The Role Of Voltage-dependent Anion Channel (VDAC) In The Transmission Of RSV By Laodelphax Striatellus And Its Response Mechanism To Several Pesticides

Laodelphax striatellus(small brown planthopper,SBPH)is one of the most common rice pests in Asia,which cause damage to rice by feeding and the transmission of rice stripe virus(RSV).The replication of RSV in SBPH needs to overcome multiple barriers,and it needs to interact with the related proteins to complete its replication.Previous studies indicated that the expression level of voltage-dependent anion channel(VDAC)in the viruliferous SBPH was significantly higher than that in the non-viruliferous SBPH.However,the interaction mechanism of VDAC in the transmission of RSV virus was still unclear.For this reason,this study uses a variety of verification methods to prove the interaction between VDAC and RSV to further investigate the role of VDAC in RSV transmission.The control of SBPH in the field mainly depends on chemical control.Pesticide treatment may change various physiological and biochemical levels in SBPH,some sublethal doses of pesticides will also cause the resurgence of SBPH,and may also stimulate the proliferation of RSV in SBPH.Chemical pesticides affect the occurrence level of SBPH and RSV in the field to some extent.Our studies have confirmed that sublethal concentration of triazophos can stimulate the oviposition of SBPH,the expression of vitellin(Vg)and vitellin receptor(vitellogenin receptor,VgR),meanwhile,the expression level of RSV in the ovary and the virus-carrying rate of offspring of SBPH also increase.The interactions between VDAC and RSV exist in the ovary of SBPH.VDAC,as the gatekeeper on the outer membrane of mitochondria,controls apoptosis In order to scientifically evaluate the effects of jinggangmycin,carbendazim,imidacloprid and triazophos on the reproduction of SBPH,whether it will cause the resurgence of SBPH in the field,we therefore analyze the changes of VDAC and apoptosis-related genes in ovary after exposure to four pesticides and the function of VDAC in resurgence.The reasons for the resurgence of some pesticides were explained from the point of view of apoptosis,which will help to guide the selection and application of pesticides in the field,and formulate corresponding countermeasures in advance,so as to control the spread of SBPH and RSV.1.The interactions betw een VDAC and RSVThe expression profiles of VDAC and RSV showed that VDAC was expressed at all developmental stages of SBPH,low expression in nymph stage and high expression in adult stage,and the expression level of viruliferous SBPH was generally higher than that of non-viruliferous SBPH,suggesting that the relative expression of VDAC increased when RSV infected SBPH.The accumulation levels of RSV coat protein CP(Capsid protein)and RNA3(RNA3 segment)mRNA in infected SBPH were analyzed and compared.It was found that CP and RNA3 were low expression in the adult stage and high expression in the nymph stage VDAC and RSV are highly expressed in the ovary of the SBPH,and VDAC and RSV can be detected in various tissues by immunofluorescence.To explore the efficiency of RNA interference(RNAi),it was found that the inhibitory effect on VDAC and RSV was the most obvious after 48 hours of injection,so the subsequent sampling time of RNAi was 48 hours qRT-PCR results showed that the mRNA expression level of VDAC decreased by 46.82%,36.51%,62.88%and 89.29%in salivary glands,midgut,ovary and hemolymph,and RSV decreased by 71.18%,42.31%,67.74%and 63.43%,respectively.The results suggested that there may exist interaction between VDAC and RSV.The results of immunofluorescence showed that the fluorescence signals of VDAC and RSV in salivary glands,midgut and ovary were significantly decreased after RNAi of VDAC,and there was co-localization in some cases Yeast two-hybrid experiment showed that there was interaction between VDAC and RSV RdRp2.Response mechanism of VDAC to several pesticide treatmentsThe highest expression of four kinds of Lscaspase in female SBPH was subtype la,which was 45.75 times of subtype Nc,3.48 times of subtype 1c and 2.67 times of subtype 8,respectively.After RNAi of VDAC,the expression level of VDAC in whole body decreased by 84.05%,Vg decreased by 18.33%and the other three apoptosis-related genes except Lscaspase-8 were significantly down-regulated.VDAC in ovary decreased by 72.68%,Vg decreased by 31.13%.The expressions of VDAC and four apoptosis-related genes in hemolymph were significantly down-regulated,indicating that inhibition of VDAC expression may affect the reproduction and apoptosis of SBPH.LC10 of imidacloprid significantly increased the oviposition of viruliferous SBPH,LC10 of imidacloprid and LC30 of triazophos could significantly stimulate the expressions of Vg and VDAC,and the downstream apoptosis-related gene caspases also increased,indicating that the increased expression of Vg in the ovary of SBPH may also increase its apoptosis.The two selected fungicides-jinggangmycin inhibited the expression of Vg at the concentration of 100 ppm,while carbendazim stimulated the expression of Vg at the concentration of 200 ppm,but the responses of the four caspases to both were not significant The number of apoptotic cells in the eclosion 4D and 10D ovaries treated with imidacloprid and triazophos was higher than that in the control group by TUNEL staining,and the apoptosis rate of 4D ovaries was significantly increased by exposure to imidacloprid.