A pilot study demonstrating the identification of Trypanosoma brucei gambiense and T. b. rhodesiense in vectors using a multiplexed high-resolution me

非洲人类锥虫病（HAT）是由锥虫亚种布氏锥虫、冈比亚锥虫和罗得西亚锥虫引起的一种潜在致命寄生虫感染，由采采蝇传播。HAT监测依赖于主动和被动病例发现，随着许多疫源地病例数接近消除，需要采用简单和可持续的筛查方法。在这里，我们描述了一种新的qPCR检测方法的原理应用证明，该方法设计用于检测采采蝇中两种人类致病性锥虫。对先前鉴定为布氏弓形虫阳性的96条野生采采蝇进行了化验评估，其中2条已知为罗得西亚弓形虫阳性。结果表明，该方法特异性强，与非靶锥虫无交叉反应，检测限为104 tryps/mL。qPCR成功鉴定出3只罗得西亚锥虫阳性蝇类，与参考pcr结果一致。该方法在筛选布氏体致病性亚种时提供了一种替代多种pcr方法，并在不到2小时内产生定量结果。该方法可作为一种简单有效的方法，在已知的HAT疫源地、有复发危险的地区或受HAT分布变化威胁的地区对大量采采蝇进行筛查。

Human African Trypanosomiasis (HAT) is a potentially fatal parasitic infection caused by the trypanosome sub-species Trypanosoma brucei gambiense and T. b. rhodesiense transmitted by tsetse flies. HAT surveillance is reliant on active and passive case detection and with case numbers nearing elimination in many foci there is a need for simple and sustainable screening methods. Here, we describe a proof of principle application of a novel qPCR assay designed to detect both human pathogenic trypanosome species in tsetse. Assay evaluation was carried out on 96 wild-caught tsetse previously identified to be positive for T. brucei s. l. of which two were known to be positive for T. b. rhodesiense. The assay was found to be highly specific with no cross-reactivity with non-target trypanosome species and the assay limit of detection was 104 tryps/mL. The qPCR successfully identified three T. b. rhodesiense positive flies, in agreement with the reference PCRs. This assay provides an alternative to running multiple PCRs when screening for pathogenic sub-species of T. brucei s. l. and produces quantitative results in less than 2 hours. This method could provide a component of a simple and efficient method of screening large numbers of tsetse flies in known HAT foci, in areas at risk of recrudescence or threatened by the changing distribution of HAT.