| Summary: | The <i>β-tubulin</i> (<i>benA</i>) gene is a promising target for the identification of <i>Aspergillus</i> species. Assessment of the clinical implementation and performance of <i>benA</i> gene-based <i>Aspergillus</i> polymerase chain reaction (PCR) remains warranted. In this study, we assessed the analytical performance of the BenA probe PCR in comparison with the Aspergenius kit. We prospectively collected bronchoalveolar lavage (BAL) fluid via diagnostic bronchoscopy from adult patients with hematologic diseases. <i>BenA</i> gene-based multiplex real-time PCR and sequential melting temperature analysis were performed to detect the azole resistance of <i>Aspergillus fumigatus</i>. In total, 76 BAL fluids in 75 patients suspicious of invasive pulmonary aspergillosis (IPA) were collected. Before the application of PCR, the prevalence of proven and probable IPA was 32.9%. However, after implementing the <i>benA</i> gene-based PCR, 15.8% (12 out of 76) of potential IPA cases were reclassified as probable IPA. The analytical performance of the BenA probe PCR in BAL samples was comparable to that of the Aspergenius kit. The diagnostic performance was as follows: sensitivity, 52.0%; specificity, 64.7%; positive predictive value, 41.9%; negative predictive value, 73.3%; positive likelihood ratio, 1.473; and negative likelihood ratio, 0.741. Moreover, <i>benA</i> gene-based <i>Aspergillus</i> PCR discriminated all major sections of <i>Aspergillus</i>, including cryptic species such as <i>Aspergillus tubingensis.</i> Sequential melting temperature analysis successfully detected 2 isolates (15.4%) of <i>A. fumigatus</i> carrying resistant mutations. <i>BenA</i> gene-based <i>Aspergillus</i> PCR with melting temperature analysis enhances diagnostic accuracy and detects not only cryptic species but also resistant mutations of <i>A. fumigatus</i>. It shows promise for clinical applications in the diagnosis of IPA.
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