A Set of Diagnostic Tests for Detection of ActiveBabesia duncaniInfection

Abstract

ABSTRACTHuman babesiosis is a rapidly emerging and potentially fatal tick-borne disease caused by intraerythrocytic apicomplexan parasites of theBabesiagenus. Among the various species ofBabesiathat infect humans,B. duncanihas been found to cause severe and life-threatening infections. Detection of activeB. duncaniinfection is critical for accurate diagnosis and effective management of the disease. While molecular assays for the detection ofB. duncaniinfection in blood are available, a reliable strategy to detect biomarkers of active infection has not yet been developed. Here, we report the development of the firstB. duncaniantigen capture assays that rely on the detection of twoB. duncani-exported immunodominant antigens, BdV234 and BdV38. The assays were validated using blood samples from cultured parasites in human erythrocytes andB. duncani-infected laboratory mice at different parasitemia levels and following therapy. The assays display high specificity with no cross-reactivity withB. microti,B. divergens,BabesiaMO1, orP. falciparum.The assay also demonstrates high sensitivity, detecting as low as 115 infected erythrocytes/µl of blood. Screening of 1,731 blood samples from diverse biorepositories, including previously identified Lyme and/orB. microtipositive human samples and new specimens from field mice, showed no evidence ofB. duncaniinfection in these samples. The assays could be useful in diverse diagnostic scenarios, including point-of-care testing for earlyB. duncaniinfection detection in patients, field tests for screening reservoir hosts, and high-throughput screening such as blood collected for transfusion.Short summaryWe developed two ELISA-based assays, BdACA38 and BdACA234, for detectingB. duncani, a potentially fatal tick-borne parasite causing human babesiosis. The assays target two immunodominant antigens, BdV234 and BdV38, demonstrating high specificity (no cross-reactivity with otherBabesiaspecies orPlasmodium falciparum) and sensitivity (detecting as low as 115 infected erythrocytes/µl). The assays were validated using in vitro-cultured parasites and infected mice. Screening diverse blood samples showed no evidence ofB. duncaniactive infection among 1,731 human and field mice blood samples collected from the north-eastern, midwestern, and western US. These assays offer potential in diverse diagnostic scenarios, including early patient detection, reservoir animal screening, and transfusion-transmitted babesiosis prevention.