Tissue- and time-dependent deployment of Toll- and Imd-associated immune responses in Manduca sexta larvae.

Little is known about how the Toll and Imd pathways are deployed across tissues and over time in insects outside the Drosophila lineage. To address this, we performed a systematic analysis of immune gene responses in Manduca sexta using deep RNA-seq of fat body, hemocytes, and midgut after aseptic wounding or microbial injection, identifying 1895 genes strongly induced by injury or infection. Immune challenge triggered extensive transcriptional reprogramming, including induction of pattern recognition receptors, serine protease cascades, cytokines, antimicrobial peptides (AMPs), and core immune signaling pathway components, whereas wounding elicited a more restricted immune response characterized primarily by AMP induction. Comparison among tissues revealed striking specialization of immune and physiological programs. Fat body and hemocytes exhibited coordinated immune activation accompanied by suppression of metabolic functions, whereas midgut displayed a distinct profile. To examine temporal patterns associated with Toll- and Imd-biased responses, we analyzed AMP induction kinetics after challenge with live or heat-killed bacteria and fungi. Strongly Toll-biased AMP genes showed delayed induction, whereas Imd-biased genes exhibited more heterogeneous temporal patterns. Moreover, Toll- and Imd-associated gene sets were enriched among early-induced genes in fat body and hemocytes but not in midgut, where early responses were dominated by Imd-associated genes. These results provide a tissue- and time-resolved analysis of Toll- and Imd-associated transcriptional responses in M. sexta and reveal substantial spatial and temporal diversification in the deployment of Toll- and Imd-associated immune programs under the conditions examined.