Expression, immunolocalization and serodiagnostic value of Tc38630 protein from Trypanosoma congolense

Abstract

Animal African trypanosomosis is a serious constraint to livestock sector development in sub-Saharan Africa. The disease, mainly caused by Trypanosoma congolense, has a limitation in its diagnosis and treatment. There is urgent need for a simple, rapid detection technique to replace the few available serological tests that are of variable sensitivity and specificity. Currently, there is a promising use of recombinant proteins to improve on the trypanosome lysate to detect antibodies. In this respect, we have identified a stage-specific gene that is relatively highly expressed in metacyclic and blood trypomastigotes of T. congolense. According to previously obtained differential protein expression data, the gene TcIL3000.0.38630 (1,236 bp) is by 8.5 times more expressed in metacyclic and blood trypomastigotes than in procyclic trypomastigotes and epimastigotes. The same stage specific expression pattern was shown in Western blot analysis. In addition, in confocal laser scanning microscopy the Tc38630 protein was present in the cytosol and on the cell surface of metacyclic and blood trypomastigotes. Through bioinformatics, the Tc38630 had N-terminal signal sequence, hydrophilic extracellular domain, single transmembrane alpha-helix and short cytoplasmic domain, which is characteristic of the Trypanosoma brucei invariant surface glycoprotein. However, unlike T. brucei invariant surface glycoprotein, the Tc38630 existed as a single copy gene with a probable allelic polymorphism at the Nar I restriction site. The recombinant Tc38630-based ELISA detected antibodies against Tc38630 as early as 7 days post infection in experimentally infected mouse model. Taken together, our results suggest that the Tc38630 is a novel potential diagnostic antigen of Animal African trypanosomosis.

Proteomic Selection of Immunodiagnostic Antigens for Trypanosoma congolense

“…It is worth noting that it has been previously reported that this antigen does not cross react with the common non-pathogenic cattle trypanosome T. theileri [11].…”

“…Nevertheless, three proteins belonging to the invariant surface glycoprotein (ISG) family (Tc38630, Tc45510 and Tc29290) were successfully identified as potential diagnostic antigens by proteomic selection followed by assessment of likely ease of expression in E. coli followed by protein expression trials. The best performing of these, Tc38630, was also recently reported as a potential diagnostic antigen for T. congolense infections using experimental mouse sera [11]. Although annotated as hypothetical in the genome database, Tc38630 has been previously identified by quantitative proteomics as being most abundant on the metacyclic and bloodstream trypomastigote forms of T. congolense [17].…”

“…The surface location, reasonable abundance and lack of similarity with any mammalian proteins most likely all contribute to the immunogenicity of ISGs in animal infections. Indeed, ISGs have been proposed and/or used as diagnostic antigens for human or animal infections with T. brucei gambiense [7], [21], T. evansi [22] and T. congolense [11]. Nevertheless, choosing which of the many (49 in the case of T. congolense ) ISG protein sequences in each trypanosome species [23] to select for protein expression and ELISA trials is a daunting task without the prioritisation provided by the proteomic approach taken in [7] for T. brucei and in this paper for T. congolense .…”

“…The two best performing antigens were both ISG family members, one of which (Tc38630) has been recently reported as a potential diagnostic antigen for T. congolense infections using mouse sera [11]. In this study, we extensively characterise a recombinant Tc38630 construct against diagnostically relevant cattle sera and demonstrated its potential for the diagnosis of T. congolense infections in bovines.…”

Proteomic Identification of Immunodiagnostic Antigens for Trypanosoma vivax Infections in Cattle and Generation of a Proof-of-Concept Lateral Flow Test Diagnostic Device

“…brucei gambiense and cattle T . congolense infections [ 10 – 12 ]. One of these antigens, a recombinant invariant surface glycoprotein (rISG65-1), has been selected by the Foundation for Innovative New Diagnostics (FIND) for development of a next-generation ‘all recombinant’ LFT for human African trypanosomiasis.…”