PDF(Pubmed)
Abstract：
The COVID-19 pandemic highlighted the necessity of fast, sensitive, and efficient methods to test large populations for respiratory viruses. The \"gold standard\" molecular assays for detecting respiratory viruses, such as quantitative polymerase chain reaction (qPCR) and reverse transcription qPCR (RT-qPCR), rely on invasive swab samples and require time-consuming and labor-intensive extraction processes. Moreover, the turnaround time for RT-qPCR-based assays is too lengthy for rapid screening. Extraction-free saliva-based methods provide a non-invasive sampling process with a fast turnaround time and are suitable for high-throughput applications. However, when used with a standard RT-qPCR system, the absence of extraction significantly reduces the assays\' sensitivity. Here, using a novel optical modulation biosensing (OMB) platform, we developed a rapid and highly sensitive extraction-free saliva-based molecular assay. We blindly tested 364 paired nasopharyngeal swabs and saliva samples from suspected SARS-CoV-2 cases in Israel. Compared with the gold standard swab-based RT-qPCR assay, the sensitivity of the extraction-free saliva-based OMB assay is 90.7%, much higher than the sensitivity of extraction-free saliva-based RT-qPCR assay (77.8%) with similar specificity (95.3% and 97.6%, respectively). Moreover, out of 12 samples identified by the OMB-based assay as positive, 8 samples were collected from hospitalized patients in a COVID-19 ward and were verified to be SARS-CoV-2-positive upon admission, indicating that the actual clinical sensitivity and specificity of the OMB assay are higher. Considering its user-friendly saliva-based protocol, short and cost-effective extraction-free process, and high clinical accuracy, the OMB-based molecular assay is very suitable for high-throughput testing of large populations for respiratory viruses.
OBJECTIVE: Three years after the SARS-CoV-2 outbreak, there are no molecular tests that combine low-cost and straightforward sample preparation, effective sample handling, minimal reagent and disposable requirements, high sensitivity, and high throughput required for mass screening. Existing rapid molecular techniques typically sacrifice certain requirements to meet others. Yet, localized outbreaks of novel viral diseases happen daily in different parts of the world. In this context, respiratory diseases are of specific importance, as they are frequently airborne and highly contagious, with the potential for a rapid global spread. The widely accepted opinion is that another pandemic is just a question of time. To ensure that the containment efforts for the upcoming \"disease X\" are successful, introducing rapid, high-throughput, and highly sensitive diagnostic methods for detecting and identifying pathogens is critical. A few months into the pandemic, saliva was suggested as a diagnostic matrix for SARS-CoV-2 detection. The collection of saliva does not require swabs and is minimally invasive. In particular, extraction-free saliva-based assays require fewer reagents and disposables, and therefore are faster and cheaper, offering an appealing alternative for low-income countries. Unfortunately, current extraction-free saliva-based detection methods, such as direct RT-qPCR or isothermal amplification, have either low sensitivity or low throughput. Therefore, we believe that the presented highly sensitive ht-OMBi platform and the extraction-free saliva-based molecular assay can become an essential tool in the infectious disease monitoring toolbox.
OBJECTIVE: Three years after the SARS-CoV-2 outbreak, there are no molecular tests that combine low-cost and straightforward sample preparation, effective sample handling, minimal reagent and disposable requirements, high sensitivity, and high throughput required for mass screening. Existing rapid molecular techniques typically sacrifice certain requirements to meet others. Yet, localized outbreaks of novel viral diseases happen daily in different parts of the world. In this context, respiratory diseases are of specific importance, as they are frequently airborne and highly contagious, with the potential for a rapid global spread. The widely accepted opinion is that another pandemic is just a question of time. To ensure that the containment efforts for the upcoming \"disease X\" are successful, introducing rapid, high-throughput, and highly sensitive diagnostic methods for detecting and identifying pathogens is critical. A few months into the pandemic, saliva was suggested as a diagnostic matrix for SARS-CoV-2 detection. The collection of saliva does not require swabs and is minimally invasive. In particular, extraction-free saliva-based assays require fewer reagents and disposables, and therefore are faster and cheaper, offering an appealing alternative for low-income countries. Unfortunately, current extraction-free saliva-based detection methods, such as direct RT-qPCR or isothermal amplification, have either low sensitivity or low throughput. Therefore, we believe that the presented highly sensitive ht-OMBi platform and the extraction-free saliva-based molecular assay can become an essential tool in the infectious disease monitoring toolbox.
摘要：
COVID-19大流行强调了快速、敏感,以及检测大量呼吸道病毒的有效方法。检测呼吸道病毒的“黄金标准”分子检测，如定量聚合酶链反应（qPCR）和逆转录qPCR（RT-qPCR），依赖于侵入性拭子样品，需要耗时和劳动密集型的提取过程。此外，基于RT-qPCR的分析的周转时间对于快速筛选来说太长。基于无提取唾液的方法提供了具有快速周转时间的非侵入性取样过程,并且适合于高通量应用。然而,当与标准RT-qPCR系统一起使用时，没有提取显著降低了测定的灵敏度。这里,使用一种新颖的光调制生物传感（OMB）平台，我们开发了一种快速且高度灵敏的无唾液提取分子检测方法.我们盲目地测试了364对来自以色列疑似SARS-CoV-2病例的鼻咽拭子和唾液样本。与基于金标准拭子的RT-qPCR检测相比，基于无提取唾液的OMB测定的灵敏度为90.7%，比基于无唾液提取的RT-qPCR检测的灵敏度（77.8%）高得多，具有相似的特异性（95.3%和97.6%，分别)。此外，在通过基于OMB的检测鉴定为阳性的12个样本中，从COVID-19病房的住院患者中收集了8个样本，在入院时被证实为SARS-CoV-2阳性，表明OMB检测的实际临床敏感性和特异性更高。考虑到其基于唾液的用户友好协议，短且具有成本效益的无提取过程，和高临床准确性，基于OMB的分子检测方法非常适合对大量呼吸道病毒进行高通量检测.
目标：SARS-CoV-2爆发三年后，没有结合低成本和简单样品制备的分子测试，有效的样品处理，最少的试剂和一次性要求，高灵敏度,和大规模筛选所需的高通量。现有的快速分子技术通常牺牲某些要求来满足其他要求。然而，新型病毒性疾病的局部爆发每天都在世界不同地区发生。在这种情况下，呼吸系统疾病特别重要，因为它们经常通过空气传播并且具有高度传染性，具有快速全球传播的潜力。广泛接受的意见是，另一场大流行只是时间问题。为了确保对即将到来的“疾病X”的遏制努力取得成功，引入快速，高通量，和高度敏感的诊断方法来检测和识别病原体是至关重要的。大流行几个月后，唾液被建议作为SARS-CoV-2检测的诊断基质。唾液的收集不需要拭子并且是微创的。特别是,基于唾液的无提取化验需要更少的试剂和一次性用品，因此更快，更便宜，为低收入国家提供了一个有吸引力的替代方案。不幸的是,当前基于无唾液提取的检测方法，如直接RT-qPCR或等温扩增，具有低灵敏度或低吞吐量。因此,我们相信，提出的高度敏感的ht-OMBi平台和无提取的基于唾液的分子检测可以成为传染病监测工具箱中的重要工具。
目标：SARS-CoV-2爆发三年后，没有结合低成本和简单样品制备的分子测试，有效的样品处理，最少的试剂和一次性要求，高灵敏度,和大规模筛选所需的高通量。现有的快速分子技术通常牺牲某些要求来满足其他要求。然而，新型病毒性疾病的局部爆发每天都在世界不同地区发生。在这种情况下，呼吸系统疾病特别重要，因为它们经常通过空气传播并且具有高度传染性，具有快速全球传播的潜力。广泛接受的意见是，另一场大流行只是时间问题。为了确保对即将到来的“疾病X”的遏制努力取得成功，引入快速，高通量，和高度敏感的诊断方法来检测和识别病原体是至关重要的。大流行几个月后，唾液被建议作为SARS-CoV-2检测的诊断基质。唾液的收集不需要拭子并且是微创的。特别是,基于唾液的无提取化验需要更少的试剂和一次性用品，因此更快，更便宜，为低收入国家提供了一个有吸引力的替代方案。不幸的是,当前基于无唾液提取的检测方法，如直接RT-qPCR或等温扩增，具有低灵敏度或低吞吐量。因此,我们相信，提出的高度敏感的ht-OMBi平台和无提取的基于唾液的分子检测可以成为传染病监测工具箱中的重要工具。
