肺孢子菌感染或定植与呼吸系统疾病的相关性探讨

目的 检测呼吸疾病患者肺内肺孢子菌基因阳性率,探讨肺孢子菌定植或感染的影响因素及其与呼吸系统疾病的相关性。方法 设计3对特异性引物,构建检测肺孢子菌16S rRNA基因的LAMP反应体系。通过对大鼠感染模型的肺孢子菌基因检测,验证其敏感性和特异性;通过对临床呼吸疾病患者痰液样本肺孢子菌基因的检测,分析肺孢子菌感染或定殖与呼吸系统疾病的相关性及其影响因素。结果 本研究建立的LAMP方法检测肺孢子菌16S rRNA基因的最低检测极限为50 copies/mL;与念珠菌等8种呼吸道病原体不发生交叉反应。共收集和检测98例呼吸疾病患者痰标本,肺孢子菌基因总检出率为63.3%(62/98)。合并呼吸道感染者63例,非合并感染者35例,肺孢子菌基因阳性率分别为60.3%和75.0%,χ²检验两者差异有统计学意义。肺部影像学有改变者肺孢子菌基因检出率为72.5%,肺部影像学无改变者检出率为53.19%,χ²检验两者具有显著性差异。COPD和非COPD患者肺孢子菌基因检出率分别为53.2%和71.2%,χ²检验两者无显著性差异;COPD急性发作期和稳定期阳性率分别为66.8%和46.9%,χ²检验两者差异无统计学意义。CD4⁺ T淋巴细胞数降低患者肺孢子菌基因检出率80.0%;CD4⁺ T 淋巴细胞数正常者的检出率为60.27%,χ²检验两者差异具有统计学意义。1,3-β-D-葡聚糖血含量升高和含量正常患者的肺孢子菌基因检测结果显示,χ²检验两者差异无统计学意义。结论 呼吸疾病患者中肺孢子菌基因检出率较高。两者是否有直接的因果关系尚需进一步研究。合并其他细菌感染者肺孢子菌基因阳性率较低,其机制有待于进一步研究。CD4⁺ T细胞数降低者肺孢子菌基因阳性率升高,符合肺孢子菌为机会感染的特点。

Correlation between Pneumosystis colonization/infection and respiratory diseases

In the study,we investigated the correlation between respiratory diseases and Pneumocystis colonization or infection.Species-specific primers targeted the 16S rRNA gene were designed,the loop mediated isothermal amplification （LAMP） for testing Pneumosystis was established and its sensitivity and specificity was identified.The LAMP was performed to detect Pneumocystis gene in induced sputum from patients suffer from respiratory diseases.The correlation of the Pneumosystis colonization and respiratory diseases was analyzed.Results showed that the minimum amount of LAMP amplified gene was 50 copies/mL; there was no cross-reaction in other 8 kinds of respiratory pathogens.A total of 98 sputum specimens were detected and the positive rate of Pneumosystis gene was 63.3％.The positive rate of Pneumocystis gene was 53.2％ in COPD patients and 71.2％ in non-COPD patients.The positive rate was 66.8％ during COPD acute phase and 46.9％ during stability course; the difference is no statistically significant.Merger respiratory tract infection was detected in 63 cases,and the positive rate was 60.3％ in co-infected patients and 75.0％ in non-co-infected patients,showing no statisticaly difference.Positive rate of Pneumocystis gene was 72.5 ％ in the patients with changed lung imaging and 53.2 ％ in those with no lung imaging change,both having significant differences.The detection rate was 80.0％ when the CD4 ＋ T lymphocyte count was lower,and 60.3％ in the patients whose CD4＋ T lymphocyte count was normal.Pneumocystis genetic testing results showed that both had significantly statistical significance （P＞0.05） in the patients with increased 1,3-beta-D-glucan content or normal.In conclusions,the detection rate of Pneumocystis gene is higher in patients with respiratory diseases.Its mechanism remains to be further research.Pneumocystis infection or colonization is influenced by the amount of CD4＋ T cells.