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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.

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^* 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.

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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.

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