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Detection of Babesia gibsoni in dogs by combining recombinase polymerase amplification (RPA) with lateral flow (LF) dipstick

  • Jie Cui
  • Yangnan Zhao
  • Yali Sun
  • Long Yu
  • Qin Liu
  • Xueyan Zhan
  • Muxiao Li
  • Lan He
  • Junlong Zhao
Original Paper

Abstract

Babesia gibsoni is a protozoan parasite responsible for the majority of reported cases of canine babesiosis in China. Currently, microscopic examination of the Giemsa-stained thin blood smears is the main diagnosis method in clinic. Here, we report the recombinase polymerase amplification-lateral flow (LF-RPA) dipstick detection method for targeting B. gibsoni cytochrome c oxidase subunit I (cox I) gene. The reaction takes only 20–30 min under isothermal temperatures between 30 and 45 °C. Specificity was evaluated using DNA from related apicomplexan parasites and their host, while the sensitivity was calculated based on the DNA from the experimental B. gibsoni-infected dogs. Results indicated that the LF-RPA method is 20 times more sensitive than the conventional PCR based on 18S rRNA and has no cross reaction with any other test DNAs. The applicability of the LF-RPA method was further evaluated using 15 samples collected from clinic. Thirteen of the 15 samples (86.67%) were detected as positive by LF-RPA, while 10 of them (66.67%) were found positive by conventional PCR. Overall, the novel LF-RPA assay is effective for the detection of B. gobsini and has considerable advantages over the conventional PCR in sensitivity, specificity, simplicity in operation, less time consumption, and visual detection. The LF-RPA method may facilitate the surveillance and early detection of B. gibsoni infection in dogs.

Keywords

Babesia gibsoni Recombinase polymerase amplification Lateral flow Diagnosis Babesiosis 

Notes

Acknowledgments

The authors would like to thank the members of our lab.

Funding information

This work was supported by the National Key Research and Development program of China (2017YFD0501201), the National Key Basic Research Program (973 program) of China (Grant No. 2015CB150300), and the Natural Science Foundation of Hubei Province (2017CFA020).

Compliance with ethical standards

This study was approved by the Scientific Ethic Committee of Huazhong Agricultural University. All pet dogs were handled in accordance with the Animal Ethics Procedures and Guidelines of the People’s Republic of China (Permit number: HZAUCA-2016-007). All blood samples were collected under the owner’s informed consent.

Conflict of interest

The authors declare that they have no conflict of interest.

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Jie Cui
    • 1
    • 2
  • Yangnan Zhao
    • 1
    • 2
  • Yali Sun
    • 1
    • 2
  • Long Yu
    • 1
    • 2
  • Qin Liu
    • 1
    • 2
  • Xueyan Zhan
    • 1
    • 2
  • Muxiao Li
    • 1
    • 2
  • Lan He
    • 1
    • 2
    • 3
  • Junlong Zhao
    • 1
    • 2
    • 3
  1. 1.State Key Laboratory of Agricultural Microbiology, College of Veterinary MedicineHuazhong Agricultural UniversityWuhanPeople’s Republic of China
  2. 2.Key Laboratory for Development of Veterinary Diagnostic Products, Ministry of AgricultureHuazhong Agricultural UniversityWuhanPeople’s Republic of China
  3. 3.Key Laboratory of Preventive Veterinary Medicine in Hubei ProvinceThe Cooperative Innovation Center for Sustainable Pig ProductionWuhanChina

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