Generation of a pdmH1N1 2018 Influenza A Reporter Virus Carrying a mCherry Fluorescent Protein in the PA Segment

Abstract

Influenza A virus (IAV) is a major human pathogen associated with significant morbidity and mortality worldwide. Through serial passage in mice, we generated a recombinant pdmH1N1 2009 IAV, A/Guangdong/GLW/2018 (GLW/18-MA), which encodes an mCherry gene fused to the C-terminal of a polymerase acidic (PA) segment and demonstrated comparable growth kinetics to the wild-type. Nine mutations were identified in the GLW/18-MA genome: PA (I61M, E351G, and G631S), NP (E292G), HA1 (T164I), HA2 (N117S and P160S), NA (W61R), and NEP (K44R). The recombinant IAV reporter expresses mCherry, a red fluorescent protein, at a high level and maintains its genetic integrity after five generations of serial passages in Madin-Darby Canine Kidney cells (MDCK) cells. Moreover, the imaging is noninvasive and permits the monitoring of infection in living mice. Treatment with oseltamivir or baicalin followed by infection with the reporter IAV led to a decrease in fluorescent protein signal in living mice. This result demonstrates that the IAV reporter virus is a powerful tool to study viral pathogenicity and transmission and to develop and evaluate novel anti-viral drugs, inhibitors, and vaccines in the future.

Generation and Characterization of an Influenza D Reporter Virus

“…They allow for targeted mutagenesis of the viral proteins for functional studies and the incorporation of reporter genes encoding fluorescent or bioluminescent proteins into the viral genome. In particular, fluorescent reporter viruses have been widely used as powerful tools for studying in vitro and in vivo dynamics of Influenza A virus (IAV) infections and evaluating treatment options [10][11][12][13][14]. Different strategies for introducing reporter genes into the segmented IAV genome have been established, including introducing eGFP into the shortest segment encoding the non-structural proteins NS1 and NS2 [10][11][12][13][14][15].…”

“…In particular, fluorescent reporter viruses have been widely used as powerful tools for studying in vitro and in vivo dynamics of Influenza A virus (IAV) infections and evaluating treatment options [10][11][12][13][14]. Different strategies for introducing reporter genes into the segmented IAV genome have been established, including introducing eGFP into the shortest segment encoding the non-structural proteins NS1 and NS2 [10][11][12][13][14][15]. During the virus replication cycle, the NS1 protein is translated directly from an unspliced mRNA transcript and functions as a suppressor of the innate immune system.…”

Unraveling Novel Strategies: Targeting Miz1 for Degradation to Enhance Antiviral Defense against Influenza A Virus

“…In 2023, Via Nova Therapeutics received clearance from the FDA for the investigational new drug application of VNT-101, a small molecule inhibitor targeting influenza A nucleoprotein [ 6 ]. However, the changing nature of viruses also leads to the emergence of drug-resistant strains, thereby making antiviral agents less effective [ 7 ]. Hence, there is an urgent need for better understanding the pathogenesis of IAV infection and developing novel therapies targeting aberrant signaling pathways.…”