Abstract:
Innate immunity has a key role in the control of microbial infections in both vertebrates and invertebrates. In insects, including vectors that transmit parasites that cause major diseases such as trypanosomosis, leishmaniasis and filariasis, antimicrobial peptides and agglutinins form an important component of innate immunity and participate in regulating parasite development. In this study, induced haemolymph peptides from a non-vector, nonheamatophagous insect, Schistocerca gregaria were assessed using one and two dimensional gel electrophoresis following Trypanosoma brucei inoculation. The pattern of protein induction was assessed by inoculating the locusts with parasites followed by haemolymph collection at 0, 6, 18, 24, 30, 42, and 48 hour. The amount of protein in each sample was quantified and found to increase with time, with the 18 hour sample having the highest protein concentration. On analysis using SDS-PAGE, five peptides were found to differ in terms of their presence and relative abundance in all the samples following T. brucei challenge. Following 2D-PAGE, some peptides were found to be induced, enhanced or suppressed while others were unaffected. In vitro assays were performed to ascertain the extent of trypanosomes lysis by incubating the parasites with various haemolymph samples obtained after challenge with the parasites. This further indicated that lysis increased with increasing protein Concentrate on, with complete lysis (100%) being attained in the 18 hour sample after 75 minutes. The effects of sugars on the induction of the proteins were determined by inoculating the insects with the parasites and introducing the sugars (500mM of Dglucosamine, D-galactose, D-glucose and N-acetylglucosamine) after 30 minutes. Samples collected 18 hours later were subjected to protein quantification followed by an in vitro lysis assay. The sugar, D-glucosamine was found to have the highest inhibition with D-galactose having the least effect on the induction. An approximated 80% lysis was observed with 0.5mg/ml of the 18 hour sample treated with D-galactose and only 10% lysis with Dglucosamine. Further analysis was carried out by subjecting the samples to immunodetection with antibodies raised against Glossina proteolytic lectin (Gpl), an induced midgut lectin found among the Glossina spp. and none of the samples collected post T. brucei challenge showed cross-reactivity. These induced proteins have the potential of being used to modulate tsetse fly vectorial competence.
