A significant study on the detection of trypanosome parasites in livestock has been published as a peer-reviewed preprint on eLife. The study utilizes a CRISPR-based diagnostic tool, SHERLOCK4AAT, to detect various trypanosome species causing Animal African Trypanosomiasis (AAT, also known as Nagana), advancing AAT diagnostics and drawing attention in the fields of epidemiology, public health, and veterinary medicine.

AAT, caused by Trypanosoma parasites, puts over one million livestock at risk across 37 African countries, with substantial economic impacts. In endemic regions, AAT causes approximately $4.75 billion in agricultural GDP losses annually. Moreover, these livestock are potential hosts for human trypanosome infections, making it critical to map and monitor disease transmission.

Current methods for diagnosing AAT suffer from insufficient sensitivity and reliability. Co-first author Roger-Junior Eloiflin, a research assistant at the INTERTRYP Research Center of the University of Montpellier (CIRAD/IRD), noted that accurate monitoring of livestock infection status using reliable diagnostic tools is crucial for achieving and sustaining the World Health Organization's disease elimination goals.

To overcome the limitations of existing methods, the research team adapted the SHERLOCK (Specific High-sensitivity Enzymatic Reporter unLOCKing) detection method, previously successful in detecting Human African Trypanosomiasis (HAT), to develop SHERLOCK4AAT, which is capable of detecting animal trypanosomes from a single dried blood spot.

During development, the researchers identified DNA targets in highly conserved and species-specific regions of trypanosomes, testing designs using 18S ribosomal RNA and GAPDH gene sequences for species identification. For pan-trypanosome detection, the initially selected single target achieved 93% specificity for detecting HAT in blood, with sensitivity improved after adding a second guide RNA for multiplex detection. For species-specific testing, although no single gene target could distinguish all species, SHERLOCK4AAT successfully differentiated closely related AAT-causing species, with a detection limit ranging from 10 to 1,000 parasites per milliliter, comparable to existing molecular tests. However, species-specific testing for Trypanosoma vivax performed less effectively, indicating the need for different gene targets.

After establishing the test's sensitivity in the laboratory, the research team applied the SHERLOCK4AAT toolkit to analyze trypanosome species in free-range and farm pigs in Côte d'Ivoire and Guinea. The results showed that 62.7% of pigs were infected with at least one trypanosome species, with Trypanosoma brucei gambiense (a trypanosome capable of infecting humans) detected in pigs at both locations, suggesting these pigs may serve as infection hosts.

Co-corresponding author Brice Rotureau, research director at the Parasitology Department of the Institut Pasteur, Paris, concluded that the findings align with limited studies on trypanosome prevalence in domestic pigs, confirming the effectiveness of the SHERLOCK4AAT toolkit in analyzing trypanosome distribution in regions where HAT remains at low levels. He added that, given pigs’ proximity to humans and ease of frequent sampling, the toolkit can use pigs as sentinels to monitor the transmission of human-infecting trypanosomes.