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Studies of inactivation mechanism of non-enveloped icosahedral virus by a visible ultrashort pulsed laser.

Tsen SW, Kingsley DH, Poweleit C, Achilefu S, Soroka DS, Wu TC, Tsen KT - Virol. J. (2014)

Bottom Line: These studies provide fundamental knowledge on photon-virus interactions on femtosecond time scales.From the analysis of the transmission electron microscope (TEM) images of viral particles before and after USP laser irradiation, the locations of weak structural links on the capsid of MNV-1 were revealed.We envision that this non-invasive, efficient viral eradication method will find applications in the disinfection of pharmaceuticals, biologicals and blood products in the near future.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physics, Arizona State University, Tempe, Arizona 85287, USA. tsen@asu.edu.

ABSTRACT

Background: Low-power ultrashort pulsed (USP) lasers operating at wavelengths of 425 nm and near infrared region have been shown to effectively inactivate viruses such as human immunodeficiency virus (HIV), M13 bacteriophage, and murine cytomegalovirus (MCMV). It was shown previously that non-enveloped, helical viruses such as M13 bacteriophage, were inactivated by a USP laser through an impulsive stimulated Raman scattering (ISRS) process. Recently, enveloped virus like MCMV has been shown to be inactivated by a USP laser via protein aggregation induced by an ISRS process. However, the inactivation mechanism for a clinically important class of viruses--non-enveloped, icosahedral viruses remains unknown.

Results and discussions: We have ruled out the following four possible inactivation mechanisms for non-enveloped, icosahedral viruses, namely, (1) inactivation due to ultraviolet C (UVC) photons produced by non-linear optical process of the intense, fundamental laser beam at 425 nm; (2) inactivation caused by thermal heating generated by the direct laser absorption/heating of the virion; (3) inactivation resulting from a one-photon absorption process via chromophores such as porphyrin molecules, or indicator dyes, potentially producing reactive oxygen or other species; (4) inactivation by the USP lasers in which the extremely intense laser pulse produces shock wave-like vibrations upon impact with the viral particle. We present data which support that the inactivation mechanism for non-enveloped, icosahedral viruses is the impulsive stimulated Raman scattering process. Real-time PCR experiments show that, within the amplicon size of 273 bp tested, there is no damage on the genome of MNV-1 caused by the USP laser irradiation.

Conclusion: We conclude that our model non-enveloped virus, MNV-1, is inactivated by the ISRS process. These studies provide fundamental knowledge on photon-virus interactions on femtosecond time scales. From the analysis of the transmission electron microscope (TEM) images of viral particles before and after USP laser irradiation, the locations of weak structural links on the capsid of MNV-1 were revealed. This important information will greatly aid our understanding of the structure of non-enveloped, icosahedral viruses. We envision that this non-invasive, efficient viral eradication method will find applications in the disinfection of pharmaceuticals, biologicals and blood products in the near future.

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Inactivation of MNV-1 by the visible ultrashort pulsed laser. (a) Graphs of plaque forming units (PFU)/ml for control (without laser irradiation) and laser irradiated MNV-1 purified samples. (b) Graphs of plaque forming units (PFU)/ml for control (without laser irradiation) and laser irradiated MNV-1 unpurified samples. The USP laser is operated at a wavelength of 425 nm with a pulse width of 100 fs, a repetition rate of 80 MHz, and an average power of 120 mWs. The laser exposure time was 2 h. S.D error bars are small to be readily observed.
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Figure 1: Inactivation of MNV-1 by the visible ultrashort pulsed laser. (a) Graphs of plaque forming units (PFU)/ml for control (without laser irradiation) and laser irradiated MNV-1 purified samples. (b) Graphs of plaque forming units (PFU)/ml for control (without laser irradiation) and laser irradiated MNV-1 unpurified samples. The USP laser is operated at a wavelength of 425 nm with a pulse width of 100 fs, a repetition rate of 80 MHz, and an average power of 120 mWs. The laser exposure time was 2 h. S.D error bars are small to be readily observed.

Mentions: Since virus stocks and samples are often propagated in complex tissue culture media that contains potentially chromogenic constituents such as neutral red indicator dye, amino acids, cellular proteins and nucleic acids, MNV-1 used in this work was propagated in indicator-free media and partially purified by sucrose gradient or Optiprep gradient ultracentrifugation, respectively. Figure 1(a) shows a bar graph of the plaque forming units (PFU) for control and laser-irradiated MNV-1 samples in buffer solution. The control represents the sample with no laser irradiation. We observed a load reduction of 3.1 ± 0.1 in log10 scale for the laser-treated MNV-1 group. Here, load reduction is defined as the ratio of the number of plaque in the control to the number of plaque in the laser-irradiated sample. For reference, Figure 1(b) shows the load reduction of 3.0 ± 0.1 in log10 scale for the unpurified laser-treated MNV-1.


Studies of inactivation mechanism of non-enveloped icosahedral virus by a visible ultrashort pulsed laser.

Tsen SW, Kingsley DH, Poweleit C, Achilefu S, Soroka DS, Wu TC, Tsen KT - Virol. J. (2014)

Inactivation of MNV-1 by the visible ultrashort pulsed laser. (a) Graphs of plaque forming units (PFU)/ml for control (without laser irradiation) and laser irradiated MNV-1 purified samples. (b) Graphs of plaque forming units (PFU)/ml for control (without laser irradiation) and laser irradiated MNV-1 unpurified samples. The USP laser is operated at a wavelength of 425 nm with a pulse width of 100 fs, a repetition rate of 80 MHz, and an average power of 120 mWs. The laser exposure time was 2 h. S.D error bars are small to be readily observed.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3924410&req=5

Figure 1: Inactivation of MNV-1 by the visible ultrashort pulsed laser. (a) Graphs of plaque forming units (PFU)/ml for control (without laser irradiation) and laser irradiated MNV-1 purified samples. (b) Graphs of plaque forming units (PFU)/ml for control (without laser irradiation) and laser irradiated MNV-1 unpurified samples. The USP laser is operated at a wavelength of 425 nm with a pulse width of 100 fs, a repetition rate of 80 MHz, and an average power of 120 mWs. The laser exposure time was 2 h. S.D error bars are small to be readily observed.
Mentions: Since virus stocks and samples are often propagated in complex tissue culture media that contains potentially chromogenic constituents such as neutral red indicator dye, amino acids, cellular proteins and nucleic acids, MNV-1 used in this work was propagated in indicator-free media and partially purified by sucrose gradient or Optiprep gradient ultracentrifugation, respectively. Figure 1(a) shows a bar graph of the plaque forming units (PFU) for control and laser-irradiated MNV-1 samples in buffer solution. The control represents the sample with no laser irradiation. We observed a load reduction of 3.1 ± 0.1 in log10 scale for the laser-treated MNV-1 group. Here, load reduction is defined as the ratio of the number of plaque in the control to the number of plaque in the laser-irradiated sample. For reference, Figure 1(b) shows the load reduction of 3.0 ± 0.1 in log10 scale for the unpurified laser-treated MNV-1.

Bottom Line: These studies provide fundamental knowledge on photon-virus interactions on femtosecond time scales.From the analysis of the transmission electron microscope (TEM) images of viral particles before and after USP laser irradiation, the locations of weak structural links on the capsid of MNV-1 were revealed.We envision that this non-invasive, efficient viral eradication method will find applications in the disinfection of pharmaceuticals, biologicals and blood products in the near future.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physics, Arizona State University, Tempe, Arizona 85287, USA. tsen@asu.edu.

ABSTRACT

Background: Low-power ultrashort pulsed (USP) lasers operating at wavelengths of 425 nm and near infrared region have been shown to effectively inactivate viruses such as human immunodeficiency virus (HIV), M13 bacteriophage, and murine cytomegalovirus (MCMV). It was shown previously that non-enveloped, helical viruses such as M13 bacteriophage, were inactivated by a USP laser through an impulsive stimulated Raman scattering (ISRS) process. Recently, enveloped virus like MCMV has been shown to be inactivated by a USP laser via protein aggregation induced by an ISRS process. However, the inactivation mechanism for a clinically important class of viruses--non-enveloped, icosahedral viruses remains unknown.

Results and discussions: We have ruled out the following four possible inactivation mechanisms for non-enveloped, icosahedral viruses, namely, (1) inactivation due to ultraviolet C (UVC) photons produced by non-linear optical process of the intense, fundamental laser beam at 425 nm; (2) inactivation caused by thermal heating generated by the direct laser absorption/heating of the virion; (3) inactivation resulting from a one-photon absorption process via chromophores such as porphyrin molecules, or indicator dyes, potentially producing reactive oxygen or other species; (4) inactivation by the USP lasers in which the extremely intense laser pulse produces shock wave-like vibrations upon impact with the viral particle. We present data which support that the inactivation mechanism for non-enveloped, icosahedral viruses is the impulsive stimulated Raman scattering process. Real-time PCR experiments show that, within the amplicon size of 273 bp tested, there is no damage on the genome of MNV-1 caused by the USP laser irradiation.

Conclusion: We conclude that our model non-enveloped virus, MNV-1, is inactivated by the ISRS process. These studies provide fundamental knowledge on photon-virus interactions on femtosecond time scales. From the analysis of the transmission electron microscope (TEM) images of viral particles before and after USP laser irradiation, the locations of weak structural links on the capsid of MNV-1 were revealed. This important information will greatly aid our understanding of the structure of non-enveloped, icosahedral viruses. We envision that this non-invasive, efficient viral eradication method will find applications in the disinfection of pharmaceuticals, biologicals and blood products in the near future.

Show MeSH
Related in: MedlinePlus