Improved Methods for the Diagnosis of African Trypanosomosis

Rebeski, Dierk E.; Winger, E.M; Rogovic, B; Robinson, M.M; Crowther, John R.; Dwinger, R.H.

doi:10.1590/s0074-02761999000200024

Cited by 19 publications

(5 citation statements)

References 12 publications

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“…While this issue is non-existent in antigen-based assays, this immunoassay type has its own requirements, especially with regards to the target antigen or biomarker. The target antigen should be (i) stable in the host environment and produced over the entire course of the infection, (ii) available at concentrations that allow for its detection, (iii) specific for a given pathogen, and (iv) the capture and detection antibodies targeting the antigen should be able to out-compete infection-induced host anti-pathogen antibodies [25,[48][49][50][51][52][53]. Previously developed antigen-based immunoassays for T. evansi detection operated via an "unbiased" methodology; i.e., raising antibodies against whole parasites or preparations of specific parasite fractions without prior knowledge of the target antigen [19,54,55].…”

Section: Discussionmentioning

confidence: 99%

An Unbiased Immunization Strategy Results in the Identification of Enolase as a Potential Marker for Nanobody-Based Detection of Trypanosoma evansi

et al. 2020

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Trypanosoma evansi is a widely spread parasite that causes the debilitating disease “surra” in several types of ungulates. This severely challenges livestock rearing and heavily weighs on the socio-economic development in the affected areas, which include countries on five continents. Active case finding requires a sensitive and specific diagnostic test. In this paper, we describe the application of an unbiased immunization strategy to identify potential biomarkers for Nanobody (Nb)-based detection of T. evansi infections. Alpaca immunization with soluble lysates from different T. evansi strains followed by panning against T. evansi secretome resulted in the selection of a single Nb (Nb11). By combining Nb11-mediated immuno-capturing with mass spectrometry, the T. evansi target antigen was identified as the glycolytic enzyme enolase. Four additional anti-enolase binders were subsequently generated by immunizing another alpaca with the recombinant target enzyme. Together with Nb11, these binders were evaluated for their potential use in a heterologous sandwich detection format. Three Nb pairs were identified as candidates for the further development of an antigen-based assay for Nb-mediated diagnosis of T. evansi infection.

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Section: Discussionmentioning

confidence: 99%

An Unbiased Immunization Strategy Results in the Identification of Enolase as a Potential Marker for Nanobody-Based Detection of Trypanosoma evansi

et al. 2020

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“…Apart from not cross-reacting in diagnostic test for the detection of other endemic parasites, the target antigen should be present in detectable quantities during an active infection [ 23 , 49 ]. An additional requirement is that the capture and detection antibodies targeting the antigen should be able to out-compete the host anti-pathogen antibodies induced during infection [ 15 , 23 ]. In this paper, we have devised a Nb-based approach that could circumvent the above-mentioned issues hampering the development of antigen-based pathogen detection.…”

Section: Discussionmentioning

confidence: 99%

“…It has been documented that host antibodies usually sequester parasite antigens [ 15 ]. Given that MAbs are conventional antibodies, the host IgG molecules might conceal the epitopes from the MAbs employed in the diagnostic test [ 23 ]. Nb-based diagnostics could circumvent this binding interference caused by the host’s antibody response.…”

Section: Introductionmentioning

confidence: 99%

An Anti-proteome Nanobody Library Approach Yields a Specific Immunoassay for Trypanosoma congolense Diagnosis Targeting Glycosomal Aldolase

et al. 2016

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BackgroundInfectious diseases pose a severe worldwide threat to human and livestock health. While early diagnosis could enable prompt preventive interventions, the majority of diseases are found in rural settings where basic laboratory facilities are scarce. Under such field conditions, point-of-care immunoassays provide an appropriate solution for rapid and reliable diagnosis. The limiting steps in the development of the assay are the identification of a suitable target antigen and the selection of appropriate high affinity capture and detection antibodies. To meet these challenges, we describe the development of a Nanobody (Nb)-based antigen detection assay generated from a Nb library directed against the soluble proteome of an infectious agent. In this study, Trypanosoma congolense was chosen as a model system.Methodology/Principal FindingsAn alpaca was vaccinated with whole-parasite soluble proteome to generate a Nb library from which the most potent T. congolense specific Nb sandwich immunoassay (Nb474H-Nb474B) was selected. First, the Nb474-homologous sandwich ELISA (Nb474-ELISA) was shown to detect experimental infections with high Positive Predictive Value (98%), Sensitivity (87%) and Specificity (94%). Second, it was demonstrated under experimental conditions that the assay serves as test-of-cure after Berenil treatment. Finally, this assay allowed target antigen identification. The latter was independently purified through immuno-capturing from (i) T. congolense soluble proteome, (ii) T. congolense secretome preparation and (iii) sera of T. congolense infected mice. Subsequent mass spectrometry analysis identified the target as T. congolense glycosomal aldolase.Conclusions/SignificanceThe results show that glycosomal aldolase is a candidate biomarker for active T. congolense infections. In addition, and by proof-of-principle, the data demonstrate that the Nb strategy devised here offers a unique approach to both diagnostic development and target discovery that could be widely applied to other infectious diseases.

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“…Only for T. b. gambiense , monitoring tools are available for both detection and staging of the disease (4, 31–33). Existing field applicable antibody-based diagnostic tests still suffer from a lack of positive predictive value and cannot differentiate between active or cured infections (32, 34, 35). Direct diagnosis aimed at parasite antigen detection is often hampered by sequestration of parasite antigens by the host’s antibodies or by concealing of epitopes from the diagnostic monoclonal antibodies (mAbs) (36).…”

Section: Introductionmentioning

confidence: 99%

Nanobodies As Tools to Understand, Diagnose, and Treat African Trypanosomiasis

et al. 2017

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African trypanosomes are strictly extracellular protozoan parasites that cause diseases in humans and livestock and significantly affect the economic development of sub-Saharan Africa. Due to an elaborate and efficient (vector)–parasite–host interplay, required to complete their life cycle/transmission, trypanosomes have evolved efficient immune escape mechanisms that manipulate the entire host immune response. So far, not a single field applicable vaccine exists, and chemotherapy is the only strategy available to treat the disease. Current therapies, however, exhibit high drug toxicity and an increased drug resistance is being reported. In addition, diagnosis is often hampered due to the inadequacy of current diagnostic procedures. In the context of tackling the shortcomings of current treatment and diagnostic approaches, nanobodies (Nbs, derived from the heavy chain-only antibodies of camels and llamas) might represent unmet advantages compared to conventional tools. Indeed, the combination of their small size, high stability, high affinity, and specificity for their target and tailorability represents a unique advantage, which is reflected by their broad use in basic and clinical research to date. In this article, we will review and discuss (i) diagnostic and therapeutic applications of Nbs that are being evaluated in the context of African trypanosomiasis, (ii) summarize new strategies that are being developed to optimize their potency for advancing their use, and (iii) document on unexpected properties of Nbs, such as inherent trypanolytic activities, that besides opening new therapeutic avenues, might offer new insight in hidden biological activities of conventional antibodies.

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Improved Methods for the Diagnosis of African Trypanosomosis

Cited by 19 publications

References 12 publications

An Unbiased Immunization Strategy Results in the Identification of Enolase as a Potential Marker for Nanobody-Based Detection of Trypanosoma evansi

An Unbiased Immunization Strategy Results in the Identification of Enolase as a Potential Marker for Nanobody-Based Detection of Trypanosoma evansi

An Anti-proteome Nanobody Library Approach Yields a Specific Immunoassay for Trypanosoma congolense Diagnosis Targeting Glycosomal Aldolase

Nanobodies As Tools to Understand, Diagnose, and Treat African Trypanosomiasis

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