Olsson M, Elvin K, Lidman C, Löfdahl S, Linder E
Swedish Institute for Infectious Disease Control and Microbiology, Karolinska Institute, Stockholm, Sweden.
Scand J Infect Dis. 1996;28(6):597-600. doi: 10.3109/00365549609037967.
Detection of Pneumocystis carinii by the polymerase chain reaction (PCR), based on the thymidylate synthase (TS) gene of rat P. carinii, is a specific and sensitive method for the detection of the parasite in respiratory samples. However, the use of the method is limited by a laborious phenol-chloroform DNA extraction method and an expensive and time-consuming hybridization procedure. For routine clinical samples, DNA preparation can be simplified and hybridization substituted by a nested PCR technique. Such a modified PCR procedure, based on the TS gene of P. carinii, was evaluated on 190 induced sputum samples from 50 immunosuppressed patients, infected with human immunodeficiency virus (HIV), with and without symptoms of P. carinii pneumonia (PCP). The PCR assay, preceded by a rapid DNA preparation (Wizard DNA Clean-up), detected P. carinii-DNA in 13/15 sputa containing parasites as seen by microscopy using immunocytochemical (IFL) staining, and in 10 additional sputum samples lacking demonstrable parasites by microscopy. These samples are to be considered as 'true' positives, since all but 2 were from patients, who developed a PCP within 1 year. We conclude that the nested PCR assay is more sensitive than IFL for the detection of P. carinii in AIDS patients, prior to the debut of PCP symptoms.
基于大鼠卡氏肺孢子虫胸苷酸合成酶（TS）基因，采用聚合酶链反应（PCR）检测卡氏肺孢子虫，是一种在呼吸道样本中检测该寄生虫的特异且灵敏的方法。然而，该方法的应用受到繁琐的酚 - 氯仿DNA提取方法以及昂贵且耗时的杂交程序的限制。对于常规临床样本，DNA制备可通过巢式PCR技术简化，杂交也可用其替代。基于卡氏肺孢子虫TS基因的这种改良PCR程序，在来自50例感染人类免疫缺陷病毒（HIV）的免疫抑制患者的190份诱导痰样本上进行了评估，这些患者有或无卡氏肺孢子虫肺炎（PCP）症状。该PCR检测在快速DNA制备（Wizard DNA Clean-up）之后，在13/15份经免疫细胞化学（IFL）染色显微镜检查发现含有寄生虫的痰样本中检测到了卡氏肺孢子虫DNA，并且在另外10份显微镜检查未发现明显寄生虫的痰样本中也检测到了。这些样本应被视为“真正”的阳性样本，因为除2份外，所有样本均来自在1年内发生PCP的患者。我们得出结论，在AIDS患者出现PCP症状之前，巢式PCR检测在检测卡氏肺孢子虫方面比IFL更灵敏。
