surface coatings that rapidly inactivate sars-cov-2
TRANSCRIPT
William DuckerSaeed Behzadinasab, Mohsen Hosseini
Dept. of Chemical Engineering and Center for Soft Matter and Biological Physics, Virginia Tech
Alex Chin and Leo Poon (SARS-CoV-2 tests)
School of Public Health, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong
Special Administrative Region, China.
Joseph Falkinham (Bacterial and Fungal Tests)
Virginia Tech, Biological Sciences
Surface Coatings that Rapidly Inactivate SARS-CoV-2, Bacteria, and Fungi
COVID-19• ~ 200 million people infected worldwide (WHO)
• ~ 4.5 million deaths worldwide (WHO)
2
100 M
50 M
cases
deaths
1 M
2 M
3 M
https://coronavirus.jhu.edu/map.html
September 2021
3
Alien Invasion
World War
Nuclear War
Climate Change
Covid-19
Prof. Ducker I need three more points on Q4
…
…
What can I doto help?
Infection RoutesSubject of Current Research
4
Main Route: via respiratory droplets or aerosols
Secondary Route: via solids “fomite”
Fomite Research:Hamster studies: Transfer via Fomites, Sia,. Nature 2020,Epidemiology: 25% via fomites, Meiksin, A. Epidemiol. Infect. 2020
“People who come into contact with potentially infectious surfaces often also have close contact with the infectious person, making the distinction between respiratory droplet and
fomite transmission difficult to discern.” WHO
Viability on a Solid
5
Higher numbers = Better at infecting cells
1. Van Doremalen et al., N Engl. J. Med., 2020 382, 1564
6
1. Chin et al., Lancet Microbe., 2020, 1, e10
SARS-CoV-2 viable on solids for up to one week
Research Concept
7
Ducker Group Research
8
HypothesisA coating can be used to speed the inactivation of
SARS-CoV-2 on solids
ApplicationIf such a coating were applied to communal objects,
then this might reduce the spread of COVID-19 and other infectious diseases
Desired features of a coating
9
• Inactivate the novel coronavirus quickly –minutes
• Ongoing or continuous “kill”
• Applicable to everyday objects regardless of material
• High durability
• Easy to Apply • Active material• Support structure• Morphology
Coating Parameters
Active Material
10
Approaches• Deliberate design• Consider what works
Design• Virus assembles spontaneously• Change solution conditions such as
assembly is no longer stable (e.g., ethanol, soap)
• Knock out a necessary function
Kim et al., 2020, Cell 181, 914–921
Changing the Environment
11
Polymer• Highly Charged
• Changes electrostatic interactions• Dangles out into solution
• Maximum contact with virus• Tether to a solid
GOPS
PAH
This coating very effective at killing bacterial
Healthy Vero E6 cells
Cytopathic effect (CPE)
12
Test of Viral ActivityUse Vero E6 cells (Green Monkey kidney cells) as a proxy for human cells
Microscope image Showing complete layer of healthy cells
incomplete layer of curled-up or detached cells
Results: Polyallylamine Films
13
Big dilution before virus No longer kills cells BAD
Little effect on SARS-CoV-2 from the tethered cationic polymer PAA.
Big dilution before virus No longer kills cells GOOD
Active Material
14
Approaches• Deliberate design – failed so far• Consider what works
Copper Stainless Steel
Van Doremalen et al., N Engl. J. Med., 2020 382, 1564
Sunada et al., J. Hazard. Mat., 2012, 235, 265
• Surface of Copper is Cu2O
• Suspension of Cu2O particles inactivate bacteriophage virus2
• Try a surface bound layer of Cu2O.
Methods1. Apply a thin film of polyurethane (PU) using sponge
2. Allow for partial curing of PU
3. Cover the film with Cu2O suspension
4. Thermally anneal the film at 120 C
5. Thoroughly wash the film
6. Ar plasma treat the surface
15Brown, red, maroon, burgundy
Cu2O Results on Glass
16
• Viral reduction: >99.98% (=3.64 log, p=5x10-4, CI95%=[99.95,100]) in 1 h
• Dramatic change in the SARS-CoV-2 viability in 1 hour
lots of active virus
not much active virus
log reduction = mean log10control titer
sample titer
• Half life = log 2
slope= 3.5 min, 95% C.I. = [2.5, 4.2] min
Cu2O on Glass: Short-term Results
17
Results: stainless steel 301
18
• Viral reduction: 99.90% (=2.97 log, p=8x10-3, CI95%=[98.51,100]) in 1 h
• Similarity of results on glass and stainless steel suggest that the coating is responsible for the inactivation of the virus (p=0.59)• Behzadinasab 2020 ACS Applied Materials and Interfaces
• Press coverage on BBC World, NPR, ABC, NBC, CBS, Fox
Results: Ongoing Kill
19
• Material: glass• 1 cycle =
exposed to SARS-CoV-2, disinfected with 70% ethanol in water.
• Viral reduction: 99.89% (=2.95 log, p=4x10-8, CI95%=[99.79,100]) in 1 h
• Coating does not lose its activity after repeated exposures to the viral particles
Durability
20
Cross-hatch test ASTM D3359-Method B
• Material: stainless steel• 3 samples were tested• Affected squares:
• avg. = 2.4• std. dev. = 0.8
Characterization: XPS
21
• Binding energy = 932.7 eV, can be assigned to either Cu metal or Cu2O
• Kinetic energy = 916.2 eV• Eb+Ek = 1848.9 eV is in excellent agreement with the value of Cu2O
After plasma treatment
Before plasma treatment
Cu 2P3/2
Cu L3M4,5M4,
5
22
First Draft: Cu2O in polyurethane
Also kills(1) Bacteria:Escherichia coliStaphylococcus aureusMethicillin-resistant S. aureus (MRSA)Pseudomonas aeruginosa, Acinetobacter baumannii,Stenotrophomonas maltophilia,: Mycobacterium avium, Mycobacterium chimaera,Mycobacterium abscessus, (2) yeasts:Candida albicansCandida auris, (3) fungus:Aspergillus niger
Prof. Joe FalkinhamMyra GordonVirginia Tech
Conclusion
23
We have developed a surface coating that:• Inactivates SARS-CoV-2 and
other organisms quickly• Is applicable to communal
objects• Has very high durability at
various conditions• Is mechanically robust
The Future I Porous Films
24
Inactivation in minutes: Imbibe the droplet into a surface film.
25
Hydrophilic Porous CoatingNo polyurethane, sinter Cu2O
Cuprous oxide Cupric oxide
26
Imbibition of water into CuO
Hydrophilic for months
5 µL droplet
Rapid loss of Infection Ability
27
28
99.7% reduction compared to glass within 1 minute (p = 0.0189)
Loss of Infection Ability in < 1 min
Peel test on CuO Coating
29
30
31
Transparent Films
32
Transparent Films
Acknowledgments
33
The Ducker lab• Saeed Behzadinasad• Mohsen Hosseini• Zac Benmamoun• Jared Arkfeld
The Poon lab• Prof. Leo Poon• Prof. Alex Chin
NSF CBET 1902364
NCFL• Steve McCartney
Surface Analysis Lab• Xu Feng
Supporting Slides
Cross-hatch test ASTM D3359-Method B
• Material: stainless steel• 3 samples were tested• Affected squares:
• avg. = 2.4• std. dev. = 0.8
35
Results: Durability
Characterization
36
• X-section SEM image of film
• Methods: the samples were coated with 5 nm of Iridium
• Film is uniformthickness = ~12 μm
• Size distribution of Cu2O particles. Mean = 5.1 μm
• Data from the manufacturer (Chemet Corporation)
2020.04.29 07:41:39
1 / 1
Cumulative % at diameter :
:
:
:
:
:
(1)0.500 (µm)- 0.411(%)
(2)5.000 (µm)- 55.097(%)
(3)425.0 (µm)- 100.000(%)
(4)300.0 (µm)- 100.000(%)
(5)212.0 (µm)- 100.000(%)
(6)150.0 (µm)- 100.000(%)
:
:
:
:
(7)106.0 (µm)- 100.000(%)
(8)75.00 (µm)- 100.000(%)
(9)45.00 (µm)- 100.000(%)
(10)38.00 (µm)- 100.000(%)
HORIBA Laser Sca tte ring Particle Size Dis tribution Ana lyzer LA-960
Diameter on cumulative % :
:
:
:
:
:
:
(2)10.00 (%)- 1.9220 (µm)
(3)20.00 (%)- 2.7327 (µm)
(5)50.00 (%)- 4.6653 (µm)
(7)70.00 (%)- 6.1218 (µm)
(8)80.00 (%)- 7.1315 (µm)
(9)90.00 (%)- 8.7096 (µm)
(10)99.99 (%)- 19.6850 (µm)
ID#
Sam p le nam e
Material
Source
Lot num b er
:
:
:
:
:
201910171252308
Chem et Ult rafine Cup rous Oxide-5
Cu2O
JG
T2696
Test o r assay num b er
Transm it t ance (R)
Transm it t ance (B)
Circulat ion sp eed
Agitat ion speed
:
:
:
:
:
117741
87.6 (%)
87.2 (%)
4
4
Ult rasound
Iterat ion m ode
:
:
Off
Manual
0
12
2
4
6
8
10
0.010 50000.100 1.000 10.00 100.0 1000
0
100
10
20
30
40
50
60
70
80
90
q (
%)
Diameter (µm)
Un
de
rsiz
e (
%)
No.
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
Unders ize (%)
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
Diameter (µm)
0.011
0.013
0.015
0.017
0.020
0.022
0.026
0.029
0.034
0.039
0.044
0.051
0.058
0.067
0.076
0.087
0.100
0.115
0.131
0.150
0.172
0.197
0.226
0.259
q (%)
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
No.
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
Unders ize (%)
0.000
0.000
0.110
0.253
0.437
0.673
0.973
1.355
1.848
2.487
3.329
4.464
5.991
8.000
10.571
13.857
18.041
23.321
29.867
37.759
46.889
56.874
67.033
76.503
Diameter (µm)
0.296
0.339
0.389
0.445
0.510
0.584
0.669
0.766
0.877
1.005
1.151
1.318
1.510
1.729
1.981
2.269
2.599
2.976
3.409
3.905
4.472
5.122
5.867
6.720
q (%)
0.000
0.000
0.110
0.143
0.184
0.236
0.300
0.382
0.493
0.639
0.842
1.135
1.527
2.009
2.570
3.286
4.185
5.280
6.546
7.892
9.130
9.984
10.159
9.470
No.
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
Unders ize (%)
84.484
90.540
94.734
97.383
98.855
99.569
99.877
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
Diameter (µm)
7.697
8.816
10.097
11.565
13.246
15.172
17.377
19.904
22.797
26.111
29.907
34.255
39.234
44.938
51.471
58.953
67.523
77.339
88.583
101.460
116.210
133.103
152.453
174.616
q (%)
7.980
6.056
4.195
2.649
1.472
0.714
0.308
0.123
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
No.
73
74
75
76
77
78
79
80
81
82
83
84
85
Undersize (%)
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
100.000
Diameter (µm)
200.000
229.075
262.376
300.518
344.206
394.244
451.556
517.200
592.387
678.504
777.141
890.116
1000.000
q (%)
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
Diam eter on %
Mean size
Median size
Mode size
Span
Variance
St. Dev.
COV
Geo. mean size
:
:
:
:
:
:
:
:
5.07892 (µm)
4.66531 (µm)
5.4632 (µm)
Off
7.5916 (µm 2)
2.7553 (µm)
54.2494 (%)
4.3166 (µm)
Chi Square
% on Diam eter
D10
D90
:
:
1.92196 (µm)
8.70960 (µm)
Convergence factor
Refractive index (R)
:
:
10
Cuprous Hoganas[copper oxide( 2.710 - 0.000i),ethanol( 1.360)]
0.01 0.10 1.0 10.0 100.0Diameter (micron)
1
2
1
0
8
6
4
2
0
q (
%)
TCID50
37
It’s the dilution necessary to see an effect
+
+
+
+
+
+
+
+
+
+
+
-
+
-
-
-
-
-
-
-
+ Cytopathic effect – Healthy cells
50% affected at dilution of 10-1.75
Well Plate
replicates
10-0.5 10-1 10-1.5 10-2 10-2.5
Dilution of virus after on test solid
1. Grow Vero E6 cells in each of many wells in a well plate2. Make series of dilutions of the virus after recovered from solid3. Expose cells to virus
Results: Polyurethane Only
38
• Viral reduction: 10% (=0.04 log, p=0.22, CI95%=[-164,100]) in 1 h
• Cuprous oxide is necessary for the virucidal activity, likely the active ingredient
Can you become infected by SARS-CoV-2 by touching a contaminated surface?
39
CDC: https://www.cdc.gov/coronavirus/2019-ncov/more/science-and-research/surface-
transmission.html#ref8
Science Brief: SARS-CoV-2 and Surface (Fomite) Transmission for Indoor Community
Environments
“The principal mode by which people are infected with SARS-CoV-2 (the virus that causes COVID-19) is through exposure to respiratory droplets carrying infectious virus. It is possible for people to be infected through contact with contaminated surfaces or objects (fomites), but the risk is generally considered to be low.”“Because of the many factors affecting the efficiency of environmental transmission, the relative risk of fomite transmission of SARS-CoV-2 is considered low compared with direct contact, droplet transmission, or airborne transmission 1, 2”
(Ref 1 is a model based on assumptions, Ref 2 is a review)
However, it is not clear what proportion of SARS-CoV-2 infections are acquired through surface transmission.” ”Hand hygiene is a barrier to fomite transmission and has been associated with lower risk of infection 6.” …“the risk of SARS-CoV-2 infection via the fomite transmission route is low, and generally less than 1 in 10,000, which means that each contact with a contaminated surface has less than a 1 in 10,000 chance of causing an infection 7, 8, 9.
Causality
40
Epidemiology
Modelling of outbreaks suggest that transmission via fomites may contribute up to 25% of deaths during periods of lockdown.
Meiksin A: Dynamics of COVID-19 transmission including indirect transmission mechanisms: a mathematical analysis. Epidemiol Infect
2020, 148:E257.
A study on Golden Hamsters showed that the virus can be indirectly transmitted through fomites. Sia et al: Pathogenesis and transmission of SARS-CoV-2 in golden
hamsters. Nature 2020, 583:834-838
• Leaching of ions, e.g., Cu2+, Cu+, • Oxidation of viral components• Disruption of spike proteins / lipid layer
• Surface catalysis to produce reactive oxygen species• Contact killing
• disassembly on adsorption• photocatalysis
Mechanism
41
Sunada et al. J. Hazard. Mat., 2012, 235, 265
Methods: viral tests1. Disinfect the solid with 70% ethanol in water
2. Dry in air
3. Apply a 5-𝜇L droplet on the surface. The droplet contains 7.8 Log unit TCID50/mL of SARS-CoV-2
4. Wait for a prescribed period of time, during which droplet evaporates (temp.: 22-23 °C, rel. hum.: 60-70%)
5. Immerse the sample with 300 𝜇L of viral transport medium to remove virus
6. Test viral solution to see if active.
7. The eluted virus is titrated by a 50% tissue culture infective dose assay in Vero E6 cells (i.e. Green Monkey kidney cells)
42
Cu2O toxicity• Abundant copper presence for thousands of years
• Toxicity Category III by the EPA: slightly toxic and slightly irritating1
• A 2009 EPA report1: “Current available literature and studies do not indicate any systemic toxicity associated with copper exposure. … There are no residential or occupational risks of concern resulting from exposure to copper products.”
• Used as marine antifoulant and it has an adverse effect on marine organisms2
• Clinical and animal studies of textiles containing copper oxide reports no effects on human skin.3
43
1. EPA Reregistration Eligibility Decision (RED) for Coppers; https:// www3.epa.gov/pesticides/…/.pdf 2009.
2. Amara et al. Environ. Toxicol. Pharmacol. 2018, 57, 115.
3. Borkow. Curr. Chem. Biol. 2012, 6, 86.
Introduction: SARS-CoV-2SARS-CoV-2 is a ssRNA virus
Spike protein • Binds with the receptor on cells
Envelope• Is consist of mainly lipids
44https://covid19.who.int
Characterization
45
Cross-hatch test ASTM D3359-Method B
• Material: stainless steel• 3 samples were tested• Affected squares:
• avg. = 2.4• std. dev. = 0.8
Results: Durability
46
• Viral reduction: 99.96% (=3.39 log, p=8x10-4, CI95%=[99.56,100]) in 1 h
• Film still active after storage under water