Visualizing real-time influenza virus infection, transmission and protection in ferrets

Karlsson, Erik A.; Meliopoulos, Victoria A.; Savage, Chandra; Livingston, Brandi; Mehle, Andrew; Schultz‐Cherry, Stacey

doi:10.1038/ncomms7378

Cited by 110 publications

(142 citation statements)

References 34 publications

Supporting

Mentioning

136

Contrasting

Order By: Relevance

“…The A/Puerto Rico/8/1934 influenza virus lacking the NS1 gene (PR8-⌬NS1) was provided by Adolfo Garcia-Sastre (Icahn School of Medicine, Mount Sinai) (21) and propagated in eggs, and virus titer in MDCK cells was determined by microneutralization assay as described previously (22). Vesicular stomatitis virus (VSV) was a kind gift from Michael Whitt, University of Tennessee Health Science Center, Memphis, TN.…”

Section: Cellsmentioning

confidence: 99%

Type I Interferon Response Limits Astrovirus Replication and Protects against Increased Barrier Permeability In Vitro and In Vivo

Marvin

Huerta

Sharp

et al. 2016

J Virol

Self Cite

View full text Add to dashboard Cite

Little is known about intrinsic epithelial cell responses against astrovirus infection. Here we show that human astrovirus type 1 (HAstV-1) infection induces type I interferon (beta interferon [IFN-␤]) production in differentiated Caco2 cells, which not only inhibits viral replication by blocking positive-strand viral RNA and capsid protein synthesis but also protects against HAstV-1-increased barrier permeability. Excitingly, we found similar results in vivo using a murine astrovirus (MuAstV) model, providing new evidence that virus-induced type I IFNs may protect against astrovirus replication and pathogenesis in vivo. IMPORTANCEHuman astroviruses are a major cause of pediatric diarrhea, yet little is known about the immune response. Here we show that type I interferon limits astrovirus infection and preserves barrier permeability both in vitro and in vivo. Importantly, we characterized a new mouse model for studying astrovirus replication and pathogenesis. The successful replication and spread of many enteric viruses depend upon modulating immune factors produced by intestinal epithelial cells (IECs) including interferons (IFNs) (1, 2). For instance, enteric adenoviruses are sensitive to IEC-produced type I IFNs, unlike respiratory adenoviruses (3), while rotavirus exploits type I IFN signaling in IECs to promote early viral replication (4). However, nothing is known about the impact of IFN on astrovirus infection.Astroviruses are small, nonenveloped, RNA viruses that are one of the most important causes of pediatric acute gastroenteritis worldwide (5-8). Infection begins by binding to an unidentified receptor(s) on epithelial cells after fecal-oral transmission followed by entry via endosomes (9). After viral uncoating, the positive-sense, single-stranded RNA genome is translated into a polyprotein precursor that is subsequently cleaved into proteins required for replication and the assembly of progeny virions. The genome contains three open reading frames: ORF1a, ORF1b, and ORF2. ORF1a and ORF1b encode nonstructural proteins involved in transcription and replication of the virus, while ORF2 encodes the capsid protein (10, 11). Negative-strand RNA is produced from the positive genomic strand, which can be detected 6 to 12 h postinfection (hpi) (12). Transcription of the negativestrand genome yields the genomic and subgenomic RNA. Human astrovirus (HAstV) proteins have been associated with membranes in infected cells likely serving as the site for replication and assembly (13-15). After assembly, the progeny virions egress from the cell, a process promoted by caspase activation (16).Recently, Guix et al. found that HAstV type 4 (HAstV-4) replication induces type I IFN production and that pretreatment of Caco2 cells with beta interferon (IFN-␤) reduced HAstV-4 capsid protein synthesis and progeny virion production (17). However, whether the IFN-␤ produced during astrovirus infection is sufficient to limit astrovirus replication, and at what step in the viral life cycle IFN-␤ affects astrovirus, remains...

show abstract

Section: Cellsmentioning

confidence: 99%

Type I Interferon Response Limits Astrovirus Replication and Protects against Increased Barrier Permeability In Vitro and In Vivo

Marvin

Huerta

Sharp

et al. 2016

J Virol

Self Cite

View full text Add to dashboard Cite

show abstract

“…Conversely, low-level detection of virus in contact ferrets in the absence of seroconversion can often be attributed to direct contact with virus-containing fomites that did not lead to productive infection in the animal and is not generally considered a true transmission event (64). Transmission studies utilizing bioluminescent influenza viruses represent an additional variable, as a transmission event can be identified by the presence of virus by in vivo imaging in the absence of infectious virus detection in nasal wash samples (69). Statistical modeling has shown that the predictive power of ferret transmission experiments can change based on the criteria used to assess transmission events, underscoring the need to be precise when describing this property (61).…”

Section: Transmission Modelsmentioning

confidence: 99%

Complexities in Ferret Influenza Virus Pathogenesis and Transmission Models

Belser

Eckert

Tumpey

et al. 2016

Microbiol Mol Biol Rev

View full text Add to dashboard Cite

SUMMARYFerrets are widely employed to study the pathogenicity, transmissibility, and tropism of influenza viruses. However, inherent variations in inoculation methods, sampling schemes, and experimental designs are often overlooked when contextualizing or aggregating data between laboratories, leading to potential confusion or misinterpretation of results. Here, we provide a comprehensive overview of parameters to consider when planning an experiment using ferrets, collecting data from the experiment, and placing results in context with previously performed studies. This review offers information that is of particular importance for researchers in the field who rely on ferret data but do not perform the experiments themselves. Furthermore, this review highlights the breadth of experimental designs and techniques currently available to study influenza viruses in this model, underscoring the wide heterogeneity of protocols currently used for ferret studies while demonstrating the wealth of information which can benefit risk assessments of emerging influenza viruses.

show abstract

“…Research into IAV has been greatly enhanced by several replication-competent reporter viruses (5-17). These reporter viruses have been crucial for identifying host factors, understanding basic viral pathogenesis, screening for antiviral compounds, and characterizing broadly reactive antibodies (6)(7)(8)(9)(10)(12)(13)(14)(15)(16)(17)(18)(19). Lack of viruses encoding reporters has delayed similar progress for IBV.…”

mentioning

confidence: 99%

“…Despite the importance of this virus, far less is known about IBV relative to influenza A virus (IAV). Research into IAV has been greatly enhanced by several replication-competent reporter viruses (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). These reporter viruses have been crucial for identifying host factors, understanding basic viral pathogenesis, screening for antiviral compounds, and characterizing broadly reactive antibodies (6)(7)(8)(9)(10)(12)(13)(14)(15)(16)(17)(18)(19).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Replication-Competent Influenza B Reporter Viruses as Tools for Screening Antivirals and Antibodies

Fulton

Palese

Heaton

2015

J Virol

View full text Add to dashboard Cite

cInfluenza B virus is a human pathogen responsible for significant health and economic burden. Research into this pathogen has been limited by the lack of reporter viruses. Here we describe the development of both a replication-competent fluorescent influenza B reporter virus and bioluminescent influenza B reporter virus. Furthermore, we demonstrate these reporter viruses can be used to quickly monitor viral growth and permit the rapid screening of antiviral compounds and neutralizing antibodies. Influenza B virus (IBV) is a common human pathogen that causes yearly epidemics and significant disease in the pediatric population (1-4). Despite the importance of this virus, far less is known about IBV relative to influenza A virus (IAV). Research into IAV has been greatly enhanced by several replication-competent reporter viruses (5-17). These reporter viruses have been crucial for identifying host factors, understanding basic viral pathogenesis, screening for antiviral compounds, and characterizing broadly reactive antibodies (6)(7)(8)(9)(10)(12)(13)(14)(15)(16)(17)(18)(19). Lack of viruses encoding reporters has delayed similar progress for IBV.Development of a fluorescent influenza B reporter virus. We have previously published a description of an IAV reporter virus that contains the Gaussia luciferase gene directly behind the PB2 open reading frame (ORF) in segment 1 of A/Puerto Rico/8/1934 virus (PR8) (7). A 2A proteolytic cleavage site inserted between the PB2 ORF and the Gaussia luciferase gene allows for the cotranslational separation of the two proteins (20).Utilizing a similar approach, an influenza virus codon-optimized ORF of mNeonGreen (mNeon), a gene encoding a bright monomeric fluorescent protein (21), was cloned behind each of the three polymerase segments of the B/Yamagata/16/ 1988 (Ya88) virus (Fig. 1A). The reporter segments were rescued individually utilizing standard protocols (22-24) to generate the PB1 mNeon, PB2 mNeon, and PA mNeon viruses. Madin-Darby canine kidney (MDCK) cells were infected at a multiplicity of infection (MOI) of 0.5 for 18 h with the recombinant wild-type (rWT) Ya88 or one of the three reporter viruses (with all experiments carried out at 33°C). Tosyl phenylalanyl chloromethyl ketone (TPCK) trypsin, which is required for spread of viral progeny, was excluded from the infection medium. The cells were imaged or subjected to flow cytometry after being labeled with Hoechst stain (Thermo Scientific) or LIVE/DEAD stain (Life Technologies), respectively. By epifluorescence microscopy, the PB1 mNeon and PB2 mNeon viruses were bright and easily observed by 18 h postinfection, but the PA mNeon virus was less detectable (Fig. 1B to E). Flow cytometry (BD LSRIIA) recapitulated the microscopy results (Fig. 1F to I). Quantification of the flow cytometry data with FlowJo indicated that both the PB1 mNeon and PB2 mNeon viruses yielded the brightest signals ( Fig. 1J and K), and the PB1 mNeon virus was chosen for further study.The PB1 mNeon virus can rapidly monitor the growth of IBV in vi...

show abstract

Visualizing real-time influenza virus infection, transmission and protection in ferrets

Cited by 110 publications

References 34 publications

Type I Interferon Response Limits Astrovirus Replication and Protects against Increased Barrier Permeability In Vitro and In Vivo

Type I Interferon Response Limits Astrovirus Replication and Protects against Increased Barrier Permeability In Vitro and In Vivo

Complexities in Ferret Influenza Virus Pathogenesis and Transmission Models

Replication-Competent Influenza B Reporter Viruses as Tools for Screening Antivirals and Antibodies

Contact Info

Product

Resources

About