Recombinase polymerase amplification lateral flow dipstick (RPA-LF) detection of Babesia orientalis in water buffalo (Bubalus babalis, Linnaeus, 1758)

Babesiosis caused by Babesia orientalis, an intraerythrocytic apicomplexan protozoan, is one of the most important diseases for water buffalo in central and southern China, leading to huge economic losses, and its main diagnostic method is microscopic examination. In this study, a recombinase polymerase amplification - lateral flow dipstick (RPA-LF) assay, targeting the mitochondrial COXI gene of B. orientalis, was developed to detect B. orientalis in water buffalo. The RPA-LF assay was carried out as an isothermal reaction at 37 degrees C within 15 min. The specificity assay showed no cross-reactivity with other protozoa, and the sensitivity assay revealed the minimum detection limit was 0.25 parasite/mu L, which was 40-fold more sensitive than that of conventional PCR (0.25 versus10 parasites/mu L blood). Moreover, the RPA-LF method was successfully applied to test clinical samples, with no significant difference being observed between RPA-LF and conventional PCR results. Compared with conventional PCR, the novel RPA-LF method had the advantages of simple operation, short time, high sensitivity, and high specificity for B. orientalis detection, indicating the potential use of RPA-LF for rapid field detection of B. orientalis.

Automated summary

A recombinase polymerase amplification - lateral flow dipstick (RPA-LF) assay targeting the mitochondrial COXI gene of Babesia orientalis was developed, showing high sensitivity and specificity for rapid detection in water buffalo. The method is simple to operate, time-efficient, and performed well in clinical sample testing.