Versage Jessica L, Severin Darlena D M, Chu May C, Petersen Jeannine M
Bacterial Zoonoses Branch, Division of Vector-Borne Infectious Diseases, National Center for Infectious Disease, Centers for Disease Control and Prevention, Ft. Collins, Colorado 80522, USA.
J Clin Microbiol. 2003 Dec;41(12):5492-9. doi: 10.1128/JCM.41.12.5492-5499.2003.
Tularemia is the zoonotic disease caused by the gram-negative coccobacillus Francisella tularensis. Its wide distribution in the environment poses a challenge for understanding the transmission, ecology, and epidemiology of the disease. F. tularensis is also considered a potential biological weapon due to its extreme infectivity. We have developed a multitarget real-time TaqMan PCR assay capable of rapidly and accurately detecting F. tularensis in complex specimens. Targeted regions included the ISFtu2 element and the 23kDa, fopA, and tul4 genes. Analysis of the four TaqMan assays demonstrated that three (ISFtu2, 23kDa, and tul4) performed within our established criterion of a detection limit of one organism. The combined use of the three assays was highly specific, displaying no cross-reactivity with the non-Francisella bacteria tested and capable of differentially diagnosing both F. tularensis and Francisella philomiragia. When the multitarget TaqMan assay (ISFtu2, 23kDa, and tul4) was compared to culturing, using environmentally contaminated specimens, the TaqMan PCR assay was significantly more sensitive than culturing (P </= 0.05). The sensitive and specific nature of this rapid multitarget TaqMan assay provides a valuable new tool that with future evaluations can be used for analyzing clinical specimens, field samples during bioterrorism threat assessment, and samples from outbreaks and for improving our understanding of the ecology and environmental prevalence of F. tularensis.
兔热病是由革兰氏阴性球杆菌土拉弗朗西斯菌引起的人畜共患病。它在环境中的广泛分布给了解该疾病的传播、生态学和流行病学带来了挑战。由于其极强的传染性，土拉弗朗西斯菌也被视为一种潜在的生物武器。我们开发了一种多靶点实时TaqMan PCR检测方法，能够快速、准确地检测复杂样本中的土拉弗朗西斯菌。靶向区域包括ISFtu2元件以及23kDa、fopA和tul4基因。对这四种TaqMan检测方法的分析表明，其中三种（ISFtu2、23kDa和tul4）在我们确定的检测限为一个生物体的标准范围内。这三种检测方法联合使用具有高度特异性，与所检测的非弗朗西斯菌属细菌无交叉反应，并且能够对土拉弗朗西斯菌和嗜肺弗朗西斯菌进行鉴别诊断。当使用受环境污染的样本将多靶点TaqMan检测方法（ISFtu2、23kDa和tul4）与培养法进行比较时，TaqMan PCR检测方法的灵敏度明显高于培养法（P≤0.05）。这种快速多靶点TaqMan检测方法的灵敏性和特异性提供了一种有价值的新工具，经过未来评估后可用于分析临床样本、生物恐怖主义威胁评估期间的现场样本、疫情爆发样本，以及增进我们对土拉弗朗西斯菌的生态学和环境流行情况的了解。
