Screening And Application Of Attractant And Its Binding Characteristics With Olfactory Proteins Of Holotrichia Parallela

Holotrichia parallela is an important subterranean insect in China because of its wide distribution and high degree of damage.The larvae(grubs)live in the soil and Stealthily harm,It is difficult to obtain the ideal effect through chemical control measures such as seed mixing and it would cause great pollution to the environment.The attractant was developed by using the tendency of adults to odor compounds to control the insect population base,Thus control insect population base.This method could be used as an important measure for ecological management of Holotrichia parallela.This study was based on the four host plant volatiles,three aggregation pheromone,one sex pheromone of Holotrichia parallela which were found and could attract Holotrichia parallela.(1)To screen the binary attractant for the control of Holotrichia parallela.(2)field trapping test in field for attractant in Xinxiang city Henan province.(3)Expression and purification of the antenna odorant binding proteins of Holotrichia parallela and binding ability of 3 proteins to 30 odor molecules was studied by fluorescence competitive binding test to explain the mechanism of attractant on Holotrichia parallela from the molecular point of view.It provides a theoretical basis for ecological management of Holotrichia parallela.The specific research results are as follows:(1)According to the result of Electro-antennogram Responses test of Holotrichia parallela to 19 formulations,A total of 7 formulations 1a,3c,4d,5e,7g,8h and 17 q could induce strong antennae potential response in male and female adults of Holotrichia parallela.The result of these 7 formulations were further screened by the behavioral response test showed,The formula with the highest selective response rate was formula 5e,up to 71.01%,and then 8h and 3c,the selective response rate were 66.09%and 65.43%,respectively.There is no significant difference among 3 selective response rate of 3formulations,and were significantly higher than another 4 formulations(p<0.05).The result of comparison of attractive effect between formulations and their single composition results show that the number of test insects attracted by 3 formulations was significantly higher than their single composition,separately.The results also showed that the three formulations had significant synergistic effects on the single component.The result of indoor cage test showed,The best solvent of three formulations were Paraffin oil,N-hexane,Dichloromethane,respectively,and the number of attracted test insects were 56,49,61,respectively.So far,a total of 3 kinds of attractants were screened out,attractant 3c(formula 3c+Paraffin oil),attractant 5e(formula 5e+ N-hexane)and attractant 8h(formula 8h+ Dichloromethane),respectively.(2)The results of 25-day field attract test showed,attractant 3c(1394)> Cis-3-hexenyl acetate(1142)> Pentadecyl acetate(748)> control(376);Attractant 5e(1345)> Cis-3-hexenol(1043)>Cis-13-docosenol(644)> control(323);Attractant 8h(1492)> Cinnamaldehyde(1056)>Cis-13-docosenol(629)> control(297).(3)3 OBPs gene sequences of Holotrichia parallela were obtained from Genbank and it were connected with the prokaryotic expression vector pet-30a(+).Finally,three OBPs were expressed andpurified,namely OBP8,OBP13 and OBP16.The three OBPs have strong binding capacity with a variety of plant volatiles,In addition,OBP13 can be well combined with pentadecyl acetate,the aggregation pheromone of Holotrichia parallela.The three OBPs have a good binding ability with some components of the three attractants screened in this study.It showed three OBPs play an important role in attracting Holotrichia parallela with three attractants.