Optical and structural properties of plasma-treated Cordyceps bassiana spores as studied by circular dichroism, absorption, and fluorescence spectroscopy

To understand the killing mechanism of fungal spores by plasma treatment, the optical, structural, and biological properties of the insect pathogenic fungus Cordyceps bassiana spores were studied. A nonthermal atmospheric-pressure plasma jet (APPJ) was used to treat the spores in aqueous solution. Optical emission spectra of the APPJ acquired in air indicated emission peaks corresponding to hydroxyl radicals and atomic oxygen. When the APPJ entered the aqueous solution, additional reactive species were derived from the interaction of plasma radicals with the aqueous solution. Fluorescence and absorption spectroscopy confirmed the generation of hydroxyl radicals and hydrogen peroxide in the plasma-activated water (PAW). Spore counting showed that plasma treatment significantly reduced spore viability. Absorption spectroscopy, circular dichroism (CD) spectroscopy, and agarose gel electrophoresis of the DNA extracted from plasma-treated spores showed a reduction in spore DNA content. The magnitude of the dip in the CD spectrum was lower in the plasma-treated spores than in the control, indicating that plasma treatment causes structural modifications and/or damage to cellular components. Tryptophan fluorescence intensity was lower in the plasma-treated spores than in the control, suggesting that plasma treatment modified cell wall proteins. Changes in spore viability and DNA content were attributed to structural modification of the cell wall by reactive species coming from the APPJ and the PAW. Our results provided evidence that the plasma radicals and the derived reactive species play critical roles in fungal spore inactivation.

[1]  K. Michaelian,et al.  A reinvestigation of the Raman spectrum of water , 1978 .

[2]  Xinpei Lu,et al.  On the Active Species Concentrations of Atmospheric Pressure Nonequilibrium Plasma Jets , 2013, IEEE Transactions on Plasma Science.

[3]  Yuan Pan,et al.  The roles of the various plasma agents in the inactivation of bacteria , 2008 .

[4]  James L. Walsh,et al.  Microplasmas: sources, particle kinetics, and biomedical applications , 2008 .

[5]  Gregory Fridman,et al.  Physical and biological mechanisms of direct plasma interaction with living tissue , 2009 .

[6]  S. Holland,et al.  Human Leukocytes Kill Aspergillus nidulans by Reactive Oxygen Species-Independent Mechanisms , 2010, Infection and Immunity.

[7]  N. N. Gessler,et al.  Reactive oxygen species in regulation of fungal development , 2007, Biochemistry (Moscow).

[8]  V. Bloomfield,et al.  Dependence of the Raman signature of genomic B-DNA on nucleotide base sequence. , 1999, Biopolymers.

[9]  Daniela Iacopino,et al.  The intrinsic fluorescence spectra of selected pollen and fungal spores , 2011 .

[10]  S. Farr,et al.  Oxidative stress responses in Escherichia coli and Salmonella typhimurium. , 1991, Microbiological reviews.

[11]  P. Pelicci,et al.  Hydrogen peroxide: a metabolic by-product or a common mediator of ageing signals? , 2007, Nature Reviews Molecular Cell Biology.

[12]  Mounir Laroussi,et al.  Low Temperature Plasma-Based Sterilization: Overview and State-of-the-Art , 2005 .

[13]  M C Manning,et al.  Tyrosine, phenylalanine, and disulfide contributions to the circular dichroism of proteins: circular dichroism spectra of wild-type and mutant bovine pancreatic trypsin inhibitor. , 1999, Biochemistry.

[14]  Steffen Brinckmann,et al.  Photons and particles emitted from cold atmospheric-pressure plasma inactivate bacteria and biomolecules independently and synergistically , 2013, Journal of The Royal Society Interface.

[15]  Zeng-zhi Li,et al.  Molecular identification of the teleomorph of Beauveria bassiana , 2002 .

[16]  E. Choi,et al.  Optical and structural properties of nanobiomaterials. , 2014, Journal of nanoscience and nanotechnology.

[17]  Christophe Leys,et al.  Atmospheric pressure plasma jet in Ar and Ar/H2O mixtures: Optical emission spectroscopy and temperature measurements , 2010 .

[18]  Mounir Laroussi,et al.  Guided ionization waves : theory and experiments , 2014 .

[19]  A. Garner,et al.  Nonthermal Atmospheric Plasma Rapidly Disinfects Multidrug-Resistant Microbes by Inducing Cell Surface Damage , 2012, Antimicrobial Agents and Chemotherapy.

[20]  K. B. Song,et al.  Measurement of hydroxyl radical density generated from the atmospheric pressure bioplasma jet , 2012 .

[21]  Sun Ja Kim,et al.  Production of intracellular reactive oxygen species and change of cell viability induced by atmospheric pressure plasma in normal and cancer cells , 2013 .

[22]  J. Zimmermann,et al.  Contact-Free Inactivation of Candida albicans Biofilms by Cold Atmospheric Air Plasma , 2012, Applied and Environmental Microbiology.

[23]  D. Mihailovic,et al.  Optical spectra of wet and dry M -DNA , 2007 .

[24]  T. Tullius,et al.  Oxidative Strand Scission of Nucleic Acids: Routes Initiated by Hydrogen Abstraction from the Sugar Moiety. , 1998, Chemical reviews.

[25]  S. Leem,et al.  Characterization of Atmospheric Pressure Microplasma Jet Source and its Application to Bacterial Inactivation , 2009 .

[26]  Satoshi Hamaguchi,et al.  Effects of pH on Bacterial Inactivation in Aqueous Solutions due to Low‐Temperature Atmospheric Pressure Plasma Application , 2010 .

[27]  Young-Wan Kwon,et al.  Raman spectroscopic study of plasma-treated salmon DNA , 2013 .

[28]  P. Chu,et al.  Characteristics of atmospheric-pressure non-thermal N2 and N2/O2 gas mixture plasma jet , 2014 .

[29]  Jue Zhang,et al.  Atmospheric pressure cold plasma as an antifungal therapy , 2011 .

[30]  N. Galeano,et al.  Transcriptome analysis of the entomopathogenic fungus Beauveria bassiana grown on cuticular extracts of the coffee berry borer (Hypothenemus hampei). , 2012, Microbiology.

[31]  Jing Fang,et al.  Inactivation of Bacteria in an Aqueous Environment by a Direct-Current, Cold-Atmospheric-Pressure Air Plasma Microjet , 2010 .

[32]  Qinggui Wang,et al.  Beauvericin, a Bioactive Compound Produced by Fungi: A Short Review , 2012, Molecules.

[33]  E. Seeberg,et al.  Mutagenicity, toxicity and repair of DNA base damage induced by oxidation. , 2003, Mutation research.

[34]  G. Sung,et al.  Successful Development of Cordyceps bassiana Stromata from Beauveria bassiana , 2010, Mycobiology.

[35]  J. Bähler,et al.  Tuning gene expression to changing environments: from rapid responses to evolutionary adaptation , 2008, Nature Reviews Genetics.

[36]  Y. Chuai,et al.  Hydrogen as a New Class of Radioprotective Agent , 2013, International journal of biological sciences.

[37]  D. M. Carey,et al.  Measurement of the Raman Spectrum of Liquid Water , 1996 .

[38]  A. Ross,et al.  Reactivity of HO2/O−2 Radicals in Aqueous Solution , 1985 .

[39]  J. Brisset,et al.  Combined Effects of Long-Living Chemical Species during Microbial Inactivation Using Atmospheric Plasma-Treated Water , 2010, Applied and Environmental Microbiology.

[40]  S. Kanazawa,et al.  Observation of OH radicals produced by pulsed discharges on the surface of a liquid , 2011 .

[41]  Jae-Hyuk Yu,et al.  Regulation of secondary metabolism in filamentous fungi. , 2005, Annual review of phytopathology.

[42]  E. Choi,et al.  Inactivation of Staphylococcus aureus on the beef jerky by radio-frequency atmospheric pressure plasma discharge treatment , 2014 .

[43]  S. Ying,et al.  A conidial protein (CP15) of Beauveria bassiana contributes to the conidial tolerance of the entomopathogenic fungus to thermal and oxidative stresses , 2011, Applied Microbiology and Biotechnology.

[44]  Jue Zhang,et al.  Inactivation of Bacillus subtilis Spores in Water by a Direct‐Current, Cold Atmospheric‐Pressure Air Plasma Microjet , 2012 .

[45]  Jue Zhang,et al.  Assessment of the roles of various inactivation agents in an argon-based direct current atmospheric pressure cold plasma jet , 2012, ICOPS 2012.

[46]  Jacek Borysow,et al.  Dynamics of OH (X2 Pi , v=0) in high-energy atmospheric pressure electrical pulsed discharge , 1995 .

[47]  E. Choi,et al.  Measurement of Reactive Hydroxyl Radical Species Inside the Biosolutions During Non-thermal Atmospheric Pressure Plasma Jet Bombardment onto the Solution , 2014, Plasma Chemistry and Plasma Processing.

[48]  M. Ammor.  Recent Advances in the Use of Intrinsic Fluorescence for Bacterial Identification and Characterization , 2007, Journal of Fluorescence.

[49]  G. Friedman,et al.  Inactivation of bacteria using dc corona discharge: role of ions and humidity , 2011, New journal of physics.

[50]  Jaroslav Kypr,et al.  Circular dichroism spectroscopy of DNA: from duplexes to quadruplexes. , 2012, Chirality.

[51]  Jaeyoung Park,et al.  The atmospheric-pressure plasma jet: a review and comparison to other plasma sources , 1998 .

[52]  David B. Graves,et al.  Long-term antibacterial efficacy of air plasma-activated water , 2011 .

[53]  S. R. Pombeiro-Sponchiado,et al.  Characterization of melanin pigment produced by Aspergillus nidulans , 2012, World journal of microbiology & biotechnology.

[54]  Aubrey D.N.J. de Grey,et al.  HO2*: the forgotten radical. , 2002 .

[55]  J. Aikens,et al.  Perhydroxyl radical (HOO.) initiated lipid peroxidation. The role of fatty acid hydroperoxides. , 1991, The Journal of biological chemistry.

[56]  P. Bruggeman,et al.  Hydrogen Peroxide Production in an Atmospheric Pressure RF Glow Discharge: Comparison of Models and Experiments , 2014, Plasma Chemistry and Plasma Processing.

[57]  Guo-Ping Zhao,et al.  Genomic perspectives on the evolution of fungal entomopathogenicity in Beauveria bassiana , 2012, Scientific Reports.

[58]  S. Reuter,et al.  Atomic oxygen formation in a radio-frequency driven micro-atmospheric pressure plasma jet , 2010 .

[59]  E. Takai,et al.  Molecular mechanism of plasma sterilization in solution with the reduced pH method: importance of permeation of HOO radicals into the cell membrane , 2013 .

[60]  Jing Fang,et al.  Reactive Oxygen Species in a Non-thermal Plasma Microjet and Water System: Generation, Conversion, and Contributions to Bacteria Inactivation—An Analysis by Electron Spin Resonance Spectroscopy† , 2012 .

[61]  Robert R. Alfano,et al.  Stokes shift emission spectroscopy of human tissue and key biomolecules , 2003 .

[62]  Seong Hwan Kim,et al.  Identification and Characterization of Eurotium rubrum Isolated from Meju in Korea , 2009, Mycobiology.

[63]  Mounir Laroussi,et al.  Nonthermal decontamination of biological media by atmospheric-pressure plasmas: review, analysis, and prospects , 2002 .

[64]  S. Moreau,et al.  Low-temperature sterilization using gas plasmas: a review of the experiments and an analysis of the inactivation mechanisms. , 2001, International journal of pharmaceutics.

[65]  David B. Graves,et al.  The emerging role of reactive oxygen and nitrogen species in redox biology and some implications for plasma applications to medicine and biology , 2012 .

[66]  George G. Khachatourians,et al.  Production, formulation and application of the entomopathogenic fungus Beauveria bassiana for insect control: current status. , 1994 .

[67]  Jue Zhang,et al.  A study of oxidative stress induced by non-thermal plasma-activated water for bacterial damage , 2013 .

[68]  W. Nelson,et al.  Steady-State and Decay Characteristics of Protein Tryptophan Fluorescence from Bacteria , 1986 .