G M Chong1, M T van der Beek2, P A von dem Borne3, J Boelens4, E Steel5, G A Kampinga6, L F R Span7, K Lagrou8, J A Maertens9, G J H Dingemans10, G R Gaajetaan10, D W E van Tegelen10, J J Cornelissen11, A G Vonk12, B J A Rijnders13. 1. Department of Internal Medicine, Section of Infectious Diseases, Erasmus University Medical Center, Rotterdam, The Netherlands g.chong@erasmusmc.nl. 2. Department of Medical Microbiology, Leiden University Medical Center, Leiden, The Netherlands. 3. Department of Haematology, Leiden University Medical Center, Leiden, The Netherlands. 4. Department of Microbiology, Ghent University Hospital, Ghent, Belgium. 5. Department of Haematology, Ghent University Hospital, Ghent, Belgium. 6. Department of Medical Microbiology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands. 7. Department of Haematology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands. 8. Department of Microbiology and Immunology, KU Leuven-University of Leuven; University Hospitals Leuven, Leuven, Belgium. 9. Department of Microbiology and Immunology, KU Leuven-University of Leuven; Department of Haematology, University Hospitals Leuven, Leuven, Belgium. 10. PathoNostics B.V., Maastricht, The Netherlands. 11. Department of Haematology, Erasmus University Medical Center, Rotterdam, The Netherlands. 12. Department of Medical Microbiology, Erasmus University Medical Center, Rotterdam, The Netherlands. 13. Department of Internal Medicine, Section of Infectious Diseases, Erasmus University Medical Center, Rotterdam, The Netherlands.
Abstract
OBJECTIVES: In patients with invasive aspergillosis (IA), fungal cultures are mostly negative. Consequently, azole resistance often remains undetected. The AsperGenius® multiplex real-time PCR assay identifies clinically relevant Aspergillus species and four resistance-associated mutations (RAMs; TR34/L98H/T289A/Y121F) in the Cyp51A gene. This multicentre study evaluated the diagnostic performance of this assay on bronchoalveolar lavage (BAL) fluid and correlated the presence of RAMs with azole treatment failure and mortality. METHODS: Stored BAL samples from patients with haematological diseases with suspected IA were used. BAL samples that were galactomannan/culture positive were considered positive controls for the presence of Aspergillus. Azole treatment failure and 6 week mortality were compared in patients with and without RAMs that had received ≥5 days of voriconazole monotherapy. RESULTS: Two hundred and one patients each contributed one BAL sample, of which 88 were positive controls and 113 were negative controls. The optimal cycle threshold cut-off value for the Aspergillus species PCR was <38. With this cut-off, the PCR was positive in 74/88 positive controls. The sensitivity, specificity, positive predictive value and negative predictive value were 84%, 80%, 76% and 87%, respectively. 32/74 BAL samples were culture negative. Azole treatment failure was observed in 6/8 patients with a RAM compared with 12/45 patients without RAMs (P = 0.01). Six week mortality was 2.7 times higher in patients with RAMs (50.0% versus 18.6%; P = 0.07). CONCLUSIONS: The AsperGenius® assay had a good diagnostic performance on BAL and differentiated WT from Aspergillus fumigatus with RAMs, including in culture-negative BAL samples. Most importantly, detection of RAMs was associated with azole treatment failure.
OBJECTIVES: In patients with invasive aspergillosis (IA), fungal cultures are mostly negative. Consequently, azole resistance often remains undetected. The AsperGenius® multiplex real-time PCR assay identifies clinically relevant Aspergillus species and four resistance-associated mutations (RAMs; TR34/L98H/T289A/Y121F) in the Cyp51A gene. This multicentre study evaluated the diagnostic performance of this assay on bronchoalveolar lavage (BAL) fluid and correlated the presence of RAMs with azole treatment failure and mortality. METHODS: Stored BAL samples from patients with haematological diseases with suspected IA were used. BAL samples that were galactomannan/culture positive were considered positive controls for the presence of Aspergillus. Azole treatment failure and 6 week mortality were compared in patients with and without RAMs that had received ≥5 days of voriconazole monotherapy. RESULTS: Two hundred and one patients each contributed one BAL sample, of which 88 were positive controls and 113 were negative controls. The optimal cycle threshold cut-off value for the Aspergillus species PCR was <38. With this cut-off, the PCR was positive in 74/88 positive controls. The sensitivity, specificity, positive predictive value and negative predictive value were 84%, 80%, 76% and 87%, respectively. 32/74 BAL samples were culture negative. Azole treatment failure was observed in 6/8 patients with a RAM compared with 12/45 patients without RAMs (P = 0.01). Six week mortality was 2.7 times higher in patients with RAMs (50.0% versus 18.6%; P = 0.07). CONCLUSIONS: The AsperGenius® assay had a good diagnostic performance on BAL and differentiated WT from Aspergillus fumigatus with RAMs, including in culture-negative BAL samples. Most importantly, detection of RAMs was associated with azole treatment failure.
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