Laboratoire Hospitalo-Universitaire de Parasitologie-Mycologie Médicale, équipe EA3593, UFR de Médecine, Université des Antilles et de la Guyane, French Guiana.
Diagn Microbiol Infect Dis. 2010 Mar;66(3):268-73. doi: 10.1016/j.diagmicrobio.2009.10.010.
The main aim of our study was to determine the added value of real-time polymerase chain reaction (PCR) for the diagnosis of Histoplasma capsulatum in routine biologic practice. No amplification signal was observed with the 18 non-H. capsulatum strains used to test the specificity of the protocol. The sensitivity threshold of the real-time PCR assay was about 10 fg of H. capsulatum DNA per microliter, tested with a 10-fold serial dilution of the positive control. We analyzed 348 human samples submitted for the routine diagnosis of systemic mycosis. Real-time PCR using the TaqMan system was evaluated against direct microscopic examination and culture. Among the 341 samples without PCR inhibition (n = 7), 66 tested positive by culture, whereas 74 tested positive by real-time PCR. Sensitivity of the real-time PCR assay was estimated at 95.4% and specificity at 96.0% with respect to culture, widely considered to be the gold standard method; however, the molecular approach in fact produced better sensitivity and specificity results. Moreover, for the 38 samples that tested negative by direct examination but positive by culture, the culture method took a mean of 31 days longer than the PCR method to generate results. The protocol presented here may be very useful for improving routine histoplasmosis diagnosis.
本研究的主要目的是确定实时聚合酶链反应（PCR）在常规生物学实践中诊断荚膜组织胞浆菌的附加值。在测试方案特异性时，未观察到 18 株非荚膜组织胞浆菌菌株的扩增信号。实时 PCR 检测的灵敏度阈值约为每微升 10 fg 的荚膜组织胞浆菌 DNA，用阳性对照的 10 倍系列稀释进行测试。我们分析了 348 个人类样本，这些样本是为了常规诊断系统性真菌感染而提交的。使用 TaqMan 系统的实时 PCR 与直接显微镜检查和培养进行了评估。在没有 PCR 抑制的 341 个样本（n = 7）中，66 个通过培养呈阳性，而 74 个通过实时 PCR 呈阳性。实时 PCR 检测的灵敏度估计为 95.4%，特异性为 96.0%，相对于被广泛认为是金标准方法的培养；然而，实际上分子方法产生了更好的敏感性和特异性结果。此外，对于 38 个直接检查阴性但培养阳性的样本，培养方法比 PCR 方法产生结果平均长 31 天。这里提出的方案对于改善常规组织胞浆菌病的诊断可能非常有用。
