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Development and characterization of mouse-adapted recombinant SARS-CoV-2 expressing reporter genes
Preprint   Open access

Development and characterization of mouse-adapted recombinant SARS-CoV-2 expressing reporter genes

Sara Mahmoud, Nathaniel Jackson, Ramya Barre, Ma Yao, Mahmoud Bayoumi, Esteban Castro, Shahrzad Ezzatpour, Richard Plemper, Stanley Perlman, Chengjin Ye, …
bioRxiv
Cold Spring Harbor Laboratory Press
02/16/2026
DOI: 10.64898/2026.02.04.703885
PMID: 41757011
url
https://doi.org/10.64898/2026.02.04.703885View
Preprint (Author's original)This preprint has not been evaluated by subject experts through peer review. Preprints may undergo extensive changes and/or become peer-reviewed journal articles. Open Access

Abstract

Transgenic K18-hACE2 mice are a standard model for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), albeit with limitations. A mouse-adapted 30 (MA30) SARS-CoV-2 has been developed to allow infection of wild-type (WT) mice strains. However, SARS-CoV-2 MA30 cannot be easily tracked in vitro, ex vivo, or in vivo. To address the problem, we developed a recombinant (r)SARS-CoV-2 based on the MA30 strain expressing fluorescent (mCherry) and luciferase (nanoluciferase, Nluc) reporter genes, alone or in combination, that enable tracking of viral infection in WT C57BL/6 and BALB/c mice. Insertion of the reporter genes resulted in minor viral attenuation in vitro, with ~0.5-1.0-log lower titers than rSARS-CoV-2 MA30 WT in A549 hACE2 cells, while maintain similar plaque morphology and replication kinetics in Vero AT cells. In vivo, reporter-expressing rSARS-CoV-2 MA30 caused transient weight loss, contrasting with lethal rSARS-CoV-2 MA30 WT infection. Bioluminescence imaging of rSARS-CoV-2 MA30 Nluc in C57BL/6 and BALB/c mice revealed peak pulmonary replication at 2 days post-infection, with resolution by day 4, and correlated with tissue viral loads. Our results demonstrate the feasibility of using rSARS-CoV-2 MA30 expressing reporter genes to track viral infection in vitro, ex vivo, and in vivo without a need for secondary approaches to monitor viral infection as are required for rSARS-CoV-2 MA30 WT. Our system is highly suitable to evaluate prophylactic vaccines and therapeutic antibodies or antiviral approaches in WT or transgenic C57BL/6 and BALB/c mice without the shortcomings of K18-hACE2 mice and with the added advantage of non-invasive monitoring of treatment efficacy.Competing Interest StatementThe authors have declared no competing interest.
 Bioluminescence Viral Infections Coronaviruses Replication Severe acute respiratory syndrome coronavirus 2 Transgenic mice

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