Thank you for visiting Nature.com. The browser version you are using has limited CSS support. For the best experience, we recommend that you use an updated browser (or disable Compatibility Mode in Internet Explorer). In the meantime, to ensure continued support, we will render the site without styles and JavaScript.
A contaminated health care environment plays an important role in the spread of multidrug-resistant (MDR) organisms and C. difficile. The purpose of this study was to evaluate the effect of ozone produced by a dielectric barrier discharge (DBD) plasma reactor on the action of vancomycin-resistant Enterococcus faecalis (VRE), carbapenem-resistant Klebsiella pneumoniae (CRE), carbapenem-resistant Antibacterial effects of different materials contaminated with Pseudomonas spp. Pseudomonas aeruginosa (CRPA), carbapenem-resistant Acinetobacter baumannii (CRAB) and Clostridium difficile spores. Various materials contaminated with VRE, CRE, CRPA, CRAB and C. difficile spores were treated with ozone at various concentrations and exposure times. Atomic force microscopy (AFM) demonstrated surface modification of bacteria after ozone treatment. When a dose of 500 ppm ozone was applied to VRE and CRAB for 15 minutes, a decrease of approximately 2 or more log10 was observed in stainless steel, fabric and wood, and a decrease of 1-2 log10 was observed in glass and plastic. C. difficile spores were found to be more resistant to ozone than all other organisms tested. On AFM, after treatment with ozone, bacterial cells swelled and deformed. The ozone produced by the DBD Plasma Reactor is a simple and valuable decontamination tool for MDRO and C. difficile spores, which are known to be common pathogens of healthcare-associated infections.
The emergence of multidrug-resistant (MDR) organisms is caused by the misuse of antibiotics in humans and animals and has been identified by the World Health Organization (WHO) as a major threat to public health1. In particular, healthcare institutions are increasingly confronted with the emergence and spread of MROs. The main MROs are methicillin-resistant Staphylococcus aureus and vancomycin-resistant enterococcus (VRE), extended-spectrum beta-lactamase-producing enterobacteria (ESBL), multidrug-resistant Pseudomonas aeruginosa, multidrug-resistant Acinetobacter baumannii, and carbapenem-resistant Enterobacter (CRE). In addition, Clostridium difficile infection is a leading cause of healthcare-associated diarrhea, placing a significant burden on the health care system. MDRO and C. difficile are transmitted through the hands of healthcare workers, contaminated environments, or directly from person to person. Recent studies have shown that contaminated environments in health care settings play an important role in the transmission of MDRO and C. difficile when health workers (HCWs) come into contact with contaminated surfaces or when patients come into direct contact with contaminated surfaces 3,4. contaminated environments in health care settings reduce the incidence of MLRO and C. difficile infection or colonization5,6,7. Given the global concern about the rise of antimicrobial resistance, it is clear that more research is needed on methods and procedures for decontamination in healthcare settings. Recently, non-contact terminal cleaning methods, especially ultraviolet (UV) equipment or hydrogen peroxide systems, have been recognized as promising methods of decontamination. However, these commercially available UV or hydrogen peroxide devices are not only expensive, UV disinfection is only effective on exposed surfaces, while hydrogen peroxide plasma disinfection requires a relatively long decontamination time before the next disinfection cycle5.
Ozone has known anti-corrosion properties and can be produced inexpensively8. It is also known to be toxic to human health, but can rapidly decompose into oxygen 8. Dielectric barrier discharge (DBD) plasma reactors are by far the most common ozone generators9. DBD equipment allows you to create low-temperature plasma in the air and produce ozone. Until now, the practical use of ozone has been mainly limited to the disinfection of swimming pool water, drinking water and sewage10. Several studies have reported its use in healthcare settings8,11.
In this study, we used a compact DBD plasma ozone generator to demonstrate its effectiveness in clearing MDRO and C. difficile, even those inoculated on various materials commonly used in medical settings. In addition, the ozone sterilization process has been elucidated using atomic force microscopy (AFM) images of ozone-treated cells.
Strains were obtained from clinical isolates of: VRE (SCH 479 and SCH 637), carbapenem-resistant Klebsiella pneumoniae (CRE; SCH CRE-14 and DKA-1), carbapenem-resistant Pseudomonas aeruginosa (CRPA; 54 and 83) and carbapenem-resistant bacteria. bacteria Pseudomonas aeruginosa (CRPA; 54 and 83). resistant Acinetobacter baumannii (CRAB; F2487 and SCH-511). C. difficile was obtained from the National Pathogen Culture Collection (NCCP 11840) of the Korea Agency for Disease Control and Prevention. It was isolated from a patient in South Korea in 2019 and found to belong to ST15 using multilocus sequence typing. Brain Heart Infusion (BHI) Broth (BD, Sparks, MD, USA) inoculated with VRE, CRE, CRPA and CRAB was mixed well and incubated at 37° C. for 24 hours.
C. difficile was streaked anaerobically on blood agar for 48 hours. Several colonies were then added to 5 ml of brain heart broth and incubated under anaerobic conditions for 48 hours. After that, the culture was shaken, 5 ml of 95% ethanol was added, shaken again and left at room temperature for 30 minutes. After centrifugation at 3000 g for 20 minutes, discard the supernatant and suspend the pellet containing spores and killed bacteria in 0.3 ml of water. Viable cells were counted by spiral seeding of the bacterial cell suspension onto blood agar plates after appropriate dilution. Gram staining confirmed that 85% to 90% of the bacterial structures were spores.
The following study was conducted to investigate the effects of ozone as a disinfectant on various surfaces contaminated with MDRO and C. difficile spores, which are known to cause healthcare-associated infections. Prepare samples of stainless steel, fabric (cotton), glass, plastic (acrylic), and wood (pine) measuring one centimeter by one centimeter. Disinfect coupons before use. All samples were sterilized by autoclaving prior to infection with bacteria.
In this study, bacterial cells were spread on various substrate surfaces as well as on agar plates. The panels are then sterilized by exposing them to ozone for a certain period of time and at a certain concentration in a sealed chamber. On fig. 1 is a photograph of ozone sterilization equipment. DBD plasma reactors were fabricated by attaching perforated and exposed stainless steel electrodes to the front and back of 1 mm thick alumina (dielectric) plates. For perforated electrodes, the aperture and hole area were 3 mm and 0.33 mm, respectively. Each electrode has a round shape with a diameter of 43 mm. A high voltage high frequency power supply (GBS Elektronik GmbH Minipuls 2.2) was used to apply a sinusoidal voltage of approximately 8 kV peak to peak at a frequency of 12.5 kHz to the perforated electrodes to generate plasma at the edges of the electrodes. perforated electrodes. Since the technology is a gas sterilization method, sterilization is carried out in a chamber divided by volume into upper and lower compartments, which contain bacterially contaminated samples and plasma generators, respectively. The top compartment has two valve ports to remove and vent residual ozone. Before using in the experiment, the change in time of the ozone concentration in the room after turning on the plasma installation was measured according to the absorption spectrum of the spectral line of 253.65 nm of a mercury lamp.
(a) Scheme of an experimental setup for sterilization of bacteria on various materials using ozone generated in the DBD plasma reactor, and (b) ozone concentration and plasma generation time in the sterilization chamber. Figure was made using OriginPro version 9.0 (OriginPro software, Northampton, MA, USA; https://www.originlab.com).
First, by sterilizing bacterial cells placed on agar plates with ozone, while changing the ozone concentration and treatment time, the appropriate ozone concentration and treatment time for decontamination of MDRO and C. difficile were determined. During the sterilization process, the chamber is first purged with ambient air and then filled with ozone by turning on the plasma unit. After the samples have been treated with ozone for a predetermined period of time, a diaphragm pump is used to remove the remaining ozone. The measurements used a sample of a complete 24-hour culture (~ 108 CFU/ml). Samples of suspensions of bacterial cells (20 μl) were first serially diluted ten times with sterile saline, and then these samples were distributed on agar plates sterilized with ozone in the chamber. After that, repeated samples, consisting of samples exposed and not exposed to ozone, were incubated at 37°C for 24 hours and counted colonies to evaluate the effectiveness of sterilization.
Further, according to the sterilization conditions defined in the above study, the decontamination effect of this technology on MDRO and C. difficile was evaluated using coupons of various materials (stainless steel, fabric, glass, plastic and wood coupons) commonly used in medical institutions. Complete 24 hour cultures (~108 cfu/ml) were used. Samples of bacterial cell suspension (20 μl) were serially diluted ten times with sterile saline, and then the coupons were immersed in these diluted broths to assess contamination. Samples removed after immersion in dilution broth were placed in sterile Petri dishes and dried at room temperature for 24 hours. Place the petri dish lid on the sample and carefully place it into the test chamber. Remove the lid from the Petri dish and expose the sample to 500 ppm ozone for 15 minutes. Control samples were placed in a biological safety cabinet and were not exposed to ozone. Immediately after exposure to ozone, samples and non-irradiated samples (ie controls) were mixed with sterile saline using a vortex mixer to isolate bacteria from the surface. The eluted suspension was serially diluted 10 times with sterile saline, after which the number of diluted bacteria was determined on blood agar plates (for aerobic bacteria) or anaerobic blood agar plates for Brucella (for Clostridium difficile) and incubated at 37°C for 24 hours . or under anaerobic conditions for 48 hours at 37°C in duplicate to determine the initial concentration of the inoculum. The difference in bacterial counts between unexposed controls and exposed samples was calculated to give a log reduction in bacterial counts (ie, sterilization efficiency) under test conditions.
Biological cells must be immobilized on an AFM imaging plate; therefore, a flat and uniformly rough mica disk with a roughness scale smaller than the cell size is used as a substrate. The diameter and thickness of the disks were 20 mm and 0.21 mm, respectively. To firmly anchor the cells to the surface, the surface of the mica is coated with poly-L-lysine (200 µl), making it positively charged and the cell membrane negatively charged. After coating with poly-L-lysine, the mica disks were washed 3 times with 1 ml deionized (DI) water and air dried overnight. Then, the bacterial cells were applied to the mica surface coated with poly-L-lysine by dosing a dilute bacterial solution, left for 30 min, and then the mica surface was washed with 1 ml of deionized water.
Half of the samples were treated with ozone and the surface morphology of mica plates loaded with VRE, CRAB and C. difficile spores was visualized using AFM (XE-7, park systems). The AFM mode of operation is set to tapping mode, which is a common method for imaging biological cells. In the experiments, a microcantilever designed for non-contact mode (OMCL-AC160TS, OLYMPUS Microscopy) was used. AFM images were recorded based on a probe scan rate of 0.5 Hz resulting in an image resolution of 2048 × 2048 pixels.
To determine the conditions under which DBD plasma reactors are effective for sterilization, we conducted a series of experiments using both MDRO (VRE, CRE, CRPA, and CRAB) and C. difficile to vary ozone concentration and exposure time. On fig. 1b shows the ozone concentration time curve for each test condition after turning on the plasma device. The concentration increased logarithmically, reaching 300 and 500 ppm after 1.5 and 2.5 minutes, respectively. Preliminary tests with VRE have shown that the minimum required to effectively decontaminate bacteria is 300 ppm ozone for 10 minutes. Thus, in the following experiments, MDRO and C. difficile were exposed to ozone at two different concentrations (300 and 500 ppm) and at two different exposure times (10 and 15 minutes). Sterilization efficiency for each ozone dose and exposure time setting was calculated and shown in Table 1. Exposure to 300 or 500 ppm ozone for 10–15 minutes resulted in an overall reduction in VRE of 2 or more log10. This high level of bacterial kill with CRE was achieved with 15 minutes of exposure to 300 or 500 ppm ozone. High reduction in CRPA (> 7 log10) were achieved with exposure to 500 ppm of ozone for 15 min. High reduction in CRPA (> 7 log10) were achieved with exposure to 500 ppm of ozone for 15 min. Высокое снижение CRPA (> 7 log10) было достигнуто при воздействии 500 частей на миллион озона в течение 15 минут. A high reduction in CRPA (> 7 log10) was achieved with exposure to 500 ppm ozone for 15 minutes.暴露于500 ppm 的臭氧15 分钟后，可大幅降低CRPA (> 7 log10)。暴露于500 ppm 的臭氧15 分钟后，可大幅降低CRPA (> 7 log10)。 Существенное снижение CRPA (> 7 log10) после 15-минутного воздействия озона с концентрацией 500 ppm. Significant reduction in CRPA (> 7 log10) after 15 minutes exposure to 500 ppm ozone. Negligible killing of CRAB bacteria at 300 ppm ozone; however, at 500 ppm ozone, there was a > 1.5 log10 reduction. however, at 500 ppm ozone, there was a > 1.5 log10 reduction. однако при концентрации озона 500 частей на миллион наблюдалось снижение > 1,5 log10. however, at an ozone concentration of 500 ppm, a decrease of >1.5 log10 was observed.然而，在500 ppm 臭氧下，减少了> 1.5 log10。然而，在500 ppm 臭氧下，减少了> 1.5 log10。 Однако при концентрации озона 500 частей на миллион наблюдалось снижение >1,5 log10. However, at an ozone concentration of 500 ppm, a decrease of >1.5 log10 was observed. Exposing C. difficile spores to 300 or 500 ppm ozone resulted in a > 2.5 log10 reduction. Exposing C. difficile spores to 300 or 500 ppm ozone resulted in a > 2.5 log10 reduction. Воздействие на споры C. difficile озона с концентрацией 300 или 500 частей на миллион приводило к снижению > 2,5 log10. Exposure of C. difficile spores to 300 or 500 ppm ozone resulted in >2.5 log10 reductions.将艰难梭菌孢子暴露于300 或500 ppm 的臭氧中导致> 2.5 log10 减少。 300 或500 ppm 的臭氧中导致> 2.5 log10 减少。 Воздействие на споры C. difficile озона с концентрацией 300 или 500 частей на миллион приводило к снижению >2,5 log10. Exposure of C. difficile spores to 300 or 500 ppm ozone resulted in >2.5 log10 reductions.
Based on the experiments above, a sufficient requirement was found to inactivate bacteria at a dose of 500 ppm ozone for 15 minutes. VRE, CRAB and C. difficile spores have been tested for the germicidal effect of ozone on a variety of materials including stainless steel, fabric, glass, plastic and wood commonly used in hospitals. Their sterilization efficiency is shown in Table 2. Test organisms were evaluated twice. In VRE and CRAB, ozone was less effective on glass and plastic surfaces, although a log10 reduction of about a factor of 2 or more was observed on stainless steel, fabric and wood surfaces. C. difficile spores were found to be more resistant to ozone treatment than all other organisms tested. To statistically study the effect of ozone on the killing effect of different materials against VRE, CRAB, and C. difficile, t-tests were used to compare differences between the number of CFU per milliliter in the control and experimental groups on different materials (Fig. 2). strains showed statistically significant differences, but more significant differences were observed for VRE and CRAB spores than for C. difficile spores.
Scatterplot of the effects of ozone on bacterial killing of various materials (a) VRE, (b) CRAB, and (c) C. difficile.
AFM imaging was performed on ozone-treated and untreated VRE, CRAB, and C. difficile spores to study in detail the ozone gas sterilization process. On fig. 3a, c and e show AFM images of untreated VRE, CRAB and C. difficile spores, respectively. As seen in the 3D images, the cells are smooth and intact. Figures 3b, d and f show VRE, CRAB and C. difficile spores after ozone treatment. Not only did they decrease in overall size for all cells tested, but their surface became noticeably rougher after exposure to ozone.
AFM images of untreated VRE, MRAB and C. difficile spores (a, c, e) and (b, d, f) treated with 500 ppm ozone for 15 min. Images were drawn using Park Systems XEI version 5.1.6 (XEI Software, Suwon, Korea; https://www.parksystems.com/102-products/park-xe-bio).
Our research shows that the ozone produced by DBD plasma equipment demonstrates the ability to effectively decontaminate MDRO and C. difficile spores, which are known to be major causes of healthcare-associated infections. In addition, in our study, given that environmental contamination with MDRO and C. difficile spores can be a source of healthcare-associated infections, the germicidal effect of ozone was found to be successful for materials primarily used in hospital settings. Decontamination tests were performed using DBD plasma equipment after artificial contamination of materials such as stainless steel, cloth, glass, plastic and wood with MDRO and C. difficile spores. As a result, although the decontamination effect varies depending on the material, the decontamination ability of ozone is remarkable.
Frequently touched objects in hospital rooms require routine, low-level disinfection. The standard method for decontaminating such objects is manual cleaning with a liquid disinfectant such as a quaternary ammonium compound 13. Even with strict adherence to the recommendations for the use of disinfectants, MPO is difficult to remove by traditional environmental cleaning (usually manual cleaning)14. Therefore, new technologies are required, such as non-contact methods. Consequently, there has been interest in gaseous disinfectants, including hydrogen peroxide and ozone10. The advantage of gaseous disinfectants is that they can reach places and objects that traditional manual methods cannot reach. Hydrogen peroxide has recently come into use in medical settings, however hydrogen peroxide itself is toxic and must be handled according to strict handling procedures. Plasma sterilization with hydrogen peroxide requires a relatively long purge time before the next sterilization cycle. In contrast, ozone acts as a broad-spectrum antibacterial agent, effective against bacteria and viruses that are resistant to other disinfectants8,11,15. In addition, ozone can be produced cheaply from atmospheric air and does not require additional toxic chemicals that can leave a harmful footprint in the environment, since it eventually breaks down into oxygen. However, the reason why ozone is not widely used as a disinfectant is as follows. Ozone is toxic to human health, so its concentration does not exceed 0.07 ppm on average for more than 8 hours16, so ozone sterilizers have been developed and put on the market, mainly for cleaning exhaust gases. It is also possible to inhale gas and produce an unpleasant odor after decontamination5,8. Ozone was not actively used in medical institutions. However, ozone can be used safely in sterilization chambers and with proper ventilation procedures, and its removal can be greatly accelerated by using a catalytic converter. In this study, we demonstrate that plasma ozone sterilizers can be used for disinfection in healthcare settings. We have developed a device with high sterilization capabilities, easy operation and fast service for hospitalized patients. In addition, we have developed a simple sterilization unit that uses ambient air at no additional cost. To date, there is insufficient information on the minimum ozone requirements for MDRO inactivation. The equipment used in our study is easy to set up and has a short run time and is expected to be useful for frequent equipment sterilization.
The mechanism of the bactericidal action of ozone is not completely clear. Several studies have shown that ozone damages bacterial cell membranes, leading to intracellular leakage and eventual cell lysis17,18. Ozone can interfere with cellular enzymatic activity by reacting with thiol groups and can modify purine and pyrimidine bases in nucleic acids. This study visualized the morphology of VRE, CRAB, and C. difficile spores before and after ozone treatment and found that not only did they decrease in size, but they also became significantly rougher on the surface, indicating damage or corrosion of the outermost membrane. and internal materials due to ozone gas has a strong oxidizing ability. This damage can lead to cell inactivation, depending on the severity of the cellular changes.
C. difficile spores are difficult to remove from hospital rooms. The spores remain in the places where they shed 10,20. In addition, in this study, although the maximum logarithmic 10-fold reduction in the number of bacteria on agar plates at 500 ppm ozone for 15 minutes was 2.73, the bactericidal effect of ozone on various materials containing C spores .difficile has been reduced. Therefore, various strategies can be considered to reduce C. difficile infection in health care settings. For use in isolated C. difficile chambers only, it may also be useful to adjust exposure time and intensity of ozone treatment. In addition, we must keep in mind that the ozone decontamination method cannot completely replace conventional manual cleaning with disinfectants and antimicrobial strategies, and can also be very effective in controlling C. difficile 5 . In this study, the effectiveness of ozone as a disinfectant varied for different types of MPO. Efficacy may depend on several factors such as growth stage, cell wall, and efficiency of repair mechanisms21,22. The reason for the different sterilizing effect of ozone on the surface of each material may be due to the formation of a biofilm. Previous studies have shown that E. faecium and E. faecium increase environmental resistance when present as biofilms23, 24, 25. However, this study shows that ozone has a significant bactericidal effect on MDRO and C. difficile spores.
A limitation of our study is that we assessed the effect of ozone retention after remediation. This can lead to an overestimation of the number of viable bacterial cells.
Although this study was conducted to evaluate the effectiveness of ozone as a disinfectant in a hospital setting, it is difficult to generalize our results to all hospital settings. Thus, more research is needed to investigate the applicability and compatibility of this DBD ozone sterilizer in a real hospital environment.
The ozone produced by DBD plasma reactors could be a simple and valuable decontamination agent for MDRO and C. difficile. Thus, ozone treatment can be considered as an effective alternative to disinfection of the hospital environment.
The datasets used and/or analyzed in the current study are available from the respective authors upon reasonable request.
WHO global strategy to contain antimicrobial resistance. https://www.who.int/drugresistance/WHO_Global_Strategy.htm/en/ Available.
Dubberke, ER & Olsen, MA Burden of Clostridium difficile on the healthcare system. Dubberke, ER & Olsen, MA Burden of Clostridium difficile on the healthcare system. Dubberke, E.R. and Olsen, M.A. Burden of Clostridium difficile in the healthcare system. Dubberke, ER & Olsen, MA 艰难梭菌对医疗保健系统的负担。 Dubberke, ER & Olsen, MA Dubberke, E.R. and Olsen, M.A. The burden of Clostridium difficile on the health care system. clinical. Infect. Dis. https://doi.org/10.1093/cid/cis335 (2012).
Boyce, JM Environmental pollution has a significant impact on nosocomial infections. J. Hospital. Infect. 65 (Annex 2), 50-54. https://doi.org/10.1016/s0195-6701(07)60015-2 (2007).
Kim, YA, Lee, H. & K L.,. Kim, YA, Lee, H. & K L.,. Kim, Y.A., Lee, H. and K.L.,. Kim, YA, Lee, H. & K L.,. Kim, YA, Lee, H. & K L.,. Kim, Y.A., Lee, H. and K.L.,. Pollution and infection control of the hospital environment by pathogenic bacteria [J. Korea J. Hospital Infection Control. 20(1), 1-6 (2015).
Dancer, S.J. The fight against nosocomial infections: attention to the role of the environment and new disinfection technologies. clinical. microorganism. open 27(4), 665–690. https://doi.org/10.1128/cmr.00020-14 (2014).
Weber, DJ et al. Effectiveness of UV devices and hydrogen peroxide systems for decontamination of terminal areas: focus on clinical trials. Yes. J. Infection control. 44 (5 additions), e77-84. https://doi.org/10.1016/j.ajic.2015.11.015 (2016).
Siani, H. & Maillard, JY Best practice in healthcare environment decontamination. Siani, H. & Maillard, JY Best practice in healthcare environment decontamination. Siani, H. & Maillard, JY Передовая практика дезактивации среды здравоохранения. Siani, H. & Maillard, JY Good practice in decontamination of healthcare environments. Siani, H. & Maillard, JY 医疗环境净化的最佳实践。 Siani, H. & Maillard, JY The best practice of medical environment purification. Siani, H. & Maillard, JY Передовой опыт обеззараживания медицинских учреждений. Siani, H. & Maillard, JY Best practice in decontamination of medical facilities. EURO. J. Clin. microorganism To infect Dis. 34(1), 1-11. https://doi.org/10.1007/s10096-014-2205-9 (2015).
Sharma, M. & Hudson, JB Ozone gas is an effective and practical antibacterial agent. Sharma, M. & Hudson, JB Ozone gas is an effective and practical antibacterial agent. Sharma, M. and Hudson, J. B. Gaseous ozone is an effective and practical antibacterial agent. Sharma, M. & Hudson, JB 臭氧气体是一种有效且实用的抗菌剂。 Sharma, M. & Hudson, JB Sharma, M. and Hudson, J. B. Gaseous ozone is an effective and practical antimicrobial agent. Yes. J. Infection. control. 36(8), 559-563. https://doi.org/10.1016/j.ajic.2007.10.021 (2008).
Seung-Lok Pak, J.-D.M., Lee, S.-H. & Shin, S.-Y. & Shin, S.-Y. and Shin, S.-Yu. & Shin, S.-Y. & Shin, S.-Y. and Shin, S.-Yu. Ozone is efficiently generated using grid plate electrodes in a discharge-type ozone generator with a dielectric barrier. J. Electrostatics. 64(5), 275-282. https://doi.org/10.1016/j.elstat.2005.06.007 (2006).
Moat, J., Cargill, J., Shone, J. & Upton, M. Application of a novel decontamination process using gaseous ozone. Moat, J., Cargill, J., Shone, J. & Upton, M. Application of a novel decontamination process using gaseous ozone. Moat J., Cargill J., Sean J. and Upton M. Application of a new decontamination process using ozone gas. Moat, J., Cargill, J., Shone, J. & Upton, M. 使用气态臭氧的新型净化工艺的应用。 Moat, J., Cargill, J., Shone, J. & Upton, M. Moat J., Cargill J., Sean J. and Upton M. Application of a new purification process using ozone gas. Can. J. Microorganisms. 55(8), 928–933. https://doi.org/10.1139/w09-046 (2009).
Zoutman, D., Shannon, M. & Mandel, A. Effectiveness of a novel ozone-based system for the rapid high-level disinfection of health care spaces and surfaces. Zoutman, D., Shannon, M. & Mandel, A. Effectiveness of a novel ozone-based system for the rapid high-level disinfection of health care spaces and surfaces. Zutman, D., Shannon, M. and Mandel, A. Efficiency of a new ozone-based system for rapid, high-level disinfection of medical environments and surfaces. Zoutman, D., Shannon, M. & Mandel, A. 新型臭氧系统对医疗保健空间和表面进行快速高水平消毒的有效性。 Zoutman, D., Shannon, M. & Mandel, A. Zutman, D., Shannon, M. and Mandel, A. Efficacy of a new ozone system for rapid, high-level disinfection of medical environments and surfaces. Yes. J. Infection control. 39(10), 873-879. https://doi.org/10.1016/j.ajic.2011.01.012 (2011).
Wullt, M., Odenholt, I. & Walder, M. Activity of three disinfectants and acidified nitrite against Clostridium difficile spores. Wullt, M., Odenholt, I. & Walder, M. Activity of three disinfectants and acidified nitrite against Clostridium difficile spores. Woollt, M., Odenholt, I. and Walder, M. Activity of three disinfectants and acidified nitrite against Clostridium difficile spores. Vullt M, Odenholt I and Walder M. Activity of three disinfectants and acidified nitrites against Clostridium difficile spores. Infection Control Hospital. Epidemiology. 24(10), 765-768. https://doi.org/10.1086/502129 (2003).
Ray, A. et al. Vaporized hydrogen peroxide decontamination during an outbreak of multidrug-resistant Acinetobacter baumannii in a long-term care hospital. Infection Control Hospital. Epidemiology. 31(12), 1236-1241. https://doi.org/10.1086/657139 (2010).
Ekshtein, B.K. et al. Reduction of contamination of environmental surfaces with Clostridium difficile and vancomycin-resistant enterococci following the adoption of measures to improve cleaning methods. Infectious disease of the Navy. 7, 61. https://doi.org/10.1186/1471-2334-7-61 (2007).
Martinelli, M., Giovannangeli, F., Rotunno, S., Trombetta, CM & Montomoli, E. Water and air ozone treatment as an alternative sanitizing technology. Martinelli, M., Giovannangeli, F., Rotunno, S., Trombetta, CM & Montomoli, E. Water and air ozone treatment as an alternative sanitizing technology. Martinelli, M., Giovannangeli, F., Rotunno, S., Trombetta, K.M. and Montomoli, E. Ozone treatment of water and air as an alternative sanitation technology. Martinelli, M., Giovannangeli, F., Rotunno, S., Trombetta, CM & Montomoli, E. 水和空气臭氧处理作为替代消毒技术。 Martinelli, M., Giovannangeli, F., Rotunno, S., Trombetta, CM & Montomoli, E. Martinelli M, Giovannangeli F, Rotunno S, Trombetta SM and Montomoli E. Ozone treatment of water and air as an alternative method of disinfection. J. Previous page. medicine. Hagrid. 58(1), E48-e52 (2017).
Korean Ministry of the Environment. https://www.me.go.kr/mamo/web/index.do?menuId=586 (2022). As of January 12, 2022
Thanomsub, B. et al. Effect of ozone treatment on bacterial cell growth and ultrastructural changes. Appendix J. Gen. microorganism. 48(4), 193-199. https://doi.org/10.2323/jgam.48.193 (2002).
Zhang, YQ, Wu, QP, Zhang, JM & Yang, XH Effects of ozone on membrane permeability and ultrastructure in Pseudomonas aeruginosa. Zhang, YQ, Wu, QP, Zhang, JM & Yang, XH Effects of ozone on membrane permeability and ultrastructure in Pseudomonas aeruginosa. Zhang, YQ, Wu, QP, Zhang, JM & Yang, XH Влияние озона на проницаемость мембран и ультраструктуру Pseudomonas aeruginosa. Zhang, YQ, Wu, QP, Zhang, JM & Yang, XH Effect of ozone on membrane permeability and ultrastructure of Pseudomonas aeruginosa. Zhang, YQ, Wu, QP, Zhang, JM & Yang, XH 臭氧对铜绿假单胞菌膜通透性和超微结构的影响。 Zhang, YQ, Wu, QP, Zhang, JM & Yang, XH Zhang, YQ, Wu, QP, Zhang, JM & Yang, XH Влияние озона на проницаемость мембран и ультраструктуру Pseudomonas aeruginosa. Zhang, YQ, Wu, QP, Zhang, JM & Yang, XH Effect of ozone on membrane permeability and ultrastructure of Pseudomonas aeruginosa. J. Application. microorganism. 111(4), 1006-1015. https://doi.org/10.1111/j.1365-2672.2011.05113.x (2011).
Russell, A.D. Similarities and differences in microbial responses to fungicides. J. Antibiotics. chemotherapy. 52(5), 750-763. https://doi.org/10.1093/jac/dkg422 (2003).
Whitaker, J., Brown, BS, Vidal, S. & Calcaterra, M. Designing a protocol that eliminates Clostridium difficile: A collaborative venture. Whitaker, J., Brown, BS, Vidal, S. & Calcaterra, M. Designing a protocol that eliminates Clostridium difficile: A collaborative venture. Whitaker J, Brown BS, Vidal S and Calcaterra M. Development of a protocol to eliminate Clostridium difficile: a joint venture. Whitaker, J., Brown, BS, Vidal, S. & Calcaterra, M. 设计一种消除艰难梭菌的方案：合作企业。 Whitaker, J., Brown, BS, Vidal, S. & Calcaterra, M. Whitaker, J., Brown, B.S., Vidal, S. and Calcaterra, M. Development of a protocol to eliminate Clostridium difficile: a joint venture. Yes. J. Infection control. 35(5), 310-314. https://doi.org/10.1016/j.ajic.2006.08.010 (2007).
Broadwater, WT, Hoehn, RC & King, PH Sensitivity of three selected bacterial species to ozone. Broadwater, WT, Hoehn, RC & King, PH Sensitivity of three selected bacterial species to ozone. Broadwater, WT, Hoehn, RC & King, PH Чувствительность трех выбранных видов бактерий к озону. Broadwater, WT, Hoehn, RC & King, PH Ozone sensitivity of three selected bacterial species. Broadwater, WT, Hoehn, RC & King, PH 三种选定细菌对臭氧的敏感性。 Broadwater, WT, Hoehn, RC & King, PH Broadwater, WT, Hoehn, RC & King, PH Чувствительность трех выбранных бактерий к озону. Broadwater, WT, Hoehn, RC & King, PH Ozone sensitivity of three selected bacteria. statement. microorganism. 26(3), 391–393. https://doi.org/10.1128/am.26.3.391-393.1973 (1973).
Patil, S., Valdramidis, VP, Karatzas, KA, Cullen, PJ & Bourke, P. Assessing the microbial oxidative stress mechanism of ozone treatment through the responses of Escherichia coli mutants. Patil, S., Valdramidis, VP, Karatzas, KA, Cullen, PJ & Bourke, P. Assessing the microbial oxidative stress mechanism of ozone treatment through the responses of Escherichia coli mutants. Patil, S., Valdramidis, V.P., Karatzas, K.A., Cullen, P.J. and Burk, P. Evaluation of the Mechanism of Microbial Oxidative Stress by Ozone Treatment from Escherichia coli Mutant Reactions. Patil, S., Valdramidis, VP, Karatzas, KA, Cullen, PJ & Bourke, P. 通过大肠杆菌突变体的反应评估臭氧处理的微生物氧化应激机制。 Patil, S., Valdramidis, VP, Karatzas, KA, Cullen, PJ & Bourke, P. Patil, S., Valdramidis, V.P., Karatsas, K.A., Cullen, P.J. and Bourque, P. Evaluation of mechanisms of microbial oxidative stress in ozone treatment through Escherichia coli mutant reactions. J. Application. microorganism. 111(1), 136-144. https://doi.org/10.1111/j.1365-2672.2011.05021.x (2011).
Greene, C., Wu, J., Rickard, AH & Xi, C. Evaluation of the ability of Acinetobacter baumannii to form biofilms on six different biomedical relevant surfaces. Greene, C., Wu, J., Rickard, AH & Xi, C. Evaluation of the ability of Acinetobacter baumannii to form biofilms on six different biomedical relevant surfaces. Green, K., Wu, J., Rickard, A. Kh. and Si, K. Evaluation of the ability of Acinetobacter baumannii to form biofilms on six different biomedically relevant surfaces. Greene, C., Wu, J., Rickard, AH & Xi, C. 评估鲍曼不动杆菌在六种不同生物医学相关表面上形成生物膜的能力。 Greene, C., Wu, J., Rickard, AH & Xi, C. Evaluation of the ability of 鲍曼不动天生在六种 to form biofilm on various biomedical relevant surfaces. Green, K., Wu, J., Rickard, A. Kh. and Si, K. Evaluation of the ability of Acinetobacter baumannii to form biofilms on six different biomedically relevant surfaces. Wright. application microorganism 63(4), 233-239. https://doi.org/10.1111/lam.12627 (2016).