定量PCR检测对侵袭性真菌感染肺组织标本病原真菌的诊断价值

Diagnostic Value of Quantitative PCR in Pathogenic Fungi of Invasive Fungal Infection in Lung Tissue

目的探讨应用定量聚合酶链反应(polymerase chain reaction,PCR)检测技术诊断侵袭性真菌感染(invasive fungal infection,IFI)肺组织标本病原真菌的临床意义。方法收集2010年6月至2016年6月间我院行手术切除且病理诊断为真菌感染的肺组织标本33例和同期非真菌感染的正常肺组织标本10例为研究对象,设计针对真菌的通用引物和针对隐球菌属和曲霉菌的特异性引物,采用PCR技术和双脱氧链终止法(Sanger法测序)检测石蜡包埋肺切除组织中的真菌。结果实验组33例真菌感染的肺组织标本PCR检测结果均为阳性,与病理诊断相符,Sanger测序可以鉴定真菌到种别。PCR检测与Sanger测序敏感性差异无统计学意义(P>0.05),PCR检测与Sanger测序特异性差异无统计学意义(P>0.05);PCR检测与镜检的种属鉴定率差异有统计学意义(P<0.05),PCR检测与Sanger测序的种属鉴定率差异无统计学意义(P>0.05)。结论定量PCR检测用于诊断侵袭性真菌感染肺组织标本敏感性高、特异性好,具有一定参考意义。

Objective To investigate the clinical significance of quantitative polymerase chain reaction（polymerase chain reaction, PCR） technique in the diagnosis of fungal pathogens of invasive fungal infection （ invasive fungal infection, IFI） in lung tis- sue. Methods 33 cases of lung tissue specimens diagnosed with IFI by pathological section and 10 cases of non-fungal infected lung tissue specimens were collected as investigated subjects from June 2010 to June 2016. Universal primers for fungi and specificprimers for Cryptococcus spp. and Aspergillus spp. were designed, and then the fungi in lung tissue embedded in paraffin were detec- ted by PCR and double strand termination（ Sanger sequencing）. Results All of the PCR test results of 33 cases specimens in experi- mental group were positive, which was coherent with pathological diagnosis. Sanger sequencing could identify the species of fungi. There were no significant difference between PCR and Sanger sequencing in sensitivity and specificity （P 〉 0. 05 ）. The species identi- fication rate of PCR was significantly better than that of microscopy（P = 0. 01 ）. The difference between PCR and Sanger sequencing in detect the fungi in lung tissue specimens was not statistically significant （ P = 0. 08 ）. Conclusion Quantitative PCR is a highly sensitive and specific method for the diagnosis of invasive fungal infection in lung tissue, and it has a certain reference value.