Interrelationship Between The Transmission Of Cucurbit Chlorotic Yellows Virus By Bemisia Tabaci And Its Resistance To Imidacloprid

Plant viruses can greatly influence biological characteristics of hosts in many ways during long-term interactions with vector insects.For example,plant viruses can directly or indirectly manipulate the behavior,physiology,and defenses of vector insects,thereby promoting the efficiency of virus transmission.The Bemisia tabaci(Gennadius)is an important invasive pest all over the world,causing serious losses to agricultural production.With high survival ability and a wide host range,the B.tabaci is known as a super vector insect that can transmit more than 400 plant viruses.The Cucurbit chlorotic yellows virus(CCYV)is specifically transmitted by the B.tabaci in a semi-persistent mode and is prevalent worldwide,which seriously affect the yield and quality of melon crops,and its damage is further increasing in scope and extent.It is well known that the strategy in B.tabaci control depends on chemical synthetic insecticides,especially neonicotinoid insecticides.While the long-term overuse of neonicotinoid insecticides has lead to high resistance in B.tabaci.And the selection pressure from insecticides may be one of the reasons why of B.tabaci Q biotypes has become the dominant population in many areas,and the resistance of B.tabaci Q biotypes is becoming higher,which in turn making it difficult in B.tabaci control.Therefore,the increased resistance of vector insects and the plant viruses they transmitted have caused huge losses to agricultural production and pose a continuous threat to human health.However,whether there is an intrinsic link between plant viruses transmitted by B.tabaci and its resistance to insecticides has not been thoroughly investigated.Therefore,in this study,we investigated the interrelationship between the CCYV,B.tabaci and the neonicotinoid insecticide imidacloprid,using techniques of insect ethology and molecular biology,to further understand the coevolution between the vector insects and the viruses,and provide theoretical references for further understanding of the mutual adaptation of vector insects and the viruses under long-term co-evolution.The results were as follows.CCYV enhances tolerance to the insecticide imidacloprid in B.tabaciIn the bio-assay of imidacloprid resistance between CCYV-viruliferous and CCYV-free whiteflies,it was found that the corrected mortality rate of CCYV-viruliferous B.tabaci for 10 d was significantly lower than that of CCYV-free whiteflies(P = 0.0445).Based on this biological phenomenon,the expression of imidacloprid resistance related gene,CYP6CM1 and its upstream regulators ERK(extracellular signal-regulated kinases),p38(p38 kinases)and CREB(c AMP-response element binding protein),were further investigated by q RT-PCR.The results showed that the expression of CYP6CM1 in the CCYV-viruliferous B.tabaci was significantly up-regulated at 10 d and 15 d(P < 0.001);the upstream regulators p38 and CREB were significantly up-regulated at 10 d(Pp38 = 0.0024,PCREB = 0.0013),but there was no difference at other time points.The activities of glutathione S-transferase(GST),carboxylesterase(Car E)and cytochrome P450 detoxification enzymes of CCYV-viruliferous B.tabaci were also measured,and the results showed that,there was no difference in Car E enzyme activity,and GST enzyme activity showed a decreasing trend in viruliferous whiteflies(P = 0.0237),but P450 enzyme activity increased significantly(P = 0.0092)compared with the control at 1 d.There was a transient increase in detoxification enzyme activity as the acquisition duration by B.tabaci increased,with all detoxification enzyme activities increasing at 5 d(PGST = 0.0224,PCar E = 0.0293,PP450 = 0.0104).This indicates that the CCYV-viruliferous B.tabaci activated related detoxification metabolism,as shown by the up-regulation of the expression of the imidacloprid resistance-related gene CYP6CM1 and the transient increase in detoxification enzyme activity,thus enhancing the tolerance of its insect vector whitefly to imidacloprid.Imidacloprid-resistant B.tabaci outperformed virus transmission abilityUsing RNAi,we investigated the ability of B.tabaci to acquire CCYV after silencing the resistance-related gene CYP6CM1,and found that the amount of CCYV acquired by B.tabaci silencing CYP6CM1 was significantly lower at 1 h(P = 0.0373) and 3 h(P = 0.0076)compared with the control.In further comparing the virulence acquisition and retention capacity of resistant and susceptible B.tabaci,our results showed that the virulence acquisition capacity of resistant was stronger than that of sensitive B.tabaci,the virulence acquired by resistant B.tabaci was significantly higher at 2 h and 6 h(P2h = 0.0257,P6 h = 0.0301),but the retention capacity of resistant B.tabaci was weaker at 1 d(P = 0.0108),3 d(P = 0.0029),and 9 d(P =0.0012),the resistant B.tabaci held significantly less CCYV copies.The feeding behavior of B.tabaci with different levels of resistance was further investigated by measuring the amount of the honeydew secreted by resistant and sensitive B.tabaci,and it was found that the amount of honeydew secreted by resistant whiteflies was significantly higher than that of sensitive B.tabaci after 24 h of feeding(P = 0.0318).We speculate that this may be due to the fact that resistant B.tabaci need to increase the uptake to balance the energy expenditure caused by elevated resistance,thus leading to rapid acquisition and reduced ability to retentate the CCYV virions.Then,for the semi-persistent transmission of CCYV,the acquisition and retention period are shorter than persistent virus,and resistant B.tabaci adults are able to acquire the virus quickly and spread it to new host plants,which facilitates the spread of the CCYV.ConclusionOur study demonstrated that CCYV was able to modify the biological characteristics of the vector B.tabaci and improve the tolerance of its insect vector to imidacloprid.Furthermore,the ability of the imidacloprid-resistant B.tabaci transmitting the CCYV was superior than that of the susceptible ones.These results indicated that there is a mutual benefit between the CCYV and the vector insect whitefly under the long-term co-evolution.