Quantitative measurement of infectious virus in SARS-CoV-2 Alpha, Delta and Epsilon variants reveals higher infectivity (viral titer:RNA ratio) in clinical samples containing the Delta and Epsilon variants

Abstract

Background Novel SARS-CoV-2 Variants of Concern (VoC) pose a challenge to controlling the COVID-19 pandemic. Previous studies indicate that clinical samples collected from individuals infected with the Delta variant may contain higher levels of RNA than previous variants, but the relationship between viral RNA and infectious virus for individual variants is unknown. Methods We measured infectious viral titer (using a micro-focus forming assay) as well as total and subgenomic viral RNA levels (using RT-PCR) in a set of 165 clinical samples containing SARS-CoV-2 Alpha, Delta and Epsilon variants that were processed within two days of collection from the patient. Results We observed a high degree of variation in the relationship between viral titers and RNA levels. Despite the variability we observed for individual samples the overall infectivity differed among the three variants. Both Delta and Epsilon had significantly higher infectivity than Alpha, as measured by the number of infectious units per quantity of viral E gene RNA (6 and 4 times as much, p=0.0002 and 0.009 respectively) or subgenomic E RNA (11 and 7 times as much, p<0.0001 and 0.006 respectively). Conclusion In addition to higher viral RNA levels reported for the Delta variant, the infectivity (amount of replication competent virus per viral genome copy) may also be increased compared to Alpha. Measuring the relationship between live virus and viral RNA is an important step in assessing the infectivity of novel SARS-CoV-2 variants. An increase in the infectivity of the Delta variant may further explain increased spread and suggests a need for increased measures to prevent viral transmission.

Comparative Sensitivity of Rapid Antigen Tests for the Delta Variant (B.1.617.2) of SARS-CoV-2

“…These four amino acid substitutions might affect the sensitivity of the four RATs. Alternatively, the reduced sensitivity might be caused by a reduction in the particle/PFU ratio of the delta variant [20] because delta variants harboring L452R, E484Q, and P681R in the S protein efficiently infect cells due to the high affinity of the S protein for ACE2 and their high fusion efficiency [21,22]. This means that test specimens prepared at a certain PFU might contain fewer virus particles of the delta variant than of the S-614D virus.…”

The Global Epidemic of the SARS-CoV-2 Delta Variant, Key Spike Mutations and Immune Escape

“…At present, Alpha B.1.1.7 (known as 20I/501Y.V1, VOC 202012/01) (9), Beta B.1.351 (known as 501Y.V2) (10), Gamma P.1 (known as 501Y.V3) (11) and Delta B.1.617.2 (known as 478K.V1) (12) are defined as "variants of concern (VOCs)" by the WHO. Several studies have indicated that the Delta variant has higher transmissibility (13)(14)(15) and immune evasion than the early original virus strain and the other three VOCs. COVID-19 patients infected with Delta have a higher risk of hospitalization, ICU admission, and mortality (16)(17)(18).…”

Genomic and Virological Characterization of SARS-CoV-2 Variants in a Subset of Unvaccinated and Vaccinated U.S. Military Personnel

“…This is likely due to the cumbersome nature of the assays used to quantify levels of infectious virus coupled with the requirement to perform these assays in a BSL-3 laboratory. However, a recent study found a significant amount of variability in the relationship between viral titers for infectious virus and viral RNA levels (13).…”

“…While some studies have described the amount of viable virus shed by individuals infected with the various SARS-CoV-2 variants (12)(13)(14), more studies are needed to better characterize infectious virus levels in vaccinated and unvaccinated individuals. This is especially important because each study uses different methods and focuses on different populations of individuals from different geographic locations.…”