Given the implications of increased transmissibility, virulence, host range, and immune escapes of emerging variants of SARS-CoV-2, developing in vitro models that allow for detection of variants and differences in infection dynamics is important. The objective of this study, was to evaluate the PrimeFlow RNA in-situ assay as a method of detection for multiple strains of SARS-CoV-2. Evaluation of detection and infection statuses included single infections with an Alpha, Delta, or Omicron variants and dual infections with Alpha/Omicron or Delta/Omicron. RNA probes specific for the Spike protein coding region, were designed (omicron or non-omicron specific). SARS-CoV-2 RNA was detected in greater frequency in the Vero E6 and minimally in the fetal deer testicle cell lines by flow cytometry using this approach for viral detection of multiple variants. Most evident in the Vero E6 cells, 24 h post infection both Alpha and Delta predominated over Omicron in dual infections. This is the first report using the PrimeFlow assay for the detection of SARS-CoV-2 at the single-cell level and as a potential model for competition of variants utilizing infection dynamics in cell culture.

考虑到 SARS-CoV-2 新兴变种的传播性、毒力、宿主范围和免疫逃逸增加的影响，开发能够检测变种和感染动态差异的体外模型非常重要。本研究的目的是评估 PrimeFlow RNA 原位测定作为多种 SARS-CoV-2 毒株的检测方法。检测和感染状态的评估包括 Alpha、Delta 或 Omicron 变体的单一感染以及 Alpha/Omicron 或 Delta/Omicron 的双重感染。设计了刺突蛋白编码区特异性的RNA探针（omicron或非omicron特异性）。通过流式细胞术使用这种方法检测多种变体的病毒，在 Vero E6 中检测到 SARS-CoV-2 RNA 的频率较高，在鹿胎睾丸细胞系中检测到的频率最低。在 Vero E6 细胞中最为明显，感染后 24 小时，在双重感染中 Alpha 和 Delta 均优于 Omicron。这是第一份使用 PrimeFlow 检测在单细胞水平检测 SARS-CoV-2 的报告，并作为利用细胞培养中的感染动力学进行变异竞争的潜在模型。