Effectiveness of the Nanosilver/TiO 2 -Chitosan Antiviral Filter on the Removal of Viral Aerosols

Abstract

Background:

The precaution of airborne transmission of viruses, such as influenza, SARS, MERS, and COVID-19, is essential for reducing infection. In this study, we applied a zero-valent nanosilver/titania-chitosan (nano-Ag⁰/TiO₂-CS) filter bed, whose broad-spectrum antimicrobial efficacy has been proven previously, for the removal of viral aerosols to minimize the risk of airborne transmission.

Methods:

The photochemical deposition method was used to synthesize the nano-Ag⁰/TiO₂-CS antiviral material. The surface morphology, elemental composition, and microstructure of the nano-Ag⁰/TiO₂-CS were analyzed by a scanning electron microscopy/energy dispersive X-ray spectroscopy and a transmission electron microscopy, respectively. The MS2 bacteriophages were used as surrogate viral aerosols. The antiviral efficacy of nano-Ag⁰/TiO₂-CS was evaluated by the MS2 plaque reduction assay (PRA) and filtration experiments. In the filtration experiments, the MS2 aerosols passed through the nano-Ag⁰/TiO₂-CS filter, and the MS2 aerosol removal efficiency was evaluated by an optical particle counter and culture method.

Results and Conclusions:

In the MS2 PRA, 3 g of nano-Ag⁰/TiO₂-CS inactivated 97% of MS2 bacteriophages in 20 mL liquid culture (2 ± 0.5 × 10¹⁶ PFU/mL) within 2 hours. The removal efficiency of nano-Ag⁰/TiO₂-CS filter (thickness: 6 cm) for MS2 aerosols reached up to 93%. Over 95% of MS2 bacteriophages on the surface of the nano-Ag⁰/TiO₂-CS filter were inactivated within 20 minutes. The Wells–Riley model predicted that when the nano-Ag⁰/TiO₂-CS filter was used in the ventilation system, airborne infection probability would reduce from 99% to 34.6%. The nano-Ag⁰/TiO₂-CS filter could remain at 50% of its original antiviral efficiency after continuous operation for 1 week, indicating its feasibility for the control of the airborne transmission.

Introduction

Airborne transmission of viruses, such as coronaviruses,^(1–3) influenza,^(4,5) measles,^(6–8) porcine epidemic diarrhea virus,⁽⁹⁾ and foot-and-mouth disease virus,⁽¹⁰⁾ plays a significant role in epidemic infection. Control of the airborne spread of viruses is challenging because aerosolized viruses may survive and remain infectious for hours^(7,11,12); airflow control through exceptional ventilation and air purification systems is required for better proficiency.^(13,14) Room ventilation and air filtration act to dilute and eliminate infectious airborne viruses, and several epidemiologic studies have underscored its substantial role in the mitigation of airborne transmission.^(13–15) Some viruses stay viable and infectious from hours to days on surfaces.^(7,11) As a result, antiviral materials for both air and surface purification have attracted extensive attention. Nanosilver has exhibited a strong antiviral potential against a broad range of virus families, including retroviridae, hepadnaviridae, paramyxoviridae, herpesviridae, poxviridae, orthomyxoviridae, and arenaviridae.⁽¹⁶⁾ Nanosilver could bind with virus coat proteins and cell membrane receptors and disturb their structure and function, which prevent viral attachment and entry into the host cells.⁽¹⁷⁾ Nanosilver could also interact with the double-stranded DNA of the viruses.⁽¹⁸⁾

Our previous study has demonstrated the excellent bactericidal efficacy of zero-valent nanosilver/titania-chitosan (nano-Ag⁰/TiO₂-CS) composite for purifying the supply air in hospital wards.⁽¹⁹⁾ However, due to the different structures and aerosol particle sizes between viruses and bacteria,^(20–23) the antimicrobial efficacy of nano-Ag⁰/TiO₂-CS against viruses and bacteria would also be different. Thus, the primary aim of this study is to evaluate the filtration efficiency and antiviral efficacy of nano-Ag⁰/TiO₂-CS filter for viral aerosols, along with the assessment of the infection risk reduction and long-term antiviral efficacy of nano-Ag⁰/TiO₂-CS.

Materials and Methods

This study was conducted in the laboratory (in vitro) using MS2 bacteriophage and its host bacterium. There is no experiment involving any human subject or cell or data.

Preparation of MS2 bacteriophage suspension

MS2 bacteriophage was used as the surrogate of the airborne virus. MS2 bacteriophages were purchased from the Bioresource Collection and Research Center (BCRC), Taiwan (BCRC No. 70235), and the host bacterium, Escherichia coli F2, was also obtained from BCRC (BCRC No. 50354). The MS2 bacteriophage suspension for the viral aerosol generation was prepared according to the following procedure. First, the MS2 bacteriophage was multiplied by the double-layer agar culture technique. The double-layer agar culture plates of MS2 were placed at −20°C for 5 hours, and then the plates were thawed at room temperature. After thawing, the liquid produced was transferred to a centrifuge tube and centrifuged at 10,000 rpm (5000 g) for 10 minutes to precipitate the host bacteria. Finally, the supernatant was filtered with a 0.22-μm pore size membrane to obtain an MS2 bacteriophage suspension free of host bacteria.

Preparation and characterization of the nano-Ag⁰/TiO₂-CS composite

The titania-chitosan (TiO₂-CS) composite was prepared according to our previously reported method.⁽¹⁹⁾ Briefly, 15 g of chitosan was completely dissolved in acetic acid solution (600 mL, 3% v/v, pH = 2.6). Then, 2.4 g of P25 TiO₂ powder (particle size 25–30 nm; Brunauer-Emmett-Teller [BET] surface area 53.3 m²/g; 80% in the anatase phase and 20% in the rutile phase) was added to the viscous chitosan solution, and then the slurry was nonstop stirred for 24 hours to obtain a viscous colloid. Afterward, 1.02 mL of 25% glutaraldehyde solution, which was used as the cross-linking agent, was added to the viscous colloid, and the mixture was continuously stirred for 8–10 hours. The resultant colloid was then dropped into a 0.5 M NaOH solution using a syringe pump to form solidified TiO₂-CS beads, and the solidification proceeded for 24 hours. Then the TiO₂-CS beads were collected and rinsed with deionized water until reaching a neutral pH. Finally, the TiO₂-CS beads were dried in an oven at 90°C for 12 hours. The 2 weight (wt) % nano-Ag⁰/TiO₂-CS beads were prepared by photochemical deposition method.⁽²⁴⁾ Concisely, appropriate amounts of AgNO₃, TiO₂-CS beads, and methanol were mixed in deionized distilled water in a Pyrex beaker. After deaerating by flowing nitrogen (30 mL/min), the sample was illuminated by 350-nm ultraviolet lamps for 6 hours in a photochemical Reactor (Consotech Corp., Taiwan). The electrons generated on the surface of TiO₂ were donated to reduce Ag⁺ to Ag⁰, while methanol served as a sacrificial agent to scavenge electron holes. Then the 2 wt% nano-Ag⁰/TiO₂-CS beads were collected and dried overnight in an oven at 120°C.

The surface morphology, elemental composition, and microstructure of the nano-Ag⁰/TiO₂-CS composite were examined by the scanning electron microscope/energy dispersive X-ray spectroscopy (SEM/EDS; JEOL JSM-7600F) and the transmission electron microscopy (TEM; JEOL JSM 1400-Plus) at 100 kV, respectively. The sample of nano-Ag⁰/TiO₂-CS for TEM analysis was scraped from the surface layer of nano-Ag⁰/TiO₂-CS beads. These surface samples were ground (in agate mortar) and dispersed in ethanol ultrasonically and then transferred to the Cu TEM grid (300 mesh with a carbon-coated film). The TEM images of TiO₂-CS beads (without coated nanosilver) were also recorded for comparison.

Evaluation of antiviral efficacy of nano-Ag⁰/TiO₂-CS composite

The plaque reduction assay (PRA) was utilized to evaluate the antiviral efficacy of nano-Ag⁰/TiO₂-CS composite. Briefly, 1, 2, and 3 g of nano-Ag⁰/TiO₂-CS beads (or 1, 2, and 3 g of TiO₂-CS beads) were added to separate test tubes with screw caps, which contained 20 mL of 0.5% peptone solution. Then 750 μL of MS2 culture was added to each test tube (titer of MS2 = 2 ± 0.6 × 10¹⁶ plaque-forming units/mL). The test tubes containing MS2 cultures and 20 mL of 0.5% peptone solution were served as the control group. Then, all the test tubes lay horizontally on the rotating rods (Fig. 4b) of a Roller Mixer (Digisystem Laboratory Instruments, Inc., New Taipei City, Taiwan). The MS2 liquid cultures and nano-Ag⁰/TiO₂-CS beads (or TiO₂-CS beads) were well mixed by continuously rotating the test tube. The MS2 bacteriophage titer in these test tubes was determined at every 2- or 1-hour interval by the double-layer agar technique. Meanwhile, the release of nanosilver in the liquid was analyzed with an Inductively Coupled Plasma Optical Emission Spectrometer (Agilent 725). Peptone solution (0.5%) and deionized water were served as negative controls (without MS2 culture). Each experiment was replicated thrice. The survival fraction (SF) of the MS2 bacteriophage was evaluated by dividing the fraction PFU/mL at each loading of nano-Ag⁰/TiO₂-CS beads (PFU_nano-Ag0) or TiO₂-CS beads (PFU_TiO2-CS) by the fraction of control groups (PFU_control):

S F = \frac{P F U_{n a n o - A g^{0}} o r P F U_{T i O_{2} - C S}}{P F U_{c o n t r o l}}

(1)

The SF of the MS2 bacteriophage was described by first-order kinetics: $ln [S F] = - k \times t$ (2)

where k is the inactivation rate constant, and t is time.

Experimental system

According to the experiment parts' function, the experimental system is divided into four parts: the zero-air supply, the viral-aerosol generation, filtration, and sampling parts, as shown in Figure 1. Before each experiment, the experimental system would be disinfected with 75% ethanol for 30 minutes. After the disinfection was completed, a control sample would be taken at the end of the experimental system by a modified AGI30 (Fig. 1) to confirm the decontamination result. Some previous studies^(25,26) have demonstrated that 70% ethanol can reduce MS2 bacteriophage titer by 0.7 log10 in 2 minutes and 3.68 log10 in 20 minutes.

FIG. 1.

Diagram of the experimental system.

The zero-air supply part produced zero air by pushing the compressed air through the dryer (containing alumina and silica gel) to remove the moisture and then through the HEPA capsule (HEPA Capsule PN 12144) to eliminate the airborne particles.

The viral-aerosol generation part includes a rotameter, a Collison nebulizer (3-jet Modified MRE type; BGI) containing the suspension of MS2 bacteriophage, a diffusion dryer, and a Kr-85 neutralizer. The zero air was passing through the Collison nebulizer to generate MS2 droplets. Then, the air carrying MS2 droplets passed through the diffusion dryer to remove moisture and the Kr-85 neutralizer to neutralize the charges, and then the MS2 aerosols were generated. The MS2 aerosol output flow rate was controlled at 3.6 liters per minute (LPM) by a rotameter.

The viral-aerosol filtration part is a bed filter, which contains an acrylic vessel (diameter: 4.65 cm) packed with nano-Ag⁰/TiO₂-CS beads at the thickness range of 0–6 cm, of which a close-up picture is shown in Figure 1. The airstream containing MS2 aerosols flew directly to the bed filter, and the total airflow rate through the nano-Ag⁰/TiO₂-CS bed filter was 12.5 LPM. The removal efficiency of the nano-Ag⁰/TiO₂-CS filter for MS2 aerosols was evaluated by the following equation: $η_{f i l t e r} = \frac{C_{0} - C_{f}}{C_{0}}$ (3)

where C₀ was the concentration of viral aerosols measured when the filter chamber was empty (no packing), while C_f was the concentration of viral aerosols quantified when the filter chamber was packed with nano-Ag⁰/TiO₂-CS (or TiO₂-CS) at the thickness of 0, 2, 4, or 6 cm.

After each experiment, 0.38 g of nano-Ag⁰/TiO₂-CS or TiO₂-CS beads in the filter was extracted with 2 mL of 1% peptone solution, and its MS2 bacteriophage titer was determined by the double-layer agar technique. The survival ratio (SR) of MS2 bacteriophage on the surface of nano-Ag⁰/TiO₂-CS ( $P F U_{n a n o - A g^{0} ∕ T i O_{2} - C S f i l t e r}$ ) to those on the surface of TiO₂-CS ( $P F U_{T i O_{2} - C S f i l t e r}$ ) was utilized to assess the effectiveness of nano-Ag⁰ on the inactivation of MS2 bacteriophages: $S R = \frac{P F U_{n a n o - A g^{0} ∕ T i O_{2} - C S f i l t e r}}{P F U_{T i O_{2} - C S f i l t e r}}$ (4)

The MS2 aerosol sampling was performed with an All Glass Impinger 30 (AGI-30) packed with glass beads. The sampling liquid was 1% peptone solution (30 mL), the sampling flow rate was 6 LPM, and the sampling time was 20 minutes. After the sampling, 1 mL of the sampling liquid was taken from the AGI-30, serially diluted, and then cultured using double-layer agar plates to evaluate the MS2 bacteriophage titer. Meanwhile, we used a six-channel (0.3, 0.5, 1.0, 2.5, 5, 10 μm) optical particle counter (OPC; 3016IAQ Lighthouse, Inc.) to real timely measure the viral-aerosol concentration in the inlet and outlet of the viral-aerosol filtration part to evaluate the removal efficiency of viral aerosols at each particle size interval.

Results and Discussion

Characteristics of nano-Ag⁰/TiO₂-CS antiviral composite

A nano-Ag⁰/TiO₂-CS bead is a rough sphere with a diameter of around 1.7 mm, as demonstrated in Figure 2a. The surface of the nano-Ag⁰/TiO₂-CS bead is very rough and porous (Fig. 2b) and composed of spherical TiO₂ nanoparticles covered by nano-Ag⁰, as shown in Figure 2c. The elemental composition analysis demonstrated a high percentage of Ag (wt% = 22%) on the surface of the nano-Ag⁰/TiO₂-CS beads, as shown in Figure 2d.

FIG. 2.

SEM images of nano-Ag⁰/TiO₂-CS bead (a) 30 × (b) 1000 × (c) 10,000 × and (d) elemental composition of nano-Ag⁰/TiO₂-CS composite. nano-Ag⁰/TiO₂-CS. nanosilver/titania-chitosan; SEM, scanning electron microscope.

The TEM images in Figure 3a and b show that the nano-Ag⁰ particles are spherical or elliptical and uniformly deposited on the surface of TiO₂ particles, and the particle sizes of nano-Ag⁰ range from 2 to 16 nm (5.94 ± 2.39 nm). The size distribution of 300 particles of nano-Ag⁰ was counted from three filters. Figure 3c and d demonstrates the TEM images of TiO₂-CS composite, and no Ag was detected in the EDS analysis (as shown in the insert image).

FIG. 3.

TEM images of nano-Ag⁰/TiO₂-CS composite and particle size distribution of nano-Ag⁰ (a) 100,000 × (b) 1,000,000 × ; TEM images and elemental composition of TiO₂-CS composite (c) 100,000 × (d) 1,000,000 × . TEM, transmission electron microscopy.

Plaque reduction assay

The result of PRA of nano-Ag⁰/TiO₂-CS against MS2 bacteriophages demonstrated the remarkable antiviral efficacy of nano-Ag⁰/TiO₂-CS, as shown in Figure 4a. However, the antiviral efficacy of TiO₂-CS beads was not significant. The release of nanosilver from nano-Ag⁰/TiO₂-CS in MS2 liquid cultures is demonstrated in Table 1. In 20 mL of MS2 liquid culture (2 ± 0.5 × 10¹⁶ PFU/mL), 3 g of nano-Ag⁰/TiO₂-CS beads can deactivate 97% of MS2 phage within 2 hours. The survival of MS2 bacteriophages decreased exponentially with the time they contacted with nano-Ag⁰/TiO₂-CS, and the inactivation rate constant for 1, 2, and 3 g of nano-Ag⁰/TiO₂-CS beads in 20 mL of MS2 liquid culture was 0.605, 0.759, and 1.219 h⁻¹, respectively. Liga et al. also investigated the virus inactivation by nano-Ag/TiO₂ prepared by photochemical reduction method for drinking water treatment. Their results showed that the inactivation rates of MS2 by nano-Ag/TiO₂ were fivefold more than those by TiO₂, and the inactivation efficiency increased with the increase of Ag content.⁽²⁷⁾

FIG. 4.

(a) PRA of TiO₂-CS and nano-Ag⁰/TiO₂-CS against MS2 bacteriophage and the inactivation rate constant; the error bars represent the standard deviations of triplicate experiments; (b) Roller Mixer for PRA. PRA, plaque reduction assay.

Table 1.

The Release of the Nanosilver in MS2 Liquid Cultures

Time, hours	Mass of nano-Ag⁰/TiO₂-CS beads; TiO₂-CS beads in 20 mL of MS2 liquid cultures
Time, hours	1 g	2 g	3 g
1	—	—	596 ± 34 ng/mL; ND
2	505 ± 32 ng/mL; ND	621 ± 38 ng/mL; ND	1005 ± 48 ng/mL; ND
4	663 ± 35 ng/mL; ND	880 ± 41 ng/mL; ND	—

LOQ = 100 ng/mL.

LOQ, limit of quantitation; nano-Ag⁰/TiO₂-CS, nanosilver/titania-chitosan.

Nano-Ag⁰ can function as a reservoir for silver ions and therefore prevent the antimicrobial silver ions from being quenched by the anion ions such as Cl⁻ and OH⁻. Conclusively, nano-Ag⁰ has better antiviral efficiency than ionic nanosilver.^(28,29)

Physical removal efficiency of nano-Ag⁰/TiO₂-CS filter for viral aerosol particles

As shown in Figure 5, the aerosols of MS2 bacteriophage were the nuclei of MS2 and media droplets and their size varied from 30 nm to 10 μm and most of the MS2 aerosol mass contributed by the particles >500 nm (in the range the OPC can cover). Therefore, the OPC was used to measure the filtration efficiency for MS2 bacteriophage aerosols. Besides, our previous study has shown that the “average size distribution of the coughed droplet nuclei was 0.58–5.42 μm, and 82% of droplet nuclei centered in 0.74–2.12 μm,⁽³⁰⁾” which was also within the range the OPC can cover. As a result, we chose the OPC to measure the filtration efficiency for MS2 bacteriophage aerosols. In addition, the OPC has the advantage of real-time measurement of airborne viral particle concentration and size distribution. However, the removal efficiency of the nano-Ag⁰/TiO₂-CS filter for viral aerosols obtained by the OPC cannot indicate the inactivation of viable viral aerosols. Thus, the culture-based method (double-layer agar culture) for the measurement of viral titer was also used to evaluate the effectiveness of the nano-Ag⁰/TiO₂-CS filter on the removal of viral aerosols.

FIG. 5.

MS2 aerosol mass-size distribution (the particles <300 nm were measured by the Scanning Mobility Particle Sizer); the error bars represent the standard deviations of triplicate experiments.

As shown in Figure 6a, the physical removal efficiency of nano-Ag⁰/TiO₂-CS filter for MS2 bacteriophage aerosols measured by the OPC was a function of the particle size of MS2 aerosols. The removal efficiency of the nano-Ag⁰/TiO₂-CS filter for MS2 aerosols increased with the increase of the MS2 aerosol particle size. The highest MS2 removal efficiency occurred at MS2 aerosol diameter of 2.5–4 μm, and the primary filtration mechanisms were inertial impaction and interception.

FIG. 6.

(a) Removal efficiency (b) penetration (c) quality factor of nano-Ag⁰/TiO₂-CS filter with different thickness (0.5, 1, 2, 4, 6 cm) for MS2 aerosol particles; the error bars represent the standard deviations of triplicate experiments.

The viral-aerosol removal efficiency increased with the increase of thickness of the nano-Ag⁰/TiO₂-CS filter. When the filter thickness was 2 cm, the removal efficiency of the nano-Ag/TiO₂-CS filter for 2.5-μm viral aerosols could reach 89%, and it increased to more than 93% when the filter thickness increased to 6 cm.

We used the model based on single-grain collection efficiency (η₀) to estimate the viral-aerosol particle removal efficiency (η_v)⁽³¹⁾: $η_{v} = 1 - e x p [- \frac{3}{2} (1 - ε) η_{0} (\frac{L}{D_{A g}})]$ (5)

where ɛ is the porosity of nano-Ag⁰/TiO₂-CS filter; L is the filter thickness; D_Ag is the diameter of nano-Ag⁰/TiO₂-CS bead. The single grain collection efficiency, η₀, was presumed to be the sum of each removal mechanism, including impaction, sedimentation, interception, and diffusion, and thus, η₀ was expressed as what follows⁽³²⁾: $η_{0} = η_{I} + η_{S} + η_{T} + η_{D}$ (6)

where η_I, η_S, η_T, and η_D represent the single-grain collection efficiency of impaction, sedimentation, interception, and diffusion, respectively. The details of these parameters can be found in Ozis et al.⁽³²⁾

Figure 7b demonstrates the penetration of viral aerosol particles through the nano-Ag⁰/TiO₂-CS filter. The lowest penetration occurred in the viral aerosol particle size range of 2.5–4 μm, while the maximum penetrating particle size for the nano-Ag⁰/TiO₂-CS filter was 0.3 μm, where the collection efficiency of inertial impaction and diffusion was insignificant. The penetration of viral aerosol particles decreased with the increase of the nano-Ag⁰/TiO₂-CS filter thickness. However, the pressure drop through the filter increased with the increase of filter thickness. A higher pressure drop (resistance) would increase the loading of the fan of the air purification system. A perfect filter requires both a lower pressure drop and lower penetration for viral aerosol particles. Herein, the filter quality factor (q_F) was applied to assess the effectiveness of the filter:

FIG. 7.

(a) The survival of MS2 bacteriophage on the surface of nano-Ag⁰/TiO₂-CS; the error bars represent the standard deviations of triplicate experiments; (b) penetration of MS2 aerosols through the nano-Ag⁰/TiO₂-CS filter and the quality factor of these filters; the error bars represent the standard deviations of triplicate experiments; (c) the probability of infection predicted by the Wells–Riley model; we suppose that the respiratory rate of a person, b = 1.25 m³/h; 600 pathogen aerosol particles are produced every hour (q = 600/h); the contact time with the infected person is 10 minutes on average (t = 10 minutes); infection-free ventilation rate (Q) is estimated by the following equation [total air exchange (assumed to be 300 m³/h) × filter bed removal efficiency = infection-free ventilation rate].

q_{F} = \frac{- l n P}{Δ p} = \frac{- l n (1 - η_{v})}{Δ p}

(7)

where q_F is the filter quality factor (1/mm-H₂O); P is the penetration of viral aerosol particles; and Δp is the pressure drop (mm-H₂O). According to Figure 6c, the filter with the best q_F was that one with a filter thickness of 4 cm, and the next three were the ones with the filter thickness of 1, 2, and 6 cm, respectively. Consequently, the nano-Ag⁰/TiO₂-CS filters with a thickness of 1, 2, 4, and 6 cm were used to test their removal efficiency on viral aerosols using the culture-based method.

The removal efficiency of nano-Ag/CS-TiO₂ filter for viable viral aerosols

In the filtration experiment for viable viral aerosols, the MS2 aerosol particles first passed through a nano-Ag⁰/TiO₂-CS filter (or TiO₂-CS filter), sampled by a modified AGI 30 sampler,⁽³³⁾ and then enumerated by the double-layer agar plate method.

The removal efficiency for MS2 aerosols increased from 0.475 to 0.979 as the filter thickness increased from 1 cm to 6 cm, and the penetration of MS2 aerosols decreased accordingly (Fig. 7a and Table 2). This result was consistent with that obtained by the OPC. However, when the filter thickness was 6 cm, the quality factor was the best, of which the result was inconsistent with that obtained by the OPC. The possible reason for this case is that the OPC measured both viable and nonviable particles, including those produced from the medium of MS2 suspension. However, the double-layer agar culture method enumerated only the viable MS2 bacteriophages. Therefore, the aerosol particles generated from the liquid medium and inactive MS2 bacteriophages could cause the bias of the experimental result.

Table 2.

Removal Efficiency and Filter Quality Factor of TiO₂-CS Filter for Viable Viral Aerosols (MS2 Bacteriophage)

Filter thickness, cm	Removal efficiency, %	Filter quality factor, KPa⁻¹
1	47.7 ± 14.7	5.45 ± 1.54
2	59.5 ± 8.5	6.95 ± 0.94
4	91.9 ± 9.4	15.62 ± 6.54
6	97.9 ± 9.7	21.91 ± 7.45

After each experiment, the nano-Ag⁰/TiO₂-CS beads (or TiO₂-CS filter) in the filter bed were extracted with 0.5% peptone (0.383 g beads were extracted with 2 mL of 0.5% peptone) to verify the MS2 bacteriophage survival on the surface of nano-Ag⁰/TiO₂-CS or TiO₂-CS (contact time was 20 minutes) and evaluate the inactivation efficacy of the filters on MS2 bacteriophages. Over 93.3% of MS2 bacteriophages deposited on nano-Ag/TiO₂-CS were inactivated within 20 minutes (survival ranged from 2.75% to 6.70%), as shown in Figure 7b.

Assessment of the efficacy of nano-Ag⁰/TiO₂-CS filter on airborne infection control

To assess the feasibility of nano-Ag⁰/TiO₂-CS filter for airborne infection control, the Wells–Riley equation^(6,34) was used to estimate the probability of infection under some scenario: $p_{i} = \frac{D}{S} = [1 - e x p (\frac{- I q b t}{Q})]$ (8)

in which p_i represents the probability of infection; D is the number of infection cases; S is the number of susceptible; I is the number of infectors; q is the quanta generation rate; b is the pulmonary ventilation rate of a person; t is the exposure duration; and Q is the infection-free ventilation rate. Figure 7c demonstrates the probability of infection predicted by the Wells–Riley model. Accordingly, when the filter thickness increased from 0 to 6 cm, the infection-free ventilation rate increased from 0 to 293 m³/h, and the probability of infection decreased from 99% to 34.6%, implying a promising potential of nano-Ag⁰/TiO₂-CS filter on the control airborne infection.

Assessment of the long-term antiviral efficiency of nano-Ag⁰/TiO₂-CS filter

Joe et al.,⁽³⁵⁾ developed a model to predict the temporal antiviral efficiency of a silver nanoparticle coated air filter, and we used this model to evaluate the long-term antiviral efficiency of nano-Ag⁰/TiO₂-CS filter:

where η_anti-vir (t) is the temporal antiviral efficiency of nano-Ag⁰/TiO₂-CS filter; ρ_Ag is the coating areal density of the 2 wt% nano-Ag⁰ on the surface of TiO₂-CS beads (∼3 × 10¹⁶ nano-Ag⁰ particles/cm²); N″_in is the flux of upstream virus aerosols flowing to the nano-Ag⁰/TiO₂-CS filter (2.08 × 10¹¹ PFU/cm²/min); t is operation time (minutes); and SR is the survival ratio [Eq. (4)] that signifies the vulnerability of the MS2 bacteriophages against nano-Ag⁰/TiO₂-CS (obtained from Fig. 8a). Assuming that SR declines with an increase in MS2 aerosol particles' coverage on the surface of nano-Ag⁰, which increases with the filter operation time, and thus, the SR can be modeled as what follows:

FIG. 8.

(a) SR of MS2 bacteriophage on the surface of nano-Ag⁰/TiO₂-CS to those on the surface of TiO₂-CS; the error bars represent the standard deviations of triplicate experiments; (b) The plot of 1/SR versus M″_d; (c) Prediction of long-term antiviral efficiency of nano-Ag⁰/TiO₂-CS filter. SR, survival ratio.

where SR₀ represents the vulnerability of the virus against “clean” nano-Ag⁰/TiO₂-CS (no covering on the surface of nano-Ag⁰); γ is a constant (cm²/μg); and M″_d is the mass flux of MS2 aerosol particles (including the media particles) deposited on the surface of nano-Ag⁰/TiO₂-CS filter (μg/cm²/min), which can be obtained by the following equation:

where η_filter-dp is particle-diameter-dependent removal efficiency of nano-Ag⁰/TiO₂-CS filter for aerosol particles (obtained from Fig. 6a); C_in-dp is the particle-diameter-dependent mass concentration of upstream MS2 and media aerosol particles flowing to the nano-Ag⁰/TiO₂-CS filter (obtained from Fig. 5); U_filter is the face velocity of the nano-Ag⁰/TiO₂-CS filter; and A_filter is the surface area of nano-Ag⁰/TiO₂-CS filter. To obtain κ₀ and γ, we rearrange Equation (10), and get:

The plot of 1/SR versus M″_d has an interception of 1/SR₀ on the y-axis and a slope of γt/SR₀, as shown in Figure 8b. Replacing the term, SR, in Equation (9) with the right side of Equation (10), we get,

We can use Equation (13) to predict the long-term antiviral efficiency of the nano-Ag⁰/TiO₂-CS filter, as shown in Figure 8c. According to the model's prediction, even after continuous operation for 10,000 minutes (around 1 week), the filter could remain at 50% of its original antiviral efficiency when the filter thickness was larger than 4 cm.

Conclusions

In this study, the nano-Ag⁰/TiO₂-CS filter was used for the removal and deactivation of airborne MS2 bacteriophage particles. In the air purification system, the nano-Ag⁰/TiO₂-CS filter could remove up to 93% of airborne MS2 particles, and more than 95% of the MS2 can be efficiently deactivated on the surface of nano-Ag⁰/TiO₂-CS within 20 minutes. According to the prediction of the Wells–Riley model, the infection-free ventilation delivered by the nano-Ag⁰/TiO₂-CS filter could reduce the infection probability from 99% to 34.6%. The filter could maintain 50% of its original antiviral efficiency after continuous operating for 1 week. Conclusively, the nano-Ag⁰/TiO₂-CS filter could be a potential air purification technique to reduce airborne transmission.

Footnotes

Acknowledgments

The authors also thank Miss Yu Wei Tsai and Hsin-Jui Lee for the help with experiments.

Author Disclosure Statement

Dr. I-Jen Wang is an employee of Taipei Hospital, Ministry of Health and Welfare of Taiwan. Dr. Yen-Chi Chen, Chien Su, Wan-Tien Shen, Chun-Hsuan Bai, Ming-Hsuan Tsai, and Dr. Kuo-Pin Yu declare that they have no conflict of interest.

Funding Information

This study is financially supported by the Ministry of Science and Technology of Taiwan (MOST 105-2221-E-010-002-MY3) and Taipei Hospital, Ministry of Health and Welfare. The authors are grateful to the Electron Microscopy Facility at National Yang-Ming University for providing the TEM and SEM images.

Reviewed by:

Tim Corcoran

C.-Y. Wu

References

, Li

, Wong

, Tam

, Chan

, Lee

, Leung

, and Ho

: Evidence of airborne transmission of the severe acute respiratory syndrome virus. N Engl J Med. 2004; 350:1731–1739.

Kim

S-H

, Chang

, Sung

, Park

, Bin Kim

, Lee

, Choi

J-P

, Choi

, and Min

J-Y

: Extensive viable Middle East respiratory syndrome (MERS) coronavirus contamination in air and surrounding environment in MERS isolation wards. Rev Infect Dis. 2016; 63:363–369.

Paules

, Marston

, and Fauci

: Coronavirus infections—More than just the common cold. JAMA. 2020; 323:707–708.

Moser

, Bender

, Margolis

, Noble

, Kendal

, and Ritter

: An outbreak of influenza aboard a commercial airliner. Am J Epidemiol. 1979; 110:1–6.

Herfst

, Schrauwen

, Linster

, Chutinimitkul

, de Wit

, Munster

, Sorrell

, Bestebroer

, Burke

, and Smith

: Airborne transmission of influenza A/H5N1 virus between ferrets. Science. 2012; 336:1534–1541.

Riley

, Murphy

, and Riley

: Airborne spread of measles in a suburban elementary school. Am J Epidemiol. 1978; 107:421–432.

Bloch

, Orenstein

, Ewing

, Spain

, Mallison

, Herrmann

, and Hinman

: Measles outbreak in a pediatric practice: Airborne transmission in an office setting. Pediatrics. 1985; 75:676–683.

Remington

, Hall

, Davis

, Herald

, and Gunn

: Airborne transmission of measles in a physician's office. JAMA. 1985; 253:1574–1577.

Alonso

, Goede

, Morrison

, Davies

, Rovira

, Marthaler

, and Torremorell

: Evidence of infectivity of airborne porcine epidemic diarrhea virus and detection of airborne viral RNA at long distances from infected herds. Vet Res. 2014; 45:73–78.

10.

Mikkelsen

, Alexandersen

, Astrup

, Donaldson

, Dunkerley

, Gloster

, Champion

, Sørensen

, and Thykier-Nielsen

: Investigation of airborne foot-and-mouth disease virus transmission during low-wind conditions in the early phase of the UK 2001 epidemic. Atmos Chem Phys Discuss. 2003; 3:677–703.

11.

van Doremalen

, Bushmaker

, Morris

, Holbrook

, Gamble

, Williamson

, Tamin

, Harcourt

, Thornburg

, Gerber

, Lloyd-Smith

, de Wit

, and Munster

: Aerosol and surface stability of SARS-CoV-2 as compared with SARS-CoV-1. N Engl J Med. 2020; 382:1564–1567.

12.

Ong

SWX

, Tan

, Chia

, Lee

, Ng

, Wong

MSY

, and Marimuthu

: Air, surface environmental, and personal protective equipment contamination by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from a symptomatic patient. JAMA. 2020; 323:1610–1612.

13.

Knibbs

, Morawska

, Bell

, and Grzybowski

: Room ventilation and the risk of airborne infection transmission in 3 health care settings within a large teaching hospital. Am J Infect Control. 2011; 39:866–872.

14.

Brankston

, Gitterman

, Hirji

, Lemieux

, and Gardam

: Transmission of influenza A in human beings. Lancet Infect Dis. 2007; 7:257–265.

15.

McClean

: The effect of ultraviolet radiation upon the transmission of epidemic influenza in long-term hospital patients. Am Rev Respir Dis. 1961; 83:36–38.

16.

Galdiero

, Falanga

, Vitiello

, Cantisani

, Marra

, and Galdiero

: Silver nanoparticles as potential antiviral agents. Molecules. 2011; 16:8894–8918.

17.

Rai

, Deshmukh

, Ingle

, Gupta

, Galdiero

, and Galdiero

: Metal nanoparticles: The protective nanoshield against virus infection. Crit Rev Microbiol. 2016; 42:46–56.

18.

, Sun

, Chen

, Hui

C-K

, Ho

C-M

, Luk

, Lau

, and Che

C-M

: Silver nanoparticles inhibit hepatitis B virus replication. Antivir Ther. 2008; 13:253–262.

19.

Chen

, Liao

, Shen

, Su

, Wu

, Tsai

, Hsiao

, Yu

, and Tseng

: Effective disinfection of airborne microbial contamination in hospital wards using a zero-valent nano-silver/TiO₂-chitosan composite. Indoor Air. 2019; 29:439–449.

20.

Lindsley

, Blachere

, Thewlis

, Vishnu

, Davis

, Cao

, Palmer

, Clark

, Fisher

, and Khakoo

: Measurements of airborne influenza virus in aerosol particles from human coughs. PLoS One. 2010; 5:e15100.

21.

Pan

, Carol

, Lednicky

, Eiguren-Fernandez

, Hering

, Fan

, and Wu

C-Y

: Determination of the distribution of infectious viruses in aerosol particles using water-based condensational growth technology and a bacteriophage MS2 model. Aerosol Sci Technol. 2019; 53:583–593.

22.

Qian

, Hospodsky

, Yamamoto

, Nazaroff

, and Peccia

: Size-resolved emission rates of airborne bacteria and fungi in an occupied classroom. Indoor Air. 2012; 22:339–351.

23.

Chmiel

, Kral

, and Lenart-Boroń

: Concentration and size distribution of microbial aerosol in the historical objects in Kraków as a potential health risk and biodeterioration factor. Aerobiologia. 2019; 35:743–758.

24.

K-P

, Yu

W-Y

, Kuo

M-C

, Liou

Y-C

, and Chien

S-H

: Pt/titania-nanotube: A potential catalyst for CO2 adsorption and hydrogenation. Appl Catal B. 2008; 84:112–118.

25.

Jones

, Bellamy

, Alcock

, and Hudson

: The use of bacteriophage MS2 as a model system to evaluate virucidal hand disinfectants. J Infect Control. 1991; 17:279–285.

26.

Maillard

J-Y

, Beggs

, Day

, Hudson

, and Russell

: Effect of biocides on MS2 and K coliphages. Appl Environ Microbiol, 1994; 60:2205–2206.

27.

Liga

, Bryant

, Colvin

, and Li

: Virus inactivation by silver doped titanium dioxide nanoparticles for drinking water treatment. Water Res. 2011; 45:535–544.

28.

Xiu

Z-m

, Zhang

Q-b

, Puppala

, Colvin

, and Alvarez

: Negligible particle-specific antibacterial activity of silver nanoparticles. Nano Lett. 2012; 12:4271–4275.

29.

Badawy

AME

, Luxton

, Silva

, Scheckel

, Suidan

, and Tolaymat

. Impact of environmental conditions (pH, ionic strength, and electrolyte type) on the surface charge and aggregation of silver nanoparticles suspensions. Environ Sci Technol. 2010; 44:1260–1266.

30.

Yang

, Lee

, Chen

, Wu

, and Yu

: The size and concentration of droplets generated by coughing in human subjects. J Aerosol Med. 2007; 20:484–494.

31.

Golshahi

, Abedi

, and Tan

: Granular filtration for airborne particles: Correlation between experiments and models. Can J Chem Eng. 2009; 87:726–731.

32.

Ozis

, Singh

, Devinny

, and Sioutas

: Removal of ultrafine and fine particulate matter from air by a granular bed filter. J Air Waste Manag Assoc. 2004; 54:935–940.

33.

, Chen

, Gong

, Chen

, and Cheng

: Improving the collection efficiency of the liquid impinger for ultrafine particles and viral aerosols by applying granular bed filtration. J Aerosol Sci. 2016; 101:133–143.

34.

Rudnick

and Milton

: Risk of indoor airborne infection transmission estimated from carbon dioxide concentration. Indoor Air. 2003; 13:237–245.

35.

Joe

, Park

, and Hwang

: Evaluation of Ag nanoparticle coated air filter against aerosolized virus: Antiviral efficiency with dust loading. J Hazard Mater. 2016; 301:547–553.

Effectiveness of the Nanosilver/TiO 2 -Chitosan Antiviral Filter on the Removal of Viral Aerosols

Abstract

Background:

Methods:

Results and Conclusions:

Introduction

Materials and Methods

Preparation of MS2 bacteriophage suspension

Preparation and characterization of the nano-Ag0/TiO2-CS composite

Evaluation of antiviral efficacy of nano-Ag0/TiO2-CS composite

Experimental system

Results and Discussion

Characteristics of nano-Ag0/TiO2-CS antiviral composite

Plaque reduction assay

Physical removal efficiency of nano-Ag0/TiO2-CS filter for viral aerosol particles

The removal efficiency of nano-Ag/CS-TiO2 filter for viable viral aerosols

Assessment of the efficacy of nano-Ag0/TiO2-CS filter on airborne infection control

Assessment of the long-term antiviral efficiency of nano-Ag0/TiO2-CS filter

Conclusions

Footnotes

Acknowledgments

Author Disclosure Statement

Funding Information

References

Preparation and characterization of the nano-Ag⁰/TiO₂-CS composite

Evaluation of antiviral efficacy of nano-Ag⁰/TiO₂-CS composite

Characteristics of nano-Ag⁰/TiO₂-CS antiviral composite

Physical removal efficiency of nano-Ag⁰/TiO₂-CS filter for viral aerosol particles

The removal efficiency of nano-Ag/CS-TiO₂ filter for viable viral aerosols

Assessment of the efficacy of nano-Ag⁰/TiO₂-CS filter on airborne infection control

Assessment of the long-term antiviral efficiency of nano-Ag⁰/TiO₂-CS filter