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Animal African trypanosomosis (AAT), a disease affecting livestock, is caused by parasites of the  Trypanosoma  genus (mainly  T .  vivax  and  T .  congolense ). AAT is widespread in Sub-Saharan Africa, where it continues to impose a heavy socio-economic burden as it renders development of sustainable livestock rearing very strenuous. Active case-finding and the identification of infected animals prior to initiation of drug treatment requires the availability of sensitive and specific diagnostic tests. In this paper, we describe the development of two heterologous sandwich assay formats (ELISA and LFA) for  T .  congolense  detection through the use of Nanobodies (Nbs). The immunisation of an alpaca with a secretome mix from two  T .  congolense  strains resulted in the identification of a Nb pair (Nb44/Nb42) that specifically targets the glycolytic enzyme pyruvate kinase. We demonstrate that the Nb44/Nb42 ELISA and LFA can be employed to detect parasitaemia in plasma samples from experimentally infected mice and cattle and, additionally, that they can serve as ‘test-of-cure’ tools. Altogether, the findings in this paper present the development and evaluation of the first Nb-based antigen detection LFA to identify active  T .  congolense  infections.

Development of a Nanobody-based lateral flow assay to detect active Trypanosoma congolense infections