Olfaction Mechanisms Of Nilaparvata Lugens(st?l) In The Host Location

Sensitive olfactory system play an important role in the behavior of herbivorous insects, such as location of the host plants. The new effective pest control and monitoring technology could be developed based on the understanding of the olfactory mechanism of insects. The brown planthopper(BPH), Nilaparvata lugens(St?l), one of imporant insect pests on rice, caused a serious threat to our country’s rice production. The attractants or repellants for management of BPH would be developed successfully, if the mechanism of host location of BPH could be learned. Here, we isolated and identified the active substances in host positioning of the BPH, cloned the OBP genes and prokaryotic expression and purification, and analyzed the characteristics of Nlug OBP4 binding properties with the odorant and the behavior activity of the BPH, which may provided the theoretical basis and technical supports for the behavior control technology and clarifying the olfactory receptor mechanism of the BPH. The major results of this paper list as below: 1 Isolation and identification of the infochemicals in the host location of the BPHThe olfactory stimulants from the rice of BPH were isolated and identified by GC-EAD coupled with GC-MS. The results showed that five compounds fom rice eliciting strong EAG responses to BPH adults were 1,6-Octadien-3-ol,3,7-dimethyl-(linalool), 2,6-dimethyl-Undecane, 1,4-dichlorobenzene, cis-Linaloloxide and Tridecane. 2 Clone and sequence analysis of the Nlug OBPsThree OBP genes, Nlug OBP8, Nlug OBP9, Nlug OBP10(Gen Bank: KC663687, KC663688, KC663689) were cloned, and all of them possess complete open reading frame. The length is 414 bp, 576 bp, 756 bp respectively, coding 137, 191, 251 amino acids respectively. And their protein molecular weights were 15.09 k Da, 20.98 k Da, 27.88 k Da respectively, isoelectric point were 4.38, 6.06, 7.97 respectively. The blast of three sequences show that Nlug OBP8 gene has six conservative cysteine sites, consistent with the characteristics of the classical OBPs, belongs to the Classic OBPs, while Nlug OBP9 and Nlug OBP10 has eight conservative cysteine sites and a proline sites, belong to Plus-C OBPs. And the prediction of signal peptide analysis of N-terminal show that only Nlug OBP8 contains 21 signal peptide sequence of amino acids, Nlug OBP9 and Nlug OBP10 are no signal peptide. 3 The construction of BPH Nlug OBPs prokaryotic expression vectorsThe prokaryotic expression vector p GEX-6p-1-Nlug OBPs, p ET-17b-Nlug OBPs, p ET-20b(+)-Nlug OBPs, p ET-22b(+)-Nlug OBPs, p ET-30a(+)-Nlug OBPs and p ET-32a(+)-Nlug OBPs recombinant plasmid were constructed successfully. The results of expression of recombinant plasmid in E.coli BL21(DE3)/BL21(DE3) Plys S showed that only p GEX-6p-1-Nlug OBP8 / Nlug OBP10 and p ET-32a(+)-Nlug OBP8 / Nlug OBP10 could be expressed successfully. 4 Olfactometer bioassay of BPHThe behavioral responses of BPH to 22 compounds which had a higher binding with Nlug OBP4 and two compounds eliciting strong EAG response to BPH were tested by H-tube olfactometer. The results showed that BPH adult showed a significant preference for the β-Myrcene and 2-Heptanone diluted by 50-fold, while showed a preference for blank control over the 2-Heptanone diluted by 10- or 100-fold. BPH showed a preference for blank control over the Linalool and 1-octen-3-ol at any concentrations tested. And BPH showed a preference for blank control over the the R-(+)-Limonene,(+)-3-carene,(-)-trans Caryophyllene, Sabinene, Henicosane, Tridecane, Heptanol,(E)-2-hexen-1-ol, Benzaldehyd, Hexanal, Lonone and Methyl benzoate at the certain concentration.