Ombrouck C, Ciceron L, Biligui S, Brown S, Marechal P, van Gool T, Datry A, Danis M, Desportes-Livage I
Centre Hospitalo-Universitaire de la Pitié-Salpêtrière, Unité INSERM 313, Paris, France.
J Clin Microbiol. 1997 Mar;35(3):652-5. doi: 10.1128/jcm.35.3.652-655.1997.
A routine assay based on the PCR was developed for the detection of Enterocytozoon bieneusi and Encephalitozoon intestinalis in fecal samples. Two oligonucleotide primer pairs from a conserved region in the small-subunit rRNA genes of E. bieneusi (primer pair V1 and EB450) and E. intestinalis (primer pair V1 and SI500) were used to amplify microsporidian DNA. We achieved specific amplification of a 382-bp DNA fragment in E. intestinalis and a 353-bp DNA fragment in E. bieneusi. Boiling of the samples appeared to be most effective for DNA extraction. Fecal samples containing fewer than 10 microsporidia gave a positive result in the PCR assay. Fecal specimens from 30 human immunodeficiency virus-infected patients with microsporidiosis and fecal specimens from 42 patients suspected of having microsporidiosis were investigated by the PCR assay. The PCR assay was validated against standard staining methods (the Uvitex 2B and Chromotrope 2R staining methods) and immunofluorescence assay specific for E. intestinalis. This comparative study has shown that PCR improved species determination and can thus be considered a fast and reliable method for the detection and identification of each intestinal species.
开发了一种基于聚合酶链反应（PCR）的常规检测方法，用于检测粪便样本中的比氏肠微孢子虫和肠脑炎微孢子虫。使用来自比氏肠微孢子虫小亚基核糖体RNA基因保守区域的两对寡核苷酸引物（引物对V1和EB450）以及肠脑炎微孢子虫的（引物对V1和SI500）来扩增微孢子虫DNA。我们成功地特异性扩增出了肠脑炎微孢子虫中的一个382碱基对的DNA片段以及比氏肠微孢子虫中的一个353碱基对的DNA片段。样本煮沸似乎是最有效的DNA提取方法。含少于10个微孢子虫的粪便样本在PCR检测中呈阳性结果。通过PCR检测对30例感染人类免疫缺陷病毒并患有微孢子虫病的患者的粪便样本以及42例疑似患有微孢子虫病的患者的粪便样本进行了研究。该PCR检测方法与标准染色方法（乌洛托品2B和变色酸2R染色方法）以及针对肠脑炎微孢子虫的免疫荧光检测方法进行了验证。这项比较研究表明，PCR改善了物种鉴定，因此可被视为一种快速且可靠的检测和鉴定每种肠道微孢子虫物种的方法。
