Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.
Parasitol Res. 2010 Mar;106(4):883-8. doi: 10.1007/s00436-010-1728-y. Epub 2010 Feb 19.
Entamoeba histolytica is known to cause intestinal and extra-intestinal disease while the other Entamoeba species are not considered to be pathogenic. However, all Entamoeba spp. should be reported when identified in clinical samples. Entamoeba polecki, Entamoeba coli, and Entamoeba hartmanii can be differentiated morphologically from E. histolytica, but some of their diagnostic morphologic features overlap. E. histolytica, Entamoeba dispar, and Entamoeba moshkovskii are morphologically identical but can be differentiated using molecular tools. We developed a polymerase chain reaction (PCR) procedure followed by DNA sequencing of specific regions of 18S rRNA gene to differentiate the Entamoeba spp. commonly found in human stools. This approach was used to analyze 45 samples from cases evaluated for the presence of Entamoeba spp. by microscopy and a real-time PCR method capable of differential detection of E. histolytica and E. dispar. Our results demonstrated an agreement of approximately 98% (45/44) between the real-time PCR for E. histolytica and E. dispar and the 18S rRNA analysis described here. Five previously negative samples by microscopy revealed the presence of E. dispar, E. hartmanii, or E. coli DNA. In addition, we were able to detect E. hartmanii in a stool sample that had been previously reported as negative for Entamoeba spp. by microscopy. Further microscopic evaluation of this sample revealed the presence of E. hartmanii cysts, which went undetected during the first microscopic evaluation. This PCR followed by DNA sequencing will be useful to refine the diagnostic detection of Entamoeba spp. in stool and other clinical specimens.
溶组织内阿米巴被认为会引起肠道和肠道外疾病，而其他内阿米巴种则不被认为具有致病性。然而，当在临床样本中发现所有内阿米巴属时，都应报告。形态上可以将 polecki 内阿米巴、大肠内阿米巴和 hartman 内阿米巴与溶组织内阿米巴区分开来，但它们的一些诊断形态特征存在重叠。溶组织内阿米巴、迪斯帕内阿米巴和 moshkovskii 内阿米巴在形态上是相同的，但可以使用分子工具进行区分。我们开发了一种聚合酶链反应（PCR）程序，随后对 18S rRNA 基因的特定区域进行 DNA 测序，以区分常见于人类粪便中的内阿米巴属。该方法用于分析 45 个来自病例的样本，这些病例通过显微镜和实时 PCR 方法评估是否存在内阿米巴属。实时 PCR 法可检测溶组织内阿米巴和迪斯帕内阿米巴，本研究建立的方法可以区分内阿米巴属，实时 PCR 法和 18S rRNA 分析的结果有 98%（45/44）的一致性。显微镜检查先前为阴性的 5 个样本显示存在迪斯帕内阿米巴、hartman 内阿米巴或大肠内阿米巴 DNA。此外，我们还能够在先前通过显微镜检查报告为内阿米巴属阴性的粪便样本中检测到 hartman 内阿米巴。对该样本进行进一步的显微镜评估显示存在未在第一次显微镜评估中检测到的 hartman 内阿米巴包囊。该 PCR 方法结合 DNA 测序将有助于提高粪便和其他临床标本中内阿米巴属的诊断检测。
