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Diagnostic Utility of DNA from Aspergillus in Whole Blood Specimens

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Abstract

For several years, PCR-based detection of Aspergillus DNA has been considered a promising tool for the diagnosis of invasive aspergillosis, but its broader implementation has been hampered by a lack of standardization and the uncertain diagnostic accuracy of the wide variety of published protocols. Recently, new attempts have been made to address this problem and define a quality standard for diagnostic Aspergillus PCR. This review summarizes technical aspects of up-to-date protocols, focusing on the recently proposed guidelines of the European Aspergillus PCR Initiative (EAPCRI) for whole blood PCR testing.

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References

Papers of particular interest, published recently, have been highlighted as: • Of importance •• Of major importance

  1. • Mengoli C, Cruciani M, Barnes RA, Loeffler J, Donnelly JP. Use of PCR for diagnosis of invasive aspergillosis: systematic review and meta-analysis. Lancet Infect Dis. 2009;9:89–96. This review summarizes relevant studies done on Aspergillus PCR and provides a meta-analysis of its current role in diagnosing invasive aspergillosis.

    Article  PubMed  CAS  Google Scholar 

  2. Klingspor L, Loeffler J. Aspergillus PCR formidable challenges and progress. Med Mycol. 2009;47 Suppl 1:S241–7.

    Article  PubMed  CAS  Google Scholar 

  3. •• De Pauw B, Walsh TJ, Donnelly JP, Stevens DA, Edwards JE, Calandra T, Pappas PG, Maertens J, Lortholary O, Kauffman CA, et al. Revised definitions of invasive fungal disease from the European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) Consensus Group. Clin Infect Dis. 2008;46:1813–21. The diagnostic criteria for invasive aspergillosis (and invasive fungal infections) proposed in this guideline are the basis of all relevant studies addressing the diagnostic potential of Aspergillus PCR.

    Article  PubMed  Google Scholar 

  4. Ramsdale M. Programmed cell death in pathogenic fungi. Biochim Biophys Acta. 2008;1783:1369–80.

    Article  PubMed  CAS  Google Scholar 

  5. Mousavi SA, Robson GD. Entry into the stationary phase is associated with a rapid loss of viability and an apoptotic-like phenotype in the opportunistic pathogen Aspergillus fumigatus. Fungal Genet Biol. 2003;39:221–9.

    Article  PubMed  CAS  Google Scholar 

  6. Morton CO, Clemons KV, Springer J, Mueller JG, Rogers TR, Stevens DA, Kurzai O, Einsele H, Loeffler J. Real-time PCR and quantitative culture for monitoring of experimental Aspergillus fumigatus intracranial infection in neutropenic mice. J Med Microbiol. 2011 Mar 24 (Epub ahead of print).

  7. Mennink-Kersten MA, Ruegebrink D, Wasei N, Melchers WJ, Verweij PE. In vitro release by Aspergillus fumigatus of galactofuranose antigens, 1,3-beta-D-glucan, and DNA, surrogate markers used for diagnosis of invasive aspergillosis. J Clin Microbiol. 2006;44:1711–8.

    Article  PubMed  CAS  Google Scholar 

  8. •• White PL, Bretagne S, Klingspor L, Melchers WJ, McCulloch E, Schulz B, Finnstrom N, Mengoli C, Barnes RA, Donnelly JP, et al. Aspergillus PCR: one step closer to standardization. J Clin Microbiol. 2010;48:1231–40. This study systematically describes the optimization steps for Aspergillus PCR and summarizes the protocol requirements proposed by EAPCRI.

    Article  PubMed  CAS  Google Scholar 

  9. Millon L, Grenouillet F, Crouzet J, Larosa F, Loewert S, Bellanger AP, et al. False-positive Aspergillus real-time PCR assay due to a nutritional supplement in a bone marrow transplant recipient with GVH disease. Med Mycol. 2010;48:661–4.

    Article  PubMed  CAS  Google Scholar 

  10. Loeffler J, Hebart H, Brauchle U, Schumacher U, Einsele H. Comparison between plasma and whole blood specimens for detection of Aspergillus DNA by PCR. J Clin Microbiol. 2000;38:3830–3.

    PubMed  CAS  Google Scholar 

  11. Suarez F, Lortholary O, Buland S, Rubio MT, Ghez D, Mahe V, et al. Detection of circulating Aspergillus fumigatus DNA by real-time PCR assay of large serum volumes improves early diagnosis of invasive aspergillosis in high-risk adult patients under hematologic surveillance. J Clin Microbiol. 2008;46:3772–7.

    Article  PubMed  CAS  Google Scholar 

  12. Einsele H, Quabeck K, Muller KD, Hebart H, Rothenhofer I, Loffler J, et al. Prediction of invasive pulmonary aspergillosis from colonisation of lower respiratory tract before marrow transplantation. Lancet. 1998;352:1443.

    Article  PubMed  CAS  Google Scholar 

  13. Bissinger AL, Einsele H, Hamprecht K, Schumacher U, Kandolf R, Loeffler J, et al. Infectious pulmonary complications after stem cell transplantation or chemotherapy: diagnostic yield of bronchoalveolar lavage. Diagn Microbiol Infect Dis. 2005;52:275–80.

    Article  PubMed  Google Scholar 

  14. Schabereiter-Gurtner C, Selitsch B, Rotter ML, Hirschl AM, Willinger B. Development of novel real-time PCR assays for detection and differentiation of eleven medically important Aspergillus and Candida species in clinical specimens. J Clin Microbiol. 2007;45:906–14.

    Article  PubMed  CAS  Google Scholar 

  15. Hummel M, Spiess B, Kentouche K, Niggemann S, Bohm C, Reuter S, et al. Detection of Aspergillus DNA in cerebrospinal fluid from patients with cerebral aspergillosis by a nested PCR assay. J Clin Microbiol. 2006;44:3989–93.

    Article  PubMed  CAS  Google Scholar 

  16. Millon L, Piarroux R, Deconinck E, Bulabois CE, Grenouillet F, Rohrlich P, et al. Use of real-time PCR to process the first galactomannan-positive serum sample in diagnosing invasive aspergillosis. J Clin Microbiol. 2005;43:5097–101.

    Article  PubMed  CAS  Google Scholar 

  17. Loeffler J, Henke N, Hebart H, Schmidt D, Hagmeyer L, Schumacher U, et al. Quantification of fungal DNA by using fluorescence resonance energy transfer and the light cycler system. J Clin Microbiol. 2000;38:586–90.

    PubMed  CAS  Google Scholar 

  18. • Springer J, Loeffler J, Heinz W, Schlossnagel H, Lehmann M, Morton O, Rogers TR, Schmitt C, Frosch M, Einsele H, et al. Pathogen-specific DNA enrichment does not increase sensitivity of PCR for diagnosis of invasive aspergillosis in neutropenic patients. J Clin Microbiol. 2011;49:1267–73. This study shows that pathogen-specific DNA enrichment, which increases the analytic sensitivity of other PCR applications, does not prove beneficial for Aspergillus PCR in neutropenic patients.

    Article  PubMed  Google Scholar 

  19. • White PL, Perry MD, Loeffler J, Melchers W, Klingspor L, Bretagne S, McCulloch E, Cuenca-Estrella M, Finnstrom N, Donnelly JP, et al. Critical stages of extracting DNA from Aspergillus fumigatus in whole-blood specimens. J Clin Microbiol. 2010;48:3753–5. This study systematically analyzes the contribution of single protocol steps to the overall efficacy of Aspergillus PCR.

    Article  PubMed  Google Scholar 

  20. Loeffler J, Schmidt K, Hebart H, Schumacher U, Einsele H. Automated extraction of genomic DNA from medically important yeast species and filamentous fungi by using the MagNA Pure LC system. J Clin Microbiol. 2002;40:2240–3.

    Article  PubMed  CAS  Google Scholar 

  21. Loeffler J, Hebart H, Bialek R, Hagmeyer L, Schmidt D, Serey FP, et al. Contaminations occurring in fungal PCR assays. J Clin Microbiol. 1999;37:1200–2.

    PubMed  CAS  Google Scholar 

  22. Herrera ML, Vallor AC, Gelfond JA, Patterson TF, Wickes BL. Strain-dependent variation in 18S ribosomal DNA copy numbers in Aspergillus fumigatus. J Clin Microbiol. 2009;47:1325–32.

    Article  PubMed  CAS  Google Scholar 

  23. Costa C, Costa JM, Desterke C, Botterel F, Cordonnier C, Bretagne S. Real-time PCR coupled with automated DNA extraction and detection of galactomannan antigen in serum by enzyme-linked immunosorbent assay for diagnosis of invasive aspergillosis. J Clin Microbiol. 2002;40:2224–7.

    Article  PubMed  CAS  Google Scholar 

  24. • Millon L, Grenouillet F, Legrand F, Loewert S, Bellanger AP, Gbaguidi-Haore H, Scherer E, Henon T, Rohrlich P, Deconinck E. Ribosomal and mitochondrial DNA target for real-time PCR diagnosis of invasive aspergillosis. J Clin Microbiol 2011;49:1058–63. This study compares two different PCR targets and suggests that there may be a benefit in combining these approaches.

    Article  PubMed  CAS  Google Scholar 

  25. Bruns T, Sachse S, Straube E, Assefa S, Herrmann A, Hagel S, et al. Identification of bacterial DNA in neutrocytic and non-neutrocytic cirrhotic ascites by means of a multiplex polymerase chain reaction. Liver Int. 2009;29:1206–14.

    Article  PubMed  CAS  Google Scholar 

  26. Sachse S, Straube E, Lehmann M, Bauer M, Russwurm S, Schmidt KH. Truncated human cytidylate-phosphate-deoxyguanylate-binding protein for improved nucleic acid amplification technique-based detection of bacterial species in human samples. J Clin Microbiol. 2009;47:1050–7.

    Article  PubMed  CAS  Google Scholar 

  27. Martino R, Viscoli C. Empirical antifungal therapy in patients with neutropenia and persistent or recurrent fever of unknown origin. Br J Haematol. 2006;132:138–54.

    Article  PubMed  CAS  Google Scholar 

  28. Hebart H, Loffler J, Reitze H, Engel A, Schumacher U, Klingebiel T, et al. Prospective screening by a panfungal polymerase chain reaction assay in patients at risk for fungal infections: implications for the management of febrile neutropenia. Br J Haematol. 2000;111:635–40.

    Article  PubMed  CAS  Google Scholar 

  29. Morrissey CO, Chen SC, Sorrell TC, Bradstock KF, Szer J, Halliday CL, et al. Design issues in a randomized controlled trial of a pre-emptive versus empiric antifungal strategy for invasive aspergillosis in patients with high-risk hematologic malignancies. Leuk Lymphoma. 2011;52:179–93.

    Article  PubMed  CAS  Google Scholar 

  30. Hebart H, Klingspor L, Klingebiel T, Loeffler J, Tollemar J, Ljungman P, et al. A prospective randomized controlled trial comparing PCR-based and empirical treatment with liposomal amphotericin B in patients after allo-SCT. Bone Marrow Transplant. 2009;43:553–61.

    Article  PubMed  CAS  Google Scholar 

  31. Garcia ME, Blanco JL, Caballero J, Gargallo-Viola D. Anticoagulants interfere with PCR used to diagnose invasive aspergillosis. J Clin Microbiol. 2002;40:1567–8.

    Article  PubMed  CAS  Google Scholar 

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Conflicts of Interest

J. Löffler: Honoraria from Gilead Sciences; O. Kurzai: Research grants from SIRS-Lab and Jena, honoraria from Abbott and Pfizer, payment for educational presentations or speaking from Gilead.

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Authors and Affiliations

  1. Septomics Research Centre, Friedrich-Schiller-University Jena and Leibniz-Institute for Natural Products Research and Infection Biology – Hans-Knoell-Institute, Beutenbergstr. 11A, 07745, Jena, Germany

    Oliver Kurzai

  2. Universitätsklinikum Würzburg, Medizinische Klinik und Poliklinik II, Forschungslabore, Haus C11, 97080, Würzburg, Germany

    Jürgen Löffler

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  1. Jürgen Löffler
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Correspondence to Oliver Kurzai.

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Löffler, J., Kurzai, O. Diagnostic Utility of DNA from Aspergillus in Whole Blood Specimens. Curr Fungal Infect Rep 5, 179–185 (2011). https://doi.org/10.1007/s12281-011-0056-8

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  • DOI: https://doi.org/10.1007/s12281-011-0056-8

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