Chan Cheryl, Loh Joanne Xin Yi, Sin Wei-Xiang, Teo Denise Bei Lin, Tan Nicholas Kwan Zen, Nagarajan Chandramouli, Chen Yunxin, Lim Francesca Lorraine Wei Inng, Birnbaum Michael E, Williams Rohan B H, Springs Stacy L
Critical Analytics for Manufacturing Personalized-Medicine, Singapore-MIT Alliance for Research and Technology, Singapore 138602, Singapore.
Department of Haematology, Singapore General Hospital, Singapore 169608, Singapore.
Mol Ther Nucleic Acids. 2024 Dec 31;36(1):102444. doi: 10.1016/j.omtn.2024.102444. eCollection 2025 Mar 11.
Nucleic acid amplification tests (NAATs) have enabled fast and sensitive detection of virus infections but are unable to discriminate between live and dead/inert viral fragments or between latent and reactivated virus infections. Here, we show that extracellular viral microRNAs (viral exmiRs) are cell-free candidate biomarkers of live, latent, and reactivated virus infections, achieving fast (under 1 day) and sensitive (30 attomolar [aM]) detection by quantitative real-time reverse transcription PCR (real-time RT-qPCR). We report that spent-media-derived Epstein-Barr virus (EBV) miR-BART10-3p and herpes simplex virus 1 (HSV-1) miR-H5 are biomarkers of live EBV-2 and HSV-1 infection of T cell cultures, respectively. We identified extracellular human herpesvirus 6 (HHV-6) miR-Ro6-4 as a biomarker of endogenous latent HHV-6 in healthy human donor T cell cultures and identified human cytomegalovirus (HCMV) miR-US5-2-5p and miR-US22-5p as plasma biomarkers of endogenous latent HCMV infection. Viral exmiR profiling of spent media from EBV- and HHV-8-reactivated B cell models revealed specific signatures of elevated EBV miR-BHRF1-2-3p and HHV-8 miR-K12-10a-3p, miR-K12-10b, and miR-K12-12-3p, respectively, during virus reactivation. Our study thus suggests the utility of viral exmiR biomarkers in enabling NAAT-based detection of live, endogenous latent, and reactivated virus infections of cells.
核酸扩增检测（NAATs）能够快速、灵敏地检测病毒感染，但无法区分活病毒片段与死亡/无活性病毒片段，也无法区分潜伏性病毒感染和再激活病毒感染。在此，我们表明细胞外病毒微小RNA（病毒外泌体微小RNA）是活病毒、潜伏病毒和再激活病毒感染的无细胞候选生物标志物，通过定量实时逆转录PCR（实时RT-qPCR）可实现快速（1天内）且灵敏（30阿托摩尔[aM]）的检测。我们报告称，来自培养上清液的爱泼斯坦-巴尔病毒（EBV）miR-BART10-3p和单纯疱疹病毒1型（HSV-1）miR-H5分别是T细胞培养物中活EBV-2和HSV-1感染的生物标志物。我们鉴定出细胞外人类疱疹病毒6型（HHV-6）miR-Ro6-4是健康人类供体T细胞培养物中内源性潜伏HHV-6的生物标志物，并鉴定出人类巨细胞病毒（HCMV）miR-US5-2-5p和miR-US22-5p是内源性潜伏HCMV感染的血浆生物标志物。对EBV和HHV-8再激活B细胞模型的培养上清液进行病毒外泌体微小RNA分析发现，在病毒再激活过程中，EBV miR-BHRF1-2-3p以及HHV-8 miR-K12-10a-3p、miR-K12-10b和miR-K12-12-3p分别出现了特异性的升高特征。因此，我们的研究表明病毒外泌体微小RNA生物标志物有助于基于NAATs检测细胞的活病毒、内源性潜伏病毒和再激活病毒感染。
