Department of Global Health and Social Medicine, Harvard Medical School, Boston, USA.
Socios En Salud Sucursal (Partners In Health), Lima, Peru.
BMC Infect Dis. 2019 Jun 27;19(1):563. doi: 10.1186/s12879-019-4188-8.
Rapid and accurate diagnosis of childhood tuberculosis (TB) is challenging because children are often unable to produce the sputum sample required for conventional tests. Stool is an alternative sample type that is easy to collect from children, and studies investigating the use of stool for molecular detection of Mycobacterium tuberculosis (Mtb) have led to promising results. Our objective was to evaluate stool as an alternative specimen to sputum for Mtb detection in children. We did so using the TruTip workstation (Akonni Biosystems), a novel automated lysis and extraction platform.
We tested stool samples from 259 children aged 0-14 years old, in Lima, Peru who presented with TB symptoms. Following extraction with TruTip, we detected the presence of Mtb DNA by IS6110 real-time PCR. We calculated assay sensitivity in two groups: (1) children with culture confirmed TB (N = 22); and (2) children with clinically-diagnosed unconfirmed TB (N = 84). We calculated specificity among children in whom TB was ruled out (N = 153). Among children who were diagnosed with TB, we examined factors associated with a positive stool test.
Assay sensitivity was 59% (95% confidence interval [CI]: 39-80%) and 1.2% (95% CI: 0.0-6.5%) in children with culture-confirmed and clinically-diagnosed unconfirmed TB, respectively, and specificity was 97% (95% CI: 93-99%). The assay detected Mtb in stool of 7/7 children with smear-positive TB (100% sensitivity; 95% CI: 59-100%), and in 6/15 of children with smear-negative, culture-confirmed TB (40% sensitivity; 95% CI: 16-68%). Older age, smear positivity, culture positivity, ability to produce sputum and cavitary disease were associated with a positive stool result.
Testing of stool samples with the TruTip workstation and IS6110 amplification yielded sensitivity and specificity estimates comparable to other tests such as Xpert. Future work should include detection of resistance using the TruTip closed amplification system and assay optimization to improve sensitivity in children with low bacillary loads.
由于儿童通常无法提供常规检测所需的痰样本，因此快速准确地诊断儿童结核病 (TB) 具有挑战性。粪便样本是一种替代样本类型，易于从儿童中采集，并且研究表明使用粪便进行分枝杆菌结核 (Mtb) 的分子检测具有很大的前景。我们的目的是评估粪便作为儿童 Mtb 检测的替代痰样本。我们使用新型自动化裂解和提取平台 TruTip 工作站来完成这项工作。
我们对来自秘鲁利马的 259 名 0-14 岁出现 TB 症状的儿童的粪便样本进行了检测。TruTip 提取后，我们通过 IS6110 实时 PCR 检测 Mtb DNA 的存在。我们在两个组中计算了检测方法的灵敏度：(1) 培养证实的 TB 患儿（N=22）；(2) 临床诊断为未经证实的 TB 患儿（N=84）。我们在排除 TB 的患儿（N=153）中计算了特异性。在诊断为 TB 的患儿中，我们研究了与粪便检测阳性相关的因素。
培养证实的和临床诊断的未经证实的 TB 患儿中，检测方法的灵敏度分别为 59%（95%CI：39-80%）和 1.2%（95%CI：0.0-6.5%），特异性分别为 97%（95%CI：93-99%）。该检测方法在 7/7 名痰涂片阳性 TB 患儿的粪便中检测到 Mtb（100%灵敏度；95%CI：59-100%），在 15 名痰涂片阴性、培养阳性的 TB 患儿中检测到 6 例（40%灵敏度；95%CI：16-68%）。年龄较大、痰涂片阳性、培养阳性、能产生痰和空洞性疾病与粪便检测结果呈阳性相关。
TruTip 工作站和 IS6110 扩增检测粪便样本的灵敏度和特异性与 Xpert 等其他检测方法相当。未来的工作应包括使用 TruTip 封闭扩增系统检测耐药性，并优化检测方法以提高低菌载量儿童的灵敏度。
