A Fluorescent Recombinase Aided Amplification Assay for Detection of Babesia microti

Abstract

<i>Babesia microti</i> is one of the most common causative agents of babesiosis. A sensitive and rapid detection is necessary for screening potentially infected individuals. In this study, <i>B. microti</i> cytochrome <i>c</i> oxidase subunit I (<i>cox</i>1) was selected as the target gene, multiple primers were designed, and optimized by a recombinase-aided amplification (RAA) assay. The optimal primers and probe were labeled with fluorescein. The sensitivity of fluorescent RAA (fRAA) was evaluated using gradient diluents of the <i>cox</i>1 recombinant plasmid and genomic DNA extracted from whole blood of <i>B. microti</i> infected mice. The specificity of fRAA was assessed by other transfusion transmitted parasites. The analytical sensitivity of the fRAA assay was 10 copies of recombinant plasmid per reaction and 10 fg/µl <i>B. microti</i> genomic DNA. No cross-reaction with any other blood-transmitted parasites was observed. Our results demonstrated that the fRAA assay would be rapid, sensitive, and specific for the detection of <i>B. microti</i>.