Singh Harsharan K, Madden Victoria, Shen You Jun, Thompson Bawana D, Nickeleit Volker
Electron Microscopy Services, UNC Hospitals, Department of Pathology and Laboratory Medicine, Campus Box 7525 UNC-CH School of Medicine, The University of North Carolina at Chapel Hill, 27599, USA.
Ultrastruct Pathol. 2006 Sep-Oct;30(5):329-38. doi: 10.1080/01913120600932347.
Negative-staining electron microscopy (EM) has played a pivotal role in diagnostic virology. It is a rapid technique for viral detection in the urine and can provide an easy means for monitoring viral activity and productive infections. EM of urine for the detection of polyomaviruses has hitherto not been systematically evaluated as a screening tool for renal transplant patients at risk for BK polyomavirus nephropathy (BKN). Here, the authors discuss technical aspects of negative-staining EM of urine (n = 76 samples) and present a simple and rapid protocol for the semiquantitative evaluation of patient samples. In two patient populations (either with (n = 15 samples) or without (n = 15 samples) an established diagnosis of BKN), EM results were compared with two previously established techniques for monitoring polyomavirus activation: (1) cytology for the quantitation of decoy cells, and (2) quantitative PCR assays for the detection of BK virus DNA load levels. In both patient groups, the dynamics of decoy cell shedding by urine cytology closely paralleled free viral particle shedding by EM, and viral load levels as measured by PCR. A trend toward higher readings was observed in patients with BKN (median values, control versus BKN groups: decoy cells 21 versus 50/slide; free virions by EM: 32 versus 66 viral particles/10 high-power fields; PCR: 3.5 x 10(8) versus 5.4 x 10(8) BK virus copies/ml; all differences not statistically significant). The authors conclude that negative-staining EM and the semiquantitative assessment of free viral particles in the urine can be a useful clinical method to identify patients at increased risk for BKN. EM can be used alone or in combination with urine cytology or PCR assays.
负染色电子显微镜（EM）在诊断病毒学中发挥了关键作用。它是一种用于尿液中病毒检测的快速技术，可为监测病毒活性和生产性感染提供一种简便方法。迄今为止，尚未系统评估用于检测多瘤病毒的尿液EM作为肾移植患者发生BK多瘤病毒肾病（BKN）风险的筛查工具。在此，作者讨论了尿液负染色EM的技术方面（n = 76个样本），并提出了一种简单快速的方案用于患者样本的半定量评估。在两个患者群体中（已确诊BKN的患者（n = 15个样本）和未确诊BKN的患者（n = 15个样本）），将EM结果与两种先前建立的监测多瘤病毒激活的技术进行了比较：（1）用于定量诱饵细胞的细胞学检查，以及（2）用于检测BK病毒DNA载量水平的定量PCR检测。在两个患者组中，尿液细胞学检查中诱饵细胞脱落的动态变化与EM检测到的游离病毒颗粒脱落以及PCR检测到的病毒载量水平密切平行。在BKN患者中观察到读数有升高趋势（中位数，对照组与BKN组：诱饵细胞分别为21个/玻片和50个/玻片；EM检测的游离病毒粒子：32个/10个高倍视野和66个/10个高倍视野；PCR：3.5×10⁸ BK病毒拷贝/ml和5.4×10⁸ BK病毒拷贝/ml；所有差异均无统计学意义）。作者得出结论，负染色EM和尿液中游离病毒颗粒的半定量评估可作为一种有用的临床方法来识别BKN风险增加的患者。EM可单独使用或与尿液细胞学检查或PCR检测联合使用。
