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The PB2-E627K mutation attenuates viruses containing the 2009 H1N1 influenza pandemic polymerase.

Jagger BW, Memoli MJ, Sheng ZM, Qi L, Hrabal RJ, Allen GL, Dugan VG, Wang R, Digard P, Kash JC, Taubenberger JK - MBio (2010)

Bottom Line: The swine-origin H1N1 influenza A virus emerged in early 2009 and caused the first influenza pandemic in 41 years.The virus has spread efficiently to both the Northern and the Southern Hemispheres and has been associated with over 16,000 deaths.Given the virus's recent zoonotic origin, there is concern that the virus could acquire signature mutations associated with the enhanced pathogenicity of previous pandemic viruses or H5N1 viruses with pandemic potential.

View Article: PubMed Central - PubMed

Affiliation: Viral Pathogenesis and Evolution Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda,Maryland, USA.

ABSTRACT
The swine-origin H1N1 influenza A virus emerged in early 2009 and caused the first influenza pandemic in 41 years. The virus has spread efficiently to both the Northern and the Southern Hemispheres and has been associated with over 16,000 deaths. Given the virus's recent zoonotic origin, there is concern that the virus could acquire signature mutations associated with the enhanced pathogenicity of previous pandemic viruses or H5N1 viruses with pandemic potential. We tested the hypothesis that mutations in the polymerase PB2 gene at residues 627 and 701 would enhance virulence but found that influenza viruses containing these mutations in the context of the pandemic virus polymerase complex are attenuated in cell culture and mice.

No MeSH data available.


Related in: MedlinePlus

Viral replication kinetics in A549 cells at 37°C. Cells were infected at a 0.01 multiplicity of infection. Supernatants from infected A549 cells were collected at 12-h intervals, and viral titer was determined by a plaque assay. Cultures and measurements were performed in triplicate. Titers are expressed as numbers of PFU per milliliter. Error bars represent standard errors of the means (SEM). See the key for the different viruses evaluated.
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f1: Viral replication kinetics in A549 cells at 37°C. Cells were infected at a 0.01 multiplicity of infection. Supernatants from infected A549 cells were collected at 12-h intervals, and viral titer was determined by a plaque assay. Cultures and measurements were performed in triplicate. Titers are expressed as numbers of PFU per milliliter. Error bars represent standard errors of the means (SEM). See the key for the different viruses evaluated.

Mentions: To test the hypothesis that introduction of PB2 mutations at residue 627 or 701 into the RNP of the 2009 pandemic H1N1 virus would enhance virulence, we evaluated replication kinetics and pathogenicity. First, we observed the growth of rescued viruses in cell culture. All rescued viruses propagated well in MDCK cells (data not shown). To characterize the chimeric viruses in cells of human origin, replication kinetics in the A549 lung cell line were assessed (Fig. 1). As expected, the 1918RNP virus grew to a significantly higher (>1 log) mean peak titer than the control rNY312 virus (P = 0.0083). The CA09RNP virus grew to high titers similar to those of the 1918RNP virus (P = 0.0088 for comparison to rNY312), as did the VN1203RNP virus (P = 0.019 for comparison to rNY312), but the low-pathogenicity avian RNP genes (S09RNP virus) did not show significantly enhanced growth compared to the NY312 parent (P = 0.050) (Fig. 1). To determine the impact of host-adaptive mutations on replication, further selected isogenic pairs of viruses in which PB2 residue 627 or 701 was mutated to the mammalian or avian consensus residue as appropriate were then created. A 1918RNP virus in which PB2 residue 627 was “back-adapted” to the avian glutamic acid (1918RNP-K627E) consistently grew to a 1- to 2-log-lower peak titer (P = 0.042) than the 1918RNP virus. Conversely, mutation of PB2 residue 627 in the avian RNP S09RNP virus to the mammalian consensus residue (S09RNP-E627K) had little effect on growth in A549 cells. Next, the impact of the PB2-E627K (CA09RNP-E627K) and D701N (CA09RNP-D701N) mutations on viruses containing the 2009 pandemic RNP was assessed. Surprisingly, rather than increasing the growth of the CA09RNP virus, both of these mutant viruses grew to a 1- to 2-log-lower peak titer than the parental CA09RNP virus (P = 0.0058 and 0.0075, respectively).


The PB2-E627K mutation attenuates viruses containing the 2009 H1N1 influenza pandemic polymerase.

Jagger BW, Memoli MJ, Sheng ZM, Qi L, Hrabal RJ, Allen GL, Dugan VG, Wang R, Digard P, Kash JC, Taubenberger JK - MBio (2010)

Viral replication kinetics in A549 cells at 37°C. Cells were infected at a 0.01 multiplicity of infection. Supernatants from infected A549 cells were collected at 12-h intervals, and viral titer was determined by a plaque assay. Cultures and measurements were performed in triplicate. Titers are expressed as numbers of PFU per milliliter. Error bars represent standard errors of the means (SEM). See the key for the different viruses evaluated.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2912665&req=5

f1: Viral replication kinetics in A549 cells at 37°C. Cells were infected at a 0.01 multiplicity of infection. Supernatants from infected A549 cells were collected at 12-h intervals, and viral titer was determined by a plaque assay. Cultures and measurements were performed in triplicate. Titers are expressed as numbers of PFU per milliliter. Error bars represent standard errors of the means (SEM). See the key for the different viruses evaluated.
Mentions: To test the hypothesis that introduction of PB2 mutations at residue 627 or 701 into the RNP of the 2009 pandemic H1N1 virus would enhance virulence, we evaluated replication kinetics and pathogenicity. First, we observed the growth of rescued viruses in cell culture. All rescued viruses propagated well in MDCK cells (data not shown). To characterize the chimeric viruses in cells of human origin, replication kinetics in the A549 lung cell line were assessed (Fig. 1). As expected, the 1918RNP virus grew to a significantly higher (>1 log) mean peak titer than the control rNY312 virus (P = 0.0083). The CA09RNP virus grew to high titers similar to those of the 1918RNP virus (P = 0.0088 for comparison to rNY312), as did the VN1203RNP virus (P = 0.019 for comparison to rNY312), but the low-pathogenicity avian RNP genes (S09RNP virus) did not show significantly enhanced growth compared to the NY312 parent (P = 0.050) (Fig. 1). To determine the impact of host-adaptive mutations on replication, further selected isogenic pairs of viruses in which PB2 residue 627 or 701 was mutated to the mammalian or avian consensus residue as appropriate were then created. A 1918RNP virus in which PB2 residue 627 was “back-adapted” to the avian glutamic acid (1918RNP-K627E) consistently grew to a 1- to 2-log-lower peak titer (P = 0.042) than the 1918RNP virus. Conversely, mutation of PB2 residue 627 in the avian RNP S09RNP virus to the mammalian consensus residue (S09RNP-E627K) had little effect on growth in A549 cells. Next, the impact of the PB2-E627K (CA09RNP-E627K) and D701N (CA09RNP-D701N) mutations on viruses containing the 2009 pandemic RNP was assessed. Surprisingly, rather than increasing the growth of the CA09RNP virus, both of these mutant viruses grew to a 1- to 2-log-lower peak titer than the parental CA09RNP virus (P = 0.0058 and 0.0075, respectively).

Bottom Line: The swine-origin H1N1 influenza A virus emerged in early 2009 and caused the first influenza pandemic in 41 years.The virus has spread efficiently to both the Northern and the Southern Hemispheres and has been associated with over 16,000 deaths.Given the virus's recent zoonotic origin, there is concern that the virus could acquire signature mutations associated with the enhanced pathogenicity of previous pandemic viruses or H5N1 viruses with pandemic potential.

View Article: PubMed Central - PubMed

Affiliation: Viral Pathogenesis and Evolution Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda,Maryland, USA.

ABSTRACT
The swine-origin H1N1 influenza A virus emerged in early 2009 and caused the first influenza pandemic in 41 years. The virus has spread efficiently to both the Northern and the Southern Hemispheres and has been associated with over 16,000 deaths. Given the virus's recent zoonotic origin, there is concern that the virus could acquire signature mutations associated with the enhanced pathogenicity of previous pandemic viruses or H5N1 viruses with pandemic potential. We tested the hypothesis that mutations in the polymerase PB2 gene at residues 627 and 701 would enhance virulence but found that influenza viruses containing these mutations in the context of the pandemic virus polymerase complex are attenuated in cell culture and mice.

No MeSH data available.


Related in: MedlinePlus