| Insects have a sensitive sense of smell,which plays an important role in their multiplying and environmental adaptation.The olfactory system of insects can specifically recognize the odor molecules in the environment,and take it as a clue for feeding,courtship,choosing host and spawning sites and avoiding natural enemies.Helicoverpa assulta,as noctuid insect,mainly damage tobacco and peppers.Elucidating the recognition mechanism of the olfactory system of Helicoverpa assulta to its main host tobacco volatiles could help us to further understand the mechanism of insect olfactory recognition on host plants and also provide a basis to screen more effective attractants to Helicoverpa assulta.In this essay,the electrophysiological and behavioral response of adult Helicoverpa assulta to the main components of the volatiles from tobacco plant were determined using the EAG and ovipositional preference tests,based on the collecting and identifying the volatile of tobacco plant,the main host of Helicoverpa assulta.Forthermore,five odorant receptor genes of Helicoverpa assulta were cloned and the ligand of these odorant receptors were screened through two-electrode voltage clamp in vitro,preliminarily identified the corresponding relationship between the odor receptors and the main constituents of tobacco volatiles.Here are main test results:Collection and identification of tobacco volatile:Collected volatiles of the whole plant of common tobacco in flowering period and analyzed it by gas chromatography-mass spectrometry(GC-MS).GC-MS results showed that the main components of tobacco volatiles are nonanal,heptaldehyde,linalool,caryophyllene,solanone,methyl salicylate,nicotine,caryophyllene,and etc.The content of solanone was the highest,the content of caryophyllene and methyl salicylate also accounted for a large proportion,and the content of nonanal was the least.The EAG and behavioral responses of Helicoverpa assulta to tobacco volatiles:All the main components of the tobacco volatiles measured by GC-MS evoked electroantennogram responses of the moth,among which the response to nonanal was the greatest.In addition,the reaction to linalool and caryophyllene showed that the female was significantly greater than the male,however,the response to other odors showed nosignificant male-female difference.In the behavioral experiments,we found that nonanal had a significant attracting effect on oviposition of Helicoverpa assulta at the concentration of 0.01 mol/L and 0.001mol/L,and had obvious repellent effect on oviposition of female moth at the concentration of 0.1mol/L.Solanone,nicotine and heptanal showed significant repellent effect on moth at a concentration of 0.1 mol/L,and solanone with an attraction effect at the concentration of 0.01mol/L.Cloning and functional identification of the odor receptor of Helicoverpa assulta:The full-length sequences of common odorant receptors(OR18,OR20,OR23,OR26 and OR67)of tobacco budworm were cloned and the function of the receptors were studied through the double-electrode voltage of Xenopus oocytes.The results showed that OR67 had a greater reaction with nonanal and heptanal,with the former reacting up to 1900 n A,and the latter to 400 n A.OR23 reacted to nerolidol,farnesene and geranyl acetate,with the reaction values of 300 n A,990 n A and 1400 n A,respectively.OR 26 reacted to eugenol,a plant volatile,with the reaction of 310 n A.Receptors of solanone and nicotine have not yet been found and will continue to be screened in the next trial.The voltage clamp test results are generally in agreement with the EAG experimental results.The response of the main olfactory receptors to the odorants of Helicoverpa assulta:Single-sensor recording(SSR)method was applied to measure the response of the short-hair sensilla and cone sensilla in antennae to odor substances evoking the electrophysiological response of OR67,and the recording site was 5-65 flagellomeres of antennae.Both types of sensilla responded to nonanal.84.4% of the sensilla basiconica responded to the test odor,among which two types of neurons were recorded,one of the neurons reacted markedly to the test odor,with the highest to nonanal,while the other did not respond to the test odor.90% of the long cone sensilla responded to the test odor,in which two types of neurons were recorded,one of which reacted to nonanal and heptaldehyde,while the other responded mainly to oxidized caryophyllene.The test results for the long cone sensilla was in accordance with the OR67 voltage clamp results,suggesting that OR67 may be present in the sensilla basiconca,and that there may be another receptor that also recognizes nonanal,but has not been found yet,and will continue to be tested and indentified in the next phase. |
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