Immunization Studies in Rabbits Using Midgut Membrane Bound Protein Derived from rhipicephalus Appendiculatus Neumann, rhipicephalus Rvertsi evertsiNeumann and Amblyomma variegatum fabricius

Abstract

The objective of this study was to immunize rabbits with midgut membrane-bound proteins derived from partially engorged Rhipicephalus appendiculatus, ~. evertsi evertsi and Amblyomma variegatum female ticks a~d assess whether the immunity elicited was protective against both homologous and heterologous tick ins tars and to isolate and identify the protective antigens. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis of the Gut Membrane-Bound Protein (GMBP) antigens demonstrated protein bands with molecular weights ranging from 14 to 140 kDa. Approximately 37 protein bands were fractionated from ~. appendiculatus GMBP antigens, approximately 45 protein bands ere fractionated from . evertsi evertsi GMBP antigens and approxi ately 39 protein bands were fractionated from !. variegatum GMBP antigens. Twentytwo of the isolated proteins were shared among the three tick species. The ability of rabbits to acquire resistance 'to ~. appendiculatus, R. evertsi evertsi and !. variegatum was determined by injecting three separate se~s of rabbits with respective GMBP antigens. Resistance was manifested by prolonged feeding, reduction in engorgement weights, egg mass weights, mou ting and percentage hatchability and increa ed xvii mortality. Cross-resistance was evaluated by dividing !. appendiculatus, !. evertsi evertsi and !. variegatum resistant rabbits into three groups each and challenging them with homologous and heterologous live stages. Considerably high cross-resistance was apparent among the three groups. Cross-protection was more pronounced in the homologous than heterologous systems. Enzyme Linked Immunosorbent Assay. (ELISA) technique detected circulating antibodies in the immune sera to GMBP from homologous and heterologous systems one week after the primary dose. Ouchterlony double immunodiffusion reactions with anti-tick GMBP sera formed 2 to 4 precipitin lines with homologous GMBP antigens and 1 to 2 precipitin line(s) with each heterologous GMBP antigens. A line of complete identity was observed when immune sera to GMBP antigens reacted with GMBP from homologous and heterologous tick species, suggesting common antigenic epitopes. Western blot analysis on GMBP of R. appendiculatus, !. evertsi evertsi and !. variegatum with sera from immunized rabbits detected protein bands specific to the homologous GMBP antigens, and revealed considerable cross-reactions in the heterologous systems. In conclusion, there was prolonged feeding periods, reduced engorged weights, egg mass weights XVlll hatchability and moulting and increased death rate of both homologous and heterologous challenge ticks which fed on resistant rabbits. This was due to the presence of common antigens. The presence of cross-reacting antigens conferred cross-protection. These results have pointed out that it is possible to protect livestock from R. appendiculatus, !. evertsi evertsi and !. variegatum using an antigen from anyone of the three tick species hence reducing the expence of having to develop an antigen to control each tick species as there are in existence.