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Specific Detection of Pneumocystis jirovecii in Clinical Samples by Flow Cytometry

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Fungal Diagnostics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 968))

Abstract

Pneumocystis jirovecii is an opportunistic pathogen responsible for severe pneumonia in immunocompromised patients. Flow cytometry (FC) is a method widely used in different areas of clinical diagnosis like hematology and immunology. Recently it has started to be used in microbiology with a great potential for diagnosis of emergent microorganisms in clinical samples, especially when present in low numbers. The detection of Pneumocystis jirovecii in respiratory samples can be performed by FC, using specific monoclonal antibodies. Considering clinical diagnosis as a reference method, we previously showed FC to be 100% sensitive and specific when compared to immunofluorescence. Being an automated method, it is faster and less subject to human error than microscopic evaluation.

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References

  1. Murray PR, Rosenthal KS, Pfaller MA (2005) Fungal classification, structure, and replication. In: Murray PR, Rosenthal KS, Pfaller MA (eds) Medical microbiology, 5th edn. Mosby, Philadelphia, Pennsylvania, pp 67–73

    Google Scholar 

  2. Su YS, Lu JJ, Perng CL, Chang FY (2008) Pneumocystis jirovecii pneumonia in patients with and without human immunodeficiency virus infection. J Microbiol Immunol Infect 41:478–4823

    PubMed  Google Scholar 

  3. Kelley CF, Checkley W, Mannino DM, Franco-Paredes C, Del Rio C, Holguin F (2009) Trends in hospitalizations for AIDS associated Pneumocystis jirovecii pneumonia in the United States (1986 to 2005). Chest 136:190–197

    Article  PubMed  Google Scholar 

  4. Procop GW, Haddad S, Quinn J, Wilson ML, Henshaw NG, Reller LB, Artymyshyn RL, Katanik MT, Weinstein MP (2004) Detection of Pneumocystis jirovecii in respiratory specimens by four staining methods. J Clin Microbiol 42:3333–3335

    Article  PubMed  CAS  Google Scholar 

  5. Nevez G, Chabé M, Rabodonirina M, Virmaux M, Dei-Cas E, Hauser PM, Totet A (2008) Nosocomial Pneumocystis jirovecii infections. Parasite 15:359–365

    PubMed  CAS  Google Scholar 

  6. Morris A, Netravali M, Kling HM, Shipley T, Ross T, Sciurba FC, Norris KA (2008) Relationship of Pneumocystis antibody response to severity of chronic obstructive pulmonary disease. Clin Infect Dis 47:e64–e68

    Article  PubMed  Google Scholar 

  7. Krajicek BJ, Thomas CF Jr, Limper AH (2009) Pneumocystis pneumonia: current concepts in pathogenesis, diagnosis, and treatment. Clin Chest Med 30:265–278

    Article  PubMed  Google Scholar 

  8. Durand-Joly I, Chabé M, Soula F, Delhaes L, Camus D, Dei-Cas E (2005) Molecular diagnosis of Pneumocystis pneumonia. FEMS Immunol Med Microbiol 45:405–410

    Article  PubMed  CAS  Google Scholar 

  9. Jiancheng W, Minjun H, Yi-jun A, Lan S, Zengzhu G, Jianrong S, Xixiong K (2009) Screening Pneumocystis carinii pneumonia in non-HIV-infected immunocompromised patients using polymerase chain reaction. Diagn Microbiol Infect Dis 64:396–401

    Article  PubMed  Google Scholar 

  10. Alvarez-Martínez MJ, Miró JM, Valls ME, Moreno A, Rivas PV, Solé M, Benito N, Domingo P, Muñoz C, Rivera E, Zar HJ, Wissmann G, Diehl AR, Prolla JC, de Anta MT, Gatell JM, Wilson PE, Meshnick SR (2006) Sensitivity and specificity of nested and real-time PCR for the detection of Pneumocystis jirovecii in clinical specimens. Diagn Microbiol Infect Dis 56:153–160

    Article  PubMed  Google Scholar 

  11. Robberts FJL, Liebowitz LD, Chalkley LJ (2007) Polymerase chain reaction detection of Pneumocystis jirovecii: evaluation of 9 assays. Diagn Microbiol Infect Dis 58:385–392

    Article  PubMed  CAS  Google Scholar 

  12. Watson JV (1991) Introduction to flow cytometry, 1st edn. Cambridge University Press, Cambridge

    Book  Google Scholar 

  13. Álvarez-Barrientos A, Arroyo J, Cantón R, Nombela C, Sánchez-Pérez M (2000) Applications of flow cytometry to clinical microbiology. Clin Microbiol Rev 13:167–195

    Article  PubMed  Google Scholar 

  14. Givan A (2005) Flow cytometry: an introduction. In: Hawley TS, Hawley RG (eds) Methods in molecular biology: flow cytometry protocols, 2nd edn. Humana Press, Totowa, NJ, pp 1–32

    Google Scholar 

  15. Shapiro HM, Nebe-von-caron G (2005) Multiparameter flow cytometry of bacteria. In: Hawley TS, Hawley RG (eds) Methods in molecular biology: flow cytometry protocols, 2nd edn. Humana Press, Totowa, NJ, pp 33–43

    Google Scholar 

  16. Montemayor M, Galofré B, Ribas F, Lucena F (2007) Comparative study between two laser scanning cytometers and epifluorescence microscopy for detection of Cryptosporidium oocysts in water. Cytometry A 71:173–179

    Google Scholar 

  17. Pina-Vaz C, Rodrigues AG (2010) Evaluation of antifungal susceptibility using flow cytometry. Methods Mol Biol 638:281–289

    Article  PubMed  CAS  Google Scholar 

  18. Pina-Vaz C, Rodrigues AG, Sansonetty F, Martinez-De-Oliveira J, Fonseca AF, Mårdh P-A (2000) Antifungal activity of local anesthetics against Candida species. Infect Dis Obstet Gynecol 8:124–137

    PubMed  CAS  Google Scholar 

  19. Pina-Vaz C, Sansonetty F, Rodrigues AG, Martinez-De-Oliveira J, Fonseca AF, Mårdh P-A (2000) Antifungal activity of ibuprofen alone and in combination with fluconazole against Candida species. J Med Microbiol 49:831–840

    PubMed  CAS  Google Scholar 

  20. Pina-Vaz C, Sansonetty F, Rodrigues AG, Costa-Oliveira S, Tavares C, Martinezde-Oliveira J (2001) Cytometric approach for a rapid evaluation of susceptibility of Candida strains to antifungals. Clin Microbiol Infect 7:609–618

    Article  PubMed  CAS  Google Scholar 

  21. Pina-Vaz C, Costa-de-Oliveira S, Rodrigues AG, Espinel-Ingroff A (2005) Comparison of two probes for testing susceptibilities of pathogenic yeasts to voriconazole, itraconazole, and caspofungin by flow cytometry. J Clin Microbiol 43:4674–4679

    Article  PubMed  CAS  Google Scholar 

  22. Pina-Vaz C, Rodrigues AG, Costa-de-Oliveira S, Ricardo E, Mårdh P-A (2005) Potent synergic effect between ibuprofen and azoles on Candida resulting from blockade of efflux pumps as determined by FUN-1 staining and flow cytometry. J Antimicrob Chemother 56:678–685

    Article  PubMed  CAS  Google Scholar 

  23. Barbosa J, Bragada C, Costa-de-Oliveira S, Ricardo E, Rodrigues AG, Pina-Vaz C (2010) A new method for the detection of Pneumocystis jirovecii using flow cytometry. Eur J Clin Microbiol Infect Dis 29:1147–1152

    Article  PubMed  CAS  Google Scholar 

  24. Pina-Vaz C, Costa-Oliveira S, Rodrigues AG, Salvador A (2004) Novel method using a laser scanning cytometer for detection of mycobacteria in clinical samples. J Clin Microbiol 42:906–908

    Article  PubMed  Google Scholar 

  25. Pina-Vaz C, Costa-de-Oliveira S, Rodrigues AG (2005) Safe susceptibility testing of Mycobacterium tuberculosis by flow cytometry with the fluorescent nucleic acid stain SYTO 16. J Med Microbiol 54:77–81

    Article  PubMed  CAS  Google Scholar 

  26. Barbosa JM, Costa-de-Oliveira S, Rodrigues AG, Hanscheid T, Shapiro H, Pina- Vaz C (2008) A flow cytometric protocol for detection of Cryptosporidium spp. Cytometry A 73:44–47

    PubMed  Google Scholar 

  27. Barbosa J, Costa-de-Oliveira S, Rodrigues AG, Pina-Vaz C (2008) Optimization of a flow cytometry protocol for detection and viability assessment of Giardia lamblia. Travel Med Infect Dis 6:234–239

    Article  PubMed  Google Scholar 

  28. Barbosa J, Rodrigues AG, Pina-Vaz C (2009) Cytometric approach for detection of Encephalitozoon intestinalis, an emergent agent. Clin Vaccine Immunol 16:1021–1024

    Article  PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Microbiology, Faculty of Medicine, Cardiovascular Research Unit, University of Porto, Porto, Portugal

    Joana Barbosa, Sofia Costa-de-Oliveira, Ana Teresa Silva, Acácio Gonçalves Rodrigues & Cidália Pina-Vaz

  2. Escola Superior de Saúde Jean Piaget/Vila Nova de Gaia, Gulpilhares, Vila Nova de Gaia, Portugal

    Joana Barbosa

Authors
  1. Joana Barbosa
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  2. Sofia Costa-de-Oliveira
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  3. Ana Teresa Silva
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  4. Acácio Gonçalves Rodrigues
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  5. Cidália Pina-Vaz
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Corresponding author

Correspondence to Joana Barbosa .

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Editors and Affiliations

  1. , NCBES, Molecular Diagnostics Research Gr, National University of Ireland, Galway, IDA Business Park, Dangan, Galway, Ireland

    Louise O'Connor

  2. , NCBES, Molecular Diagnostics Research Gr, National University of Ireland, Galway, IDA Business Park, Dangan, Galway, Ireland

    Barry Glynn

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Barbosa, J., Costa-de-Oliveira, S., Silva, A.T., Rodrigues, A.G., Pina-Vaz, C. (2013). Specific Detection of Pneumocystis jirovecii in Clinical Samples by Flow Cytometry. In: O'Connor, L., Glynn, B. (eds) Fungal Diagnostics. Methods in Molecular Biology, vol 968. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-257-5_16

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  • DOI: https://doi.org/10.1007/978-1-62703-257-5_16

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-256-8

  • Online ISBN: 978-1-62703-257-5

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