Development of insect resistance to biopesticides is a current and pertinent global issue. Earlier, it was established that lepidopteran larvae can recover from Bt intoxication via a midgut regenerative response and subsequently generate resistance. Molecular aspects of restoration of the midgut integrity following toxin exposure are emerging recently. In the present study, we bring out the pivotal role of gut arylphorin in mediating the midgut regenerative response following sublethal Bt exposure in Achaea janata. Bt-induced midgut damage was characterized by microscopic analysis using differential interference contrast (DIC) and immunofluorescence (IF). Extensive disruption of brush-border membrane, associated with the underlying cytoskeletal alterations including F-actin, α-actin and β-tubulin was observed. Single-photon fluorescence microscopy combined with fluorescence lifetime imaging (FLIM) established the metabolic state associated with enhanced stem cell proliferation and migration from the basal side towards the luminal side following the damage. In-silico analysis revealed the phylogenetic relationship of gut arylphorin with closely related insect species and indicated the presence of two different subunits. Transient RNAi knockdown of the arylphorin resulted in diminished expression of mitotic Cyclin B mRNA levels. Human monoclonal Cyclin B antibody cross-reactivity with the Cyclin B located in the stem cells further validate the role of arylphorin as the mitogenic factor responsible for stem cell proliferation and epithelial regeneration. An in-depth understanding of resistance mechanisms will aid in the design of new strategies for the long-term usage and efficacy of Bt technology against pest control.

昆虫对生物农药的抗药性的发展是当前和相关的全球性问题。早些时候，已经确定鳞翅目幼虫可以通过中肠再生反应从 Bt 中毒中恢复，并随后产生抗性。最近出现了毒素暴露后中肠完整性恢复的分子方面。在本研究中，我们揭示了Achaea janata亚致死 Bt 暴露后肠道 arylphorin 在介导中肠再生反应中的关键作用. Bt 诱导的中肠损伤的特征是使用微分干涉对比 (DIC) 和免疫荧光 (IF) 进行显微镜分析。观察到与包括 F-肌动蛋白、α-肌动蛋白和 β-微管蛋白在内的潜在细胞骨架改变相关的刷状缘膜的广泛破坏。单光子荧光显微镜结合荧光寿命成像 (FLIM) 建立了与增强的干细胞增殖和损伤后从基底侧向管腔侧迁移相关的代谢状态。硅片分析揭示了肠道 arylphorin 与密切相关的昆虫物种的系统发育关系，并表明存在两个不同的亚基。对 arylphorin 的瞬时 RNAi 组合式导致有丝分裂细胞周期蛋白 B mRNA 水平的表达减少。人单克隆细胞周期蛋白 B 抗体与位于干细胞中的细胞周期蛋白 B 的交叉反应进一步验证了芳基福林作为负责干细胞增殖和上皮再生的有丝分裂因子的作用。深入了解抗性机制将有助于设计长期使用 Bt 技术对抗害虫防治的新策略和有效性。