Targeting the tsetse-trypanosome interplay using genetically engineered Sodalis glossinidius

Author summaryTsetse flies are the main vectors of African trypanosomiasis, a group of diseases caused by the protozoan Trypanosoma parasites which poses a severe burden on human/animal health and agricultural development in sub-Saharan Africa. The lack of prophylactic drugs and limitations of the existing control programs urge the need for developing alternative strategies to complement the existing control programs. Paratransgenesis, the genetic manipulation of insect symbiotic microorganisms to block pathogen transmission, is a promising strategy for controlling vector-borne diseases. In this experimental study we successfully genetically modified the tsetse fly gut symbiont S. glossinidius to express trypanosome-interfering proteins (i.e. Nanobodies) thereby impairing trypanosome development in the fly. The application of the concept of using pathogen-targeting Nbs delivered by insect symbiotic bacteria could be extended to other vector-pathogen systems. Furthermore, our symbiont-based Nb delivery system can also be applied as a powerful laboratory tool to unravel the molecular determinants of the vector-pathogen association. Sodalis glossinidius, a secondary bacterial symbiont of the tsetse fly, is currently considered as a potential delivery system for anti-trypanosomal components interfering with African trypanosome transmission (i.e. paratransgenesis). Nanobodies (Nbs) have been proposed as potential candidates to target the parasite during development in the tsetse fly. In this study, we have generated an immune Nb-library and developed a panning strategy to select Nbs against the Trypanosoma brucei brucei procyclic developmental stage present in the tsetse fly midgut. Selected Nbs were expressed, purified, assessed for binding and tested for their impact on the survival and growth of in vitro cultured procyclic T. b. brucei parasites. Next, we engineered S. glossinidius to express the selected Nbs and validated their ability to block T. brucei development in the tsetse fly midgut. Genetically engineered S. glossinidius expressing Nb_88 significantly compromised parasite development in the tsetse fly midgut both at the level of infection rate and parasite load. Interestingly, expression of Nb_19 by S. glossinidius resulted in a significantly enhanced midgut establishment. These data are the first to show in situ delivery by S. glossinidius of effector molecules that can target the trypanosome-tsetse fly crosstalk, interfering with parasite development in the fly. These proof-of-principle data represent a major step forward in the development of a control strategy based on paratransgenic tsetse flies. Finally, S. glossinidius-based Nb delivery can also be applied as a powerful laboratory tool to unravel the molecular determinants of the parasite-vector association.