Superior real‐time polymerase chain reaction detection of Babesia microti parasites in whole blood utilizing high‐copy BMN antigens as amplification targets

Abstract

Cumulatively, our data demonstrate that RT-PCR detection of the BMN family of seroreactive antigens reflects a sensitive and superior assay for the detection of B. microti in whole blood samples.

Persistence of Babesia microti Infection in Humans

“…Nucleic acid detection offers a better correlate of active infection. Nucleic acid detection based tests (NAT), such as polymerase chain reaction (PCR) and transcription-mediated amplification (TMA), more effectively identify low-level infections than other laboratory tests [15,16,17,18,19]. The molecular methods for Babesia detection generally rely on amplification of the 18S gene, which encodes the small subunit ribosomal RNA gene.…”

“…The molecular methods for Babesia detection generally rely on amplification of the 18S gene, which encodes the small subunit ribosomal RNA gene. Publication of the full B. microti genome in 2012 has enabled identification of higher copy number detection targets, allowing the routine detection of less than 10 B. microti parasites per ml of blood [18,20]. Additional technological advances such as sample concentration, high copy detection targets such as BMN multigene family members, and bead-based target capture may further enhance the sensitivity of NAT assays [21,22].…”

“…Detection of Babesia antigen(s) offers additional markers of active infection. Efforts should be made to identify excreted/secreted B. microti antigen to develop high throughput detection assays for diagnostic and donor screening purposes [18,22]. Inoculation of patient blood into small mammals such as hamsters, gerbils, or SCID mice for detection of Babesia is less sensitive than PCR and impractical but is useful for investigation of rare variants/strains of Babesia and for blood donor studies [15].…”

“…The guidance also allows for requalification of those donors who are deferred for a history of babesiosis or positive Babesia test result after two years of negative testing. Experimental molecular [17,18] and commercial antibody-based assays [55] have also been developed that could have utility in donor screening.…”

Combined Immunofluorescence (IFA) and Fluorescence In Situ Hybridization (FISH) Assays for Diagnosing Babesiosis in Patients from the USA, Europe and Australia

“…Alternatively, Babesia parasite nucleic acid is detected by qPCR on blood samples [ 3 , 4 ] and the detection of ribosomal RNA within infected red blood cells (iRBCs) by fluorescence in situ hybridization (FISH) [ 9 ]. Several qPCR tests have been developed for B. microti and are reported to detect <10 parasites per μL of blood [ 10 , 11 ]. They are the preferred methods for screening blood for transfusion [ 12 , 13 ].…”