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
. 2024 Jul 22;16(14):2377.
doi: 10.3390/nu16142377.

Selaginella tamariscina Ethanol Extract Attenuates Influenza A Virus Infection by Inhibiting Hemagglutinin and Neuraminidase

Won-Kyung Cho  1 , Hee-Jeong Choi  1 , Jin Yeul Ma  1
Affiliations

Selaginella tamariscina Ethanol Extract Attenuates Influenza A Virus Infection by Inhibiting Hemagglutinin and Neuraminidase

Won-Kyung Cho et al. Nutrients. .

Abstract

Selaginella tamariscina is a perennial plant that is used for diverse diseases. This study investigated whether Selaginella tamariscina has an antiviral effect against influenza A virus (IAV) infection. We used green fluorescent protein (GFP)-tagged influenza A virus (IAV) to examine the effect of Selaginella tamariscina ethanol extract (STE) on influenza viral infection. Fluorescence microscopy and flow cytometry showed that STE potently represses GFP expression by the virus, dose-dependently. STE significantly inhibited the expression of the IAV M2, NP, HA, NA, NS1, and PB2 proteins. Time-of-addition and hemagglutination inhibition assays showed that STE has an inhibitory effect on hemagglutinin and viral binding on the cells at an early infection time. In addition, STE exerted a suppressive effect on the neuraminidase activity of the H1N1 and H3N2 IAVs. Furthermore, dose-dependently, STE inhibited the cytopathic effect induced by H3N2, as well as by H1N1 IAV. Especially in the presence of 200 µg/mL STE, the cytopathic effect was completely blocked. Our findings suggest that STE has antiviral efficacy against IAV infection; thus, it could be developed as a natural IAV inhibitor.

Keywords: Selaginella tamariscina; cytopathic effect; hemagglutinin; influenza A virus; neuraminidase.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Cytotoxicity and antiviral effect of Selaginella tamariscina ethanol extract (STE) in RAW 264.7 cells. (A) The toxicity of STE in the cells was evaluated using a CCK-8 assay. The data represent the mean ± SD from three independent experiments. The unpaired Student t-test was used to assess the statistical significance. * P < 0.5, ** P < 0.05, and *** P < 0.005, compared with the untreated control. (B,C) The cells were cotreated with PR8-GFP IAV and STE. The effect of STE on PR8-GFP IAV infection was evaluated by comparing GFP expression using a fluorescence microscope (B) and FACS analysis (C). The data represent the mean ± SD from three independent experiments. The unpaired Student t-test was used to assess the statistical significance. ** P < 0.005, and *** P < 0.0005, compared with the virus-infected control.
Figure 2
Figure 2
STE protects the cells from the cytopathic effect induced by H1N1 (A) and H3N2 (B) influenza virus infection. STE at the indicated concentrations or medium (mock) was mixed with H1N1 or H3N2 IAV before infection of the cells. The cells infected with the mixture were incubated until the cytopathic effect formed. The cell viability was determined via a CCK-8 assay. The data represent the mean ± SD from three independent experiments. The unpaired Student t-test was used to assess the statistical significance. * P < 0.05, ** P < 0.005; NS, no significance.
Figure 3
Figure 3
STE suppresses IAV protein expression. STE and PR8-GFP IAV were incubated for 1 h at 4 °C. The cells were coinfected with the mixture for 24 h at 37 °C. The cells were fixed and detected with antibodies against IAV proteins, including M2, NP, NS1, NA, HA, and PB2 (red color). To detect the nuclei, the cells were stained with Hoechst 33342 (blue color). The co-localization images of red viral proteins and blue nuclei were captured using a fluorescence microscope.
Figure 4
Figure 4
STE affects the virus attachment and entry and directly kills the virus at an early phase. The cells were cotreated with STE (100 µg/mL) and PR8-GFP IAV (10 MOI) to examine the effect of STE on viral attachment, entry, or virucidal stage. The detailed time-of-addition methods were described in the Materials and Methods Section. The images of GFP-expressing cells were obtained using brightfield and fluorescence microscopy (A). The cells fixed with paraformaldehyde were analyzed by flow cytometry (B). The data represent the mean ± SD from three independent experiments. The unpaired Student t-test was used to assess the statistical significance. ** P < 0.005, *** P < 0.0005; NS, no significance.
Figure 5
Figure 5
STE reduces the HA unit of influenza viruses (A,B). The cells were cotreated with STE at the indicated concentrations and H1N1 IAV for 24 h at 37 °C. The 2-fold serially diluted supernatants and chicken RBC cells were mixed in round 96-well plates for 1 h at room temperature. The red circle indicates hemagglutination (HA) units.
Figure 6
Figure 6
STE dose-dependently represses the neuraminidase activities of H1N1 and H3N2 IAVs. Serially diluted STE (A) or oseltamivir carboxylate (B) was mixed with H1N1 or H3N2 IAV in 96-well black plates. The detailed neuraminidase activity assay was described in materials and methods. The data represent the mean ± SD from three independent experiments. The unpaired Student t-test was used to assess the statistical significance. * P < 0.05, ** P < 0.005, and *** P < 0.0005, compared with the H1N1 virus-infected group. # P < 0.05, ## P < 0.005, and ### P < 0.0005, compared with the H3N2 virus-infected group.
See this image and copyright information in PMC

Similar articles

See all similar articles

References

    1. Atkins N., Harikar M., Duggan K., Zawiejska A., Vardhan V., Vokey L., Dozier M., de Los Godos E.F., McSwiggan E., McQuillan R., et al. What are the characteristics of participatory surveillance systems for influenza-like-illness? J. Glob. Health. 2023;13:04130. doi: 10.7189/jogh.13.04130. - DOI - PMC - PubMed
    1. Paules C., Subbarao K. Influenza. Lancet. 2017;390:697–708. doi: 10.1016/S0140-6736(17)30129-0. - DOI - PubMed
    1. Gerber M., Isel C., Moules V., Marquet R. Selective packaging of the influenza A genome and consequences for genetic reassortment. Trends Microbiol. 2014;22:446–455. doi: 10.1016/j.tim.2014.04.001. - DOI - PubMed
    1. Bouvier N.M., Palese P. The biology of influenza viruses. Vaccine. 2008;26((Suppl. S4)):D49–D53. doi: 10.1016/j.vaccine.2008.07.039. - DOI - PMC - PubMed
    1. Sautto G.A., Kirchenbaum G.A., Ross T.M. Towards a universal influenza vaccine: Different approaches for one goal. Virol. J. 2018;15:17. doi: 10.1186/s12985-017-0918-y. - DOI - PMC - PubMed

MeSH terms

LinkOut - more resources