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Efficacy of an antimicrobial surface coating against human coronavirus 229E and SARS-CoV-2

Published:September 06, 2021DOI:https://doi.org/10.1016/j.ajic.2021.08.031

      Highlights

      • A quaternary ammonium polymer coating demonstrates efficacy against coronaviruses.
      • Human coronavirus 229E and SARS-CoV-2 are reduced by >99.9% within 2 hours.
      • Carrier wash method more effective for harvesting viruses than the swab method.

      Abstract

      The COVID-19 pandemic has accelerated the demand for alternatives to standard cleaning and disinfection practices. Antiviral coatingsmay provide an alternative to common surface treatments. A newly developed quaternary ammonium polymer coating was applied to stainless steel coupons and evaluated for efficacy against human coronavirus 229E and SARS-CoV-2. The polymer coating reduced levels of both test viruses by greater than 99.9% relative to non-coated stainless steel coupons during a 2-hour contact time.

      Key Words

      Background

      Respiratory viruses remain viable on inanimate, environmental surfaces (ie, fomites) for extended periods of time depending on temperature, humidity, and other factors.
      • Boone SA
      • Gerba CP
      Significance of fomites in the spread of respiratory disease and enteric viral disease.
      Effective cleaning and surface disinfection practices reduce levels of infectious viruses and the potential for their spread.
      • Sassi HP
      • Sifuentes LY
      • Konig DW
      • et al.
      Control of the spread of viruses in a long-term care facility using hygiene protocols.
      Nevertheless, recontamination can occur at any time following the use of liquid disinfectants, which are typically wiped dry from surfaces after application. A number of self-disinfecting surface formulations have been developed during recent years.
      • Querido MM
      • Neves L
      • Neves P
      • Pereira CS
      • Teixeria JP.
      Self-disinfecting surfaces and infection control.
      A continuously-active quaternary ammonium polymer applied to high-touch surfaces by certified technicians using electrostatic sprayers previously resulted in the reduced spread of antibiotic-resistant bacterial infections in clinical settings.
      • Ellingson KD
      • Pogreba-Brown K
      • Gerba CP
      • Elliott SP
      Impact of a novel antimicrobial surface coating on health care-associated infections and environmental, at 2 urban hospitals.
      The formulation was subsequently modified to increase cationic functionality for more efficient disruption of the lipid bilayers that comprise bacterial membranes and viral envelopes. Human coronavirus 229E (HCoV-229E), a seasonal enveloped respiratory pathogen of humans, has been employed as a surrogate for emergent coronaviruses to assess persistence, survival, and antiviral efficacy.
      • Carraturo F
      • Del Giudice C
      • Morelli M
      • et al.
      Persistence of SARS-CoV-2 in the environment and COVID-19 transmission risk from environmental matrices and surfaces.
      The objective of this study was to assess the antiviral effectiveness of the new formulation following application to stainless steel coupons against SARS-related Coronavirus 2 (SARS-CoV-2) and HCoV-229E.

      Methods

      Acetone-washed, pre-autoclaved stainless-steel coupons (Type 304, unidirectional finish) coated with the reformulated quaternary ammonium polymer (SurfaceWise2) were supplied by Allied BioScience, Inc (Dallas, TX). An electrostatic sprayer mounted onto a robotic slide was used to coat the coupons, with coverage uniformity monitored by an x-ray fluorescence spectrometer. The coating was cured overnight under ambient conditions. Non-coated stainless-steel coupons were used as control surfaces. The carriers were not subjected to wear and abrasion testing prior to experimentation.
      Human coronavirus 229E and the two cell lines used in the study, MRC-5 and Vero E6, were procured from the American Type Culture Collection (ATCC; Manassas, VA). SARS-CoV-2 Isolate USA-WA1/2020 was deposited by the Centers for Disease Control and Prevention and obtained through BEI Resources, The National Institute of Allergies and Infectious Disease (NIAID), and The National Institutes of Health (NIH). All laboratory work involving SARS-CoV-2 was conducted within a Biosafety Level 3 laboratory.
      HCoV-229E (ATCC VR-740) and SARS-CoV-2 (BEI NR-52281) stocks were prepared according to the coronavirus propagation protocol described by Gundy et al. (2009)
      • Gundy PM
      • Gerba CP
      • Pepper IL
      Survival of coronaviruses in water and wastewater.
      using the MRC-5 (ATCC CCL-171) and Vero E6 (ATCC CRL-1586) cell lines, respectively. Stock virus concentrations were determined using the TCID50 (tissue culture infectious dose at the 50% endpoint) assay technique as described by Hierholzer and Killington (1996)
      • Hierholzer JC
      • Killington RA
      • Stokes A
      Virology Methods Manual.
      , and quantified using the Spearman-Karber algorithm.
      Antiviral efficacy of the coating was assessed using the American Society of Testing and Materials E1053-20 Method.

      ASTM International. ASTM E1053-20. Standard practice to assess virucidal activity of chemicals intended for disinfection of inanimate, nonporous environmental surfaces. 2020. Accessed April 15, 2020. https://www.astm.org/Standards/E1053.htm

      Modifications to the method were 1) the use of stainless-steel coupons, and 2) preapplication of the polymer coating to the coupons followed by inoculation of virus. Test 1 against HCoV-229E (Table 1) was performed by inoculating 0.10 mL of stock virus onto polymer-coated and noncoated control coupons (25.81 cm2) mounted into sterile Petri dishes, with spreading over the surface to within ∼0.32 cm of the perimeter edge. The coupons were incubated (22-23°C; 30-50% relative humidity) for 2 hours with the Petri dish lids on. Swabs predipped in Letheen Broth Base (LBB; Neogen Corp., Lansing, MI) were used for product neutralization and to harvest viruses from carriers. The swabs were placed into tubes containing 1 mL of LBB and vortexed vigorously. The suspensions were immediately passed through Sephadex G-10 gel columns by centrifugation (5 minutes at 3,500 × g). Subsequent tests against HCoV-229E (Test 2) and SARS-CoV-2 were modified by the addition of 5% soil in the form of fetal bovine serum (FBS), and by decreasing the carrier size and viral inoculum volume (Table 1). A wash method was also employed by rinsing the carriers 4-5 times using 1 mL of LBB, with further virus detachment using a cell scraper. Gel filtration followed as described. Neutralized suspensions underwent 1:10 serial dilutions using minimal essential media (Mediatech, Inc., Manassas, VA) followed by plating onto host cell monolayers in 96-well trays (6 replicates per dilution). Following incubation at 35°C in a 5% CO2 atmosphere, wells were scored for the presence of cytopathogenic effects and cytotoxicity. The method of neutralization was also validated per ASTM E1053-20 (data not shown). Additional assay parameters are listed in Table 1.
      Table 1Experimental parameters for human coronavirus 229E and SARS-CoV-2 antiviral efficacy testing
      FBS, fetal bovine serum.
      ParameterHCoV-229E, Test 1HCoV-229E, Test 2SARS-CoV-2
      Carrier dimensions25.81 cm26.54 cm26.54 cm2
      Inoculum volume0.10 mL0.05 mL0.05 mL
      Soil loadNone5% FBS5% FBS
      Virus harvest methodSwab methodWash methodWash method
      Per well assay volume0.05 mL0.10 mL0.10 mL
      Assay incubation period9 days9 days10 days
      low asterisk FBS, fetal bovine serum.

      Results

      The reformulated quaternary ammonium coating achieved >99.9% reduction of HCoV-229E and SARS-CoV-2 during the 2-hour contact time in the absence and presence of 5% organic soil (Table 2). The wash method for harvesting the carriers facilitated greater recovery of viruses compared to the swab method. Cytotoxicity was observed in the lowest plated dilution (100) of Vero E6 cells, while no toxic effects were evident on MRC-5 cells at the 100 dilution.
      Table 2Viral recovery (log10) and reduction of HCoV-229E and SARS-CoV-2 on quaternary ammonium polymer-coated coupons relative to noncoated stainless steel carriers at 2 hours
      CPE, cytopathogenic effects; N.A., Not applicable; N.D., Not determined.
      ,
      All noncoated control and coated coupons tested in triplicate (n = 3) per contact time, except for coated test coupons against SARS-CoV-2 (n = 6).
      ,
      FBS, fetal bovine serum.
      Coupon typeContact time (hours)HCoV-229E (No soil load, swab method)HCoV-229E (5% FBS soil load, wash method)SARS-CoV-2 (5% FBS soil load, wash method)
      Noncoated control04.63 ± 0.29N.D.5.50 ± 0.17
      Noncoated control24.75 ± 0.255.72 ± 0.255.06 ± 0.10
      Coated test2≤ 0.80 ± 0.00≤ 0.56 ± 0.10≤ 1.53 ± 0.07
      # of CPE (+) coated test carriersN.A.0/31/31/6
      Detection limit (log10)N.A.≤ 0.80≤ 0.50≤ 1.50
      Log10 ReductionN.A.≥ 3.94≥ 5.16≥ 3.53
      Percent ReductionN.A.≥ 99.98%≥ 99.9994%≥ 99.97%
      low asterisk CPE, cytopathogenic effects; N.A., Not applicable; N.D., Not determined.
      All noncoated control and coated coupons tested in triplicate (n = 3) per contact time, except for coated test coupons against SARS-CoV-2 (n = 6).
      FBS, fetal bovine serum.

      Discussion

      The results demonstrate similar reductions of >99.9% (>3-log10) for HCoV-229E and SARS-CoV-2 following 2 hours of exposure to the reformulated quaternary ammonium coating applied to stainless steel coupons. However, further studies are warranted to assess effectiveness following application to a variety of nonporous and porous surface types. Recovery levels of both viruses from noncoated study controls after 2 hours suggest comparative rates of inactivation on Type 304 stainless steel under the environmental conditions implemented, although a larger sample size would be preferable to confirm statistical significance. Cytotoxic effects were observed after plating undiluted (100) neutralized filtrates from coated carriers onto Vero E6 host cells, while none were observed on MRC-5 cells. Widespread cell monolayer damage in the absence of infectious viruses typifies cytotoxicity; therefore, dilutions that display such effects are not utilized to calculate log10 reductions. However, they are determinative in establishing assay detection limits which can vary by orders of magnitude as observed for the HCoV-229E and SARS-CoV-2 assays (≤0.50 log10 and ≤1.50 log10, respectively).
      Surface-active coatings that have antiviral capabilities are not meant to substitute for regular cleaning and disinfection practices, but rather serve as an additional barrier for reducing human exposure to infectious viruses that may be present on fomites.
      • Rutala WA
      • Gergen MF
      • Emily-Sick-Bennett
      • Anderson DJ
      • Weber DJ
      Antimicrobial activity of a continuously active disinfectant against healthcare pathogens.
      With the recent detection of infectious SARS-CoV-2 from the bedside table, remote control, bed rails, and flooring in the hospital room of an infected patient,
      • Ahn JY
      • An S
      • Sohn Y
      • et al.
      Environmental contamination in the isolation rooms of COVID-19 patients with severe pneumonia requiring mechanical ventilation or high-flow oxygen therapy.
      the importance of effective hygiene protocols for environmental surfaces remains imperative.

      References

        • Boone SA
        • Gerba CP
        Significance of fomites in the spread of respiratory disease and enteric viral disease.
        Appl Environ Microbiol. 2007; 73: 1687-1696
        • Sassi HP
        • Sifuentes LY
        • Konig DW
        • et al.
        Control of the spread of viruses in a long-term care facility using hygiene protocols.
        Am J Infect Contr. 2015; 43: 702-706
        • Querido MM
        • Neves L
        • Neves P
        • Pereira CS
        • Teixeria JP.
        Self-disinfecting surfaces and infection control.
        Colloids Surfaces B: Biointerfaces. 2019; 178: 8-21
        • Ellingson KD
        • Pogreba-Brown K
        • Gerba CP
        • Elliott SP
        Impact of a novel antimicrobial surface coating on health care-associated infections and environmental, at 2 urban hospitals.
        Clin Infect Dis. 2020; 71: 1807-1813
        • Carraturo F
        • Del Giudice C
        • Morelli M
        • et al.
        Persistence of SARS-CoV-2 in the environment and COVID-19 transmission risk from environmental matrices and surfaces.
        Environ Pollut. 2020; 265, Part B115010
        • Gundy PM
        • Gerba CP
        • Pepper IL
        Survival of coronaviruses in water and wastewater.
        Food Environ Virol. 2009; 1: 10-14
        • Hierholzer JC
        • Killington RA
        • Stokes A
        Virology Methods Manual.
        Academic Press, Inc., San Diego, CA1996
      1. ASTM International. ASTM E1053-20. Standard practice to assess virucidal activity of chemicals intended for disinfection of inanimate, nonporous environmental surfaces. 2020. Accessed April 15, 2020. https://www.astm.org/Standards/E1053.htm

        • Rutala WA
        • Gergen MF
        • Emily-Sick-Bennett
        • Anderson DJ
        • Weber DJ
        Antimicrobial activity of a continuously active disinfectant against healthcare pathogens.
        Infect Control Hosp Epidemiol. 2019; 40: 1284-1286
        • Ahn JY
        • An S
        • Sohn Y
        • et al.
        Environmental contamination in the isolation rooms of COVID-19 patients with severe pneumonia requiring mechanical ventilation or high-flow oxygen therapy.
        J Hosp Infect. 2020; 106: 570-576