of sensitivity of targets. The last two are the main mechanisms.Taking the soybean aphid as the research object, and revealing its resistant mechanism to lambda-cyhalothrin by the biochemistry, molecular biology and comparative proteomic methods, are of the practical singnificance of management to prevent and treat the soybean aphid harm.The same genetic background sensitive(CSS) and resistant(CRR) strain of soybean aphid was established in the laboratory, as the effect of continuous selection with lambda-cyhalothrin,the resistance ratio of CSS and CRR strain is 43.42-fold. Cross-resistance spectrum of CRR strain with other insecticides was established, so that the experiment basis for the rational use of pesticide was provided in the field. The synergism adding the enzyme inhibitors to the CRR strain, the adding fuctions of synergist which help the soybean aphid to resist the lambda-cyhalothrin was studied. Basing on the stable CRR and CSS strains, the specific activities and correlation parameters of two carboxylesterase were analyzed and compared; The changes of the CRR strain carboxylesterase was revealed on the m RNA transcription level through the q RT-PCR. The results showed that carboxylesterase excessive expression was associated with the resistance and the esterase activity and the cytochrome P450 enzyme activity was the main mechanism of the resistant. Through the analysis of the expression of cytochrome P450 quantity has been cleared that the increasing amount of cytochrome P450 enzyme expression was an important factor on Soybean aphid resistant to lambda-cyhalothrin. Basing on its genetic IIS4- S6 cloning and sequence analysis, the genes about sodium channel relevant to the Soybean aphid resistance as the pyrethrins insecticide targetsin-depth exploration were discussed. On this basis of Proteomic technology analysis of differences in protein expression with the CSS and the CRR strain, the resistance response protein was analyzed.The results of cross-resistance of CRR strain with other insecticides revealed that the resistant strain showed a medium cross-resistance with chlorpyrifos(11.66-fold),a lower cross-resistance with acephate(8.20-fold),a medium cross-resistance with esfenvalerate(13.83-fold),a medium cross-resistance with cyfluthrin(9.64-fold),a higher cross-resistance with cypermethrin(37.23-fold),a lower cross-resistance with bifenthrin(4.81-fold)a medium cross-resistance with methomyl(9.32-fold),a medium cross-resistance with carbofuran(14.60-fold),scarely cross-resistance with insecticides from chlorfenapyr, imidacloprid, acetamiprid, Diafenthiuron and abamectin. Synergist results revealed that adding the TPP DEF PBO to the lambda-cyhalothrin, the CRR strain synergistic coefficient reached 5.85, 23.00 and 40.59 separately; for the CSS strain Synergistic coefficient reached 0.26, 0.35 and 3.00 separately,the results showed that the synergistic agent has certain inhibition confrontational.Esterase kinetics determination results indicated that the comparation the CRR strain with the CSS the ratio of the carboxylesterase activity was 1.405-fold, that activities of carboxylesterases were significantly greater in the resistant strain than that in the susceptible strain( P< 0.01). Analysis of the soybean aphid rboxylesterase expressed quantitily. Using the q RT- PCR technology the CRR strain with the CSS was 5.87-fold, the results showed that the resistant strains and sensitive strains carboxylesterase gene m RNA transcription had significantly difference. In addition, soybean aphid cytochrome P450 oxidase gene were determined in the CRR strain CYP6A13-like, CYP6A2-like,CYP6A14-like and Cytochrome b-c1 over expression. Sodium channel gene IIS4, IIS5, iis 6 cloning and sequencing, nucleotide sequence contains the kdr and super – kdr site. If the soybean aphid resistant strain sodium channel gene site mutation happened in the sequences corresponding, the results illustrated that the two sites have relationship with soybean aphid kdr and super-kdr resistant to lambda-cyhalothrin, lay a theoretical foundation for studying the sodium channel Related to the soybean aphid resistant to lambda-cyhalothrin.Lambda-cyhalothrin is now widely used in China to control the soybean aphid Aphis glycines. To dissect the resistance mechanism, a laboratory-selected resistant soybean aphid strain(CRR) was established with a 43.42-fold resistance ratio to ?-cyhalothrin than the susceptible strain(CSS) in adult aphids. In this study, a comparative proteomic analysis between the CRR and CSS strains revealed the molecular resistance mechanism of soybean aphids for ?-cyhalothrin. Approximately 493 protein spots were detected in two-dimensional polyacrylamide gel electrophoresis(2-DE). Thirty-six protein spots displayed differential expression of more than 2-fold in the CRR strain compared to the CSS strain. Out of these 36 protein spots, 21 had elevated and 15 had decreased expression. Twenty-four differentially expressed proteins were identified by MALDI TOF MS/MS and categorized into the functional groups cytoskeleton-related protein, carbohydrate and energy metabolism, protein folding, antioxidant system, and nucleotide and amino acid metabolism. Function analysis revealed that cytoskeleton-related proteins and energy metabolism proteins had central roles in the ?-cyhalothrin resistance of A. glycines. The differential expression of ?-cyhalothrin responsive proteins reflected the overall change in cellular structure and metabolism after insecticide treatment in aphids. |
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