Validation of a new Aspergillus real-time PCR assay for direct detection of Aspergillus and azole resistance of Aspergillus fumigatus on bronchoalveolar lavage fluid.
نویسندگان
چکیده
Azole resistance in Aspergillus fumigatus is increasingly reported. Here, we describe the validation of the AsperGenius, a new multiplex real-time PCR assay consisting of two multiplex real-time PCRs, one that identifies the clinically relevant Aspergillus species, and one that detects the TR34, L98H, T289A, and Y121F mutations in CYP51A and differentiates susceptible from resistant A. fumigatus strains. The diagnostic performance of the AsperGenius assay was tested on 37 bronchoalveolar lavage (BAL) fluid samples from hematology patients and 40 BAL fluid samples from intensive care unit (ICU) patients using a BAL fluid galactomannan level of ≥1.0 or positive culture as the gold standard for detecting the presence of Aspergillus. In the hematology and ICU groups combined, there were 22 BAL fluid samples from patients with invasive aspergillosis (IA) (2 proven, 9 probable, and 11 nonclassifiable). Nineteen of the 22 BAL fluid samples were positive, according to the gold standard. The optimal cycle threshold value for the presence of Aspergillus was <36. Sixteen of the 19 BAL fluid samples had a positive PCR (2 Aspergillus species and 14 A. fumigatus samples). This resulted in a sensitivity, specificity, and positive and negative predictive values of 88.9%, 89.3%, 72.7%, and 96.2%, respectively, for the hematology group and 80.0%, 93.3%, 80.0%, and 93.3%, respectively, in the ICU group. The CYP51A real-time PCR confirmed 12 wild-type and 2 resistant strains (1 TR34-L98H and 1 TR46-Y121F-T289A mutant). Voriconazole therapy failed for both patients. The AsperGenius multiplex real-time PCR assay allows for sensitive and fast detection of Aspergillus species directly from BAL fluid samples. More importantly, this assay detects and differentiates wild-type from resistant strains, even if BAL fluid cultures remain negative.
منابع مشابه
Azole resistance in Aspergillus fumigatus from bronchoalveolar lavage fluid samples of patients with chronic diseases.
OBJECTIVES Triazole resistance in Aspergillus fumigatus has been increasing. We explored the A. fumigatus azole resistance profiles in bronchoalveolar lavage (BAL) fluid samples from Danish patients examined for aspergillosis. METHODS A total of 94 BAL samples from 87 patients were evaluated by galactomannan (GM) test and A. fumigatus CYP51A profiling by PCR. RESULTS Aspergillus spp. were i...
متن کاملComparison of an Aspergillus real-time polymerase chain reaction assay with galactomannan testing of bronchoalvelolar lavage fluid for the diagnosis of invasive pulmonary aspergillosis in lung transplant recipients.
BACKGROUND Early diagnosis and treatment of invasive pulmonary aspergillosis (IPA) improves outcome. METHODS We compared the performance of publicly available pan-Aspergillus, Aspergillus fumigatus-, and Aspergillus terreus-specific real-time polymerase chain reaction (PCR) assays with the Platelia galactomannan (GM) assay in 150 bronchoalveolar lavage (BAL) samples from lung transplant recip...
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Bronchoalveolar lavage (BAL) is widely used for evaluation of patients with suspected invasive pulmonary aspergillosis (IPA). However, the diagnostic yield of BAL for detection of IPA by culture and direct examination is limited. Earlier diagnosis may be facilitated by assays that can detect Aspergillus galactomannan antigen or DNA in BAL fluid. We therefore characterized and compared the diagn...
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Aspergillus diseases are often caused by Aspergillus fumigatus. Azoles are the mainstay of therapy, but the management of aspergillosis is hampered by the emergence of azole resistance. Rapid detection of azole resistance might benefit treatment outcome by early treatment modifications. However, the yield of fungal culture in invasive aspergillosis is low and susceptibility testing requires sev...
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ورودعنوان ژورنال:
- Journal of clinical microbiology
دوره 53 3 شماره
صفحات -
تاریخ انتشار 2015