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Enhanced Detection of Coxiella burnetii with a Complementary Locked Primer-Based Real-Time PCR Method

Molecular Diagnosis & Therapy volume 15, pages 103–107 (2011)Cite this article

Abstract

Background and Objective: Coxiella burnetii is the bacterial causative agent of Q fever in humans. Because Q fever can establish itself with an initial inoculation of fewer than ten C. burnetii cells, a sensitive detection method for C. burnetii infection is needed for early detection. We aimed to evaluate the effectiveness of a complementary locked primer (CLP)-based real-time PCR method for sensitive detection of C. burnetii infection.

Methods: To evaluate the ability of CLPs to enhance the efficiency of the real-time PCR assay for the C. burnetii IS1111 insertion sequence, the mean threshold cycle values from 20 real-time PCR replicates with either CLPs or conventional primers were determined using tenfold serial dilutions ( 102−108) of purified C. burnetii Nine Mile genomic DNA. In addition, the cross-reactivity between C. burnetii and 31 non-Coxiella species was examined.

Results: The CLP-based real-time PCR allowed specific and reliable detection of as few as 59 copies of the IS111 element present in the genome of C. burnetii, which represents approximately 2.96 genome equivalents or three cells of C. burnetii. These results demonstrate the effectiveness of CLP-based real-time PCR for sensitive detection of C. burnetii infection.

Conclusion: It can be concluded that the CLP-based real-time PCR assay is a more appropriate method for sensitive detection and quantification of C. burnetii than previously reported methods.

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Acknowledgments

Eun-Ju Kim and Hong Yong Kang contributed equally to this work. The conduct of this study and the preparation of this manuscript were supported by an intramural grant from the National Institute of Health, Korea. The authors have no conflicts of interest that are directly relevant to the content of this manuscript.

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Affiliations

  1. Division of Zoonoses, Center for Immunology and Pathology, National Institute of Health, Korea Centers for Disease Control & Prevention, Osong Health Technology Administration Complex, 187, Osongsaengmyeong 2-ro, Gangoe-myeon, Chengwon-gun, Chungcheongbuk-do, 363-951, Korea

    Eun-Ju Kim, Hong Yong Kang, Kyu-Jam Hwang, Sang-Hee Park, Mi-Yeoun Park, Sungdo Park &  Hyuk Chu

  2. iNtRON Biotechnology, Seongnam, Gyeonggi-do, Korea

    Jin Seok Yu, Ji Sung Park & Sang Hyeon Kang

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  1. Eun-Ju Kim
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Kim, EJ., Kang, H.Y., Hwang, KJ. et al. Enhanced Detection of Coxiella burnetii with a Complementary Locked Primer-Based Real-Time PCR Method. Mol Diag Ther 15, 103–107 (2011). https://doi.org/10.1007/BF03256399

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Keywords

  • Tularemia
  • Francisella Tularensis
  • Coxiella Burnetii
  • Brucella Abortus
  • IS1111 Element