Rapid Detection of Babesia motasi Responsible for Human Babesiosis by Cross-priming Amplification combined with a vertical flow

Abstract

Background: Babesia motasi is known as an etiological agent of human and ovine babesiosis. Diagnosis of babesiosis is traditionally performed by microscopy, examining Giemsa-stained thin peripheral blood smears. Rapid detection and accurate identification of species are desirable for clinical care and epidemiological studies. Methods: An easy to operate molecular method, which requires less capital equipment and incorporates cross priming amplification combined with a vertical flow (CPA-VF) visualization strip for rapid detection and identification of B. motasi. Results: The CPA-VF targets the 18S rRNA gene and has a detection limit of 50 fg per reaction; no cross reaction was observed with other piroplasms infective to sheep or Babesia infective to humans. CPA-VF and real-time (RT)-PCR had sensitivities of 95.2% (95% confidence interval [CI], 78.1-99.4%) and 90.5% (72-97.6%) and specificities of 95.8 (80.5-99.5%) and 97.9 (83.5-99.9%), respectively, versus microscopy and nested (n) PCR combined with gene sequencing. The clinical performance of the CPA-VF assay was evaluated with field blood samples from sheep (n = 240) in Jintai county, Gansu Province, and clinical specimens (n = 492) obtained from patients bitten by ticks. Conclusions: Our results indicate that the CPA-VF is a rapid, accurate, nearly instrument-free molecular diagnostic approach for identification of B. motasi. Therefore, it could be an alternative technique for epidemiological investigations and diagnoses of ovine and/or human babesiosis caused by B. motasi, especially in resource-limited regions.Keywords: Ovine babesiosis, human babesiosis, Babesia motasi, cross priming amplification, vertical flow visualization strip, detection, identification