NS-based live attenuated H1N1 pandemic vaccines protect mice and ferrets

Zhou, Bin; Li, Yan; Belser, Jessica A.; Pearce, Melissa B.; Schmolke, Mirco; Subba, Anju; Z, Shi; Zaki, Sherif R.; Blau, Dianna M.; Garcı́a-Sastre, Adolfo; Tumpey, Terrence M.; Wentworth, David E.

doi:10.1016/j.vaccine.2010.08.106

Cited by 48 publications

(40 citation statements)

References 45 publications

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“…Recombinant caAA60 and various mutant viruses were generated by cotransfection of eight reverse-genetics plasmids carrying the cDNA of each gene segment into a 293T-MDCK coculture monolayer as previously described (52). Briefly, 0.6 g of each plasmid was mixed and incubated with 15 l of TransIt-LT1 transfection reagent (Mirus Bio, Madison, WI) at 20°C for 20 min and then added to 80% confluent 293T-MDCK cocultures in 6-well plates.…”

Section: Methodsmentioning

confidence: 99%

Reversion of Cold-Adapted Live Attenuated Influenza Vaccine into a Pathogenic Virus

Zhou

Meliopoulos

Wang

et al. 2016

J Virol

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The only licensed live attenuated influenza A virus vaccines (LAIVs) in the United States (FluMist) are created using internal protein-coding gene segments from the cold-adapted temperature-sensitive master donor virus A/Ann Arbor/6/1960 and HA/NA gene segments from circulating viruses. During serial passage of A/Ann Arbor/6/1960 at low temperatures to select the desired attenuating phenotypes, multiple cold-adaptive mutations and temperature-sensitive mutations arose. A substantial amount of scientific and clinical evidence has proven that FluMist is safe and effective. Nevertheless, no study has been conducted specifically to determine if the attenuating temperature-sensitive phenotype can revert and, if so, the types of substitutions that will emerge (i.e., compensatory substitutions versus reversion of existing attenuating mutations). Serial passage of the monovalent FluMist 2009 H1N1 pandemic vaccine at increasing temperatures in vitro generated a variant that replicated efficiently at higher temperatures. Sequencing of the variant identified seven nonsynonymous mutations, PB1-E51K, PB1-I171V, PA-N350K, PA-L366I, NP-N125Y, NP-V186I, and NS2-G63E. None occurred at positions previously reported to affect the temperature sensitivity of influenza A viruses. Synthetic genomics technology was used to synthesize the whole genome of the virus, and the roles of individual mutations were characterized by assessing their effects on RNA polymerase activity and virus replication kinetics at various temperatures. The revertant also regained virulence and caused significant disease in mice, with severity comparable to that caused by a wild-type 2009 H1N1 pandemic virus. IMPORTANCEThe live attenuated influenza vaccine FluMist has been proven safe and effective and is widely used in the United States. The phenotype and genotype of the vaccine virus are believed to be very stable, and mutants that cause disease in animals or humans have never been reported. By propagating the virus under well-controlled laboratory conditions, we found that the FluMist vaccine backbone could regain virulence to cause severe disease in mice. The identification of the responsible substitutions and elucidation of the underlying mechanisms provide unique insights into the attenuation of influenza virus, which is important to basic research on vaccines, attenuation reversion, and replication. In addition, this study suggests that the safety of LAIVs should be closely monitored after mass vaccination and that novel strategies to continue to improve LAIV vaccine safety should be investigated.A nnual influenza epidemics are caused by influenza A (H1N1 and H3N2) and influenza B viruses that infect 5% to 10% of adults and 20% to 30% of children (1). Disease outcomes range from subclinical infections to severe infections that result in very high mortality. The estimated 3 to 5 million cases of severe illness and 250,000 to 500,000 deaths each year, as well as significant socioeconomic losses, warrant continuous improvements in influenza preventi...

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Section: Methodsmentioning

confidence: 99%

Reversion of Cold-Adapted Live Attenuated Influenza Vaccine into a Pathogenic Virus

Zhou

Meliopoulos

Wang

et al. 2016

J Virol

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“…For example, bioengineered IFV strains with mutations in the nonstructural (NS) genes [8,9] and other strains attenuated by codon-pair de-optimization [10] have been developed as alternatives to the licensed FluMist. These new LAIV particles can be mass-produced in cultured mammalian cells, with less chance to shed virus postimmunization and less likelihood to generate revertants within a vaccinee when compared with FluMist.…”

Section: Discussion and Significance Of The Resultsmentioning

confidence: 99%

Perspectives on replication-incompetent nasal influenza virus vaccines

Tang¹

2012

Expert Review of Vaccines

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Influenza is an emerging as well as resurging contagious disease with a worldwide impact on public health. Although broad administration of the licensed influenza virus (IFV) vaccines has mitigated the disease in many countries over the years, there are intrinsic problems associated with them. The study under evaluation reports that a novel PB2-knockout nonreplicating nasal IFV vaccine has been generated with the capacity to confer protection of mice against live IFV challenges. Moreover, an exogenous gene expressed from the bioengineered PB2-knockout IFV could elicit an immune response against the exogenous protein, showing its potential to deliver transgenes as a vector. The risk-benefit ratio of this new influenza vaccine vector is discussed.

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“…Recombinant viruses rhuH1N1 HA, NA, PA, and HNP were generated by amplification of the genomes using RT-PCR and cloning into modified reverse genetics plasmids (50). Sequence-verified reverse genetics plasmids representing each of the 8 gene segments were cotransfected into HEK-293T/MDCK cocultured monolayers as described previously (50,51). Briefly, 0.6 g of plasmid for each gene segment was mixed and incubated with 15 l of Lipofectamine 2000 (Invitrogen) at 20°C for 20 min.…”

Section: Ethics Statementmentioning

confidence: 99%

Swine Influenza Virus PA and Neuraminidase Gene Reassortment into Human H1N1 Influenza Virus Is Associated with an Altered Pathogenic Phenotype Linked to Increased MIP-2 Expression

Dlugolenski

Jones

Howerth

et al. 2015

J Virol

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Swine are susceptible to infection by both avian and human influenza viruses, and this feature is thought to contribute to novel reassortant influenza viruses. In this study, the influenza virus reassortment rate in swine and human cells was determined. Coinfection of swine cells with 2009 pandemic H1N1 virus (huH1N1) and an endemic swine H1N2 (A/swine/Illinois/02860/09) virus (swH1N2) resulted in a 23% reassortment rate that was independent of ␣2,3-or ␣2,6-sialic acid distribution on the cells. The reassortants had altered pathogenic phenotypes linked to introduction of the swine virus PA and neuraminidase (NA) into huH1N1. In mice, the huH1N1 PA and NA mediated increased MIP-2 expression early postinfection, resulting in substantial pulmonary neutrophilia with enhanced lung pathology and disease. The findings support the notion that swine are a mixing vessel for influenza virus reassortants independent of sialic acid distribution. These results show the potential for continued reassortment of the 2009 pandemic H1N1 virus with endemic swine viruses and for reassortants to have increased pathogenicity linked to the swine virus NA and PA genes which are associated with increased pulmonary neutrophil trafficking that is related to MIP-2 expression. IMPORTANCEInfluenza A viruses can change rapidly via reassortment to create a novel virus, and reassortment can result in possible pandemics. Reassortments among subtypes from avian and human viruses led to the 1957 (H2N2 subtype) and 1968 (H3N2 subtype) human influenza pandemics. Recent analyses of circulating isolates have shown that multiple genes can be recombined from human, avian, and swine influenza viruses, leading to triple reassortants. Understanding the factors that can affect influenza A virus reassortment is needed for the establishment of disease intervention strategies that may reduce or preclude pandemics. The findings from this study show that swine cells provide a mixing vessel for influenza virus reassortment independent of differential sialic acid distribution. The findings also establish that circulating neuraminidase (NA) and PA genes could alter the pathogenic phenotype of the pandemic H1N1 virus, resulting in enhanced disease. The identification of such factors provides a framework for pandemic modeling and surveillance. E ach year, influenza A viruses (IAVs) cause epidemics with high morbidity and some mortality in humans (1). In avian species, IAV infection generally is associated with mild disease, and it is only when the virus crosses species barriers that pathogenic traits are attributed to infection (2). IAV is frequently associated with spillover events, as numerous species are susceptible to IAV. Often, reassortment of IAV leads to a subtype or variant that has not previously infected humans, and as the virus adapts and is transmitted more easily, a pandemic event may loom. IAV is the cause of Ͼ30,000 deaths in the United States and Ͼ500,000 deaths worldwide annually (3). In the past century, four IAV pandemics have occurred, i.e....

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NS-based live attenuated H1N1 pandemic vaccines protect mice and ferrets

Cited by 48 publications

References 45 publications

Reversion of Cold-Adapted Live Attenuated Influenza Vaccine into a Pathogenic Virus

Reversion of Cold-Adapted Live Attenuated Influenza Vaccine into a Pathogenic Virus

Perspectives on replication-incompetent nasal influenza virus vaccines

Swine Influenza Virus PA and Neuraminidase Gene Reassortment into Human H1N1 Influenza Virus Is Associated with an Altered Pathogenic Phenotype Linked to Increased MIP-2 Expression

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