College of Veterinary Medicine, Northwest A&F University, Yangling, 712100, Shaanxi Province, People's Republic of China.
College of Veterinary Medicine, Henan Agricultural University, Zhengzhou, 450046, Henan Province, People's Republic of China.
Parasitol Res. 2021 May;120(5):1837-1844. doi: 10.1007/s00436-021-07101-2. Epub 2021 Mar 2.
Cryptosporidium is an important intestinal protozoan parasite that causes diarrhoea in humans and animals. To rapidly and specifically detect Cryptosporidium spp., we designed a pair of primers based on the small subunit ribosomal RNA (SSU rRNA) gene of Cryptosporidium spp. to be used in a new nanoparticle-assisted PCR (nano-PCR) assay. The minimum detectable concentration (1.02 pg) of this nano-PCR was 10 times more sensitive than conventional PCR using the same primer pair. The DNA samples of C. parvum, C. baileyi, C. xiaoi, C. ryanae, and C. andersoni were successfully detected by the nano-PCR. No amplifications were evident with DNA samples of some common intestinal pathogens, including Eimeria tenella, Blastocystis sp., Giardia lamblia, Enterocytozoon bieneusi, and Balantidium coli. To validate the clinical usefulness of the novel nano-PCR, a total of 40 faecal samples from goats, camels, calves, and chickens were examined. The positive rate of Cryptosporidium spp. was 27.5% (11/40), which was consistent with that of an established nested PCR. These results indicate that the novel nano-PCR assay enables the rapid, specific, and accurate detection of Cryptosporidium infection in animals. The findings provide a technical basis for the clinical diagnosis, prevention, and control of cryptosporidiosis.
隐孢子虫是一种重要的肠道原生动物寄生虫，可引起人类和动物腹泻。为了快速且特异性地检测隐孢子虫属，我们基于隐孢子虫属的小亚基核糖体 RNA (SSU rRNA) 基因设计了一对引物，用于新的纳米颗粒辅助 PCR (nano-PCR) 检测。与使用相同引物对的常规 PCR 相比，该 nano-PCR 的最小可检测浓度（1.02pg）灵敏 10 倍。nano-PCR 成功检测到 C. parvum、C. baileyi、C. xiaoi、C. ryanae 和 C. andersoni 的 DNA 样本。用一些常见的肠道病原体的 DNA 样本进行检测时，没有明显的扩增，包括柔嫩艾美耳球虫、蓝氏贾第鞭毛虫、隐孢子虫、肠上皮细胞内原虫和结肠小袋纤毛虫。为了验证新型 nano-PCR 的临床实用性，对来自山羊、骆驼、小牛和鸡的 40 份粪便样本进行了检测。隐孢子虫属的阳性率为 27.5%（11/40），与已建立的巢式 PCR 一致。这些结果表明，新型 nano-PCR 检测可快速、特异性和准确地检测动物中的隐孢子虫感染。该研究结果为隐孢子虫病的临床诊断、预防和控制提供了技术基础。
