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Quantitative TaqMan PCR for detection of Pneumocystis jiroveci.
J Microbiol Methods. 2005 Jun; 61(3):381-7.JM

Abstract

We developed a quantitative real-time PCR assay for detection and quantification of Pneumocystis jiroveci in bronchoalveolar lavage (BAL) specimens based on primers and probe targeting the gene encoding beta-tubulin. The assay was able to detect 50 DNA copies per ml of a standard plasmid containing the target sequence. The intra- and interassay coefficients of variation were 0.46%-4.27% and 0.05-2.00% over 5 log(10) values. Fifty-seven controls of human, viruses, bacteria and fungi DNA samples were amplified and found negative. Fifty-three BAL samples sent to the laboratory for diagnosis of pneumocystosis were prospectively investigated by real-time PCR and direct microscopic examinations (DME) using Giemsa stain and direct immunofluorescence. All PCR negative samples were negative by microscopy. Among the 24 (45%) BAL found PCR positive, 8 were positive by microscopy (35%). The copy numbers of the target gene were between 4.4 x 10(3) and 2.8 x 10(6) per ml for the microscopically positive samples and between 8 and 9.2 x 10(3) per ml for the microscopically negative samples. In conclusion, we developed a rapid, sensitive and specific real time PCR for the diagnosis and quantification of Pneumocystis jiroveci in BAL samples.

Authors+Show Affiliations

Brancart F
Laboratory of Microbiology, Erasme Hospital-ULB, 808 Route de Lennik, 1070 Brussels, Belgium. frbranca@ulb.ac.be
Rodriguez-Villalobos H
No affiliation info available
Fonteyne PA
No affiliation info available
Peres-Bota D
No affiliation info available
Liesnard C
No affiliation info available

MeSH

Base SequenceBronchoalveolar Lavage FluidDNA, FungalHumansMycologyPneumocystis cariniiPneumonia, PneumocystisPolymerase Chain ReactionSensitivity and SpecificityTubulin

Pub Type(s)

Journal Article

Language

eng

PubMed ID

15767014

Citation

Brancart, Françoise, et al. "Quantitative TaqMan PCR for Detection of Pneumocystis Jiroveci." Journal of Microbiological Methods, vol. 61, no. 3, 2005, pp. 381-7.
Brancart F, Rodriguez-Villalobos H, Fonteyne PA, et al. Quantitative TaqMan PCR for detection of Pneumocystis jiroveci. J Microbiol Methods. 2005;61(3):381-7.
Brancart, F., Rodriguez-Villalobos, H., Fonteyne, P. A., Peres-Bota, D., & Liesnard, C. (2005). Quantitative TaqMan PCR for detection of Pneumocystis jiroveci. Journal of Microbiological Methods, 61(3), 381-7.
Brancart F, et al. Quantitative TaqMan PCR for Detection of Pneumocystis Jiroveci. J Microbiol Methods. 2005;61(3):381-7. PubMed PMID: 15767014.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Quantitative TaqMan PCR for detection of Pneumocystis jiroveci. AU - Brancart,Françoise, AU - Rodriguez-Villalobos,Hector, AU - Fonteyne,Pierre-Alain, AU - Peres-Bota,Daliana, AU - Liesnard,Corinne, PY - 2004/11/02/received PY - 2004/12/30/revised PY - 2005/01/04/accepted PY - 2005/3/16/pubmed PY - 2005/5/4/medline PY - 2005/3/16/entrez SP - 381 EP - 7 JF - Journal of microbiological methods JO - J Microbiol Methods VL - 61 IS - 3 N2 - We developed a quantitative real-time PCR assay for detection and quantification of Pneumocystis jiroveci in bronchoalveolar lavage (BAL) specimens based on primers and probe targeting the gene encoding beta-tubulin. The assay was able to detect 50 DNA copies per ml of a standard plasmid containing the target sequence. The intra- and interassay coefficients of variation were 0.46%-4.27% and 0.05-2.00% over 5 log(10) values. Fifty-seven controls of human, viruses, bacteria and fungi DNA samples were amplified and found negative. Fifty-three BAL samples sent to the laboratory for diagnosis of pneumocystosis were prospectively investigated by real-time PCR and direct microscopic examinations (DME) using Giemsa stain and direct immunofluorescence. All PCR negative samples were negative by microscopy. Among the 24 (45%) BAL found PCR positive, 8 were positive by microscopy (35%). The copy numbers of the target gene were between 4.4 x 10(3) and 2.8 x 10(6) per ml for the microscopically positive samples and between 8 and 9.2 x 10(3) per ml for the microscopically negative samples. In conclusion, we developed a rapid, sensitive and specific real time PCR for the diagnosis and quantification of Pneumocystis jiroveci in BAL samples. SN - 0167-7012 UR - https://www.unboundmedicine.com/medline/citation/15767014/Quantitative_TaqMan_PCR_for_detection_of_Pneumocystis_jiroveci_ DB - PRIME DP - Unbound Medicine ER -