Tsetse salivary glycoproteins are modified with paucimannosidic N-glycans, are recognised by C-type lectins and bind to trypanosomes

African sleeping sickness is caused by Trypanosoma brucei , a parasite transmitted by the bite of a tsetse fly. Trypanosome infection induces a severe transcriptional downregulation of tsetse genes encoding for salivary proteins, which reduces its anti-hemostatic and anti-clotting properties. To better understand trypanosome transmission and the possible role of glycans in insect bloodfeeding, we characterized the N -glycome of tsetse saliva glycoproteins. Tsetse salivary N -glycans were enzymatically released, tagged with either 2-aminobenzamide (2-AB) or procainamide, and analyzed by HILIC-UHPLC-FLR coupled online with positive-ion ESI-LC-MS/MS. We found that the N -glycan profiles of T . brucei -infected and naïve tsetse salivary glycoproteins are almost identical, consisting mainly (>50%) of highly processed Man ₃ GlcNAc ₂ in addition to several other paucimannose, high mannose, and few hybrid-type N -glycans. In overlay assays, these sugars were differentially recognized by the mannose receptor and DC-SIGN C-type lectins. We also show that salivary glycoproteins bind strongly to the surface of transmissible metacyclic trypanosomes. We suggest that although the repertoire of tsetse salivary N -glycans does not change during a trypanosome infection, the interactions with mannosylated glycoproteins may influence parasite transmission into the vertebrate host.