Analytical Performance of Four Polymerase Chain Reaction (PCR) and Real Time PCR (qPCR) Assays for the Detection of Six Leishmania Species DNA in Colombia.
Analytical Performance of Four Polymerase Chain Reaction (PCR) and Real Time PCR (qPCR) Assays for the Detection of Six Leishmania Species DNA in Colombia.
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四种聚合酶链式反应和实时定量聚合酶链式反应检测哥伦比亚6种利什曼原虫DNA的分析性能
DOI：
10.3389/fmicb.2017.01907
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发表时间：
2017
影响因子：
5.2
通讯作者：
Ramírez JD
中科院分区：
文献类型：
--
作者：
León CM;Muñoz M;Hernández C;Ayala MS;Flórez C;Teherán A;Cubides JR;Ramírez JD
利什曼病包括由利什曼属原生动物引起的一系列寄生虫病。由于利什曼原虫的高灵敏度和特异性，分子工具已被广泛应用于利什曼原虫的检测。然而，分子平台的分析性能，如PCR和实时PCR (qPCR)，包括各种各样的分子标记从未被评估过。在此，目的是评估4种基于PCR的分析方法（针对4种不同的目标设计），并在传统和实时PCR平台上应用的分析性能。我们评估了传统PCR和实时PCR的分析性能，确定了专有性和包容性，预期报告范围（ARR），检测限（LoD）和准确性，使用针对kDNA， HSP70, 18S和ITS-1目标的引物。我们观察到kDNA是最敏感的，但不符合排他性标准。与其他标记相比，HSP70在常规PCR和qPCR中具有较高的LoD（分别为1 × 101和1 × 10-1个等效寄生虫/mL）， qPCR的变异系数较高。四种分子标记在测试的天数之间没有统计学上的显著差异。本研究表明，18S标记在qPCR检测物种（哥伦比亚流行物种）的分析敏感性和特异性方面表现最好。因此，我们建议在未来的研究中使用更广泛的物种来探索分析和诊断性能。
Leishmaniasis comprises a spectrum of parasitic diseases caused by protozoans of the genus Leishmania. Molecular tools have been widely employed for the detection of Leishmania due to its high sensitivity and specificity. However, the analytical performance of molecular platforms as PCR and real time PCR (qPCR) including a wide variety of molecular markers has never been evaluated. Herein, the aim was to evaluate the analytical performance of 4 PCR-based assays (designed on four different targets) and applied on conventional and real-time PCR platforms. We evaluated the analytical performance of conventional PCR and real time PCR, determining exclusivity and inclusivity, Anticipated Reportable Range (ARR), limit of detection (LoD) and accuracy using primers directed to kDNA, HSP70, 18S and ITS-1 targets. We observed that the kDNA was the most sensitive but does not meet the criterion of exclusivity. The HSP70 presented a higher LoD in conventional PCR and qPCR in comparison with the other markers (1 × 101 and 1 × 10-1 equivalent parasites/mL respectively) and had a higher coefficient of variation in qPCR. No statistically significant differences were found between the days of the test with the four molecular markers. The present study revealed that the 18S marker presented the best performance in terms of analytical sensitivity and specificity for the qPCR in the species tested (species circulating in Colombia). Therefore, we recommend to explore the analytical and diagnostic performance in future studies using a broader number of species across America.
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影响因子：
2.7
作者：
Abbasi I;Kirstein OD;Hailu A;Warburg A
通讯作者：
Warburg A
影响因子：
1.5
作者：
BOCK, JH;LANGER, PJ
通讯作者：
LANGER, PJ
DOI：
10.4269/ajtmh.2010.09-0414
发表时间：
2010-08-01
影响因子：
3.3
作者：
Boggild, Andrea K.;Ramos, Ana P.;Llanos-Cuentas, Alejandro
通讯作者：
Llanos-Cuentas, Alejandro
影响因子：
3.4
作者：
Oliveira, Diego Molina de;Lonardoni, Maria Valdrinez Campana;Silveira, Thais Gomes Verzignassi
通讯作者：
Silveira, Thais Gomes Verzignassi
影响因子：
2.1
作者：
Bezerra-Vasconcelos, Diana R.;Melo, Luciana M.;Bevilaqua, Claudia M. L.
通讯作者：
Bevilaqua, Claudia M. L.