Peripheral Olfactory Sensation Mechanism Of Holotrichia Oblita On Host Plant Volatiles

Holotrichia oblita Faldermann(Coleoptera)is an underground pest that seriously harms crops in the northern region of China.The insect has the characteristics of wide distribution,miscellaneous feeding,and serious damage.For a long time the traditional methods such as pesticides which have been used to prevent and control it have poor effects,even it has also caused a variety of potential threats to soil environment and natural enemy.Therefore,it is imminent to find a green and efficient method of prevention and control of Holotrichia oblita.In this study,Holotrichia oblita adults were used as the research object.First,the types of volatiles of its host plants were determined,and the compounds that attracted the behavior of Holotrichia oblita were preliminarily screened by EAG experiment and Y tube behavior experiments which provide candidate compounds for the development of effective behavior modifiers.Then the role of odorant receptors(OR)and sensory neuron membrane proteins(SNMP)in the recognition of functional compounds in Holotrichia oblita was investigated,so as to clarify the peripheral olfactory sensation mechanism of the functional host plant volatiles.This has guiding significance for the prevention and development of effective behavior regulators for Holotrichia oblita.The main results are as follows:(1)EAG experiment was conducted on 150 volatiles of 25 host plants of Holotrichia oblita.The results showed that the adult antennae of Holotrichia oblita had a different response to 150 plant-derived volatiles under the stimulation of a compound concentration of 1 μg/μL.Compared with solvent paraffin oil control group,leaf acetate,trans-2-hexene-1-ol,cis-3-hexenyl isobutyrate,6-Methyl-5-heptenone,benzyl alcohol,6-methyl-5-hepten-2-one these six compounds have strong antenna reactions.(2)Y-tube behavioral experiment tests on three plants volatiles of Holotrichia oblita: leaf acetate,trans-2-hexene-1-ol,cis-3-hexenyl isobutyrate.The results of the behavior selection of these three volatiles show that,leaf acetate has a significant attraction effect on Holotrichia oblita adults at concentrations of 0.1μg/μL and 1μg/μL,trans-2-hexene-1-ol and cis-3-hexenyl isobutyrate at a concentration of 0.1μg/μL have a significant attraction to Holotrichia oblita adults.(3)According to the published sequence information of Holotrichia oblita odorant receptor gene family and sensory neuron membrane protein gene family design specific primers.A total of ORF sequences of HoblOR1,HoblOR5,HoblOR6,HoblOR15,HoblORco and HoblSNMP1 these six genes were obtained.And successfully cloned it into p-GEMHE expression vector,which laid the foundation for subsequent in vitro expression experiments.(4)Through two electrode voltage clamp system to study the function of four olfactory receptors of Holotrichia oblita,and a ligand compound that could obviously activate the HoblOR15 receptor was screened.Leaf acetate has a significant attracting effect on Holotrichia oblita and as a sex attractant synergist for various insects,it can improve the trapping effect of sex attractants on insects.(5)Co-express HoblSNMP1 and HoblOR15/HoblORco in Xenopus oocytes,compared with oocytes expressing HoblSNMP1/HoblOR15/HoblORco to oocytes expressing HoblOR15/HoblORco,has greater reaction to leaf acetate compound.It is believed that HoblSNMP1 can enhance the recognition of HoblOR15 to leaf acetate.In summary,this research preliminarily screened host plant volatiles that have a significant attraction to Holotrichia oblita.Among them,the attraction effect of leaf acetate is the most obvious.It is speculated that the compound may have an important effect on the behavior of the adult.Later,it was determined that HoblOR15 can specifically bind to leaf acetate and be activated.HoblOR15 participated in the perception process of leaf acetate in Holotrichia oblita.At the same time,HoblSNMP1 can significantly enhance the recognition effect of HoblOR15 on leaf acetate.This research completes the study of the peripheral olfactory sensation mechanism of leaf acetate in Holotrichia oblita.At the same time,leaf acetate can be used as a candidate compound,which is of great significance for the future prevention and development of an effective attractant for Holotrichia oblita.