Effect Of Citrus Sinensis Infected With CTV On The Behavioral Response Of Toxoptera Citricida

Tristeza caused by Citrus tristeza virus?CTV?,is an important citrus virus disease and induce economic losses to the citrus industry worldwide.CTV can be transmitted in a semi-persistent manner by aphids,of which Toxoptera citricida?Kirkaldy?is one of the most effective vector insects.In China,virulent CTV strains and T.citricidaare widely distributed.Besides,the prevalence of tristeza is rapidly emerging due to the domestic citrus planting pattern optimizing,which threatens the safe production of sweet orange varieties.In the host-virus-vector insect tripartite interaction system,the virus affects the host plant through altering the host's secondary metabolism and other physiological and biochemical processes,which further changes the host-selection behaviors,population growth and other biological characteristics of the vector.Finally,to a certain extent,it affects the transmission of the virus by vector insects.The influence of the virus on the vector insects closely related to the mode of insect transmission.By systematically studying the interactions amongthe host-virus-vector insect tripartite party,it is helpful to propose new ideas and strategies for the effective prevention and control of vector transmitted plant virus diseases.In this study,CTV-Citrus sinensis-brown citrus aphid were used as the research objects.The biological characteristics of the host-selection behavior of the brown citrus aphid on CTV-infested sweet orange were evaluated by gas chromatography-mass spectrometry?GC-MS?and RT-q PCR.Meanwhile,the host plants infected by CTV were also assayed through analyzing the content and composition of the volatile organic compounds?VOCs?and the expression levels of related genes in terpenoid biosynthesis pathway.Besides,the aphids' host selection and population growth on terpineoids?thymol,terpinolene,citral,?-terpineol and myrcene?treated plants were also evaluated.From the perspective of plant defense compounds,the molecular mechanism of the interaction between CTV-affected host and brown citrus aphid was analyzed.It provided a theoretical basis for the virus-host-vector tripartite interaction.The main results are as follows:1.Through the optimization of key reaction factors such as the concentration(Mg²⁺,d NTPs and primers)and annealing temperature,a One-step duplex RT-PCR detection system for CTV and CYVCV was established.The One-step duplex RT-PCR system was used to detect both CTV and CYVCV from samples.2.The host-selection behavior of the brown citrus aphid on healthy and three CTV isolates affected sweet oranges was analyzed with the Y-type of olfactory instrument.We found that plants infected with CTV had attraction effect to brown citrus aphid.Among them,the severe isolate CT14 affected sweet orange plants were the most attractive to the CTV-free brown citrus aphid.The selection rate was as high as 69%,which was significantly higher than that of the healthy sweet orange.3.The components and total amount of volatile compounds in the leaves were significantly affected by aphid feeding and CTV invasion alone or together.The total content of volatiles was the highest with the relative content of 12259.09?g.g-1 FW in healthy sweet orange leaves,followed by CTV and aphid co-infected plants?9865.91?g.g-1 FW?and to a less extent aphid infected plants?8864.21?g.g-1 FW?.The total content in CTV-infected sweet orange leaves?8704.20?g.g-1 FW?was the lowest.4.Monoterpenes were the richest components of total volatiles content in sweet oranges,followed by monoterpene aldehydes and monoterpene alcohols.CTV infection significantly reduced the effects of thirteen monoterpenes and four monoterpene aldehydes.The proportion of sesquiterpenoids,monoterpene phenols,monoterpene esters,and sesquiterpene oxides were significantly increased in the host with CTV infection.However,aphid infection,CTV infection,and co-infestation of sweet orange both caused a significant decrease in the proportion of GLVs.The mock sweet orange does not contain thymol and methyl geranate,but they were contained both in CTV-infected,aphid-infected,and co-infected plants.5.RT-q PCR was used to analyze the expression levels of genes related to terpenoids.Relative to mock plants,the expression of genes associated with MVA pathway in CTV-infected plants or aphid-feeding plants was greatly changed.The expression level of AACT genes was significantly increased,while HMGR,HMGS,MK and FPPS were all sharply down-regulated.The expression levels of DXS,CMK,ISPF, HDR,and GGPS genes related to the MEP pathway were significantly down-regulated in CTV-infected sweet oranges.However,the expression of FNS and GMD3 genes was significantly increased in CTV-infected plants.6.The external treatment with ?-terpineol to the host showed a positive effect,while the treatment with thymol,terpinolene,and myrcene displayed a repellent effect on the brown citrus.The effect of exogenous application of terpinolene and myrcene on the host selectivity of brown citrus aphid was mutually correlated with the indirect effects of CTV infection induced host sweet orange on brown citrus aphid,suggesting that the change of the content and proportion of volatile components in the host sweet orange infected by CTV may be one of the important factors in the selection behavior of CTV infection for the behavior selection of brown citrus aphid.