Effects Of Hexaflumuron On Trehalase Activity And Expression Of Trehalase Gene From Spodoptera Litura Fabircius

Cotton leaf worm, Spodoptera litura Fabricius (Lepidoptera, Noctuidae), is a gluttonousagricultural pest that causes a massive reduction in the word. Due to good insecticidal effectagainst S. litura, benzoylphenylurea insecticides, such as hexaflumuron, are widely accepted.But with extensive use, insect resistance becomes more and more serious. In order to delaythe resistance of S. litura to hexaflumuron, and to investigate the mechanism ofhexaflumuronm, we studied effects of hexaflumuron on trehalase activity and trehalase geneexpression in chitin synthesis in this paper. The results are summarized as follows.1. Effects of hexaflumuron on trehalase activity and trehalase gene expression of larvaeand prepupa of S. lituraWe determined trehalase activity by hexaflumuron directly reacted with trehalase in vitroand hexaflumuron treated5thinstar larvae with DNS method. The results of hexaflumurondirectly reacted with trehalase indicated that trehalase activity decreased when hexaflumuronconcentrations were below1153mg·L-1. With its dosage increasing, the inhibitory effect totrehalase became stronger, while the rate is slowing. After the5thinstar larvaes werecontinuously treated with three sublethal concentrations (46mg·L-1,92mg·L-1and138mg·L-1)of hexaflumuron for different lenth of times, the trehalase activity was determined. The resultsshowed when the hexaflumuron at46mg·L-1concentration, trehalase activity of the larvaerised to some extent; with hexaflumuron-treated time increasing, activation effect graduallydecreased, and turned to inhibitory effect at prepupa stage. When the hexaflumuron at92mg·L-1and138mg·L-1concentration, trehalase activity of the larvae and prepupa decreasedpartly; and inhibition rate of138mg·L-1was significant(P<0.05) and higher than that of92mg·L-1at the same length of hexaflumuron treated times．With improved anthrone method, trehalose content of the5thinstar larvae continuouslytreated with hexaflumuron for different lenth of times was determined. The results indicatedthat trehalose content had a certain degree of rise when the hexaflumuron at46mg·L-1concentration, while it decreased significantly(P<0.05) when the hexaflumuron at92mg·L-1and138mg·L-1concentration.2. Trehalase gene expression was analyzed every12hours after the1thinstar larvae bythe relative quantitative PCR. Treh-1mRNA was highly expressed in the molting period for2th,3thand5thinstar larvae, and3thhour-12instar larvae; while Treh-2mRNA was highlyexpressed in2thinstar hour-48larvae to4thinstar hour-24larvae and the molting period for5th instar larvae. The trehalase gene of the larvae in the molting period was highly expressed.This may be for more trehalase to decompose trehalose, which provide energy for molting andthe substrate for the epidermis.3. The effects of flufenoxuron on the mRNA expression of trehalase genes were analyzedusing real-time quantitative PCR, with the5thinstar larvae continuously treated in threesublethal concentrations (46mg·L-1,92mg·L-1and138mg·L-1) of hexaflumuron for24h,48h and72h. The results indicated that lower dose of hexaflumuron promoted gene mRNAexpression, while higher dose inhibited. With hexaflumuron-treated time increasing, theactivation of hexaflumuron on Treh-1mRNA expression showed a trend of rising to a certaindegree, while that of hexaflumuron on Treh-2expression decreased and turned to inhibitionafter72h. The inhibition of higher dose of hexaflumuron on trehalase genes mRNAexpression gradually strengthened, and there were significant differences after48h and72h(P<0.05).