Abstract
BackgroundThe only available diagnostic method for East African trypanosomiasis is light microscopy of blood samples. A simple immunodiagnostic would greatly aid trypanosomiasis control.Methodology and Principal FindingsTo find trypanosome proteins that are specifically recognised by sera from human sleeping sickness patients, we have screened the Trypanosoma brucei brucei proteome by Western blotting. Using cytosolic, cytoskeletal and glycosomal fractions, we found that the vast majority of abundant trypanosome proteins is not specifically recognised by patient sera. We identified phosphoglycerate kinase (PGKC), heat shock protein (HSP70), and histones H2B and H3 as possible candidate diagnostic antigens. These proteins, plus paraflagellar rod protein 1, rhodesain (a cysteine protease), and an extracellular fragment of the Trypanosoma brucei nucleoside transporter TbNT10, were expressed in E. coli and tested for reactivity with patient and control sera. Only TbHSP70 was preferentially recognized by patient sera, but the sensitivity and specificity were insufficient for use of TbHSP70 alone as a diagnostic. Immunoprecipitation using a native protein extract revealed no specifically reacting proteins.ConclusionsNo abundant T. brucei soluble, glycosomal or cytoskeletal protein is likely to be useful in diagnosis. To find useful diagnostic antigens it will therefore be necessary to use more sophisticated proteomic methods, or to test a very large panel of candidate proteins.
Highlights
Human African trypanosomiasis is transmitted by Tsetse flies and occurs in a broad band from south of the Sahara to Zimbabwe
African trypanosomes are coated by Variant Surface Glycoprotein (VSG), and have many VSG genes and pseudogenes which are sequentially expressed during infection
Immunoblotting of Sub-Cellular Fractions Our aim was to find a set of trypanosome proteins that are recognised by sera from patients with African sleeping sickness, but not by control sera from the same region
Summary
It occurs in two major forms, caused by Trypanosoma brucei gambiense A low-cost, simple and rapid diagnosis method with high sensitivity is required: microscopic confirmation and a DNA-based subspecies determination could be restricted to positive cases. Gambiense isolates possess and express at least one of three particular VSG genes. This enabled the development, for T. b. There is no immediate prospect of the use of amplification methods for surveillance of trypanosomiasis in Africa. They are too expensive and timeconsuming, and require too much training for implementing personnel.
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