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Applied and Environmental Microbiology
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Food Microbiology

New Real-Time Quantitative PCR Procedure for Quantification of Bifidobacteria in Human Fecal Samples

Miguel Gueimonde, Satu Tölkkö, Teemu Korpimäki, Seppo Salminen
Miguel Gueimonde
1Department of Biochemistry and Food Chemistry
2Functional Foods Forum, University of Turku, Turku, Finland
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  • For correspondence: miguel.gueimonde@utu.fi
Satu Tölkkö
1Department of Biochemistry and Food Chemistry
2Functional Foods Forum, University of Turku, Turku, Finland
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Teemu Korpimäki
1Department of Biochemistry and Food Chemistry
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Seppo Salminen
1Department of Biochemistry and Food Chemistry
2Functional Foods Forum, University of Turku, Turku, Finland
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DOI: 10.1128/AEM.70.7.4165-4169.2004
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ABSTRACT

The application of a real-time quantitative PCR method (5′ nuclease assay), based on the use of a probe labeled at its 5′ end with a stable, fluorescent lanthanide chelate, for the quantification of human fecal bifidobacteria was evaluated. The specificities of the primers and the primer-probe combination were evaluated by conventional PCR and real-time PCR, respectively. The results obtained by real-time PCR were compared with those obtained by fluorescent in situ hybridization, the current gold standard for intestinal microbiota quantification. In general, a good correlation between the two methods was observed. In order to determine the detection limit and the accuracy of the real-time PCR procedure, germfree rat feces were spiked with known amounts of bifidobacteria and analyzed by both methods. The detection limit of the method used in this study was found to be about 5 × 104 cells per g of feces. Both methods, real-time PCR and fluorescent in situ hybridization, led to an accurate quantification of the spiked samples with high levels of bifidobacteria, but real-time PCR was more accurate for samples with low levels. We conclude that the real-time PCR procedure described here is a specific, accurate, rapid, and easy method for the quantification of bifidobacteria in feces.

  • Copyright © 2004 American Society for Microbiology
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New Real-Time Quantitative PCR Procedure for Quantification of Bifidobacteria in Human Fecal Samples
Miguel Gueimonde, Satu Tölkkö, Teemu Korpimäki, Seppo Salminen
Applied and Environmental Microbiology Jul 2004, 70 (7) 4165-4169; DOI: 10.1128/AEM.70.7.4165-4169.2004

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New Real-Time Quantitative PCR Procedure for Quantification of Bifidobacteria in Human Fecal Samples
Miguel Gueimonde, Satu Tölkkö, Teemu Korpimäki, Seppo Salminen
Applied and Environmental Microbiology Jul 2004, 70 (7) 4165-4169; DOI: 10.1128/AEM.70.7.4165-4169.2004
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KEYWORDS

Bifidobacterium
Colony Count, Microbial
feces
Polymerase Chain Reaction

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