FTA®卡作为支气管肺泡灌洗样本中肺孢子虫DNA提取方法的实用性。

Usefulness of FTA® cards as a Pneumocystis-DNA extraction method in bronchoalveolar lavage samples.

作者信息

Rodiño Jenniffer M, Aguilar Yudy A, Rueda Zulma Vanessa, Vélez Lázaro A

机构信息

a Grupo Investigador de Problemas en Enfermedades Infecciosas (GRIPE), Facultad de Medicina ;

b Corporación de Ciencias Básicas Biomédicas, Universidad de Antioquia , Medellín , Colombia ;

出版信息

Infect Dis (Lond). 2016;48(5):367-72. doi: 10.3109/23744235.2015.1129550. Epub 2016 Jan 21.

DOI:10.3109/23744235.2015.1129550

PMID:26950684

Abstract

BACKGROUND

FTA® cards (Fast Technology for Analysis of Nucleic Acids) are an alternative DNA extraction method in bronchoalveolar lavage (BAL) samples for Pneumocystis jirovecii molecular analyses. The goal was to evaluate the usefulness of FTA® cards to detect P. jirovecii-DNA by PCR in BAL samples compared to silica adsorption chromatography (SAC).

METHODS

This study used 134 BAL samples from immunocompromised patients previously studied to establish microbiological aetiology of pneumonia, among them 15 cases of Pneumocystis pneumonia (PCP) documented by staining and 119 with other alternative diagnoses. The FTA® system and SAC were used for DNA extraction and then amplified by nested PCR to detect P. jirovecii. Performance and concordance of the two DNA extraction methods compared to P. jirovecii microscopy were calculated. The influence of the macroscopic characteristics, transportation of samples and the duration of the FTA® card storage (1, 7, 10 or 12 months) were also evaluated.

RESULTS

Among 134 BAL samples, 56% were positive for P. jirovecii-DNA by SAC and 27% by FTA®. All 15 diagnosed by microscopy were detected by FTA® and SAC. Specificity of the FTA® system and SAC were 82.4% and 49.6%, respectively. Compared to SAC, positivity by FTA® decreased with the presence of blood in BAL (62% vs 13.5%). The agreement between samples at 7, 10 and 12 months was 92.5% for FTA®. Positive cases by FTA® remained the same after shipment by mail.

CONCLUSIONS

Results suggest that FTA® is a practical, safe and economical method to preserve P. jirovecii-DNA in BAL samples for molecular studies.

摘要

背景

FTA®卡（核酸快速分析技术）是支气管肺泡灌洗（BAL）样本中用于耶氏肺孢子菌分子分析的另一种DNA提取方法。目的是评估与硅胶吸附色谱法（SAC）相比，FTA®卡在通过PCR检测BAL样本中耶氏肺孢子菌DNA方面的实用性。

方法

本研究使用了134份来自免疫功能低下患者的BAL样本，这些样本之前已用于确定肺炎的微生物病因，其中15例经染色确诊为肺孢子菌肺炎（PCP），119例为其他替代诊断。使用FTA®系统和SAC进行DNA提取，然后通过巢式PCR扩增以检测耶氏肺孢子菌。计算了两种DNA提取方法与耶氏肺孢子菌显微镜检查相比的性能和一致性。还评估了宏观特征、样本运输以及FTA®卡储存时间（1、7、10或12个月）的影响。

结果

在134份BAL样本中，SAC法检测耶氏肺孢子菌DNA的阳性率为56%，FTA®法为27%。所有15例经显微镜诊断的病例均被FTA®和SAC检测到。FTA®系统和SAC的特异性分别为82.4%和49.6%。与SAC相比，BAL中存在血液时FTA®的阳性率降低（62%对13.5%）。FTA®在7、10和12个月时样本之间的一致性为92.5%。FTA®阳性病例经邮寄运输后保持不变。