Rapid diagnosis of Theileria annulata by recombinase polymerase amplification combined with a lateral flow strip (LF-RPA) in epidemic regions

- A LF-RPA was firstly developed to detect Theileria annulata infections.
- The sensitivity of LF-RPA is similar to PCR for detecting the field samples.
- The LF-RPA could be used to surveil and control of Theileria annulata.

Rapid and accurate diagnosis of Theileria annulata infection contributes to the formulation of strategies to eradicate this parasite. A simple and efficient diagnostic tool, recombinase polymerase amplification (RPA) combined with a lateral flow (LF) strip, was used in detection of Theileria and compared to other methods that require expensive instruments and skilled personnel. Herein, we established and optimized an LF-RPA method to detect the cytochrome b gene of T. annulata mitochondrial DNA from experimentally infected and field-collected blood samples. This method has many unparalleled characteristics, including that it is rapid (clear detection in 5 min at constant temperature), sensitive (the limitation of detection is at least 2 pg genomic DNA), and specific (no cross-reaction with other piroplasms that infect cattle). The LF-RPA assay was evaluated via testing 17 field blood samples and comparing the results of that of a PCR, showing 100% agreement, which demonstrates the ability of the LF-RPA assay to detect T. annulata infections in small number of samples (n = 17). Taken together, the results indicate that this method could be used as an ideal diagnostic tool for detecting T. annulata in endemic regions with limited to fewer and local resources and could also be a potential technique for the surveillance and control of blood protozoa.