Antiviral Characterization of Advanced Materials: Use of Bacteriophage Phi 6 as Surrogate of Enveloped Viruses Such as SARS-CoV-2

Abstract

The bacteriophage phi 6 is a virus that belongs to a different Baltimore group than SARS-CoV-2 (group III instead of IV). However, it has a round-like shape and a lipid envelope like SARS-CoV-2, which render it very useful to be used as a surrogate of this infectious pathogen for biosafety reasons. Thus, recent antiviral studies have demonstrated that antiviral materials such as calcium alginate hydrogels, polyester-based fabrics coated with benzalkonium chloride (BAK), polyethylene terephthalate (PET) coated with BAK and polyester-based fabrics coated with cranberry extracts or solidified hand soap produce similar log reductions in viral titers of both types of enveloped viruses after similar viral contact times. Therefore, researchers with no access to biosafety level 3 facilities can perform antiviral tests of a broad range of biomaterials, composites, nanomaterials, nanocomposites, coatings and compounds against the bacteriophage phi 6 as a biosafe viral model of SARS-CoV-2. In fact, this bacteriophage has been used as a surrogate of SARS-CoV-2 to test a broad range of antiviral materials and compounds of different chemical natures (polymers, metals, alloys, ceramics, composites, etc.) and forms (films, coatings, nanomaterials, extracts, porous supports produced by additive manufacturing, etc.) during the current pandemic. Furthermore, this biosafe viral model has also been used as a surrogate of SARS-CoV-2 and other highly pathogenic enveloped viruses such as Ebola and influenza in a wide range of biotechnological applications.

Keywords: SARS-CoV-2; antiviral characterization; antiviral materials; bacteriophage phi 6; biosafety conditions; coatings; composites; extracts; films; nanomaterials; porous supports.

Figure 1

Bacteriophage phi 6 and SARS-CoV-2. Created by Ángel Serrano-Aroca with Biorender.

Figure 2

Microscopic images of the bacteriophage phi 6 and SARS-CoV-2: (A) Cryo-electron microscopy image of the nucleocapsids (NCs) of the bacteriophage phi 6 highlighted with black arrows. A partially disrupted NC is pointed out by a white arrow, where the core can be appreciated with a clear angular inner layer. Reprinted in part with permission from [52]. Copyright 1997 JOHN WILEY AND SONS. (B) Diameter of SARS-CoV-2 viral particles attached in cell membrane (white arrow) (helium ion microscopy image). The number of measured particles (N), mean (M), standard deviation (SD) and virus particles (arrow) are indicated [51].

Figure 3

Antiviral properties of biocompatible calcium alginate films against enveloped viruses such as the bacteriophage phi 6 and SAS-CoV-2. Calcium alginate swollen structure in viral aqueous solution. Cell viability results in human keratinocytes after performing ANOVA with subsequent Tukey’s post hoc test: *** p > 0.001; ns, not significant [5].

Figure 4

Antimicrobial face mask (FFPCOVID MASK) that inactivates enveloped viruses such as the bacteriophage phi 6 and SARS-CoV-2, and MRSA and MRSE multidrug-resistant bacteria, from UCV Research-Visormed [63] (left); protective face masks: difference between conventional face masks and antimicrobial face masks (right). Created by Ángel Serrano-Aroca with Biorender.

Figure 5

Antiviral characterization of an antimicrobial face shield using the bacteriophage phi 6 as a viral model of SARS-CoV-2 for biosafety reasons: (a) Antimicrobial face shield developed by the Serrano BBlab (www.serranobblab.com, accessed on 19 March 2022): next generation of preventive equipment against infections caused by enveloped viruses such as SARS-CoV-2 and multidrug-resistant bacteria. The material is composed of polyethylene terephthalate (PET) coated with benzalkonium chloride (BAK). The double-layer method was used to determine the loss of viral viability after 1 min of viral contact: (b) Bacteriophage phi 6 titration images of undiluted samples for the materials. The reduction in infection capacity can be observed by the reduction in white spots. (c) Decrease in infection titers expressed in plaque-forming units per mL (PFU/mL). CONTROL: bacteriophages without being in contact with any material; U Plastic: untreated PET; S plastic: PET treated with solvent; BAK plastic: PET treated with solvent and BAK [20].

Figure 6

Antiviral characterization of non-woven fabrics coated with two types of commercial cranberry extracts against the bacteriophage phi 6 for biosafety reasons: (A) The double-layer method was used to determine the viral viability after 1 min of viral contact (titration images of undiluted samples). These images show the reduction in infection capacity (reduction in white spots). (B) Reduction in infection titers of the bacteriophage phi 6 in a logarithm of plaque-forming units per mL (log(PFU/mL)) measured by the double-layer method at 1 min of viral contact. Statistical analysis: *** p > 0.001; ** p > 0.01; ns: not significant. (C) High-resolution field-emission scanning electron microscopy (HR-FESEM) of the non-woven fabrics, at two different magnifications (×100 and ×1000), before (a,b) and after the treatment with the VITAFAIR cranberry extract (E10V) (c,d) or the NUTRIBIOLITE cranberry extract (E10N) (e,f). CONTROL: bacteriophages without being in contact with any material; Control S: uncoated non-woven fabric [10].

Figure 7

Cobalt-chromium-molybdenum porous superalloy with superior antiviral activity fabricated by additive manufacturing. Antiviral filters were tested using the bacteriophage phi 6 as a surrogate of SARS-CoV-2 for biosafety reasons [13].