Department of Chemistry, Colorado State University, 1801 Campus Delivery, Fort Collins, CO, 80523, USA.
Department of Microbiology, Immunology, and Pathology, Colorado State University, 1601 Campus Delivery, Fort Collins, CO, 80523, USA.
Anal Bioanal Chem. 2021 May;413(11):2933-2941. doi: 10.1007/s00216-021-03225-7. Epub 2021 Feb 22.
Several species of fungus from the genus Aspergillus are implicated in pulmonary infections in immunocompromised patients. Broad screening methods for fungal infections are desirable, as cultures require a considerable amount of time to provide results. Herein, we developed degradation and detection methods to produce and detect D-glucosamine (GlcN) from Aspergillus niger, a species of filamentous fungus. Ultimately, these techniques hold the potential to contribute to the diagnosis of pulmonary fungal infections in immunocompromised patients. In the following studies, we produced GlcN from fungal-derived chitin to serve as a marker for Aspergillus niger. To accomplish this, A. niger cells were lysed and subjected to a hydrochloric acid degradation protocol. Products were isolated, reconstituted in aqueous solutions, and analyzed using hydrophilic interaction liquid chromatography (HILIC) in tandem with electrospray ionization time-of-flight mass spectrometry. Our results indicated that GlcN was produced from A. niger. To validate these results, products obtained via fungal degradation were compared to products obtained from the degradation of two chitin polymers. The observed retention times and mass spectral extractions provided a two-step validation confirming that GlcN was produced from fungal-derived chitin. Our studies qualitatively illustrate that GlcN can be produced from A. niger; applying these methods to a more diverse range of fungi offers the potential to render a broad screening method for fungal detection pertinent to diagnosis of fungal infections.
几种曲霉菌属真菌与免疫功能低下患者的肺部感染有关。由于培养需要相当长的时间才能得出结果，因此需要广泛的真菌感染筛查方法。在此，我们开发了从黑曲霉（Aspergillus niger）产生和检测 D-葡糖胺（GlcN）的降解和检测方法，黑曲霉是一种丝状真菌。这些技术最终可能有助于诊断免疫功能低下患者的肺部真菌感染。在随后的研究中，我们从真菌来源的几丁质中产生 GlcN 作为黑曲霉的标志物。为此，我们裂解黑曲霉细胞并进行盐酸降解方案。将产物分离，在水溶液中重构，并使用亲水相互作用液相色谱（HILIC）与电喷雾电离飞行时间质谱联用进行分析。我们的结果表明从黑曲霉中产生了 GlcN。为了验证这些结果，将通过真菌降解获得的产物与通过两种几丁质聚合物降解获得的产物进行了比较。观察到的保留时间和质谱提取提供了两步验证，确认 GlcN 是由真菌来源的几丁质产生的。我们的研究定性地表明 GlcN 可以从黑曲霉中产生；将这些方法应用于更广泛的真菌范围可能提供一种广泛的真菌检测筛选方法，有助于诊断真菌感染。
