Identification Of Host Plant Volatiles And Pheromones And Their Electroantennogram Responses Of Bactrocera Minax

The Chinese citrus fly,Bactrocera(Tetradacus)minax(Enderlein),is a destructive quarantine pest of citrus fruits in China,including Guizhou Province,Hubei Province,Hunan Province,Sichuan Province,Chongqing city,and so on.Oversea distribution areas include Bhutan,Japan,India,Vietnam,and so on.Citrus volatile organic compounds(VOCs)and insect pheromones play a critical role in regulating pest insects' behaviors and chemical communication.and have the advantages of high efficiency and pollution-free control.However,the active citrus volatile compounds,especially those emitted by immature fruits and pheromones affecting B.minax,are still unknown.and the citrus fruit volatiles among different citrus varieties is different.Therefore,it is necessary to identify and analyze the host volatiles and pheromones,using headspace solid-phase microextraction combined with gas chromatography-mass spectrometry(HS-SPME-GC-MS)to collect and identify the volatiles of different varieties of citrus fruits and the pheromone components of B.minax,Furthermore,the gas chromatography-electroantennogram detection(GC-EAD)technique has been used to screen candidate compounds from volatiles through the olfactory receptors on B.minax antennae.The age-and sex-specific electroantennogram(EAG)responses of B.minax adults to different doses of active volatiles and pheromones were determined.The main results of this study are described as follows: 1.Identification of VOCs of different citrus fruitsOwing to differences in the preference of B.minax to different citrus cultivars,we compared the volatiles of immature fruits from five cultivars,including Hongjiang sweet orange(Citrus sinensis),Navel orange(Citrus sinensis),Satsuma mandarin(Citrus unshiu)with strong preference of B.minax,and Eureka lemon(Citrus limon),Xiangshui lemon(Citrus limon)with weak preference of B.minax.A total of 72 compounds were identified,including 32 hydrocarbons,19 alcohols,4 aldehydes,11 ketones,4 esters,and 2 ethers.The distribution and content of each volatiles in the five citrus varieties were different and correlated.Among them,the main volatiles were hydrocarbons.Our principal component analysis(PCA)results were carried out on the factor loading diagram,showing that the five cultivars could be distinguished and linked.2.Study on the active components of the volatiles from unmature Hongjiang sweet orangeThe active components of the volatiles from the fruits of Hongjiang sweet orange,which can cause the antennae reaction of the adults of B.minax,were analyzed.The results of the GC-EAD revealed that B.minax males responded to d-limonene,terpinen-4-ol and citral,whereas the females responded to d-limonene and nonanal.The age-and sex-specific electroantennogram(EAG)responses of B.minax adults to different doses of active compounds were determined.EAG responses significantly uaffected by the age or interaction with dose of B.minax,and the volatiles at doses of 100 or 1000 ?g had maximum dose-EAG responses.3.Identification and electroantennographical response of pheromone in B.minaxThe results of HS-SPME-GC-MS analysis showed that N-(3-methyl-butyl)acetamide was one of the main volatiles of 15-day-old B.minax,which is released by both male and female adults at different times,and might be the pheromone of B.minax.Furthermore,the electroantennogram responses of 3-days-old and 15-days-old B.minax to 1 ?g,10 ?g,100 ?g and 1,000 ?g of N-(3-methyl-butyl)acetamide were measured.The results showed that there was a strong electroantennogram response to N-(3-methyl-butyl)acetamide in adults of B.minax.The optimum content of electroantennogram was from 100 ?g to 1,000 ?g.The difference of electroantennogram response could be affected by dose and dose ×sex.In conclusion,The antennal connection method and GC-EAD technology of B.minax were established and optimized for the first time in this study,which provided a new research tool for screening compounds with insects' antennae.Among them,the analysis of volatiles from different varieties of citrus fruits provides guidance for the rational use of host plants for biological control of B.minax.GC-EAD technology was used to screen the active components of the volatiles from the host plants of B.minax,which makes it possible for further development of attractants or repellents of B.minax.The identification of the components and understanding the behavior of the pheromone provide a new stage for biological control of the B.minax.