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Journal of Clinical Microbiology, December 1999, p. 3865-3871, Vol. 37, No. 12
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Specific Detection of Aspergillus Species in Blood and
Bronchoalveolar Lavage Samples of Immunocompromised Patients by
Two-Step PCR
Heyko
Skladny,1
Dieter
Buchheidt,1,*
Corinna
Baust,1
Frank
Krieg-Schneider,1,
Wolfgang
Seifarth,1
Christine
Leib-Mösch,1,2 and
Rüdiger
Hehlmann1
III. Medizinische Klink, Klinikum Mannheim,
University of Heidelberg, D-68305 Mannheim,1
and Institute of Molecular Virology, GSF-National Research
Center for Environment and Health, D-85764
Neuherberg,2 Germany
Received 22 February 1999/Returned for modification 12 July
1999/Accepted 9 August 1999
The increasing incidence of aspergillosis, a life-threatening
infection in immunocompromised patients, emphasizes the need to improve
the currently limited diagnostic tools. We developed a two-step PCR
assay that specifically amplifies a region of the 18S rRNA gene that is
highly conserved in Aspergillus species. A number of
primers with the least homology to equivalent human or
Candida gene sequences were screened for the pairs that
gave the highest sensitivity and specificity. No cross-reaction with the wide range of fungal and bacterial pathogens so far tested was
observed. This assay allows direct and rapid detection of down to 10 fg
of Aspergillus DNA corresponding to 1 to 5 CFU per ml of
blood. A total of 315 blood and bronchoalveolar lavage samples from 140 subjects, including 93 patients at risk for invasive fungal disease,
were screened. The result was a 100% correlation between positive
histology, culture, or high-resolution computed tomography findings and
PCR results. The test specificity was 89%. Our data point to the
considerable potential clinical value of this simple, specific, rapid,
and inexpensive PCR assay for improving the means of early diagnosis of
systemic aspergillosis in high-risk patients.
*
Corresponding author. Mailing address: III.
Medizinische Klinik, Klinikum Mannheim der Universität
Heidelberg, Wiesbadenerstraße 7-11, D-68305 Mannheim, Germany. Phone:
49-621-383-4115. Fax: 49-621-383-4201. E-mail:
dieter.buchheidt{at}urz.uni-heidelberg.de.
Present address: QIAGEN GmbH, Hilden, Germany.
Journal of Clinical Microbiology, December 1999, p. 3865-3871, Vol. 37, No. 12
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
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