Development Of Deltamethrin Resistance In Small Brown Planthopper,Laodelphax Striatellus(Fallén) And The Mechanisms

The small brown planthopper(SBPH),Laodelphax striatellus(Fallén),is an economically important phytophagous species in the family of Delphacidae.It has a wide distribution range from south-east Asia to Siberia and to Europe.This insect attacks several important agricultural crops including rice,corn.wheat,oat and barley.L.striatellus was controlled mainly by using insecticides.However,long-term and extensive use of insecticides led insect to develop serious resistance.To prolong the use life of insecticides and effectiveness the resistance development role should be studied for establishment of logical resistance management tactics.In this study,based on the ready chlorpyrifos resistant strain and recovery strains,the deltamethrin-resistant strain was developed.The resistance development during selection and the expression level of the resistance-related P450 genes,esterase gene,and ABC transpoter genes was monitored in parallel.The full-length cDNA of sodium channel gene of L.striatellus was cloned and compared between susceptible and resistant strains.These studies will provide a theoretical basis to clarify the evolution mechanism of deltamethrin-resistance in L.striatellus.1.Infleence of different insecticide treatments on the resistance development in L.StriatellusIn this study,chlorpyrifos-resistant strain of L.striatellus was used as original to develop deltamethrin-resistant strain by spraying the insects on rice seedlings with insecticides.The resistance monitoring results showed that the deltamethrin resistance development in this strain started at the exponential increasing period without the normal slow increasing period.In the first six generations deltamethrin resistance increased to 185.31 times,and further selection for another 5 generations led to even higher resistance that the dose of 5000mg/L caused only 4.35%mortality in the eleventh generation.Then,the selected L.striatellus strain with high resistance to deltamethrin was rotationally treated with chlorpyrifos,imidacloprid and natural recession and monitoring was performed to check the resistance dynamics.With natural recession,the sensitivities of the insects to deltamethrin,chlorpyrifos,imidacloprid were all recovered,but with different speed and the sensitivity to imidacloprid recovered fastest.The susceptibility to the same insecticide recovered with different speed because of various treatments.The recovery speed of susceptibility to deltamethrin with different treatments varied from the highest to the lowest treatments were imidacloprid,natural recession and chlorpyrifos;the recovery speed of susceptibility to chlorpyrifos from the highest to the lowest treatments were imidacloprid,natural recession and chlorpyrifos;the recovery speed of susceptibility to imidacloprid from the highest to the lowest treatments were natural recession chlorpyrifos and imidacloprid;the recovery speed of susceptibility to beta-cypermethrin also differed with treatments,and the imidacloprid treatment was the highest and then the chlorpyrifos treatment and natural recession.In conclusion,the resistance developed rapidly when the resistant strains were used as original and treated with high selection pressure;L.striatellus can produce high levels of resistance to deltamethrin and made the treatment invalided;the sensitivity recover speeds of the L.striatellus strain with high resistance to deltamethrin varied with rotational treatments or insecticides used.These results could provide theoretical basis for rotational use of insecticide for control of L.striatellus in the field.2.The detoxification genes for resistance to various insecticides in L.striatellusEight P450 genes,two carboxylesterase and seventeen ABC genes had been identified previously in our laboratory as resistance-related genes.In the present work,we test all related genes in various strains produced during resistance selection and recovery.The results showed that the expression levels of CYP439A1v3,CYP6AY3v2,CYP306A2v2,CYP6FU1,CYP353D1v2 significantly up-regulated in deltamethrin-resistant strain,and the expression ratios were 2.66-,2.65-,4.42-,2.36-,and 2.37 fold.Of which,CYP353D1v2 was found also significantly up-regulated in chlorpyrifos selection and imidacloprid selection strains with the expression ratios 1.97-and 2.13 fold.Thus,the over-expression of CYP353D1v2 might contribute to deltamethrin,chlorpyrifos and imidacloprid resistance in L.striatellus.The expression levels of LsCE12 and LsCE36 significantly up-regulated in deltamethrin resistant strain.The expression ratio was 2.45-and 14.93 fold for LsCE12 and LsCE36 in the ninth selection generation(C9),and The expression ratios for LsCE36 was 2.08 in the seventh generation(C7).Thus it can be seen that the gene may play an important role on deltamethrin-resistance,and chlorpyrifos-resistance as well.The 10 ABC genes ABCA1,ABCA3,ABCB2,ABCG2,ABCG3,ABCG7,ABCG10,ABCH4,ABCH5,ABCH6 significantly up-regulated in deltamethrin resistant strain,and the expression ratios were 2.09-,2.10-,2.06-,2.65-,2.34-,2.36-,2.37-,3.95-,5.69-and 3.12 fold in in the ninth generation(C9).However,in the recovery strain treated in different ways,the 10 ABC recovery strain treated in different ways,the 10 ABC genes showed no significant difference as compared with the susceptible strain.Thus,these genes may be only related with deltamethrin-resistance.3.Cloning and sequence analysis of sodium channel gene in susceptible and resistance strains of L.Striatellus.In this study,we cloned the full-length cDNA of sodium channel gene(Lspara),open reading frame(ORF)was 6171 bp and encoded 2075 amino acids.Lspara shares common structure and sequence features with the sodium genes of other insect species,has a gene structure typical for sodium channels:four homologous domains(Ⅰ-Ⅳ),each domain possessing six transmembrane segments(S1-S6),and a MFM motif.Sequence analysis showed that sodium channel gene of L.Striatellus has a close genetic relationship with Apolygus lucorum and Cimex lectularius,belong to the same clade.Sequence analysis between susceptible and resistance strains showed that no previous reported resistance mutations existed in the LsparaⅡ region M827,M918,M929,L932,L1011,L1014,V1016,LsparaⅢ area A1410,A1494,F1534,F1538,or within LsparaⅣ area A1752,However,deltamethrin-resistant strain as compared to susceptible strains has a amino acid fragment insertion VSIYYFPT within Lsparal-LsparaⅡ area.Whether the mutation relates to the resistance to deltamethrin of L.Striatellus needs further studied.