Detection of Balamuthia mitochondrial 16S rRNA gene DNA in clinical specimens by PCR
Detection of Balamuthia mitochondrial 16S rRNA gene DNA in clinical specimens by PCR
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DOI：
10.1128/jcm.43.7.3192-3197.2005
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发表时间：
2005-07-01
影响因子：
9.4
通讯作者：
Schuster, FL
中科院分区：
文献类型：
--
作者：
Yagi, S;Booton, GC;Schuster, FL
曼氏Balamathia mandrillaris是一种自由生活的阿米巴，在免疫功能低下和免疫功能正常的个体中引起肉芽肿性阿米巴脑炎。由于缺乏特异性症状和难以识别活检组织中的阿米巴，大多数病例未得到诊断或有效治疗，导致> 95%的死亡率。我们在此报告5例经间接免疫荧光（IIF）染色血清抗Balamuthia抗体（滴度>= 1：128）诊断的Balamuthia病，并通过IIF未染色脑组织切片和/或检测苏木精-伊红染色切片中的阿米巴确认。此外，我们已经使用PCR检测线粒体16 S rRNA基因DNA的阿米巴在临床标本，如脑组织和脑脊液（CSF）从个人Balamuthia脑炎。通过PCR在来自所有五个个体的临床样品中成功地检测到Balamuthia DNA。在3例脑组织、3例脑脊液和2例肺组织样本中的1例中检测到，但在2例肾组织样本中未检测到。一份未固定的脑组织样本的Balamuthia培养呈阳性。为了测试PCR检测Balamuthia DNA的灵敏度，将来自两个阿米巴感染阴性个体的CSF标本加标Balamuthia阿米巴。我们发现，它是可能的检测Balamuthia DNA的PCR混合物中含有线粒体DNA从1到少至0.2阿米巴每反应混合物。单个Balamuthia变形虫含有多个线粒体靶标;因此，0.2个变形虫代表用于扩增的多个靶标，并且不等同于0.2个变形虫作为靶标。
Balamathia mandrillaris is a free-living ameba that causes granulomatous amebic encephalitis in both immunocompromised and immunocompetent individuals. Because of a lack of pathognomonic symptoms and the difficulty in recognizing amebas in biopsied tissues, most cases are not diagnosed or effectively treated, leading to a > 95% mortality. We report here on five cases of balamuthiasis that were diagnosed by indirect immunofluorescence (IIF) staining of serum for anti-Balamuthia antibodies (titer >= 1:128) and confirmed by IIF of unstained brain tissue sections and/or detection of amebas in hematoxylin-eosin-stained slides. Additionally, we have used the PCR for the detection of mitochondrial 16S rRNA gene DNA from the ameba in clinical specimens such as brain tissue and cerebrospinal fluid (CSF) from individuals with Balamuthia encephalitis. Balamuthia DNA was successfully detected by the PCR in clinical samples from all five individuals. It was detected in brain tissue from three cases, in CSF from three cases, and in one of two samples of lung tissue from two individuals, but not in two samples of kidney tissue tested. One sample of unfixed brain tissue was culture positive for Balamuthia. In order to test the sensitivity of the PCR for detection of Balamuthia DNA, CSF specimens from two individuals negative for amebic infection were spiked with Balamuthia amebas. We found that it was possible to detect Balamuthia DNA in the PCR mixtures containing mitochondrial DNA from 1 to as little as 0.2 ameba per reaction mixture. A single Balamuthia ameba contains multiple mitochondrial targets; thus, 0.2 ameba represents multiple targets for amplification and is not equivalent to 0.2 of an ameba as a target.