Lu J J, Chen C H, Bartlett M S, Smith J W, Lee C H
Department of Pathology, Tri-Service General Hospital and National Defense Medical Center, Taipei, Taiwan, Republic of China.
J Clin Microbiol. 1995 Oct;33(10):2785-8. doi: 10.1128/jcm.33.10.2785-2788.1995.
We have recently developed a nested PCR method which amplifies internal transcribed spacers (ITS) of the ribosomal RNA genes of Pneumocystis carinii. To determine whether this PCR method can be used to diagnose P. carinii infections, we examined 30 bronchoalveolar lavage (BAL) specimens that were shown microscopically to contain P. carinii organisms by the P. carinii ITS PCR (Pc-ITS-PCR) and five other PCR methods that have been described for detecting P. carinii in clinical specimens. The targets of these PCR methods are portions of 18S rRNA, mitochondrial (mt) rRNA, 5S rRNA, thymidylate synthase (TS), and dihydrofolate reductase (DHFR). We also examined five different fungi, including Saccharomyces cerevisiae, Candida albicans, Histoplasma capsulatum, Cryptococcus neoformans, and Aspergillus fumigatus to determine the specificity of these six PCR methods for P. carinii. All 30 BAL specimens were positive by both the Pc-ITS-PCR and the 18S rRNA gene PCR, whereas only 26 (87%), 18 (60%), 10 (33%), and 7 (23%) of 30 BAL specimens were positive by mt rRNA gene PCR, TS gene PCR, 5S rRNA gene PCR, and DHFR gene PCR, respectively. Although the 18S rRNA gene PCR had the same sensitivity as the Pc-ITS-PCR, it nonspecifically amplified S. cerevisiae and C. albicans. The TS gene PCR also produced false-positive PCR results with C. albicans and C. neoformans. None of the other PCR methods (Pc-ITS-PCR, mt rRNA gene, 5S rRNA gene, and DHFR gene PCR) amplified the control fungal DNA. Considering both sensitivity and specificity, we conclude that Pc-ITS-PCR is the most effective of the six PCR methods evaluated in this study for the detection of P. carinii in BAL specimens.
我们最近开发了一种巢式聚合酶链反应（PCR）方法，该方法可扩增卡氏肺孢子虫核糖体RNA基因的内部转录间隔区（ITS）。为了确定这种PCR方法是否可用于诊断卡氏肺孢子虫感染，我们检测了30份支气管肺泡灌洗（BAL）标本，这些标本经显微镜检查显示含有卡氏肺孢子虫病原体，采用卡氏肺孢子虫ITS PCR（Pc-ITS-PCR）以及已描述的用于检测临床标本中卡氏肺孢子虫的其他五种PCR方法。这些PCR方法的靶标是18S rRNA、线粒体（mt）rRNA、5S rRNA、胸苷酸合成酶（TS）和二氢叶酸还原酶（DHFR）的部分片段。我们还检测了五种不同的真菌，包括酿酒酵母、白色念珠菌、荚膜组织胞浆菌、新型隐球菌和烟曲霉，以确定这六种PCR方法对卡氏肺孢子虫的特异性。所有30份BAL标本经Pc-ITS-PCR和18S rRNA基因PCR检测均为阳性，而30份BAL标本中分别只有26份（87%）、18份（60%）、10份（33%）和7份（23%）经mt rRNA基因PCR、TS基因PCR、5S rRNA基因PCR和DHFR基因PCR检测为阳性。尽管18S rRNA基因PCR与Pc-ITS-PCR具有相同的敏感性，但它非特异性地扩增了酿酒酵母和白色念珠菌。TS基因PCR对白色念珠菌和新型隐球菌也产生了假阳性PCR结果。其他PCR方法（Pc-ITS-PCR、mt rRNA基因、5S rRNA基因和DHFR基因PCR）均未扩增对照真菌DNA。综合考虑敏感性和特异性，我们得出结论，在本研究评估的六种PCR方法中，Pc-ITS-PCR是检测BAL标本中卡氏肺孢子虫最有效的方法。
