Evolution Of Resistance To Bt Toxins In Mythimna Separata(Walker)

The Oriental Armyworm?OAW?,Mythimna separata?Walker?,is a destructive pest of agricultural crops in Asia and Australia.It is the main pest that periodical outbreak on maize.Transgenic Bt maize high-expressing Bt toxins season long provides a new approach for control of OAW.While the commercialization of Bt maize in more than 20 countries there are evidence to suggest that the development of resistance in target pests will seriously threaten the sustainable use of transgenic Bt maize.Effective resistance management strategy such as high dose refuge strategy,pyramid strategy can delay or avoid resistance development in targets.Understanding the evolution and inheritance of resistance to Bt toxins in insects will provide scientific bases for establishing a successful insect resistance management program.In this work,we evaluated the toxicity of 7 Cry toxins,i.e.Cry1Ab,Cry1Ac,Cry1Ah,Cry1Fa,Cry2Aa,Cry2Ab and Cry1Ie,and 3 Vip toxins,i.e.Vip3Aa19,Vip3Aa16and Vip3Ca against OAW neonate larvae using diet incorporated bioassay.In addition,the interactions between Cry and Vip toxins were examined through evaluating the toxicity of bi-mixed of Cry with Vip toxins.We assessed the evolution of resistance to Cry1Ab,Cry1F,Vip3Aa19 toxins in OAW by exposing neonates to high-concentration of those toxins for 7 to 10 days,respectively.Finally,the fitness cost and inheritance of resistance in the Vip3Aa19-selected strain?OAW-VipR?were investigated.The bioassays results revealed that LC₅₀?lethal concentration for 50%mortality?values ranged from 1.6 to 78.6?g/g?toxin/diet?for these toxins.Among them,Vip3 proteins along with Cry1A proteins and Cry2Aa,were the ones with the highest potency,with LC₅₀ values ranging from 1.6 to 7.4?g/g.Synergism between Cry and Vip toxins was observed,being high in the combination of Vip3Aa16with Cry1 toxins,with synergetic factors ranging from 2.2 to 9.2.The Vip3Ca toxin did not show any synergistic effect with any of the toxins tested.Resistance evolution experiments were conducted under different selection pressure through larvae exposed to Cry1Ab,Cry1F,Vip3Aa19,respectively.The selection pressures were 75-80%and 99%for Cry1Ab,65-72%for Cry1F,and 95-99%for Vip3Aa19,respectively.The newly hatched larvae were fed with selecting diet for 7 to 10 days and then transferred to normal diet.Vip3Aa19 selected strain developed 278.10 folds resistance after 2 generations of selection.The resistance reached more than3,000 folds after 8 generations of selection.However,the susceptibility of Cry1F selected strain to Cry1F toxin was not significantly different from susceptible strain after 10 generations of selection.Also,the susceptibility of Cry1Ab selected strains to Cry1Ab toxin were not significantly different from susceptible strain after 5 or 8 generations of selection with Cry1Ab under selection pressures of 99%or75-80%of mortalities.The susceptibility of OAW-VipR strain to Cry1Ab was decline in 2.79-fold,i.e.Vip3Aa19selection resulted in a lower level of cross-resistance to Cry1Ab.However,the susceptibility of OAW-VipR strain to Cry1F was not significantly different comparing to the susceptible strain?0.58-fold?,i.e.no cross-resistance to Cry1F.Compared to susceptible strain,the larval stage of the OAW-VipR strain was 2.8 days longer and the emergence rate was reduced by 34%.The average of egg masses laid per female of OAW-VipR strain was decreased by 29.5%compared with the sustainable strain.While OAW-VipR strain was deselected at generation 8 of selection,the average of egg masses laid per female was decreased significantly by 39.1%,which suggested a significant fitness cost.The effective dominance?h?of F₁ offspring for Vip3Aa19 selected strain was calculated at different concentration of Vip3Aa19 toxin,showing that h decreased as concentration of Vip3Aa19increased.The degree of dominant of reciprocal F₁ crosses progenies were 0.68 and 0.33,respectively,which indicated that the resistance inheritance of Vip3Aa19 was partly dominant inheritance with maternal effect.