| Diamondback moth(Plutella xylostella L.)is an important pest of cruciferous vegetables.Due to its short life cycle,severe generation overlap,strong reproductive capacity,and improper application methods in production,it has caused most insecticides resistance.Flufenapram has been widely used for its novel targets,good fast-acting persistence,and low toxicity to mammals.However,the resistance of P.xylostella to flufenapram has become increasingly prominent.In order to explore the relationship between the resistance mechanism of P.xylostella to flubendiamide and reproductive development,three strains/populations of the sensitive strain S,the flubendimid resistant strain R_f and the field resistant population R_b were screened in this project.As the research object,the experimental research was carried out by various methods such as real-time quantitative PCR,enzyme activity determination,and ultra-high performance liquid tandem mass spectrometry,and the results obtained were as follows:1.On the basis of previous resistance selection,the immersion method was used to determine the resistance level of P.xylostella to flubendiamide,and the diamondback moth,which has a multiple of p-flubendiamide resistance,reached a high level of resistance.P.xylostella resistant flubenzimid line(R_f)with a genotype of GAG of 946.95 times.The resistance multiple of P.xylostella(R_b)in the field to flubendiamide was 1115.06 times.2.To further explain the mechanism of action between reproduction and resistance of P.xylostella,a quantitative quantitative PCR analysis was used to compare the vitellogenin(PxVg)gene and vitellogenin receptor(vitellogenin)of P.xylostella and the sensitive strains(PxVg)gene,methoprene-tolerant(PxMet)gene and trehalase(PxTre)gene,and the results showed that the expression levels of each gene in resistant strains/populations were more sensitive than those in different periods.The strains have been improved to varying degrees.This indicates that the reason for the resistance of P.xylostella to flubendiamide insecticides may be related to the increase in the relative expression of the above-mentioned reproduction-related genes.3.Enzyme-linked immunosorbent assay was used to determine the yolk protein content of female moths of P.xylostella at different emergence times.The results showed that the resistant strains had different degrees of improvement compared to the sensitive strains,and the yolk protein content decreased 12 hours after emergence.It is taken up by the ovaries,and then gradually begins to increase in content.It further illustrates that the increase in yolk protein content may be related to the development of insect ovaries.4.UPLC-MS/MS method was used to detect the content of juvenile hormone and ecdysone in the serum of 4th instar resistant and sensitive diamondback moth adults.The content differences between the resistant and sensitive strains were compared,and two hormones in the resistant strain/population were found.Compared with the sensitive strains,the content of succinate was increased to varying degrees,and the differences were significant.It was positively correlated with the expression of yolk protein and yolk protein receptor genes,and the spawning rate of resistant strains was significantly higher than that of sensitive strains.It is speculated that molting hormone and juvenile hormone of P.xylostella can affect yolk Protein and yolk protein receptor genes regulate insect reproduction.5.This experiment measured the relative expression of trehalase gene(PxTre)and trehalase activity in different strains/populations of P.xylostella,and found that the expression level and enzyme activity of resistant strains/populations were higher than those of sensitive strains at various stages.The increase of trehalase activity in sexual strains/populations leads to an increase in spawning and a decrease in trehalose content,which affects the capacity energy supply and shortens the life span of P.xylostella adults.The specific mechanism of action needs further study. |
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