背景:毛霉菌病是一种罕见的疾病,缺乏早期干预的诊断方法。采用使用calcofluorwhite-KOH的直接显微镜的可用策略,文化,放射学,和组织病理学检查通常是耗时的,需要复杂的方案。核酸扩增测试(NAAT)具有很高的灵敏度和快速检测能力。基于环介导等温扩增(LAMP)的检测提供了一种超灵敏的技术,不需要像PCR那样复杂的热循环仪,提供了一种直接的方法来改善诊断作为近护理点测试。
方法:该研究引入了一种新型的基于磁性纳米颗粒的LAMP测定法,用于捕获残留污染物以减少假阳性。解决了基于LAMP的诊断技术的主要缺点。该测定法针对cotH基因,这总是特定于Mucorales。
结果:该测定用各种Mucorales物种进行了测试,和小根霉的LOD,大黄鱼,阿耳根霉,同型根霉和伯氏Cunninghamella为1fg,1fg,0.1pg,0.1pg,和0.01ng。随后进行了一项临床蒙眼研究,该研究使用了30例诊断为毛霉菌病的患者的全血和尿液样本。该测定具有高度的可重复性并且具有>83%的总体灵敏度。
结论:毛霉菌病早期检测至关重要,因为目前的血液和尿液实验室检查缺乏敏感性,尽管进展迅速和严重的并发症,但需要几天的时间才能确认。我们开发的技术可以在不到45分钟的时间内确认毛霉菌病感染,特别关注RT-LAMP过程。因此,这项研究提供了一种可行的技术,可以从血液和尿液样本的分离DNA而不是侵入性组织样本中快速鉴定毛霉菌病。
毛霉菌病是一种难以早期诊断的疾病。本研究介绍了一种使用LAMP技术的灵敏快速诊断方法。对30名患者的血液和尿液样本进行检测,显示出良好的敏感性和可重复性。表明其非侵入性诊断的潜力。
Mucormycosis is a rare disease with scarce diagnostic methods for early intervention. Available strategies employing direct microscopy using calcofluor white-KOH, culture, radiologic, and histopathologic testing often are time-intensive and demand intricate protocols. Nucleic Acid Amplification Test holds promise due to its high sensitivity combined with rapid detection. Loop-mediated isothermal amplification (LAMP) based detection offers an ultrasensitive technique that does not require complicated thermocyclers like in polymerase chain reaction, offering a straightforward means for improving diagnoses as a near-point-of-care test. The study introduces a novel magnetic nanoparticle-based LAMP assay for carryover contaminant capture to reduce false positives. Solving the main drawback of LAMP-based diagnosis techniques. The assay targets the cotH gene, which is invariably specific to Mucorales. The assay was tested with various species of Mucorales, and the limit of detections for Rhizopus microsporus, Lichtheimia corymbifera, Rhizopus arrhizus, Rhizopus homothallicus, and Cunninghamella bertholletiae were 1 fg, 1 fg, 0.1 pg, 0.1 pg, and 0.01 ng, respectively. This was followed by a clinical blindfolded study using whole blood and urine samples from 30 patients diagnosed with Mucormycosis. The assay has a high degree of repeatability and had an overall sensitivity of > 83%. Early Mucormycosis detection is crucial, as current lab tests from blood and urine lack sensitivity and take days for confirmation despite rapid progression and severe complications. Our developed technique enables the confirmation of Mucormycosis infection in < 45 min, focusing specifically on the RT-LAMP process. Consequently, this research offers a viable technique for quickly identifying Mucormycosis from isolated DNA of blood and urine samples instead of invasive tissue samples.
Mucormycosis is a challenging disease to diagnose early. This study introduces a sensitive and rapid diagnostic approach using Loop-mediated isothermal amplification technology. Testing blood and urine samples from 30 patients revealed promising sensitivity and repeatability, indicating its potential for non-invasive diagnosis.