The humoral response plays a crucial role in insect defense against parasites and pathogens, typically producing antimicrobial peptides through the Toll, IMD, and Jak-STAT signaling pathways, as well as melanization via phenoloxidases. However, many studies use nonpathogenic or opportunistic organisms and often infect insects in nonnatural ways, such as piercing or injecting the pathogen into the hemocoel. The objective of this study was to examine the modulation of the main humoral pathway genes involved in the interaction between the nonmodel organism Diatraea saccharalis (the sugarcane borer) and different biological control agents. We identified and evaluated the expression of DsDorsal (Toll pathway), DsRelish (IMD pathway), DsSTAT (JAK/STAT pathway), DsPPO1, and DsPPO2 (PO pathway) in larvae and pupae of D. saccharalis exposed or not to different biological control agents. The biocontrol agents used were: (i) the bacterium Bacillus thuringiensis var. aizawai GC-91, which is pathogenic to D. saccharalis
(ii) the fungus Metarhizium anisopliae ESALQE9 strain, which is employed to control the froghoppers of the genus Mahanarva in sugarcane fields, though it exhibits low virulence to D. saccharalis
and (iii) the generalist parasitoid Tetrastichus howardi. Our results demonstrate that B. thuringiensis at LC