Infectious Disease Clinical Research Program, Department of Preventive Medicine and Biostatistics, Uniformed Services University of the Health Sciences, Rockville, MD, United States of America.
Henry M Jackson Foundation for the Advancement of Military Medicine, Bethesda, MD, United States of America.
PLoS One. 2018 Aug 30;13(8):e0202178. doi: 10.1371/journal.pone.0202178. eCollection 2018.
The use of Polymerase Chain Reaction (PCR) assays for pathogen detection in travelers' diarrhea (TD) field studies is limited by the on-site processing and storage requirements for fecal specimens. The objectives of this investigation were to i) characterize the pathogen distribution in deployed military personnel with TD using the TaqMan® Array Card PCR (TAC) on frozen stool and diarrheal smears on Whatman FTA Elute cards (FTA cards), and to ii) compare TAC detection of enteropathogen targets using smeared FTA cards and frozen stool, using TAC on frozen stool as the 'reference standard'. Stool samples, obtained from active duty personnel with acute TD enrolled in a field trial, were smeared onto FTA cards and stored at room temperature. A corresponding aliquot of stool was frozen in a cryovial. FTA cards and frozen stool samples were tested at a central lab, using a customized TAC for detection of TD pathogens. 187 paired frozen stool samples and smeared FTA cards were stored for a median of 712 days (IQR 396-750) before testing. Overall detection rates were 78.6% for frozen stool and 73.2% for FTA cards. Diarrheagenic Escherichia coli were the most common bacteria identified. Using the TAC results on frozen stool as the reference, the overall sensitivity and specificity of TAC on FTA cards was 72.9% and 98.0% respectively. TAC on FTA cards demonstrated a decrease in sensitivity with increasing frozen stool quantification cycle (Cq) (90.0% in FTA cards with a corresponding frozen stool Cq < 30, and 72.9% in samples with a corresponding frozen stool Cq < 35). Our findings support the use and further development of FTA cards in combination with a quantitative PCR assay for enteropathogen detection in TD field studies.
聚合酶链反应（PCR）检测旅行者腹泻（TD）病原体的应用受到粪便标本现场处理和储存要求的限制。本研究的目的是：i）使用 TaqMan®Array Card PCR（TAC）对冷冻粪便和在 Whatman FTA Elute 卡（FTA 卡）上的粪便涂片检测部署军人腹泻病患者的病原体分布情况，ii）使用 FTA 卡上的涂片和冷冻粪便比较 TAC 检测肠道病原体靶标，以 TAC 检测冷冻粪便为“参考标准”。从参加现场试验的急性 TD 现役人员中获得粪便样本，涂抹在 FTA 卡上并在室温下储存。相应的粪便等分试样在冷冻管中冷冻。在中央实验室使用针对 TD 病原体定制的 TAC 对 FTA 卡和冷冻粪便样本进行检测。187 对冷冻粪便样本和涂抹 FTA 卡储存中位数为 712 天（IQR 396-750），然后进行检测。冷冻粪便的总检测率为 78.6％，FTA 卡的总检测率为 73.2％。腹泻性大肠埃希菌是最常见的鉴定细菌。以冷冻粪便上的 TAC 结果为参考，FTA 卡上 TAC 的总灵敏度和特异性分别为 72.9％和 98.0％。FTA 卡上的 TAC 随冷冻粪便定量循环（Cq）的增加而灵敏度降低（对应冷冻粪便 Cq <30 的 FTA 卡中为 90.0％，对应冷冻粪便 Cq <35 的样本中为 72.9％）。我们的研究结果支持在 TD 现场研究中使用 FTA 卡结合定量 PCR 检测肠道病原体，并进一步开发该方法。
