无症状结核接触者外周血单个核细胞中复杂 DNA 的检测：一项观察性研究。

Detection of complex DNA in CD34-positive peripheral blood mononuclear cells of asymptomatic tuberculosis contacts: an observational study.

机构信息

Barts and the London School of Medicine and Dentistry, Queen Mary University of London, London, UK.

Armauer Hansen Research Institute, Addis Ababa, Ethiopia.

出版信息

Lancet Microbe. 2021 Jun;2(6):e267-e275. doi: 10.1016/S2666-5247(21)00043-4.

DOI:10.1016/S2666-5247(21)00043-4

PMID:34100007

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8172384/

Abstract

BACKGROUND

Haematopoietic stem cells expressing the CD34 surface marker have been posited as a niche for complex bacilli during latent tuberculosis infection. Our aim was to determine whether complex DNA is detectable in CD34-positive peripheral blood mononuclear cells (PBMCs) isolated from asymptomatic adults living in a setting with a high tuberculosis burden.

METHODS

We did a cross-sectional study in Ethiopia between Nov 22, 2017, and Jan 10, 2019. Digital PCR (dPCR) was used to determine whether complex DNA was detectable in PBMCs isolated from 100 mL blood taken from asymptomatic adults with HIV infection or a history of recent household or occupational exposure to an index case of human or bovine tuberculosis. Participants were recruited from HIV clinics, tuberculosis clinics, and cattle farms in and around Addis Ababa. A nested prospective study was done in a subset of HIV-infected individuals to evaluate whether administration of isoniazid preventive therapy was effective in clearing complex DNA from PBMCs. Follow-up was done between July 20, 2018, and Feb 13, 2019. QuantiFERON-TB Gold assays were also done on all baseline and follow-up samples.

FINDINGS

Valid dPCR data (ie, droplet counts >10 000 per well) were available for paired CD34-positive and CD34-negative PBMC fractions from 197 (70%) of 284 participants who contributed data to cross-sectional analyses. complex DNA was detected in PBMCs of 156 of 197 participants with valid dPCR data (79%, 95% CI 74-85). It was more commonly present in CD34-positive than in CD34-negative fractions (154 [73%] of 197 46 [23%] of 197; p<0·0001). Prevalence of dPCR-detected complex DNA did not differ between QuantiFERON-negative and QuantiFERON-positive participants (77 [78%] of 99 79 [81%] of 98; p=0·73), but it was higher in HIV-infected than in HIV-uninfected participants (67 [89%] of 75 89 [73%] of 122, p=0·0065). By contrast, the proportion of QuantiFERON-positive participants was lower in HIV-infected than in HIV-uninfected participants (25 [33%] of 75 73 [60%] of 122; p<0·0001). Administration of isoniazid preventive therapy reduced the prevalence of dPCR-detected complex DNA from 41 (95%) of 43 HIV-infected individuals at baseline to 23 (53%) of 43 after treatment (p<0·0001), but it did not affect the prevalence of QuantiFERON positivity (17 [40%] of 43 at baseline 13 [30%] of 43 after treatment; p=0·13).

INTERPRETATION

We report a novel molecular microbiological biomarker of latent tuberculosis infection with properties that are distinct from those of a commercial interferon-γ release assay. Our findings implicate the bone marrow as a niche for in latently infected individuals. Detection of complex DNA in PBMCs has potential applications in the diagnosis of latent tuberculosis infection, in monitoring response to preventive therapy, and as an outcome measure in clinical trials of interventions to prevent or treat latent tuberculosis infection.

FUNDING

UK Medical Research Council.

摘要

背景

在潜伏性结核感染期间，表达 CD34 表面标志物的造血干细胞被认为是复杂细菌的一个生态位。我们的目的是确定在来自高结核负担地区的无症状成年人的外周血单个核细胞（PBMC）中是否可以检测到复杂 DNA。

方法

我们在 2017 年 11 月 22 日至 2019 年 1 月 10 日期间在埃塞俄比亚进行了一项横断面研究。使用数字 PCR（dPCR）来确定从感染 HIV 或近期有家庭或职业接触人或牛结核的索引病例的无症状成年人中采集的 100 mL 血液中分离的 PBMC 中是否可以检测到复杂 DNA。参与者是从 HIV 诊所、结核诊所和亚的斯亚贝巴及其周边的奶牛场招募的。在一组 HIV 感染个体中进行了嵌套前瞻性研究，以评估异烟肼预防治疗是否有效清除 PBMC 中的复杂 DNA。随访时间为 2018 年 7 月 20 日至 2019 年 2 月 13 日。所有基线和随访样本均进行了 QuantiFERON-TB Gold 检测。

发现

来自 284 名参与者的数据可用于进行横断面分析，其中 197 名（70%）参与者的 CD34-阳性和 CD34-阴性 PBMC 分数具有有效的 dPCR 数据（即液滴计数>每孔 10000 个）。在 197 名具有有效 dPCR 数据的参与者中，有 156 名（79%，95%CI 74-85）检测到 PBMC 中的复杂 DNA。它在 CD34-阳性分数中比在 CD34-阴性分数中更常见（197 名参与者中的 154 名[73%]比 197 名参与者中的 46 名[23%]；p<0·0001）。dPCR 检测到的复杂 DNA 的患病率在 QuantiFERON 阴性和 QuantiFERON 阳性参与者之间没有差异（99 名参与者中的 77 名[78%]比 98 名参与者中的 79 名[81%]；p=0·73），但在 HIV 感染参与者中比在 HIV 未感染参与者中更高（75 名参与者中的 67 名[89%]比 122 名参与者中的 89 名[73%]；p=0·0065）。相比之下，HIV 感染参与者中 QuantiFERON 阳性参与者的比例低于 HIV 未感染参与者（75 名参与者中的 25 名[33%]比 122 名参与者中的 73 名[60%]；p<0·0001）。异烟肼预防治疗降低了从基线时 43 名 HIV 感染个体中的 41 名（95%）到治疗后 43 名中的 23 名（53%）的 dPCR 检测到的复杂 DNA 的患病率（p<0·0001），但它并没有影响 QuantiFERON 阳性的患病率（43 名参与者中的 17 名[40%]在基线时 43 名参与者中的 13 名[30%]在治疗后；p=0·13）。

解释

我们报告了潜伏性结核感染的一种新的分子微生物学生物标志物，其性质与商业干扰素-γ释放试验不同。我们的发现暗示骨髓是潜伏感染个体中细菌的一个生态位。在 PBMC 中检测到复杂 DNA 可能在潜伏性结核感染的诊断、监测预防性治疗的反应以及作为预防或治疗潜伏性结核感染的临床试验的结局指标方面具有应用潜力。