Effects Of The Inhibition Of Juvenile Hormone Synthesis On The Development And Reproduction Of Plutella Xylostella

Juvenile hormone(JH)belongs to sesquiterpenoids and is a kind of small molecule lipophilic physiological active substance.It is synthesized and secreted by a couple of endocrine organs,corpora allata(CA),and released into the hemolymph.JH controls all aspects of insect life,and plays a switch role in the process of growth and development,including that regulates embryonic development and imaginal diskdifferentiation,involves in regulating diapause,vitellogenin synthesis and absorption,and oocyte maturationin insects.However,the physiological functions of JH varywith the insect species.Plutella xylostella(L.)is one of the most destructive pests to cruciferous vegetables all over the world.Due to its resistant to all kinds of pesticides caused by the widespread and unreasonable use,and its strong reproductive ability,there is still a lack of safe and effective methods to control the P.xylostella population.In this paper,we used the methods of bioinformatics,molecular biology,and anatomy etc to focus on: 1)the molecular structure and phylogenetic relationship ofjuvenile hormone acid methyltransferase(JHAMT)and farnesoic acid epoxidase(CYP15C1),which is the key genes of JH synthesis pathway,2)Effects of JHAMT and CYP15C1 silence on JH synthesis,pupae growth and reproductive development of P.xylostella,3)Effects of JH inhibitors on JH synthesis,pupae growth and reproductive development of P.xylostella.The main results are as follows:Px JHAMT with the gene ID Px009591 contained an open reading frame(ORF)of825 bp encoding 274 amino acids.The theoretical molecular weight was estimated at31.74 k Da with the isoelectric point of 5.66.The JHAMT sequence containeda highly conserved Motif I region,where existed S-adenosine-L-methionine(SAM),indicating that JHAMT belongs to SAM-dependent methyltransferase family.The phylogenetic analysis showed that Px JHAMT was clustered with all other Lepidoptera species JHAMTs,but it existed in a single sub-branch.The CYP15C1 with the gene ID of Px011481 contained an ORF of 1353 bp encoding 450 amino acids.The theoretical molecular weight was estimated at 50.90 k Da with the isoelectric point of 6.71.CYP15C1 contained the conserved domains of cytochrome P450 superfamily.Px CYP15C1 was also clustered with all other Lepidoptera species CYP15 s,and is most closely related to CYP15C1 of Cydia pomonella.Px JHAMT was mainly expressed in the larval stage,and Px CYP15C1 was expressed in each stage,but both of them was highly expressed in 3rd instar larvae.Additionally,both Px JHAMT and Px CYP15C1 were significantly highly expressed in the ovary of P.xylostella.The one-day old pupae were selected for RNAinterference experiment to detect the JH titer.It was found that the JH titer of P.xylostella decreased significantly at 8,16 and 24 hours after Px JHAMT and Px CYP15C1 RNAi,but there was no significant difference at 36 h.The expression of Px JHAMT and Px CYP15C1 were consistent with the variation trend of JH titer.There was no significant difference in ovariolet length,the number of laid eggs and egg hatcing rate.The one-day old pupae were treated with JH inhibitor to detect the JH titer,epidermal gene expression,pupal development,and fecundity.The JH titer of P.xylostella in the treatment group decreased significantly at each time point.The expression of epidermal gene Px013419 was significantly increased at 8,16 and 24 h,but Px002031 expression significantly increased at 24 h,and Px009558 expression was significantly decreased at 24 h,then significantly increased at 36 h,Px011256 expression was decreased significantly at 8 and 24 h.Px011149 expression was decreased significantly at 8 and 24 h,and Px011257 decreased significantly at 8 h,16 and 36 h,but decreased significantly at 16 h.The deformity rate of pupae in the treatment group was as high as 18.3%,speculating that JH might be related to the pupal morphogenesis of P.xylostella.JH inhibitor treatement resulted in shorter ovarioles of newly emerged females,less vitelline depositon,as well as lower Vg and Vg R expression.Furthermore,there was lower egg laid and hatching rate of P.xylostellain the treatment group than that in control group.In this study,two methods were used to investigate the effects of the inhibition of JH synthesis on the development and reproduction of P.xylostella.One is to inhibit the Px JHAMT and Px CYP15C1 by RNAi,which is the key genes in JH synthesis pathway.The other is to treatwith JH inhibitor.The results confirmed that interfering with the expression of Px JHAMT and Px CYP15C1 could significantly inhibit JH synthesis,and the JH inhibitor experiment showed that JH played an important role in the development and reproduction of P.xylostella.The results will establish the theoretical foundations on which it will be possible to use the hormonal regulatory mechanisms in P.xylostella to screen potential genes as ways to better control P.xylostella populations.