Objectives

The development of a rapid diagnostic test for viable SARS-CoV-2 is important for infection control. Real-time RT-PCR assays detect non-viable virus, and cell culture differentiates viable virus but it takes several weeks and is labour-intensive. Subgenomic RNAs may reflect replication-competent virus. We therefore evaluated the usefulness of subgenomic RNAs for diagnosing viable SARS-CoV-2 in patients with COVID-19.

Methods

Patients with various severities of confirmed COVID-19 were enrolled at a tertiary hospital between February and December 2020. RT-PCR assay results for genomic and subgenomic RNA of SARS-CoV-2 from nasopharyngeal swab, sputum and saliva specimens were compared with cell culture results.

Results

A total 189 specimens from 20 COVID-19 patients were tested in genomic and subgenomic PCR assays and cultured on Vero cells. Of these 189 samples, 62 (33%) gave positive culture results, 93 (49%) negative results and the remaining 34 (18%) indeterminate results. Compared with cell culture results, the sensitivities of genomic RNA and subgenomic RNA of the N and S genes were comparable at 100%, but the specificity of subgenomic RNA (N, 65% and S, 68%) was higher than that of genomic RNA (N, 23% and S, 17%, p < 0.001). The mean durations of positive culture and subgenomic RNA were 11.39 ± 10.34 and 13.75 ± 11.22 days after symptom onset (p 0.437), respectively, while that of genomic RNA was 22.85 ± 11.83 days after symptom onset (p < 0.001).

Discussion

Our comparison of subgenomic RNA detection with symptom duration and SARS-CoV-2 culture positivity provides a significant advancement on the transmissibility-based approach beyond the detection of SARS-CoV-2 genomic RNA, and warrants further studies on the development of better diagnostic strategy.

中文翻译：

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在 COVID-19 患者中检测活 SARS-CoV-2 的亚基因组 RNA 的诊断效用

目标

开发活 SARS-CoV-2 的快速诊断测试对于感染控制很重要。实时 RT-PCR 检测可检测非活病毒，细胞培养可区分活病毒，但需要数周时间且劳动密集型。亚基因组 RNA 可能反映具有复制能力的病毒。因此，我们评估了亚基因组 RNA 在 COVID-19 患者中诊断可行的 SARS-CoV-2 的有用性。

方法

2020 年 2 月至 2020 年 12 月期间，在一家三级医院招募了确诊为 COVID-19 的各种严重程度的患者。 将鼻咽拭子、痰液和唾液标本中 SARS-CoV-2 的基因组和亚基因组 RNA 的 RT-PCR 检测结果与细胞培养进行了比较结果。

结果

对来自 20 名 COVID-19 患者的总共 189 份样本进行了基因组和亚基因组 PCR 检测，并在 Vero 细胞上培养。在这 189 个样本中，62 个 (33%) 的培养结果呈阳性，93 个 (49%) 呈阴性，其余 34 个 (18%) 的结果不确定。与细胞培养结果相比，N和S基因的基因组RNA和亚基因组RNA的敏感性在100%相当，但亚基因组RNA（N，65%和S，68%）的特异性高于基因组RNA (N, 23% 和 S, 17%, p < 0.001)。阳性培养和亚基因组 RNA 的平均持续时间分别为症状发作后 11.39 ± 10.34 和 13.75 ± 11.22 天（p = 0.437），而基因组 RNA 的平均持续时间为症状发作后 22.85 ± 11.83 天（p < 0.001）。

讨论

我们将亚基因组 RNA 检测与症状持续时间和 SARS-CoV-2 培养阳性进行比较，为基于传染性的方法提供了超越 SARS-CoV-2 基因组 RNA 检测的重大进展，并需要进一步研究开发更好的诊断策略.