丝状真菌对全球免疫功能低下的个体存在重大的健康危害;然而,在感染期间迅速和精确地识别它们仍然具有挑战性。在这项研究中,建立了一种基于TaqMan探针的多重实时荧光定量PCR(M-qPCR)方法,用于同时检测四种重要病原丝状真菌的目标基因:烟曲霉ANXC4基因,镰刀菌EF1-α基因.,Mucorales的线粒体rnl基因,和hcp100基因。在这个M-qPCR检测中,4种真菌的检测限(LoD)均为100份,相关系数(R2)均大于0.99。该测定的特异性为100%,最低检测限为100拷贝/反应。总之,建立了一种M-qPCR检测方法,具有很高的特异性和敏感性,可在临床上快速同时检测四种重要的丝状真菌。
目的:世界卫生组织于2022年制定了第一个真菌优先病原体清单(WHOFPPL)。烟曲霉,Mucorales,镰刀菌属。,和组织支原体。是WHOFPPL的关键优先组和高度优先组中具有丝状形态的四种病原真菌。近几十年来,随着易感人群的增加,这四种丝状真菌感染在免疫功能低下患者中变得更加常见和严重。这给公共卫生系统带来了沉重负担。然而,在感染期间迅速和精确地识别它们仍然具有挑战性。我们的研究成功建立了基于TaqMan探针的多重实时qPCR检测四种临床重要的丝状真菌,A.烟,镰刀菌属。,Mucorales,和组织胞浆,具有高灵敏度和特异性,这显示了迅速和精确诊断真菌感染的潜力。
Filamentous fungi present significant health hazards to immunocompromised individuals globally; however, the prompt and precise identification of them during infection remains challenging. In this study, a TaqMan probe-based multiplex real-time PCR (M-qPCR) assay was developed to detect simultaneously the target genes of four important pathogenic filamentous fungi: ANXC4 gene of Aspergillus fumigatus, EF1-α gene of Fusarium spp., mitochondrial rnl gene of Mucorales, and hcp100 gene of Histoplasma capsulatum. In this M-qPCR assay, the limit of detection (LoD) to all four kinds of fungi was 100 copies and the correlation coefficients (R2) were above 0.99. The specificity of this assay is 100%, and the minimum detection limit is 100 copies/reaction. In conclusion, an M-qPCR detection assay was well established with high specificity and sensitivity for rapid and simultaneous detection on four important filamentous fungi in the clinic.
OBJECTIVE: World Health Organization developed the first fungal priority pathogens list (WHO FPPL) in 2022. Aspergillus fumigatus, Mucorales, Fusarium spp., and Histoplasma spp. are the four types of pathogenic fungi with filamentous morphology in the critical priority group and high priority group of WHO FPPL. These four filamentous fungal infections have become more common and severe in immunocompromised patients with the increase in susceptible populations in recent decades, which resulted in a substantial burden on the public health system. However, prompt and precise identification of them during infection remains challenging. Our study established successfully a TaqMan probe-based multiplex real-time qPCR assay for four clinically important filamentous fungi, A. fumigatus, Fusarium spp., Mucorales, and Histoplasma capsulatum, with high sensitivity and specificity, which shows promising potential for prompt and precise diagnosis against fungal infection.