Gut bacteria induce heterologous immune priming in Rhodnius prolixus encompassing both humoral and cellular immune responses

Insects lack the adaptive, antibody mediated responses of vertebrates, yet they possess a robust innate immune system capable of defending themselves against pathogens. Immune priming has been observed in multiple insect species, wherein exposure to a pathogen provides protection against subsequent infections by the pathogen. Heterologous immune priming has also been described, where presence of one bacterial species provides protection against another. We determined that Rhodococcus rhodnii, a gut symbiont of the kissing bug Rhodnius prolixus, induces strong heterologous immune priming, while axenic bugs lacking gut bacteria are highly susceptible to pathogens. Commensal Escherichia coli provides less robust protection. R. rhodnii must be alive within the insect as dead bacteria do not stimulate immune priming and pathogen resistance. Removal of R. rhodnii from the gut reduces resistance to pathogens while restoring it to axenic bugs improves pathogen resistance, though not completely. Unlike most other examples of symbiont-mediated immune priming, we find no evidence that R. rhodnii ever leaves the gut, despite activating a potent immune response in the hemocoel and fat body. R. rhodnii and E. coli activate both the IMD and Toll pathways indicating cross-activation of the pathways, while silencing of either pathway leads to a loss of the protective effect. Several antimicrobial peptides are induced in the fat body by presence of gut bacteria. When E. coli is in the gut, expression of antimicrobial peptides is often higher than when R. rhodnii is present, while R. rhodnii induces proliferation of hemocytes and induces a stronger melanization response than E. coli. Hemolymph from R. rhodnii bugs has a greater ability to convert the melanin precursor DOPA to melanization products than axenic or E. coli-harboring bugs. These results demonstrate that R. rhodnii's benefits to its host extend beyond nutritional provisioning, playing an important role in the host immune system.