Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2011 Jun;92(Pt 6):1410-1415.
doi: 10.1099/vir.0.030379-0. Epub 2011 Feb 23.

Insertion of a multibasic cleavage site in the haemagglutinin of human influenza H3N2 virus does not increase pathogenicity in ferrets

Eefje J A Schrauwen  1 , Theo M Bestebroer  1 , Vincent J Munster  2   1 , Emmie de Wit  2   1 , Sander Herfst  1 , Guus F Rimmelzwaan  1 , Albert D M E Osterhaus  1 , Ron A M Fouchier  1
Affiliations

Insertion of a multibasic cleavage site in the haemagglutinin of human influenza H3N2 virus does not increase pathogenicity in ferrets

Eefje J A Schrauwen et al. J Gen Virol. 2011 Jun.

Abstract

A multibasic cleavage site (MBCS) in the haemagglutinin (HA) protein of influenza A virus is a key determinant of pathogenicity in chickens, and distinguishes highly pathogenic avian influenza (HPAI) viruses from low pathogenic avian influenza viruses (LPAI). An MBCS has only been detected in viruses of the H5 and H7 subtypes. Here we investigated the phenotype of a human H3N2 virus with an MBCS in HA. Insertion of an MBCS in the H3N2 virus resulted in cleavage of HA and efficient replication in Madin-Darby canine kidney cells in the absence of exogenous trypsin in vitro, similar to HPAI H5N1 virus. However, studies in ferrets demonstrated that insertion of the MBCS into HA did not result in increased virus shedding, cellular host range, systemic replication or pathogenicity, as compared with wild-type virus. This study indicates that acquisition of an MBCS alone is insufficient to increase pathogenicity of a prototypical seasonal human H3N2 virus.

PubMed Disclaimer

Figures

Fig. 1.
Fig. 1.
In vitro phenotype of H3N2wt, H3N2MBCS, H5N1wt and H5N1ΔMBCS. (a and b) Western blots of lysates of 293T cells transfected with HAs of (a) H3N2wt and H3N2MBCS, (b) H5N1wt and H5N1ΔMBCS upon treatment with (T) or without (−) trypsin. (c) Replication of H3N2wt (▪ or ▪) and H3N2MBCS (○ or ○) in the presence (black lines) or absence (grey lines) of trypsin. (d) Replication of H5N1wt (○ or ○) and H5N1ΔMBCS (▪ or ▪) in the presence (black lines) or absence (grey lines) of trypsin. MDCK cells were inoculated with 0.01 TCID50 per cell and supernatant samples were harvested 6, 12, 24 and 48 h later. Supernatant samples were titrated in MDCK cells. Geometric mean titres and sd were calculated from two independent experiments. Data for Fig. 1(b and d) were taken with permission from Munster et al. (2010) (Copyright, American Society for Microbiology).
Fig. 2.
Fig. 2.
Weight loss and virus replication in ferrets inoculated with H3N2wt and H3N2MBCS virus. (a) Weight loss of ferrets inoculated with H3N2wt (○) and H3N2MBCS (▪) virus. Mean body weight and sd were calculated as percentages of body weight at time of inoculation for each group. After day 3, only three animals remained in each group. (b and c) Virus shedding from the nose (b) and throat (c) of ferrets inoculated with H3N2wt (black bars) and H3N2MBCS (grey bars). (d) Virus detection in tissues of ferrets inoculated with H3N2wt (black bars) and H3N2MBCS (grey bars) virus at day 3. Geometric mean titres are shown, with error bars indicating the sd. The lower limit of detection is indicated by the dotted line. The number of positive tissues from the ferrets are shown.
Fig. 3.
Fig. 3.
Tissue distribution of the H3N2wt and H3N2MBCS virus in ferrets. Expression of viral antigen in nasal turbinates, trachea and brain of ferrets inoculated with H3N2wt and H3N2MBCS at 3 days p.i. Tissue sections were stained with a mAb against influenza A virus nucleoprotein, visible as red-brown staining.
See this image and copyright information in PMC

References

    1. Belser J. A., Lu X., Maines T. R., Smith C., Li Y., Donis R. O., Katz J. M., Tumpey T. M. (2007). Pathogenesis of avian influenza (H7) virus infection in mice and ferrets: enhanced virulence of Eurasian H7N7 viruses isolated from humans. J Virol 81, 11139–11147 10.1128/JVI.01235-07. - DOI - PMC - PubMed
    1. Bertram S., Glowacka I., Steffen I., Kühl A., Pöhlmann S. (2010). Novel insights into proteolytic cleavage of influenza virus hemagglutinin. Rev Med Virol 20, 298–310 10.1002/rmv.657. - DOI - PMC - PubMed
    1. Bodewes R., Rimmelzwaan G. F., Osterhaus A. D. (2010). Animal models for the preclinical evaluation of candidate influenza vaccines. Expert Rev Vaccines 9, 59–72 10.1586/erv.09.148. - DOI - PubMed
    1. Bogs J., Veits J., Gohrbandt S., Hundt J., Stech O., Breithaupt A., Teifke J. P., Mettenleiter T. C., Stech J. (2010). Highly pathogenic H5N1 influenza viruses carry virulence determinants beyond the polybasic hemagglutinin cleavage site. PLoS ONE 5, e11826 10.1371/journal.pone.0011826. - DOI - PMC - PubMed
    1. Chen J., Lee K. H., Steinhauer D. A., Stevens D. J., Skehel J. J., Wiley D. C. (1998). Structure of the hemagglutinin precursor cleavage site, a determinant of influenza pathogenicity and the origin of the labile conformation. Cell 95, 409–417 10.1016/S0092-8674(00)81771-7. - DOI - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources