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Evaluation of a bipolar ionization device in inactivation of antimicrobial-resistant bacteria, yeast, Aspergillus spp. and human coronavirus

Published:April 19, 2022DOI:https://doi.org/10.1016/j.jhin.2022.04.004

      Summary

      Background

      The efficacy of bipolar ionization in the healthcare setting has yet to be proven. A major limitation of studies sponsored by industry has been the assessment of efficiency within test chambers in which ozone levels are not adequately controlled.

      Aim

      To assess the effectiveness of bipolar ionization against antimicrobial-resistant bacteria, fungi and human coronavirus within a controlled test chamber designed to mitigate the effect of ozone.

      Methods

      Bacteria- and fungi-inoculated gauze pads, and human coronavirus 229E-inoculated stainless steel plates were placed within the vicinity of the AIO-2 bipolar ionizer and left at room temperature (2 h for coronavirus and 4 h for bacteria and fungi).

      Findings

      Four hours of exposure to bipolar ionization showed a 1.23–4.76 log reduction, corresponding to a 94.2–>99.9% colony-forming units/gauze reduction, in Clostridioides difficile, Klebsiella pneumoniae carbapenemase-producing K. pneumoniae, meticillin-resistant Staphylococcus aureus and multi-drug-resistant S. aureus. A 1.2 log 50% tissue culture infectious dose reduction in human coronavirus was observed after 2 h.

      Conclusion

      The assessment of bipolar ionization systems merits further investigation as an infection control measure.

      Keywords

      Introduction

      Since the occurrence of the coronavirus disease 2019 (COVID-19) global pandemic, there has been a growing market for air purification and reduction of surface contamination using air ionization devices [
      • Haiken M.
      Used in large-scale ventilation systems worldwide, bipolar ionization could be a secret weapon in the war against COVID-19.
      ]. The antimicrobial effectiveness of ions has long been controversial. A review by Krueger and Reed concluded that negative and positive ions inhibit growth of micro-organisms [
      • Kreuger A.P.
      • Reed E.J.
      Biological impact of small air ions.
      ]. A study focusing on static charges on fomitic surfaces and deposition of bacteria showed that bipolar ionization resulted in a reduction of bacterial deposition [
      • Meschke S.
      • Smith B.D.
      • Yost M.
      • Miksch R.R.
      • Gefer P.
      • Gehlke S.
      • et al.
      The effect of surface charge, negative and bipolar ionization on the deposition of airborne bacteria.
      ]. In a trial conducted in an intensive care unit, Kerr et al. found that negative air ionizers were associated with a significant decrease in acinetobacter infections as well as patient colonization [
      • Kerr K.G.
      • Beggs C.B.
      • Dean S.G.
      • Thornton J.
      • Donnelly J.K.
      • Todd N.J.
      • et al.
      Air ionisation and colonisation/infection with methicillin-resistant Staphylococcus aureus and Acinetobacter species in an intensive care unit.
      ]. In a recent real-world hospital setting, the implementation of a technology using charged particles reduced healthcare-associated infections (HAIs) by 45% [
      • Ereth M.H.
      • Fine J.
      • Stamattatos F.
      • Mathew B.
      • Hess D.
      • Simpser E.
      Healthcare-associated infection impact with bioaerosol treatment and COVID-19 mitigation measures.
      ].
      Of the various technologies claiming to deactivate bacteria, viruses and fungi, variants of bipolar ionization technology (needle-point, corona discharge, plasma cluster etc.) have enjoyed renewed commercial popularity as a result of the COVID-19 pandemic. In bipolar ionization, positive (H+) and negative (O2-) ions are generated when water molecules are exposed to high-voltage electrodes. The mechanism(s) associated with the biocidal effect of positive and negative ions have not been clearly established. The purported mechanism of the inactivation of micro-organisms and viruses is the clustering of these ions around viruses and micro-organisms, resulting in the formation of OH radicals, which remove hydrogen, and the formation of water vapour, leading to inactivation [].
      While droplet and airborne transmission are considered to be the main routes of exposure in the ongoing COVID-19 pandemic, disinfection of contaminated or potentially contaminated surfaces is also one of the strategies for controlling COVID-19, as it has been shown that severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) can remain viable on different surfaces from hours to a few days [
      • Chin A.W.H.
      • Chu J.T.S.
      • Perea M.R.A.
      • Hui K.P.Y.
      • Yen H.-L.
      • Chan M.C.W.
      • et al.
      Stability of SARS-CoV-2 in different environmental conditions.
      ]. In this study, an independent evaluation of a bipolar ionization technology device against antimicrobial-resistant bacteria, Candida albicans, Aspergillus fumigatus and human coronavirus was performed in a controlled laboratory environment, in which ozone and the concentration of ions were not allowed to accumulate in an enclosed environment. The objective was to assess the potential utility of this technology within a healthcare environment as an adjunct to existing protocols for minimizing HAIs.

      Methods

      Bacteria, fungi and virus tested

      The organisms tested were ATCC strains of meticillin-resistant Staphylococcus aureus (MRSA, ATCC 43300), Clostridioides difficile (ATCC 17857), C. albicans (ATCC 10231), A. fumigatus (ATCC 13073) and human coronavirus 229E (ATCC VR-740), as well as clinical isolates of multi-drug-resistant Pseudomonas aeruginosa (MDRP), multi-drug-resistant Acinetobacter baumannii (MDRAB) and Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae (KPC-KP).

      Bipolar ionizer

      An H-ION CLUSTER Industrial Air Purifier Module (AIO2) (Sudo Premium Engineering, Seoul, Korea) was used in this study.

      Preparation of organisms to be tested

      MRSA, MDRP, MDRAP and KPC-KP were plated on trypticase soy agar and incubated for 18 h at 35 °C. C. difficile was cultured using an anaerobic blood agar plate (Centers for Disease Control and Prevention formulation) with 5% horse blood, and incubated using an AnaeroPak rectangular jar and Anaeropack gas generator (Mitsubishi Gas Chemical, Tokyo, Japan) for 48 h at 35 °C. A. fumigatus was inoculated on potato dextrose agar and cultured at room temperature for 1 week. Human coronavirus 229E was propagated for 3–5 days on MRC-5 cells (JCRB No. B0521) in E-MEM supplemented with 10% fetal bovine serum (FBS). Suspensions of bacterial equivalent to McFarland 0.5 were prepared and diluted 1:150 using physiological saline for a concentration of approximately 1.0 x 106 colony-forming units (cfu)/mL. For A. fumigatus, a spore suspension was prepared and diluted using physiological saline for a spore suspension of approximately 1 x 105 spores/mL. Similarly, a suspension of C. albicans equivalent to approximately 1.0 x 106 cfu/mL was prepared. One millilitre of the prepared bacterial and fungal suspensions was applied to a sterile, 5 cm x 5 cm gauze pad (Kawamoto Corporation, Osaka, Japan) and left at room temperature [
      • Kobayashi I.
      • Katsuse A.
      • Kanesaka I.
      • Enomoto M.
      • Ando H.
      • Kobayashi N.
      • et al.
      Evaluating the microbiocidal effectiveness of a microplasma discharge device against various bacteria including antimicrobial-resistant bacteria and fungi.
      ]. Sterile 1.5 cm x 1.5 cm SUS304 (Japanese SIS standard) stainless steel squares were loaded into sterile Petri dishes using sterile forceps. A 4.3 log10 50% tissue culture infectious dose (TCID50)/mL suspension of human coronavirus 229E tissue culture was applied to the surface of the stainless steel squares and left at room temperature [
      • Pottage T.
      • Garratt I.
      • Onianwa O.
      • Spencer A.
      • Paton S.
      • Verlander N.Q.
      • et al.
      A comparison of persistence of SARS-CoV-2 variants on stainless steel.
      ].

      Test environment

      Based on several other publications on the evaluation of air purification systems, an acrylic chamber was selected as the test environment [
      • Oritz-Grisales P.
      • Patino-Murillo J.
      • Duque-Grisales E.
      Computational models for reducing air pollution through the generation of negative ions.
      ,]. Testing was performed within a 240-L acrylic chamber that measured 100 cm (W) x 60 cm (D) x 40 cm (H). The acrylic chamber was placed within a Class II (Type A1) biological safety cabinet (BSC).
      As ozone is heavier than air and therefore tends to sink, the inoculated gauze and steel plates were placed on a platform 30 cm higher than the floor of the acrylic chamber. In order to prevent the accumulation of ozone and ions within the acrylic chamber, the acrylic chamber was raised 2 cm above the BSC workspace to pull air out of the chamber. A Model 1200 ozone counter (Dairec, Inc., Kurashiki City, Japan) was placed within the acrylic chamber to monitor ozone levels within the negative pressure environment. The temperature within the test chamber was 21–22 °C and 38–50% relative (Figure 1).

      Measuring the effectiveness of bipolar ionization

      The bacteria- and fungi-inoculated gauze pads, and the human coronavirus 229E-inoculated stainless steel plates were placed within the vicinity of the AIO-2 bipolar ionizer and left at room temperature for up to 4 h. The distance between the gauze pads or stainless steel plates and the AIO-2 bipolar ionizer was 35 cm. As a control, inoculated gauze pads and stainless steel plates were placed within another acrylic chamber without the AIO-2 ionizer.
      After 4 h, bacteria and fungi from each of the gauze pads were extracted by immersion in tubes containing 10 mL of sterile physiological saline and vortexed. Ten-fold serial dilutions of the suspensions were performed using sterile physiological saline, after which 0.1 mL of each of the dilutions was inoculated using a spreader on to either trypticase soy agar, anaerobic blood agar plate with 5% horse blood or potato dextrose agar depending on the organism. Following incubation, colony counts of three replicates were performed to determine the mean colony counts.
      As the infectivity of human coronavirus 229E decreases significantly after drying on various surfaces, exposure to bipolar ionization was for 2 h [
      • Sizun J.
      • Yu M.W.N.
      • Talbot P.J.
      Survival of human coronaviruses 229E and OC43 in suspension and after drying on surfaces: a possible source of hospital-acquired infections.
      ]. The inoculated stainless steel squares were then retrieved, immersed immediately in 2 mL of E-MEM without 10% FBS and vortexed, followed by serial 10-fold dilution using E-MEM without 10% FBS. Quantitation of viable human coronavirus 229E was performed using 96-well tissue culture microtitre plates that were seeded with MRC-5 cells. After washing the tissue culture microtitre plates with E-MEM without 10% FBS, 100 μL of the E-MEM used to elute inoculated virus on stainless steel plates was inoculated on to MRC-5 cells. After allowing for absorption for 1 h at 37 °C under 5% CO2, E-MEM without 10% FBS was removed from the tissue culture microtitre plates, and 100 μL of E-MEM containing 2% FBS, 100 U/mL of penicillin and 100 U/mL streptomycin was dispensed into each well and incubated for 7 days at 37 °C at 5% CO2. Following incubation, the cell culture media was removed from the microtitre trays, and MRC-5 cells were exposed to 70% ethanol for 20 min. Following fixation, MRC-5 cells were observed for cytopathic effect to determine TCID50 using the Behrens–Karber method.
      Measurement of the effectiveness of bipolar ionization was performed in triplicate.

      Log10 cfu/gauze and TCID50/mL reduction following exposure to bipolar ionization

      Log reduction between controls and post-exposure was calculated using the following equation: Log10 (A/B), where A is cfu/mL or TCID50/mL after treatment and B is cfu/mL or TCID50/mL before treatment [

      Microchem Laboratory. Log and percent reductions in microbiology and antimicrobial testing. Round Rock, TX: Microchem Laboratory. Available at:https://microchemlab.com/information/log-and-percent-reductions-microbiology-and-antimicrobial-testing [last accessed March 2022].

      ]. To convert the log reduction to a percentage reduction, % reduction = (1 - 10-L) x 100 was utilized, where L represents log reduction.

      Results

      As shown in Table I, 4 h of exposure to bipolar ionization showed a 1.23–4.76 log reduction, corresponding to a 94–>99.9% cfu/gauze reduction, in C. difficile, KPC-KP, MRSA and MDRP. MDRAB was the only bacterium tested that showed <1 log or 90% cfu/gauze reduction after 4 h. In addition, a <1 log reduction was observed with no significant inactivation of A. fumigatus spores. C. albicans also showed <1 log reduction, as exposure to bipolar ionization led to an 87.3% reduction in cfu/gauze.
      Table ILog reduction of drug-resistant bacteria, Clostridioides difficile and fungi after 4 h of exposure to bipolar ionization
      OrganismOzone (ppm)Ion exposurecfu/gauze (mean of three samples)Log reduction compared with control at 4 h% reduction
      01 h2 h4 h
      MRSA0.044–0.055+1.7 × 1061.5 × 1052.3 × 1042.2199.4
      -3.7 × 1064.7 × 1063.3 × 1063.7 × 106
      MDRP0.055–0.066+2.7 × 1062.0 × 104<1.0 × 1024.76>99.9
      -8.7 × 1061.3 × 1074.0 × 1065.7 × 106
      Clostridioides difficile0.044+1.3 × 1041.4 × 1039.3 × 1021.2394.5
      -7 × 1055.7 × 1042.7 × 1041.7 × 104
      KPC-producing Klebsiella pneumoniae0.022–0.044+8.7 × 1063.7 x1062.8 × 1051.6998.0
      -1.1 × 1077.0 × 1078.0 × 1061.4 × 107
      Multi-drug-resistant Acinetobacter baumannii0.022–0.044+4.3 × 1061.0 × 1066.3 × 1050.6376.7
      -6.7 × 1064.3 × 1064.3 × 1062.7 × 106
      Candida albicans0.022+5.7 × 1063.2 × 1064.7 × 1050.8987.3
      -5 × 1066.0 × 1063.3 × 1063.7 × 106
      Aspergillus fumigatus0.033–0.055+Not tested1.7 × 1062.3 × 1060.1932.4
      -3.6 × 106Not tested2.7 × 1063.4 × 106
      cfu, colony-forming units; MRSA, meticillin-resistant Staphylococcus aureus; MDRP, multi-drug-resistant Pseudomonas aeruginosa; KPC, Klebsiella pneumoniae carbapenemase.
      Two hours of exposure to positive and negative ions led to a 1.2 log reduction in human coronavirus 229E, corresponding to a 94.0% TCID50 reduction (Table 2).
      Table IILog10 50% tissue culture infectious dose (TCID50) reduction of human coronavirus 229E after 2 h of exposure to bipolar ionization
      Human coronavirus 229E (ATCC VR-740)Ozone (ppm)Log10 TCID50/mLLog10 reduction%TCID50/mL reduction compared with control
      ControlAfter 2 h of exposure
      0.0442.71.51.294.0
      During the triplicate testing of the organisms, ozone ppm was <0.055 except for one experiment in which the ozone ppm was 0.066. The ozone concentration was monitored continuously during bipolar ionization exposure, and did not show fluctuation as the air within the acrylic chamber was pulled out continuously. Ozone also served as a surrogate for positive and negative ions to ensure that the concentration of ions within the chamber was constant.

      Discussion

      Since the beginning of the COVID-19 pandemic, a number of hospitals, schools, casinos and sports arenas have implemented variants of bipolar ionization as a technology to disinfect air and surfaces [

      Weir K. Stark Country schools invest in air cleaning technology to prevent the spread of COVID-19. The Repository. Available at: https://www.cantonrep.com/story/news/2022/02/06/stark-schools-invest-bipolar-ionization-air-ventilation-technology/9298983002/ [last accessed March 2022].

      ]. However, one manufacturer is currently the subject of a class action lawsuit which claims that the manufacturer falsely claimed that its technology was effective against SARS-CoV-2 [

      In the United States District Court for the District of Delaware. Plaintiff v. Global Plasma Solutions. Available at: https://www.classaction.org/media/garner-v-global-plasma-solutions-inc.pdf [last accessed March 2022].

      ]. The suit cites Boeing's technical assessment of bipolar ionization, which concluded that the manufacturer's technology cannot clean the air at the level claimed by independent testing.
      Although bipolar ionization technology has been around for decades, the lack of many rigorous peer-reviewed studies makes it difficult to assess the effectiveness of this technology in air and surface disinfection. Many of the claims of manufacturers are based on either in-house studies or external studies designed and guided by the manufacturer. A major confounding variable has been the performance of evaluations within an enclosed environment, in which the concentration of ions as well as ozone increases significantly, making it difficult to determine whether the decrease in viability of micro-organisms and viruses was due to the effect of ozone or ion concentrations that are far in excess of achievable levels in a real-world setting. A study used by an electronics company to promote the effectiveness of its plasma cluster bipolar ionization technology in reducing the concentration of aerosolized SARS-CoV-2 was conducted in a 3-L enclosed chamber [
      Sharp
      Press release: Plasmacluster technology demonstrates effectiveness in reducing airborne novel coronavirus (SARS-CoV-2), a world first.
      ].
      The Occupational Safety and Health Administration has set official exposure limits for ozone at ≤0.1 ppm average over an 8-h period, which is somewhat higher than the US Environmental Protection Agency's (EPA) position that ozone output of indoor medical devices should be ≤0.05 ppm [

      Centers for Disease Control and Prevention, National Institute for Occupational Safety and Health. Ozone. Atlanta, GA: CDC. Available at: https://www.cdc.gov/niosh/idlh/10028156.html [last accessed March 2022].

      ]. In this study, ozone levels were well below 0.1 ppm and, with the exception of one reading of 0.066 ppm, ozone levels were within EPA guidelines. Furthermore, exposure of the samples was minimized by elevating the testing platform 30 cm above the acrylic chamber floor, as ozone tends to sink rather than rise. Effective ozone concentrations for micro-organisms have been reported to be 0.23–2.29 ppm for bacteria, 3–5 ppm for moulds, 0.02–0.26 for fungi, and 0.2–4.1 ppm for viruses [
      • Cristiano L.
      Could ozone be an effective disinfection measure against the novel coronavirus (SARS-CoV-2)?.
      ,
      • Kim J.G.
      • Yousef A.E.
      • Dave S.
      Application of ozone for enhancing the microbiological safety and quality of foods: a review.
      ,
      • Kowalski W.J.
      • Bahnfleth W.P.
      • Whittam T.S.
      Bactericidal effects of high airborne ozone concentrations on Escherichia coli and Staphylococcus aureus.
      ]. While the effect of ozone on the micro-organisms tested in this study cannot be completely ruled out, the ozone levels measured were below levels reported to inactivate micro-organisms.
      The study results showed a 94.4–99.9% log cfu/gauze decrease within 4 h for C. difficile, MDRP, MRSA and KPC-KP. As these bacteria are important pathogens associated with HAIs and are found in the healthcare environment, bipolar ionization merits further examination as a technology to minimize transmission of infections. The relatively low inactivation of MDRA conflicts with a previous study that showed a significant decrease in acinetobacter infections as well as patient colonization in an intensive care unit during a 5.5-month period using a negative air ionization technology [
      • Kerr K.G.
      • Beggs C.B.
      • Dean S.G.
      • Thornton J.
      • Donnelly J.K.
      • Todd N.J.
      • et al.
      Air ionisation and colonisation/infection with methicillin-resistant Staphylococcus aureus and Acinetobacter species in an intensive care unit.
      ]. A longer exposure to bipolar ionization in the present study may have demonstrated higher inactivation of MDRA. The testing environment in this study does not reflect a hospital ward or room; however, it would be difficult to conduct an experiment in the general environment, which would introduce other variables such as contamination from environmental micro-organisms. The ultimate usefulness of this technology may need to be evaluated in a pre- and post-intervention study to compare HAI rates.
      It has been estimated that one in 25 hospitalized patients in the USA develops an infection associated with hospital care; furthermore, additional infections are seen in other healthcare settings [

      Centers for Disease Control and Prevention. Winnable battles. Atlanta, GA: CDC. Available at: https://www.cdc.gov/winnablebattles/report/hais.html [last accessed March 2022].

      ]. According to the World Health Organization, seven and 15 patients, respectively, develop at least one HAI in developed and developing countries [

      World Health Organization. WHO Launches first ever global report on infection prevention and control. Geneva: WHO. Available at: https://www.who.int/news/item/06-05-2022-who-launches-first-ever-global-report-on-infection-prevention-and-control [accessed 12 May 2022].

      ]. The relative contribution of fomites and droplets or aerosals in disease transmission in the healthcare setting is not clear. Beyond currently established protocols, such as personal protective equipment, aseptic technique, hand hygiene, environmental cleanliness, etc. to minimize HAIs, bipolar ionization systems to further reduce the risk of HAIs merit assessment of effectiveness as HAIs continue to occur despite the implementation of these infection control measures.

      Acknowledgements

      The authors wish to acknowledge the support of Mr. Morimitsu Tachibana of BioNavi Corporation, Ltd in facilitating this research in the Department of Infection Control and Prevention laboratory.

      Author contributions

      IK: preparation, review and submission of the manuscript.
      IK, AK and HT: data collection and data analysis.
      IK and HT: data analysis, and review and submission of the manuscript.

      Conflict of interest statement

      None declared.

      Funding sources

      None.

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