Maksimov P, Isaksson M, Schares G, Romig T, Conraths F J
Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Epidemiology, Südufer 10, 17493 Greifswald-Insel Riems, Germany.
Department of Virology Immunobiology and Parasitology, National Veterinary Institute, Uppsala, Sweden.
Food Waterborne Parasitol. 2019 Mar 10;15:e00044. doi: 10.1016/j.fawpar.2019.e00044. eCollection 2019 Jun.
Oral uptake of infectious eggs shed by canids with their faeces may lead to development of alveolar echinococcosis in humans, which is clinically similar to a malignant infiltrative tumor and may be fatal if left untreated. is therefore regarded as one of the most important and neglected metazoan parasites in the Northern hemisphere. The diagnosis of this tapeworm in the final host plays a key role in the epidemiology of . The diagnostic performance of a magnetic-capture (MC) DNA extraction protocol in combination with a minor groove-binder real time PCR (MC-MGBqPCR) for the detection of eggs was determined relative to a highly sensitive variant of the Intestinal Scraping Technique (IST) using faecal samples of foxes. In addition, we compared results obtained by MC-MGBqPCR with those of a previously validated protocol (QIAamp Fast DNA Stool Mini Kit (QT) combined with a TaqMan qPCR). Furthermore, a workflow using the NucleoMagVet DNA extraction kit (NM) in combination with MGBqPCR and TaqMan-qPCR was also included in the comparisons. To estimate the analytical sensitivity, phosphate-buffered saline and fox faecal samples were spiked with different numbers of eggs and tested in defined combinations of DNA extraction and PCR protocols. To assess the diagnostic sensitivity of the different workflows, samples were used that had been collected from the ampulla recti or the rectum of 120 foxes hunted in Brandenburg, Germany. The samples represented five IST categories formed according to the worm burden of the foxes. For DNA extraction by MC or using two other commercial extraction kits, the supernatants obtained from 3 g of bead-beaten faecal samples were used. The extracted DNAs were then processed in the respective PCR protocols. The MC-MGBqPCR showed the highest diagnostic sensitivity (93%; 95% Confidence Interval (CI): 86-97%) relative to IST. The QT extraction protocol in combination with TaqMan-qPCR had the second highest sensitivity (89%; 95% CI: 80-94%), followed by NM with MGBqPCR (86%; 95% CI: 77-93%) in comparison to IST. The lowest diagnostic sensitivity was found for the NM combined with the TaqMan-qPCR protocol (72%; 95% CI: 62-82%). In conclusion, the MC-MGBqPCR seems to represent a suitable alternative to IST. However, applied to 3 g faecal samples, the less costly QT-TaqMan-qPCR workflow yielded a similar diagnostic sensitivity relative to IST. However, differences between these two workflows were not statistically significant.
人类经口摄入犬科动物粪便中排出的感染性虫卵可能会引发肺泡型棘球蚴病，该病在临床上类似于恶性浸润性肿瘤，若不治疗可能会致命。因此，它被视为北半球最重要且最易被忽视的后生动物寄生虫之一。在终末宿主体内诊断这种绦虫对其流行病学具有关键作用。相对于使用狐狸粪便样本的肠道刮取技术（IST）的高灵敏度变体，确定了磁捕获（MC）DNA提取方案与小沟结合剂实时PCR（MC-MGBqPCR）相结合用于检测虫卵的诊断性能。此外，我们将MC-MGBqPCR获得的结果与先前验证的方案（QIAamp Fast DNA Stool Mini Kit（QT）结合TaqMan qPCR）的结果进行了比较。此外，使用NucleoMagVet DNA提取试剂盒（NM）结合MGBqPCR和TaqMan-qPCR的工作流程也纳入了比较。为了估计分析灵敏度，在磷酸盐缓冲盐水和狐狸粪便样本中加入不同数量的虫卵，并在特定的DNA提取和PCR方案组合中进行测试。为了评估不同工作流程的诊断灵敏度，使用了从德国勃兰登堡捕杀的120只狐狸的直肠壶腹或直肠收集的样本。这些样本代表了根据狐狸的蠕虫负担形成的五个IST类别。对于通过MC或使用其他两种商业提取试剂盒进行DNA提取，使用从3克经珠磨的粪便样本中获得的上清液。然后将提取的DNA在各自的PCR方案中进行处理。相对于IST，MC-MGBqPCR显示出最高的诊断灵敏度（93%；95%置信区间（CI）：86-97%）。QT提取方案与TaqMan-qPCR相结合的灵敏度次之（89%；95%CI：80-94%），其次是NM与MGBqPCR（86%；95%CI：77-93%）与IST相比。发现NM与TaqMan-qPCR方案结合的诊断灵敏度最低（72%；95%CI：62-82%）。总之，MC-MGBqPCR似乎是IST的合适替代方法。然而，应用于3克粪便样本时，成本较低的QT-TaqMan-qPCR工作流程相对于IST产生了相似的诊断灵敏度。然而，这两种工作流程之间的差异没有统计学意义。
