Evaluating the impact of ultraviolet C exposure conditions on coliphage MS2 inactivation on surfaces

Abstract

The COVID-19 pandemic has raised interest in using devices that generate ultraviolet C (UVC) radiation as an alternative approach for reducing or eliminating microorganisms on surfaces. Studies investigating the efficacy of UVC radiation against pathogens use a wide range of laboratory methods and experimental conditions that can make cross-comparison of results and extrapolation of findings to real-world settings difficult. Here, we use three different UVC-generating sources – a broad-spectrum pulsed xenon light, a continuous light-emitting diode (LED), and a low-pressure mercury vapour lamp – to evaluate the impact of different experimental conditions on UVC efficacy against the coliphage MS2 on surfaces. We find that a nonlinear dose–response relationship exists for all three light sources, meaning that linear extrapolation of doses resulting in a 1-log₁₀ (90%) reduction does not accurately predict the dose required for higher (e.g. 3-log₁₀ or 99.9%) log₁₀ reductions. In addition, our results show that the inoculum characteristics and underlying substrate play an important role in determining UVC efficacy. Variations in microscopic surface topography may shield MS2 from UVC radiation to different degrees, which impacts UVC device efficacy. These findings are important to consider in comparing results from different UVC studies and in estimating device performance in field conditions.

Introduction

The COVID-19 pandemic has raised interest in alternative approaches for cleaning and disinfecting surfaces, particularly in high-touch or high-traffic areas, such as public transportation, schools and businesses. Devices that emit ultraviolet (UV) radiation provide many potential advantages over traditional liquid disinfectants in that they can be activated and controlled remotely and automatically, leave no chemical residue, and can be more compatible with sensitive materials (Ma et al. 2021). As a result, the use of UV devices, particularly those that emit UVC radiation (200–280 nm), has gained increasing attention, although UVC has been used for antimicrobial purposes in water, air and on surfaces for over 100 years (Kowalski 2010).

UVC radiation can be effective against all types of microorganisms and viruses, damaging proteins and nucleic acids (DNA and RNA) as it is absorbed (Bintsis et al. 2000; Hadi et al. 2020; Ma et al. 2021). Efficacy of UVC radiation against a particular pathogen is a function of the UVC dose (J m⁻², calculated by multiplying the UVC irradiance at the exposed surface by the time of exposure) in addition to the UVC wavelength. UVC irradiance follows the inverse-square law of light, such that doubling the distance between a UVC radiation and a surface will quadruple the amount of time needed to achieve the same dose on that surface (Kowalski 2010). Nucleic acid damage tends to peak around a UVC wavelength of 260–265 nm. As such, UVC efficacy against nonenveloped viruses (including coliphage MS2, a member of the family Leviviridae that infects the bacterium Escherichia coli) peaks at these wavelengths (Hull and Linden 2018), although enveloped viruses may respond differently to UVC radiation given the structure of their outer lipid layer (Ma et al. 2021). These wavelengths are close to the emissions peak of low-pressure mercury vapour lamps (254 nm), which have historically been the most widely used UVC sources (Heßling et al. 2020; Ma et al. 2021) for antimicrobial purposes.

Many recent studies have focused on quantifying the efficacy of UVC radiation against SARS-CoV-2 and other coronaviruses on surfaces (Hadi et al. 2020; Heßling et al. 2020; Gidari et al. 2021). Of high interest has been using UVC radiation for the treatment of personal protective equipment (PPE), including N95 respirators (Fischer et al. 2020; Simmons et al. 2020; Smith et al. 2020), so that these items can be reused. These studies have utilized different UVC sources, including far UVC devices, which emit UVC radiation in the 207–222 nm wavelength range and have been proposed to be safe for human exposure (Buonanno et al. 2020; Kitagawa et al. 2021) (traditional UVC sources present a health hazard to human eyes and skin). Moreover, these studies often utilize different laboratory methodologies and materials, which can make comparing results across different investigations difficult and leads to a wide range in reported effective doses, even for the same microorganism (Heßling et al. 2020).

The U.S. Environmental Protection Agency (EPA) considers UVC light sources that are marketed for the purpose of reducing microorganisms to be pesticide devices, which work by using physical means (e.g., light) to inactivate microorganisms and viruses, in contrast to pesticide products, which contain a substance or mixture of substances that are intended to inactivate pests. In contrast to pesticide products, which require a pre-market evaluation process and are registered by the EPA, pesticide devices do not currently require registration. While pesticide devices do not require pre-market review, false or misleading claims cannot be made about their safety or efficacy against pathogens. In addition, manufacturers of devices must have scientific data on their specific device to support any claims made regarding device efficacy. For a pesticide product to make virucidal claims, the product must demonstrate a ≥3 log₁₀ (99.9%) reduction against viruses (U.S. EPA 2018). Although UVC devices are not registered as pesticide products, the ≥3 log₁₀ inactivation is a standard of efficacy for pesticide devices making disinfection claims against viruses and can be used for comparing device performance to EPA-registered virucidal products. However, because laboratory studies use a wide range of methodologies and there is no pre-defined efficacy evaluation method for pesticide devices, meaningful comparison of device performance relative to product efficacy can be difficult, even when similar efficacies are reported because of the conditions that give rise to the reported efficacies vary widely.

Here, we investigate how different experimental variables impact UVC efficacy on surface inactivation of the coliphage MS2 using three different UVC sources (a broad-spectrum pulsed xenon UVC source, a light-emitting diode (LED) with a peak UVC wavelength of 275 nm, and a low-pressure mercury vapour lamp), different inoculum compositions and conditions (varying organic load and salt content, as well as wet vs dried inoculum) and inoculation methods (spread on the material surface vs left as a droplet), and four different material types (304 stainless steel, 301 stainless steel, glass and ABS plastic). A better understanding of how these various factors influence UVC efficacy is necessary for intercomparison of UVC efficacy studies utilizing different laboratory techniques and for extrapolating the findings of laboratory studies to predict how UVC devices will perform in real-world conditions. This research was conducted iteratively; instead of evaluating a full test matrix across all the parameters explored, which would be extensive and beyond the scope of this study, the experiments and results described below were designed in a targeted way to address specific questions related to the impact of experimental conditions as each set of tests were being conducted.

Results and discussion

Inactivation of MS2 at different UVC doses (measured by either the ILT SED270C or SED270 sensor, see Light Sources and Measurements in Materials and Methods) from three different UVC generating sources is shown in Fig. 1. Efficacy (log₁₀ reduction) is calculated as the difference in mean log₁₀ recoveries of positive control samples (samples not exposed to UVC radiation) and UVC-exposed test samples. For the data shown in Fig. 1, UVC exposures were conducted with MS2 inoculated and dried (see description of inoculation methods in Materials and Methods) onto 304 stainless steel. In these experiments, MS2 inoculum was prepared in 1X Phosphate-Buffered Saline (PBS) with 5% Foetal bovine serum (FBS, see ‘Materials and Methods’ section). Exposures were conducted over a range of distances and times for each light source in order to achieve a range of dose conditions: at 15–200 cm and 7.75–60 min for the pulsed xenon light, at 30–64 cm and 5–120 min for the LED light, and at 60 cm and 0.5–60 min for the mercury lamp. A nonlinear relationship between UVC dose and MS2 activation is present for all three light sources under these test conditions, and the trendlines that result from a logarithmic regression are shown in Fig. 1 (note that dose is plotted on a logarithmic scale). The slopes for each source’s dose–response curve are similar; the corresponding logarithmic fit slopes are 0.54 (R² = 0.96) for the low-pressure mercury vapour lamp, 0.43 (R² = 0.82) for the pulsed xenon light, and 0.61 (R² = 0.95) for the LED. Here, we do not intend to compare how effective the different UVC sources are (e.g., action spectrum study), as the focus of this study is to explore how different environmental and experimental factors impact UVC treatment on surfaces, but instead, we demonstrate that the exposures conducted with the different UVC sources used in this research show a similar, nonlinear dose–response relationship.

Figure 1.

MS2 inactivation at different UVC doses in dried, spread inoculum on 304 stainless steel from three light sources ((a) a low-pressure mercury vapour (Hg) lamp, (b) broad spectrum pulsed xenon source, and (c) an LED). UVC dose (plotted on a logarithmic scale) for each exposure with all light sources is reported as measured by the ILT 2500 metre and either the SED270C (Hg lamp and pulsed xenon) or the SED270 (LED) sensors. Each data point represents efficacy calculated by subtracting the mean log of MS2 recoveries from three replicate test (exposed) coupons from three replicate positive control (unexposed) coupons. Error bars represent pooled standard deviation from both the positive control and test samples. The trendlines plotted on each panel are the result of logarithmic regression for each respective dataset.

Although none of the exposures conducted with the three UVC sources on 304 stainless steel resulted in a 3-log₁₀ (99.9%) reduction in MS2 (Fig. 1), which is the target efficacy for virucidal disinfectant products (U.S. EPA 2018), Fig. 2 reveals that higher efficacy was observed in exposures conducted on glass coupons. Data shown in Fig. 2 were collected during exposures using the LED source with wet, spread inoculum (prepared in PBS with 5% FBS) on both 304 stainless steel and glass. Above a UVC dose of approximately 60 mJ cm⁻², inactivation of MS2 on glass occurred at consistently higher rates than on 304 stainless steel under the same dose and experimental conditions. The resulting equations from logarithmic regressions and their corresponding coefficients of determination are also shown in Fig. 2. The slope of the fitted dose–response curve generated from the exposures conducted on glass is greater than twice that of the dose–response curves generated on stainless steel (Fig. 2).

Figure 2.

Dose-inactivation response for MS2 on both 304 stainless steel (○) and glass () using the LED source with wet, manually-spread inoculum. Each data point represents efficacy calculated by subtracting the mean log of MS2 recoveries from three replicate test (exposed) coupons from three replicate positive control (unexposed) coupons. Error bars represent pooled standard deviation from both the positive control and test samples. The plotted trendlines and displayed equations are the result of logarithmic regressions of data from the two materials.

Testing with additional materials shows that material type can lead to a range of efficacies that spans >2-log₁₀ between exposures conducted with otherwise the same experimental conditions. Figure 3 shows these observed differences in efficacy with material type, with all exposures conducted at a distance of 38 cm between the pulsed xenon light source and inoculated coupons with a 30-min exposure time (~43 mJ cm⁻² measured by the SED270C sensor). Two different inoculation methods – one in which the MS2 inoculum (prepared in PBS with 5% FBS) is spread across the coupon surface and a second in which the inoculum is left as a single, intact droplet – were also used to investigate the potential impact of the inoculation method on efficacy (Fig. 3). Two-way anova showed no statistically significant differences between the inoculation methods (P = 0.25) or material type (P = 0.19) for this particular experiment (in part due to the small sample size), but the spreading of the inoculum consistently leads to greater UVC efficacies compared to exposures with intact droplets. These results are in agreement with previous studies, which have demonstrated that spreading inoculum over a greater surface area increases UVC device efficacy against microorganisms (Cadnum et al. 2016), as the act of spreading inoculum can potentially disaggregate clumped or stacked organisms, exposing a greater fraction to the UVC radiation (Raguse et al. 2016; Boyce and Donskey 2019).

Figure 3.

Efficay of the pulsed xenon source against MS2 on three different material types: 301 stainless steel (301 SS), 304 stainless steel (304 SS), and ABS plastic (ABS). 10 μl of inoculum was either manually spread across the coupon surface () or left as a single droplet () and was dried at ambient conditions prior to exposure. The test was conducted with a 30-min exposure time at a distance of 38 cm between the pulsed xenon light source and the coupons (UVC dose ~43 mJ cm⁻²). Error bars represent pooled standard deviation from both the positive control and test samples.

The composition of the inoculum also impacts UVC efficacy against MS2. The presence of mucin and/or other salts, both of which are present in saliva, can affect how UV radiation is absorbed and transmitted (Sagripanti and Lytle 2011; Barancheshme et al. 2021). To assess this effect, MS2 was inoculated onto two materials, 301 stainless steel and ABS plastic, in inoculum composed of either 1X minimum essential medium (MEM) and 5% FBS or a simulated saliva (see ‘Materials and Methods’ section) as single 10 μl droplets that were either exposed wet (immediately following inoculation) or after the droplets had dried at ambient temperature (approximately 1 h). The resulting log₁₀ reductions following a 30-min exposure to the pulsed xenon source at 38 cm distance are shown in Fig. 4. Generally, exposures conducted with wet inoculum led to greater log₁₀ redutions than dry inoculum, and MS2 was more difficult to inactive in simluated saliva than in the tissue culture media. Two-way anova demonstrated a statistical significance for both the inoculum composition (P = 0.0012) and the material type (P = 0.023). On both materials, calculated efficacies range 2 log₁₀ reductions (99%) across the inoculum compositions and conditions, highlighting the substantial impact that these factors can have on UVC efficacy. The variability in inoculum composition and conditions across different experimental studies likely accounts for a considerable portion of the variability in results from different laboratory studies, even those conducted with the same microorganism (Heßling et al. 2020). Moreover, this marked impact of inoculum composition on UVC efficacy should be considered when extrapolating laboratory studies to applied settings, where the target pathogens would likely be found in the presence of respiratory fluids that have dried onto surfaces, which proved to be the most difficult to inactivate inoculum composition and condition of those evaluated here.

Figure 4.

Efficacy of the pulsed xenon source against MS2 on two different material types (301 stainless steel (301 SS) and ABS plastic (ABS)) in two different inoculum compositions (tissue culture media (grey bars) or simulated saliva (black bars)), with inoculum exposed either wet (immediately following inoculation, solid bars) or after the inoculum had dried at ambient conditions (diagonal striped bars). In all cases, 10 μl of inoculum was left on the material surfaces as a single droplet. The test was conducted with a 30-min exposure time at a distance of 38 cm between the pulsed xenon light source and the coupons. Error bars represent pooled standard deviation from both the positive control and test samples. Legend: tissue culture media, wet (); simulated saliva, wet (), tissue culture media, dry (), simulated saliva, dry ().

Even though all four materials used in this study are considered smooth, non-porous surfaces, high magnification scanning electron microscope images reveal different surface characteristics that vary across all material types, including the 301 and 304 stainless steels (Fig. 5). The microtopography of both stainless-steel types contains crevices that may contribute to shadowing of UVC light on these surfaces. Similarly, the ABS surface contains cracks, scratches and other types of significant relief in microtopography that could contribute to shielding of MS2 from the UVC radiation. In contrast, the glass surface appears the smoothest, with the least amount of relief available to potentially shadow MS2 (and other microorganisms) from UVC radiation. These microscopic material surface observations are consistent with the differences in resulting log₁₀ reductions on the various material types (Figs 2–4), with the highest efficacies observed on the smoothest surface (glass). These material features could also contribute to the presence of a small but significant portion of virions that are shielded from UVC light, in addition to inoculum components (e.g., salts, celluar debris) that also protect some virions from UV radiation, leading to logarithmic (Figs 1 and 2) and biphasic dose–response curves (Sagripanti and Lytle 2011).

Figure 5.

Scanning electron micrograph of (a) 301 stainless steel, (b) 304 stainless steel, (c) glass and (d) ABS plastic used as carriers in the UVC exposures. Images were generated using a TESCAN MIRA3 Field Emission Scanning Electron Microscope using an accelerating voltage of 15 kV (a–c) and 5 kV (d).

The results presented here demonstrate that the experimental and laboratory conditions for UVC efficacy testing have an important impact on inactivation efficacy against microorganisms and viruses across different types of UVC-emitting devices. A nonlinear relationship exists between applied UVC dose and log₁₀ reduction for all three UVC sources tested with MS2 on stainless steel, which indicates that results from studies reporting doses that achieve 90% or 1-log₁₀ reductions on surfaces should not be linearly extrapolated to estimate the dose needed to achieve a 99.9% (3-log₁₀) reduction. Our results are also consistent with previous studies that show the efficacy of UVC devices on surfaces is dependent on the material type (Boyce and Donskey 2019; Fischer et al. 2020; Wood et al. 2020; Criscuolo et al. 2021), as well as the inoculum composition and condition (Cadnum et al. 2016; Heßling et al. 2020; Barancheshme et al. 2021). UVC efficacy will also vary depending on the target microorganism (Sigstam et al. 2013; Ye et al. 2018; Boyce and Donskey 2019; Ma et al. 2021), as well as its concentration on surfaces (Biasin et al. 2021). Although all of the materials used in this research appear smooth to the unassisted eye, we show that microscopic features on these smooth materials (Fig. 5) likely contribute to the shielding of microorganisms and viruses from UVC light, leading to lower efficacies from UVC exposures on stainless steel and plastic compared to glass. These experimental factors are necessary to consider when comparing effective dose values reported from UVC studies conducted with different experimental protocols and in evaluating how effective different UVC devices will perform in specific applied field settings.

Materials and methods

Light sources and measurements

Three different UVC-generating light sources were used in this study, selected based on research stakeholder input. The first is a pulsed xenon light source (manufacturer information withheld upon request) that emits light in a broad spectrum, from approximately 220 nm into the visible spectrum (>500 nm), by pulsing on the order of a millisecond every 6 s. See Szary et al. (2020) for the pulsed xenon source spectrum. The second is a prototype 24 Volt DC LED unit from Transit Design Group (Mississauga, ON, Canada) that consists of an array of four 24 V 275 nm LEDs and six 24 V 405 nm LEDs that emits light continuously at an effective total output power of 190 mW at 275 nm and 34 mW at 405 nm. The full width, half maximum (FWHM) of the 275 nm peak is 10 nm, and the FWHM for the 405 nm peak is 13 nm. See Fig. S1 for the wavelength spectrum of the LED source. MS2 inactivation at the 405 nm wavelength is expected to be negligible compared to radiation in the UVC range (Hull and Linden 2018). The third is a traditional low-pressure mercury vapour lamp (G30T8, Sankyo Denki, Kanagawa, Japan; 30 W power) with a peak wavelength at 253.7 nm. Light from this source was collimated for the UVC exposures using a cardboard tube 51 cm in length with an inner diameter of 7.6 cm.

Radiation from all three sources was measured using an ILT 2500 hand-held flash optical metre and a SED270 or SED270C sensor (both International Light Technologies, Peabody, MA) at the location of coupon exposures. The SED270C sensor detects light between 230 and 280 nm, with a peak response at 265 nm. It was used to record the intensity of the pulsed xenon light and mercury vapour lamp. The SED270 sensor detects light between 200 and 360 nm with a peak response at 265 nm, and it was used to record the intensity of the LED light source at 275 nm because the SED270C sensor underestimates UVC emitted by the LED based on its detection range. The metre’s custom Flash App was utilized for measuring UVC dose at the inoculated surface from the pulsed xenon light source, and the continuous metre measurement app was used for measuring UVC intensities at the inoculated surface from the LED and mercury vapour lamp sources, which were used to calculate dose based on the length of time of each exposure.

All UVC exposures reported in this study were conducted with material coupons lying on flat surfaces, except for the pulsed xenon tests shown in Fig. 1c, which were conducted with coupons oriented nearly vertical.

Materials

Four different materials were used in the UVC exposures: 304 stainless steel (unpolished 2B mill finish, 0.91 mm thick; McMaster-Carr, Douglasville, GA), 301 stainless steel (cold worked and hardened, 0.79 mm thick; McMaster-Carr), ABS plastic (1.59 mm thick; McMaster-Carr) and glass (soda-lime plate, 3 mm thick; Prism Research Glass, Research Triangle Park, NC). The stainless steel and plastic materials were cut into individual 2-cm × 4-cm coupons that are screened for uniformity (glass coupons were purchased pre-cut into 2-cm × 4-cm coupons). The coupons were washed in Liquinox anionic liquid detergent (Alconox, White Plains, NY) according to the manufacturer’s instructions, then rinsed, dried and sterilized. Stainless steel coupons were also soaked in 100% ethanol and rinsed in deionized water between the washing step and sterilization. Stainless steel and glass coupons were sterilized using a 121°C gravity cycle autoclave (SV 120 scientific pre-vacuum sterilizer; STERIS Amsco, Mentor, OH). ABS plastic coupons were sterilized using an ethylene oxide sterilizer (EOGas Sterilizers-Series 3 + Model 333.01, Andersen Sterilizers Inc., Haw River, NC). No coupons were reused during the course of this study. Scanning electron microscope (SEM) images were taken of the materials using a TESCAN MIRA3 Field Emission SEM (TESCAN, Brno, Czech Republic) at an accelerating voltage of either 15 or 5 kV (as noted in the caption for Fig. 5). Glass was imaged with a thin layer of carbon coating, ABS plastic was imaged with a thin layer of gold coating, and both stainless steel materials were imaged without any coating.

Coliphage preparation, inoculation and enumeration

The test organism used in this study was MS2, a nonenveloped bacteriophage with an icosahedral capsid and a positive-sense, single-stranded RNA genome (Valegård et al. 1990). MS2 is a member of the family Leviviridae that infects the bacterium Escherichia coli. Working stocks of E. coli, the MS-2 bacterial host, were prepared in 2 ml cryovials (10–500-26; Fisher Scientific, Hampton, NH) by adding 0.5 ml of log-phase E. coli to 0.5 ml of sterile 50% (w/v) glycerol (BP229-1; Fisher Scientific). Working stocks were stored at − 80°C until use. A bacterial culture of E. coli was prepared by thawing a frozen bacterial stock and transferring the 1 ml thawed bacterial stock into 99 ml of lysogeny broth (LB), then incubating the culture overnight at 35 ± 2°C on an orbital shaker (The Belly Dancer^™; part no. 12–453-211, Vernon Hills, IL). On the day of use, a fresh E. coli culture was started by adding an aliquot of the overnight culture to fresh LB media, then incubating at 35 ± 2°C on an orbital shaker until the OD₆₀₀ measured between ~0.4 and ~0.9, indicating readiness for use in sample plating. Working stocks of MS2 (ATCC 15597-B1) were prepared using a top agar overlay technique (Kropinski et al. 2009), where solid agar plates made with lysogeny broth (LB) agar (BD Difco 240110; Thermo Fisher Scientific, Waltham, MA) are coated with ~6 ml of molten LB top agar containing 100μl of MS2 stock and 100 μl of a log-phase E. coli C-3000 (ATCC-15597) bacterial culture. After overnight incubation at 35 ± 2°C, a sterile cell spreader was used to gently scrape the soft agar overlay from three 100 mm plates into a sterile 50-ml conical tube containing 15 ml of SM buffer (S0249; Teknova Inc., Hollister, CA). Tubes were vortexed to break up agar clumps, then centrifuged at 7000 g for 15 min. The supernatant was removed and filtered through a 0.2 μm syringe filter (PES syringe filters, 431229; Corning Inc., Corning, NY). MS2 stocks in SM buffer were stored in 1 ml volumes at −80°C until use.

For each test, coupons were inoculated with thawed MS2 stock prepared in either 1X phosphate-buffered saline (PBS) (P0196; Teknova, Hollister, CA) with 5% fetal bovine serum (FBS, Gibco 10082139; Thermo Fisher Scientific), Corning 1X Minimum Essential Medium (Product #10-010-CV, Corning, NY) with 5% FBS or in simulated saliva prepared following Heimbuch et al. (2011) (Table 1) at a target concentration of ~2 × 10⁶ virions per coupon. Each coupon was inoculated with a 10 μl droplet of the prepared inoculum, which was either left as a single droplet or spread over at least 75% of the surface of the coupon using the pipette tip. Coupons were either allowed to dry at ambient temperature before exposure or exposed while the inoculum was still wet, as denoted in the text. All test conditions (test coupons and positive controls) for each exposure were conducted in triplicate.

Table 1.

Composition of simulated saliva. Mucin is porcine gastric mucin Type III (Millipore Sigma, Burlington, MA)

Composition	Concentration
MgCl2·6H2O	196.75 μmol l−1
CaCl2·H2O	1.17 mmol l−1
NaHCO3	5.00 mmol l−1
KH2PO4	1.54 mmol l−1
K2HPO4	2.46 mmol l−1
NH4Cl	2.06 mmol l−1
KSCN	1.96 mmol l−1
(NH2)2CO	2.00 mmol l−1
NaCl	15.06 mmol l−1
KCl	13.95 mmol l−1
Mucin	0.3% (wt/vol)
Distilled water	To volume

Following the completion of each UVC exposure, coupons were aseptically placed into sterile 50-ml conical tubes containing 10 ml of 10% Dey-Engley Broth (BD Difco 281910; Thermo Fisher Scientific) and vortexed for 5 s. After the completion of all exposure conditions during each test, sample tubes were vortexed continuously for an additional 2 min. Tenfold serial dilutions were prepared in 1X PBS for each sample, and each dilution was plated in triplicate using a conventional soft agar overlay method with E. coli as the bacterial host (Kropinski et al. 2009). After overnight incubation at 35 ± 2°C, plaques were manually enumerated. Efficacy (log₁₀ reduction) is calculated as the difference in mean log₁₀ recoveries of positive control samples (samples not exposed to UVC light, but present on coupons for the same amount of time as the UVC-exposed test samples) and UVC-exposed test samples.

Statistics

All statistical analyses (logarithmic regressions and two-factor anova without replication) were performed using the Data Analysis ToolPak in Microsoft Excel for Microsoft 365 Version 2 108.

Supplementary Material

Supplementary Material

Figure S1. Wavelength spectrum of the LED source, recorded using a spectral radiometer (UVpadE Model #670027, Opsytec Dr. Groebel, Ettlingen, Germany).

Significance and Impact of the Study:

The COVID-19 pandemic has raised interest in devices that emit ultraviolet C (UVC) radiation for reducing or eliminating microorganisms on surfaces, but it is difficult to compare performance across UVC devices because their efficacy is evaluated using different methods. Here, we compare how different experimental factors affect the efficacy of UVC light for treating surfaces. We find that both differences in the fluid that contains the virus and microscopic differences in materials impact the efficacy of UVC devices and that the UVC dose reported to achieve a 90% reduction cannot always be linearly extrapolated to predict the 99.9% reduction dose.

Data availability statement

The data that support the findings of this study are openly available at https://doi.org/10.23719/1522675.