SARS-CoV-2 Reverse Genetics Reveals a Variable Infection Gradient in the Respiratory Tract

Hou, Yixuan J.; Okuda, Kenichi; Edwards, Caitlin E.; Martinez, David R.; Asakura, Takanori; Dinnon, Kenneth H.; Kato, Takafumi; Re, Lee; Yount, Boyd; Mascenik, Teresa; Chen, Gang; Olivier, Kenneth N.; Ghio, Andrew J.; Tse, Longping V.; Leist, Sarah R.; Gralinski, Lisa E.; Schäfer, Alexandra; Dang, Hong; Gilmore, Rodney C.; Nakano, Satoko; Sun, Ling; Fulcher, M. Leslie; Livraghi‐Butrico, Alessandra; Nicely, Nathan I.; Cameron, Mark J.; Cameron, Cheryl M.; Kelvin, David J.; Silva, Aravinda M. de; Margolis, David M.; Markmann, Alena J.; Bartelt, Luther A.; Zumwalt, Ross E.; Martínez, Fernando J.; Salvatore, Steven; Borczuk, Alain C.; Tata, Purushothama Rao; Sontake, Vishwaraj; Kimple, Adam J.; Jaspers, Ilona; O’Neal, Wanda K.; Randell, Scott H.; Boucher, Richard C.; Baric, Ralph S.

doi:10.1016/j.cell.2020.05.042

Cited by 1,404 publications

(1,848 citation statements)

References 104 publications

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“…SARS-CoV-2 receptors and coreceptors have been shown to be highly expressed in nasal epithelial cells (Sungnak et al, 2020). This finding is consistent with the virus infectivity or replication pattern along the respiratory tract, which peaks proximally (nasal cavity) and is relatively minimal in the distal alveolar regions (Hou et al, 2020). These findings suggest that nasal carriage of the virus could be a key feature of transmission, and that nasally administered therapeutic modalities could be potentially effective in helping to prevent spread of infection.…”

Section: Introductionsupporting

confidence: 76%

Virucidal and antiviral activity of astodrimer sodium against SARS-CoV-2 in vitro

Paull

Heery

Bobardt

et al. 2020

Preprint

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An effective response to the ongoing coronavirus disease (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) will require a range of complementary preventive modalities. The current studies were conducted to evaluate the in vitro SARS-CoV-2 antiviral activity of astodrimer sodium, a dendrimer with broad spectrum antimicrobial activity, including against enveloped viruses in in vitro and in vivo models, that is marketed for antiviral and antibacterial applications. We report that astodrimer sodium inhibits replication of SARS-CoV-2 in Vero E6 cells when added to cells 1-hour prior to or 1-hour post infection, with 50% effective concentrations reducing virus-induced cytopathic effect (EC50) ranging from 0.090 to 0.742 μM (0.002 to 0.012 mg/mL). The selectivity index (SI) in these assays was as high as 2197. Astodrimer sodium was also effective in a virucidal evaluation when mixed with virus for 1 hour prior to infection of cells (EC50 1.83 μM [0.030 mg/mL]). Results from a time of addition study, which showed infectious virus was below the lower limit of detection at all time points tested, were consistent with the compound inhibiting early virus entry steps. The data were similar for all investigations and were consistent with the potent antiviral activity of astodrimer sodium being due to inhibition of virus-host cell interactions, as previously demonstrated for other viruses. Further studies will confirm if astodrimer sodium binds to SARS-CoV-2 spike protein and physically blocks initial association of the virus with heparan sulfate proteoglycans on the host cell. Given the in vitro effectiveness and significantly high SI, astodrimer sodium warrants further investigation for potential as a nasally administered or inhaled antiviral agent for SARS-CoV-2 prevention and treatment applications.

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Section: Introductionsupporting

confidence: 76%

Virucidal and antiviral activity of astodrimer sodium against SARS-CoV-2 in vitro

Paull

Heery

Bobardt

et al. 2020

Preprint

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“…The higher viral loads of G614 in the upper airway of COVID-19 patients 26 and infected hamsters support the role of D614G mutation in viral transmissibility. The robust replication of SARS-CoV-2 in the human upper airway may be partially conferred by a higher ACE2 receptor expression in the nasal cavity compared to that in the lower respiratory tract 27,28 . Patients infected with G614 virus developed higher levels of viral RNA in the nasopharyngeal swabs than those infected with D614 virus, but disease severity is not associated with the D614G mutation 6,7,9 .…”

Section: Discussionmentioning

confidence: 99%

Spike mutation D614G alters SARS-CoV-2 fitness and neutralization susceptibility

Shi

Plante

Liu

et al. 2020

Preprint

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A spike protein mutation D614G became dominant in SARS-CoV-2 during the COVID-19 pandemic. However, the mutational impact on viral spread and vaccine efficacy remains to be defined. Here we engineer the D614G mutation in the SARS-CoV-2 USA-WA1/2020 strain and characterize its effect on viral replication, pathogenesis, and antibody neutralization. The D614G mutation significantly enhances SARS-CoV-2 replication on human lung epithelial cells and primary human airway tissues, through an improved infectivity of virions with the spike receptor-binding domain in an “up” conformation for binding to ACE2 receptor. Hamsters infected with D614 or G614 variants developed similar levels of weight loss. However, the G614 virus produced higher infectious titers in the nasal washes and trachea, but not lungs, than the D614 virus. The hamster results confirm clinical evidence that the D614G mutation enhances viral loads in the upper respiratory tract of COVID-19 patients and may increases transmission. For antibody neutralization, sera from D614 virus-infected hamsters consistently exhibit higher neutralization titers against G614 virus than those against D614 virus, indicating that (i) the mutation may not reduce the ability of vaccines in clinical trials to protect against COVID-19 and (ii) therapeutic antibodies should be tested against the circulating G614 virus before clinical development.

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“…A recent study has identified the ability of SARS-CoV-2 to replicate to higher titers in the upper respiratory tract, including nasal cells (27). Hou et al .…”

Section: Discussionmentioning

confidence: 99%

“…Hou et al . have shown that high levels of virus replication in nasal cells is associated with high levels of angiotensin-converting enzyme 2 (ACE2) receptor expression in these cells, relative to cells in the lower respiratory tract (27). Studies have also shown that rhinovirus (common cold virus) replicates to higher titers in nasal cells due to diminished temperature-dependent innate antiviral responses in these cells (28).…”

Section: Discussionmentioning

confidence: 99%

Experimental and natural evidence of SARS-CoV-2 infection-induced activation of type I interferon responses

Banerjee

El-Sayes

Budylowski

et al. 2020

Preprint

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Two highly pathogenic human coronaviruses that cause severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS) have evolved proteins that can inhibit host antiviral responses, likely contributing to disease progression and high case-fatality rates. SARS-CoV-2 emerged in December 2019 resulting in a global pandemic. Recent studies have shown that SARS-CoV-2 is unable to induce a robust type I interferon (IFN) response in human cells, leading to speculation about the ability of SARS-CoV-2 to inhibit innate antiviral responses. However, innate antiviral responses are dynamic in nature and gene expression levels rapidly change within minutes to hours. In this study, we have performed a time series RNA-seq and selective immunoblot analysis of SARS-CoV-2 infected lung (Calu-3) cells to characterize early virus-host processes. SARS-CoV-2 infection upregulated transcripts for type I IFNs and interferon stimulated genes (ISGs) after 12 hours. Furthermore, we analyzed the ability of SARS-CoV-2 to inhibit type I IFN production and downstream antiviral signaling in human cells. Using exogenous stimuli, we discovered that SARS-CoV-2 is unable to modulate IFNβ production and downstream expression of ISGs, such as IRF7 and IFIT1. Thus, data from our study indicate that SARS-CoV-2 may have evolved additional mechanisms, such as masking of viral nucleic acid sensing by host cells to mount a dampened innate antiviral response. Further studies are required to fully identify the range of immune-modulatory strategies of SARS-CoV-2.SignificanceHighly pathogenic coronaviruses that cause SARS and MERS have evolved proteins to shutdown antiviral responses. The emergence and rapid spread of SARS-CoV-2, along with its relatively low case-fatality rate have led to speculation about its ability to modulate antiviral responses. We show that SARS-CoV-2 is unable to block antiviral responses that are mounted by exogenous stimuli. Data from our study provide promising support for the use of recombinant type I IFN as combination therapy to treat COVID-19 patients. Furthermore, our data also suggest that the inability of SARS-CoV-2 to efficiently modulate antiviral responses may be associated with its low case-fatality rate compared to other pathogenic CoVs that cause SARS and MERS.

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SARS-CoV-2 Reverse Genetics Reveals a Variable Infection Gradient in the Respiratory Tract

Cited by 1,404 publications

References 104 publications

Virucidal and antiviral activity of astodrimer sodium against SARS-CoV-2 in vitro

Virucidal and antiviral activity of astodrimer sodium against SARS-CoV-2 in vitro

Spike mutation D614G alters SARS-CoV-2 fitness and neutralization susceptibility

Experimental and natural evidence of SARS-CoV-2 infection-induced activation of type I interferon responses

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