Tracking The Behaviour Of Utilizing Mutiple Host Plants Of Apolygus Lucorm By DNA-Based Methods

Area-wide cultivation of transgenic Bt cotton has effectively controlled the cotton bollworm Helicoverpa armigera with resulting reductions in the amount of broad-spectrum insecticides used in cotton production.This lowered insesticide use has caused the mirids to be major pest on Bt cotton in China.Furthermore,Apolygus lucor?m(Meyer-Dur.)have spread to a range of other crops,e.g.Chinese date Zizyphus jujuba,grape Vitis vinifera.A.lucorumhas a wide range of hosts and its feeding,courtship,spawning and shelter and other complex behavior,the feeding characteristics of A.lucor?mare not obvious,it is difficult to accurately track its use of the host habits in the field by the traditional research methods.In this study,the field population density survey and DNA-based method were used to study the host utilization habits of A.lucor?m.In order to clarify the multi-host habit and its ecological significance of the A.lucor?madult in North China,which provides the theoretical basis for the comprehensive management of A.lucor?m.The main results were summarized as follows.To optimize an extraction method of host plant DNA in A.lucorum,which is available for further developing DNA-based technology to explore host plant utilization pattern of this mirid bug in open field.Firstly,we extracted the cotton DNA in A.lucorum gut by using CTAB-based method and two others,and compared the detection rates of cotton plant DNA among three extraction methods.Secondly,we washed A.lucorum adults in the solution of 1-1.5%sodium hypochlorite for different periods,and compared the effect on removing plant DNA contaminations on adult surface and on detection rates of cotton plant DNA in A.lucorum.A CTAB-based DNA extraction protocol yielded significantly higher detection rates of cotton plant DNA than the other two by using commercial kits.Washing in 1-1.5%sodium hypochlorite for 5s successfully removed plant DNA contaminations on A.lucorum surface,and did not destroy cotton plant DNA in A.lucorum.The detection rate of cotton plant DNA decreased significantly after washing for 30s.This study established a feasible method to remove external plant DNA contaminations on A.lucorum surface,and confirmed that DNA could be extracted from the whole adult of A.lucorum instead of its gut.To clarify the selection of host plants of A.lucorum in the field.The results of sampling revealed relative higher population abundance in mungbean compared to that in cotton,confirming mungbean as a more preferred host plant species of A.lucorumadults.Then the results of feeding experiment showed that significant differences were observed between calculateddetectability half-life(DS50)of these two host plants.Molecular detecting results showed that cotton DNA can be clearly detected in30%of adultA.lucorumcollectedin mungbean plots and detection rates decreased successively along with distance from 5 to 15 m away from the mungbean-cotton midline.At the same time,15.5%of the adults contains cotton and mung bean DNA.A.lucorum adults prefer mungbeans,transferred from the cotton field to mungbeans for feeding.To clarify the selection of host plants at different stages of A.lucorum in the field.The field-plot samples showed that A.lucorum density was significantly higher on flowering H.scandens than on seedlings of M.sativa,and,similarly,mirid bug density was also higher on flowering M.sativa than on seedlings of H.scandens.In mirid bugs exposed toseedlings of H.scandens and flowering M.sativa,the detection rate of M.sativa DNA in A.lucorum guts was evidently higher than that of H.scandens.Meanwhile,in mirid bugs exposed to seedlings ofM.Sativa and flowering H.scandens,a higher detection rate of H.scandens DNA was found.Our current finding provides evidence of a strong positive relationship between population abundance and feeding preference of A.lucorum between different plants.The A.lucorumadults were trapped in Langfang experimental station and then were sequencing.Firstly,we compared the amplification efficiency and resolution of three different DNA molecular markers of rbcL,ITS2 and trnL.It was found that ITS2 had high resolution,98.9%ITS2 plant sequences could be identified in the family,54.7%could be identified in species.There were 17 different families of host plants DNA in A.lucorum adults,22.0%of the A.lucorum adults contain a variety of host plant DNA.In July,cotton,H.scandens,soybean and Helianthus were dominant,accounting for 65.0%;August,cotton,H.scandens,Artemisia,Ricinus communis,Agastache rugosa,accounting for 58.0%.At the same time,it was also found a ball of Platanus and cypresses and other plants DNA in A.lucorum adults.The A.lucorumadults were trapped in the Bohai sea and then were sequencing.It was found 20 different families of host plants DNA in A.lucorum adults,34.0%of the A.lucorumadult contain a variety of host plant DNA.In June,cotton?Humulus and Suaeda glauca were dominant,accounting for 67.0%;in July,cotton and Asteraceae were in 72.0%;in August,cotton,Artemisia annua,Artemisia scoparia and other plants mainly accounted for 40.0%.It were also found a number of plants have not been recorded as host DNA,such as Populus trichocarpa,Ulmus parvifolia,Fraxinus chinensis,Flueggea leucopyra,Ulmus pumilaand so on.This study clarified the multi-host use behavior and habits of A.lucorum,which helped to analyze the interaction between A.lucorum and host plants,and clarify the transfer and diffusion of A.lucorum.At the same time,it further clarified the utilization pattern of seasonal host plants of A.lucorum in North China,and provided important scientific basis for constructing regional population management strategy of A.lucorum.