INTRODUCTION A test comparison of in-house and commercial real-time PCR (qPCR) kits for the detection of human parasites and microsporidia in stool samples was conducted without a gold standard. Three different commercial kits were included in the comparison, with a range of 3-15 different PCR targets, while 14 targets were covered by in-house testing, so not all 16 target pathogens were covered by all assays. METHODS Residual materials from nucleic acid extractions of stool samples with very high likelihood of being colonized or infected by at least one enteric parasite species or microsporidia were tested. In all, 500 DNA samples were analyzed, but due to limited sample volume, only 250 of the 500 samples were tested per assay. Each sample was assessed with the qPCR platforms being compared and cycle threshold (Ct) values were included in a descriptive comparison. RESULTS Depending on the assay applied, qPCR detected per 250 tested samples Giardia duodenalis (184-205), Blastocystis spp. (174-183), Trichuris trichiura (118-120), Ascaris lumbricoides (79-96), Necator americanus (78-106), Hymenolepis nana (40-42), Cryptosporidium spp. (27-36), Dientamoeba fragilis (26-28), Schistosoma spp. (13-23), Enterobius vermicularis (8-14), Entamoeba histolytica (7-16), Strongyloides stercoralis (6-38), Cyclospora spp. (6-13), Taenia spp. (1-4), microsporidia (1-5), and Ancylostoma spp. (1-2). Inter-assay agreement kappa was almost perfect (0.81-1) for Dientamoeba fragilis, Hymenolepis nana, Cryptosporidium spp., and Ascaris lumbricoides, substantial (0.61-0.8) for Necator americanus, Blastocystis spp., Ancylostoma spp., Giardia duodenalis, Schistosoma spp., Trichuris trichiura, and Enterobius vermicularis, moderate (0.41-0.6) for Entamoeba histolytica, fair (0.21-0.4) for microsporidia, slight (0-0.2) for Cyclospora spp. and Strongyloides stercoralis, and poor (<0) for Taenia spp. CONCLUSIONS Varying inter-assay agreement makes interpretation of microsporidia and parasite PCR in stool samples challenging. Intra-assay agreement had been controlled during the developing of the assays. Future studies, e.g., with optimized nucleic acid procedures and including microscopically characterized samples, are advisable.

中文翻译：

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没有黄金标准的人类粪便样品中15种寄生虫和微孢子虫的商业和内部实时PCR平台的比较。

引言在没有黄金标准的情况下，进行了室内和商业实时PCR（qPCR）试剂​​盒的检测比较，以检测粪便样本中的人寄生虫和微孢子虫。比较中包括三种不同的商业试剂盒，具有3-15种不同的PCR靶标，而内部测试涵盖了14种靶标，因此并非所有测定都涵盖了全部16种靶标病原体。方法测试了粪便样本核酸提取物中的残留物质，这些残留物质极有可能被至少一种肠寄生虫物种或微孢子虫感染或感染。总共分析了500个DNA样品，但由于样品量有限，每次测定仅测试500个样品中的250个。通过比较qPCR平台评估每个样品，并将循环阈值（Ct）值包括在描述性比较中。结果根据所应用的测定，每250个测试样品中的贾第鞭毛虫（Giardia duodenalis）（184-205），Blastocystis spp检测到qPCR。（174-183），Trichuris trichiura（118-120），Ascaris lumbricoides（79-96），American Necator（78-106），Hymenolepis nana（40-42），隐孢子虫属。（27-36），脆弱的Dientamoeba（26-28），血吸虫。（13-23），Enterobius vermicularis（8-14），组织变形虫（Enttamoeba histolytica）（7-16），steryloides stercoralis（6-38），Cyclospora spp。（6-13），Ta虫属。（1-4），小孢子虫（1-5）和Ancylostoma spp。（1-2）。对于脆弱的Dientamoeba脆弱菌，Nymenolepis nana，隐孢子虫和虫（Ascaris lumbricoides），批间测定一致性kappa几乎完美（0.81-1）。8）美洲轮虫，囊虫，十二指肠虫，血吸虫，Trichiris trichiura和Vermicularis肠球菌，中度（0.41-0.6），小孢子虫（0.21-0.4），轻度（0） -0.2），用于Cyclospora spp。和硬核龙眼，以及对Ta虫（Taenia spp）不良（<0）。结论不同的分析方法之间的协议使粪便样品中小孢子虫和寄生虫PCR的解释具有挑战性。在试验开发过程中已控制了试验内协议。建议进行进一步的研究，例如使用优化的核酸程序并包括显微表征的样品。4）对于小孢子虫，对环孢菌属的孢子为轻微（0-0.2）。和硬核龙眼，以及对Ta虫（Taenia spp）不良（<0）。结论不同的分析方法之间的协议使粪便样品中微孢子虫和寄生虫PCR的解释具有挑战性。在试验开发过程中已控制了试验内协议。建议进行进一步的研究，例如使用优化的核酸程序并包括显微表征的样品。4）对于小孢子虫，对环孢菌属的孢子为轻微（0-0.2）。和硬核龙眼，以及对Ta虫（Taenia spp）不良（<0）。结论不同的分析方法之间的协议使粪便样品中小孢子虫和寄生虫PCR的解释具有挑战性。在试验开发过程中已控制了试验内协议。建议进行进一步的研究，例如采用优化的核酸程序并包括显微表征的样品。