淡足侧沟茧蜂寄生对草地贪夜蛾幼虫血淋巴黑化反应及酚氧化酶的影响

Effects of the parasitoid Microplitis pallidipes on the menanization and phenoloxidase activity of the hemolymph of Spodoptera frugiperda larvae

【目的】明确淡足侧沟茧蜂Microplitis pallidipes寄生对草地贪夜蛾Spodoptera frugiperda血淋巴黑化率、酚氧化酶原(Prophenoloxidase,PPO)基因表达量和酚氧化酶(Phenoloxidase,PO)活性的影响,为更有效的利用淡足侧沟茧蜂防控草地贪夜蛾提供理论依据。【方法】通过室内寄生实验确定草地贪夜蛾被淡足侧沟茧蜂寄生的最佳龄期;通过RT-qPCR分析草地贪夜蛾两个PPO基因(SfPPO1和SfPPO2)的时空表达谱;通过薄膜法统计草地贪夜蛾被寄生后的血淋巴黑化率;采用吸光光度法测定草地贪夜蛾被寄生后的血淋巴PO活性;采用RT-qPCR测定草地贪夜蛾被寄生后血淋巴SfPPO1和SfPPO2的相对基因表达量。【结果】草地贪夜蛾被淡足侧沟茧蜂寄生的最佳龄期为3龄幼虫,SfPPO1和SfPPO2均在卵块和4龄幼虫中基因表达量较高,且主要在幼虫血淋巴中表达。草地贪夜蛾被寄生后12、24、48和72 h的血淋巴黑化率显著低于未寄生组(P<0.05),SfPPO1和SfPPO2的相对基因表达量和PO活性也低于未寄生组。相关关系分析表明,SfPPO1的相对基因表达量与SfPPO2的表达量呈现线性关系(R2=0.9124),PO活性与两个PPO基因相对表达量之间、黑化率与PO活性和PPO基因相对表达量之间均呈现正相关。【结论】淡足侧沟茧蜂寄生会抑制草地贪夜蛾血淋巴的黑化率,下调血淋巴中酚氧化酶原基因的表达量和PO活性,且PO活性、PPO基因相对表达量及血淋巴黑化率3个指标之中,两两之间均呈现正相关性。

[Objectives]To investigate the effects of Microplitis pallidipes on the menanization,prophenoloxidase(PPO)gene expression and phenoloxidase(PO)activity,of the hemolymph of Spodoptera frugiperda larvae,and thereby provide a foundation for the more effective use of this endoparasitoid to control this pest.[Methods]The S.frugiperda larval instar most susceptible to parasitism by M.pallidipes was determined experimentally and RT-qPCR was then used to analyze the temporal and spatial expression profiles of two S.frugiperda PPO genes.The menanization rate of larvae was counted using the film method,and the PO activity of larvae was measured using the absorbance method.The expression of SfPPO1 and SfPPO2 in the hemolymph of S.frugiperda was determined with RT-qPCR.[Results]3rd instar S.frugiperda larvae were found to be the most susceptible to being parasitized by M.pallidipes.The relative expression of the two SfPPO genes in eggs and 4th instar larvae was relatively high,and both genes were mainly expressed in the hemolymph.Hemolymph menanization rates of parasitized larvae were significantly lower than those of the non-parasitized larvae after 12,24,48 and 72 hours(P<0.05).PPO expression and PO activity of parasitized larvae were also lower than those of non-parasitized larvae.There was a strong,positive linear correlation between the expression of PPO1 and PPO2(R2=0.9124),and there were also positive correlations among PO activity,PPO expression and the menanization rate.[Conclusion]Parasitism of M.pallidipes inhibited the menanization of S.frugiperda larvae and down-regulated the expression of two PPO genes and PO activity.PO activity,PPO gene expression and menanization rate were all positively correlated.