Evaluation of respiratory virus transmissibility and resilience from fomites: the case of 11 SARS-CoV-2 clinical isolates
Sofia Sisti, Elena Criscuolo, Benedetta Giuliani, Mattia Cavallaro, Michela Sampaolo, Matteo Castelli, Roberto Burioni, Massimo Locatelli, Nicola Clementi
Applied and Environmental Microbiology, doi:10.1128/aem.00774-25
The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused a global pandemic, significantly impacting social and economic aspects of society. While belonging to the Coronaviridae family, SARS-CoV-2 exhibited unique characteristics that facilitated its widespread transmission and pathogenicity. This study aimed to investigate the molecular characterization and infection kinetics of emerging SARS-CoV-2 variants, focusing on four key virological aspects: viral entry mechanisms, replication dynamics, immune evasion strategies, and surface persistence. We isolated 11 SARS-CoV-2 variants from clinical samples and conducted in vitro studies using Calu-3 cell lines and pseudovirus systems to explore viral entry. Wholegenome sequencing identified mutations linked to increased replication and enhanced immune evasion, particularly through the inhibition of interferon responses. Additionally, persistence studies on common environmental surfaces (copper, aluminum, and plastic) revealed that certain variants, such as G446S, contributed to increased viral stability, suggesting potential for indirect transmission beyond direct person-to-person contact. Furthermore, we evaluated the effectiveness of "green" disinfectants, including tea tree oil and quercetin-based compounds, which demonstrated viral load reductions of over 95% within minutes, outperforming conventional alcohol-and UV-based disinfection methods. Notably, two influenza virus isolates belonging to highly divergent subtypes are sanitized as well while showing different persistence surface profiles compared to SARS-CoV-2. These findings highlight the evolutionary strategies employed by SARS-CoV-2 to ensure transmission, its ability to adapt under selective pressure, and the importance of ongoing surveillance of emerging variants. Moreover, the efficacy of environmentally sustainable disinfectants presents a promising strategy for reducing viral transmission in both clinical and community settings. IMPORTANCE In this study, we evaluated the molecular profile of 11 SARS-CoV-2 variants, focusing on four key virological aspects: viral entry mechanisms, replica tion dynamics, immune evasion strategies, and surface persistence. Through wholegenome sequencing, we identified mutations linked to enhanced replication and immune evasion, notably through the suppression of interferon responses. Additionally, persistence studies on common environmental surfaces (copper, aluminum, and plastic) demonstrated that certain mutations, such as G446S, exhibited increased stability, suggesting a potential role for indirect transmission. This study underscores the need for continuous monitoring and the potential of eco-friendly disinfection approaches in controlling the spread of SARS-CoV-2 and possibly of other respiratory viruses.
AUTHOR AFFILIATIONS 1 Laboratory of Microbiology and Virology, Vita-Salute San Raffaele University, Milan, Italy
ADDITIONAL FILES The following material is available online.
Supplemental Material Supplemental material (AEM00774-25-S0001.docx). Supplemental methods; Fig. S1 to S6. Graphical abstract (AEM00774-25-S0002.tif). Visual depiction of study.
References
Aboubakr, Sharafeldin, Goyal, Stability of SARS-CoV-2 and other coronaviruses in the environment and on common touch surfaces and the influence of climatic conditions: a review, Transbound Emerg Dis,
doi:10.1111/tbed.13707Bailey, Curcic, Biros, Erdogmuş, Bac et al., Essential oil disinfectant efficacy against SARS-CoV-2 microbial surrogates, Front Public Health,
doi:10.3389/fpubh.2021.783832Castelli, Baj, Criscuolo, Ferrarese, Diotti et al., Characterization of a lineage C.36 SARS-CoV-2 isolate with reduced susceptibility to neutralization circulating in Lombardy, Italy, Viruses,
doi:10.3390/v13081514Criscuolo, Diotti, Ferrarese, Alippi, Viscardi et al., Fast inactivation of SARS-CoV-2 by UV-C and ozone exposure on different materials, Emerging Microbes & Infections,
doi:10.1080/22221751.2021.1872354Harvey, Carabelli, Jackson, Gupta, Thomson et al., (COG-UK) Consortium. 2021. SARS-CoV-2 variants, spike mutations and immune escape, Nat Rev Microbiol,
doi:10.1038/s41579-021-00573-0Hoffmann, Kleine-Weber, Schroeder, Krüger, Herrler et al., SARS-CoV-2 cell entry depends on ACE2 and TMPRSS2 and is blocked by a clinically proven protease inhibitor, Cell,
doi:10.1038/s41590-021-01091-0Inagaki, Saito, Sugiyama, Okabayashi, Fujimoto, Rapid inactivation of SARS-CoV-2 with deep-UV LED irradiation, Emerging Microbes & Infections,
doi:10.1080/22221751.2020.1796529Jaimes, André, Chappie, Millet, Whittaker, Phylogenetic analysis and structural modeling of SARS-CoV-2 spike protein reveals an evolutionary distinct and proteolytically sensitive activation loop, J Mol Biol,
doi:10.1016/j.jmb.2020.04.009Koch, Uckeley, Doldan, Stanifer, Boulant et al., TMPRSS2 expression dictates the entry route used by SARS-CoV-2 to infect host cells, EMBO J,
doi:10.15252/embj.2021107821Li, Wu, Nie, Zhang, Hao et al., The impact of mutations in SARS-CoV-2 spike on viral infectivity and antigenicity, Cell,
doi:10.1016/j.cell.2020.07.012Manjunathan, Periyaswami, Rosita, Pandya, Selvaraj et al., Molecular docking analysis reveals Full-Length Text, Applied and Environmental Microbiology Month
Mohammad, Choudhury, Habib, Asrani, Mathur et al., Genomic variations in the structural proteins of SARS-CoV-2 and their deleterious impact on pathogenesis: a comparative genomics approach, Front Cell Infect Microbiol,
doi:10.3389/fcimb.2021.765039Park, Khan, Chiliveri, Hu, Irvin et al., SARS-CoV-2 omicron variants harbor spike protein mutations responsible for their attenuated fusogenic pheno type, Commun Biol,
doi:10.15252/embj.2021107821Qu, Evans, Zheng, Carlin, Saif et al., Evasion of neutralizing antibody response by the SARS-CoV-2 BA.2.75 variant, bioRxiv,
doi:10.1101/2022.08.14.503921Rashid, Xie, Suleman, Shah, Khan et al., Roles and functions of SARS-CoV-2 proteins in host immune evasion, Front Immunol,
doi:10.3389/fimmu.2022.940756Romeo, Iacovelli, Scagnolari, Scordio, Frasca et al., Potential use of tea tree oil as a disinfectant agent against coronaviruses: a combined experimental and simulation study, Molecules,
doi:10.3390/molecules27123786Roy, Chan, Banadyga, He, Zhu et al., Quercetin inhibits SARS-CoV-2 infection and prevents syncytium formation by cells co-expressing the viral spike protein and human ACE2, Virol J,
doi:10.1186/s12985-024-02299-wShekunov, Zlodeeva, Efimova, Muryleva, Zarubaev et al., Cyclic lipopeptides as membrane fusion inhibitors against SARS-CoV-2: new tricks for old dogs, Applied and Environmental Microbiology Month
Shrestha, Foster, Rawlinson, Tedla, Bull, Evolution of the SARS-CoV-2 omicron variants BA.1 to BA.5: implications for immune escape and transmission, Rev Med Virol,
doi:10.1002/rmv.2381Solanki, Rajpoot, Kumar, Zhang, Ohishi et al., Structural analysis of spike proteins from SARS-CoV-2 variants of concern highlighting their functional alterations, Future Virol,
doi:10.2217/fvl-2022-0003Sun, Xie, Bu, Zhong, Zeng, Molecular characteristics, immune evasion, and impact of SARS-CoV-2 variants, Signal Transduct Target Ther,
doi:10.1038/s41392-022-01039-2Trivellin, Piva, Fiorimonte, Buffolo, Santi et al., UV-based technologies for SARS-CoV2 inactivation: status and perspectives, Electronics,
doi:10.3390/electronics10141703Van Doremalen, Bushmaker, Morris, Holbrook, Gamble et al., Aerosol and surface stability of SARS-CoV-2 as compared with SARS-CoV-1, N Engl J Med,
doi:10.1056/NEJMc2004973Zhou, Xu, Wang, Anashkina, Jiang, Impact of mutations in SARS-COV-2 spike on viral infectivity and antigenicity, Brief Bioinformatics,
doi:10.1093/bib/bbab375DOI record:
{
"DOI": "10.1128/aem.00774-25",
"ISSN": [
"0099-2240",
"1098-5336"
],
"URL": "http://dx.doi.org/10.1128/aem.00774-25",
"abstract": "<jats:title>ABSTRACT</jats:title>\n <jats:sec>\n <jats:title/>\n <jats:p>\n The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused a global pandemic, significantly impacting social and economic aspects of society. While belonging to the\n <jats:italic toggle=\"yes\">Coronaviridae</jats:italic>\n family, SARS-CoV-2 exhibited unique characteristics that facilitated its widespread transmission and pathogenicity. This study aimed to investigate the molecular characterization and infection kinetics of emerging SARS-CoV-2 variants, focusing on four key virological aspects: viral entry mechanisms, replication dynamics, immune evasion strategies, and surface persistence. We isolated 11 SARS-CoV-2 variants from clinical samples and conducted\n <jats:italic toggle=\"yes\">in vitro</jats:italic>\n studies using Calu-3 cell lines and pseudovirus systems to explore viral entry. Whole-genome sequencing identified mutations linked to increased replication and enhanced immune evasion, particularly through the inhibition of interferon responses. Additionally, persistence studies on common environmental surfaces (copper, aluminum, and plastic) revealed that certain variants, such as G446S, contributed to increased viral stability, suggesting potential for indirect transmission beyond direct person-to-person contact. Furthermore, we evaluated the effectiveness of “green” disinfectants, including tea tree oil and quercetin-based compounds, which demonstrated viral load reductions of over 95% within minutes, outperforming conventional alcohol- and UV-based disinfection methods. Notably, two influenza virus isolates belonging to highly divergent subtypes are sanitized as well while showing different persistence surface profiles compared to SARS-CoV-2. These findings highlight the evolutionary strategies employed by SARS-CoV-2 to ensure transmission, its ability to adapt under selective pressure, and the importance of ongoing surveillance of emerging variants. Moreover, the efficacy of environmentally sustainable disinfectants presents a promising strategy for reducing viral transmission in both clinical and community settings.\n </jats:p>\n <jats:sec>\n <jats:title>IMPORTANCE</jats:title>\n <jats:p>In this study, we evaluated the molecular profile of 11 SARS-CoV-2 variants, focusing on four key virological aspects: viral entry mechanisms, replication dynamics, immune evasion strategies, and surface persistence. Through whole-genome sequencing, we identified mutations linked to enhanced replication and immune evasion, notably through the suppression of interferon responses. Additionally, persistence studies on common environmental surfaces (copper, aluminum, and plastic) demonstrated that certain mutations, such as G446S, exhibited increased stability, suggesting a potential role for indirect transmission. This study underscores the need for continuous monitoring and the potential of eco-friendly disinfection approaches in controlling the spread of SARS-CoV-2 and possibly of other respiratory viruses.</jats:p>\n </jats:sec>\n </jats:sec>",
"alternative-id": [
"10.1128/aem.00774-25"
],
"article-number": "e00774-25",
"assertion": [
{
"group": {
"label": "Publication History",
"name": "publication_history"
},
"label": "Received",
"name": "received",
"order": 0,
"value": "2025-04-10"
},
{
"group": {
"label": "Publication History",
"name": "publication_history"
},
"label": "Accepted",
"name": "accepted",
"order": 2,
"value": "2025-07-30"
},
{
"group": {
"label": "Publication History",
"name": "publication_history"
},
"label": "Published",
"name": "published",
"order": 3,
"value": "2025-08-27"
}
],
"author": [
{
"ORCID": "https://orcid.org/0000-0002-8006-7218",
"affiliation": [
{
"id": [
{
"asserted-by": "publisher",
"id": "https://ror.org/01gmqr298",
"id-type": "ROR"
}
],
"name": "Laboratory of Microbiology and Virology, Vita-Salute San Raffaele University",
"place": [
"Milan, Italy"
]
}
],
"authenticated-orcid": true,
"family": "Sisti",
"given": "Sofia",
"sequence": "first"
},
{
"ORCID": "https://orcid.org/0000-0003-0185-3147",
"affiliation": [
{
"id": [
{
"asserted-by": "publisher",
"id": "https://ror.org/01gmqr298",
"id-type": "ROR"
}
],
"name": "Laboratory of Microbiology and Virology, Vita-Salute San Raffaele University",
"place": [
"Milan, Italy"
]
}
],
"authenticated-orcid": true,
"family": "Criscuolo",
"given": "Elena",
"sequence": "additional"
},
{
"ORCID": "https://orcid.org/0000-0003-4337-5024",
"affiliation": [
{
"id": [
{
"asserted-by": "publisher",
"id": "https://ror.org/01gmqr298",
"id-type": "ROR"
}
],
"name": "Laboratory of Microbiology and Virology, Vita-Salute San Raffaele University",
"place": [
"Milan, Italy"
]
}
],
"authenticated-orcid": true,
"family": "Giuliani",
"given": "Benedetta",
"sequence": "additional"
},
{
"ORCID": "https://orcid.org/0009-0009-4683-6202",
"affiliation": [
{
"id": [
{
"asserted-by": "publisher",
"id": "https://ror.org/01gmqr298",
"id-type": "ROR"
}
],
"name": "Laboratory of Microbiology and Virology, Vita-Salute San Raffaele University",
"place": [
"Milan, Italy"
]
}
],
"authenticated-orcid": true,
"family": "Cavallaro",
"given": "Mattia",
"sequence": "additional"
},
{
"ORCID": "https://orcid.org/0000-0002-5063-0129",
"affiliation": [
{
"id": [
{
"asserted-by": "publisher",
"id": "https://ror.org/039zxt351",
"id-type": "ROR"
}
],
"name": "IRCCS San Raffaele Scientific Institute",
"place": [
"Milan, Italy"
]
}
],
"authenticated-orcid": true,
"family": "Sampaolo",
"given": "Michela",
"sequence": "additional"
},
{
"ORCID": "https://orcid.org/0000-0003-0295-7941",
"affiliation": [
{
"id": [
{
"asserted-by": "publisher",
"id": "https://ror.org/01gmqr298",
"id-type": "ROR"
}
],
"name": "Laboratory of Microbiology and Virology, Vita-Salute San Raffaele University",
"place": [
"Milan, Italy"
]
}
],
"authenticated-orcid": true,
"family": "Castelli",
"given": "Matteo",
"sequence": "additional"
},
{
"affiliation": [
{
"id": [
{
"asserted-by": "publisher",
"id": "https://ror.org/01gmqr298",
"id-type": "ROR"
}
],
"name": "Laboratory of Microbiology and Virology, Vita-Salute San Raffaele University",
"place": [
"Milan, Italy"
]
}
],
"family": "Burioni",
"given": "Roberto",
"sequence": "additional"
},
{
"affiliation": [
{
"id": [
{
"asserted-by": "publisher",
"id": "https://ror.org/039zxt351",
"id-type": "ROR"
}
],
"name": "IRCCS San Raffaele Scientific Institute",
"place": [
"Milan, Italy"
]
}
],
"family": "Locatelli",
"given": "Massimo",
"sequence": "additional"
},
{
"ORCID": "https://orcid.org/0000-0002-1822-9861",
"affiliation": [
{
"id": [
{
"asserted-by": "publisher",
"id": "https://ror.org/01gmqr298",
"id-type": "ROR"
}
],
"name": "Laboratory of Microbiology and Virology, Vita-Salute San Raffaele University",
"place": [
"Milan, Italy"
]
},
{
"id": [
{
"asserted-by": "publisher",
"id": "https://ror.org/039zxt351",
"id-type": "ROR"
}
],
"name": "IRCCS San Raffaele Scientific Institute",
"place": [
"Milan, Italy"
]
}
],
"authenticated-orcid": true,
"family": "Clementi",
"given": "Nicola",
"sequence": "additional"
}
],
"container-title": "Applied and Environmental Microbiology",
"container-title-short": "Appl Environ Microbiol",
"content-domain": {
"crossmark-restriction": true,
"domain": [
"journals.asm.org"
]
},
"created": {
"date-parts": [
[
2025,
8,
27
]
],
"date-time": "2025-08-27T12:40:22Z",
"timestamp": 1756298422000
},
"deposited": {
"date-parts": [
[
2025,
8,
27
]
],
"date-time": "2025-08-27T12:40:26Z",
"timestamp": 1756298426000
},
"editor": [
{
"affiliation": [],
"family": "Buan",
"given": "Nicole R.",
"sequence": "additional"
}
],
"funder": [
{
"award": [
"PE_00000007"
],
"name": "Ministero dell Istruzione, dell Universita e della Ricerca"
}
],
"indexed": {
"date-parts": [
[
2025,
8,
27
]
],
"date-time": "2025-08-27T13:10:16Z",
"timestamp": 1756300216174,
"version": "3.44.0"
},
"is-referenced-by-count": 0,
"issued": {
"date-parts": [
[
2025,
8,
27
]
]
},
"language": "en",
"license": [
{
"URL": "https://creativecommons.org/licenses/by/4.0/",
"content-version": "vor",
"delay-in-days": 0,
"start": {
"date-parts": [
[
2025,
8,
27
]
],
"date-time": "2025-08-27T00:00:00Z",
"timestamp": 1756252800000
}
},
{
"URL": "https://journals.asm.org/non-commercial-tdm-license",
"content-version": "tdm",
"delay-in-days": 0,
"start": {
"date-parts": [
[
2025,
8,
27
]
],
"date-time": "2025-08-27T00:00:00Z",
"timestamp": 1756252800000
}
}
],
"link": [
{
"URL": "https://journals.asm.org/doi/pdf/10.1128/aem.00774-25",
"content-type": "application/pdf",
"content-version": "vor",
"intended-application": "text-mining"
},
{
"URL": "https://journals.asm.org/doi/pdf/10.1128/aem.00774-25",
"content-type": "unspecified",
"content-version": "vor",
"intended-application": "similarity-checking"
}
],
"member": "235",
"original-title": [],
"prefix": "10.1128",
"published": {
"date-parts": [
[
2025,
8,
27
]
]
},
"published-online": {
"date-parts": [
[
2025,
8,
27
]
]
},
"publisher": "American Society for Microbiology",
"reference": [
{
"DOI": "10.1038/s41579-023-00878-2",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_2_2"
},
{
"DOI": "10.1016/j.jmb.2020.04.009",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_3_2"
},
{
"DOI": "10.1038/s41591-021-01347-0",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_4_2"
},
{
"DOI": "10.2217/fvl-2022-0003",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_5_2"
},
{
"DOI": "10.1002/rmv.2381",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_6_2"
},
{
"DOI": "10.1080/22221751.2021.1872354",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_7_2"
},
{
"DOI": "10.3390/electronics10141703",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_8_2"
},
{
"DOI": "10.3390/molecules27123786",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_9_2"
},
{
"DOI": "10.3389/fpubh.2021.783832",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_10_2"
},
{
"DOI": "10.1080/22221751.2020.1796529",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_11_2"
},
{
"DOI": "10.3390/v13081514",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_12_2"
},
{
"DOI": "10.3389/fcimb.2021.765039",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_13_2"
},
{
"DOI": "10.1101/2022.08.14.503921",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_14_2",
"unstructured": "Qu P Evans JP Zheng YM Carlin C Saif LJ Oltz EM Xu K Gumina RJ Liu S-L. 2022. Evasion of neutralizing antibody response by the SARS-CoV-2 BA.2.75 variant. bioRxiv. doi:10.1101/2022.08.14.503921"
},
{
"DOI": "10.1016/j.it.2020.10.004",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_15_2"
},
{
"DOI": "10.1093/bib/bbab375",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_16_2"
},
{
"DOI": "10.1038/s41586-021-03944-y",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_17_2"
},
{
"DOI": "10.1016/j.cell.2020.07.012",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_18_2"
},
{
"DOI": "10.1038/s42003-023-04923-x",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_19_2"
},
{
"DOI": "10.15252/embj.2021107821",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_20_2"
},
{
"DOI": "10.1016/j.cell.2020.02.052",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_21_2"
},
{
"DOI": "10.1146/annurev-immunol-032713-120231",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_22_2"
},
{
"DOI": "10.1038/s41590-021-01091-0",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_23_2"
},
{
"DOI": "10.1038/s41579-022-00839-1",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_24_2"
},
{
"DOI": "10.1038/s41392-022-01039-2",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_25_2"
},
{
"DOI": "10.1038/s41579-021-00573-0",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_26_2"
},
{
"DOI": "10.3389/fimmu.2022.940756",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_27_2"
},
{
"DOI": "10.1056/NEJMc2004973",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_28_2"
},
{
"DOI": "10.1111/tbed.13707",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_29_2"
},
{
"DOI": "10.1186/s12985-024-02299-w",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_30_2"
},
{
"DOI": "10.1186/s12859-022-04724-9",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_31_2"
},
{
"DOI": "10.1016/j.antiviral.2023.105575",
"doi-asserted-by": "publisher",
"key": "e_1_3_3_32_2"
}
],
"reference-count": 31,
"references-count": 31,
"relation": {},
"resource": {
"primary": {
"URL": "https://journals.asm.org/doi/10.1128/aem.00774-25"
}
},
"score": 1,
"short-title": [],
"source": "Crossref",
"subject": [],
"subtitle": [],
"title": "Evaluation of respiratory virus transmissibility and resilience from fomites: the case of 11 SARS-CoV-2 clinical isolates",
"type": "journal-article",
"update-policy": "https://doi.org/10.1128/asmj-crossmark-policy-page"
}
