key: cord-0805532-ncau27zc authors: Meister, T. L.; Dreismeier, M.; Vidal Blanco, E.; Brueggemann, Y.; Heinen, N.; Kampf, G.; Todt, D.; Nguyen, H. P.; Steinmann, J.; Schmidt, W. E.; Steinmann, E.; Quast, D. R.; Pfaender, S. title: Risk of SARS-CoV-2 transmission by fomites: a clinical observational study in highly infectious COVID-19 patients date: 2022-03-23 journal: nan DOI: 10.1101/2022.03.22.22272773 sha: 5aae5160cdaeb710b216b6c84a936a24a303e6f6 doc_id: 805532 cord_uid: ncau27zc Background: The contribution of droplet-contaminated surfaces for virus transmission has been discussed controversially in the context of the current Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) pandemic. Importantly, the risk of fomite-based transmission has not been systematically addressed. Methods: We initiated this single-center observational study to evaluate whether hospitalized COVID-19 patients can contaminate stainless steel carriers by coughing or intensive moistening with saliva and to assess the risk of SARS-CoV-2 transmission upon detection of viral loads and infectious virus in cell culture. Fifteen hospitalized patients with a high baseline viral load (CT value [≤] 25) shortly after admission were included. We documented clinical and laboratory parameters and used patient samples to perform virus culture, quantitative PCR and virus sequencing. Results: Nasopharyngeal and oropharyngeal swabs of all patients were positive for viral RNA on the day of the study. Infectious SARS-CoV-2 could be isolated from 6 patient swabs (46.2 %). While after coughing, no infectious virus could be recovered, intensive moistening with saliva resulted in successful viral recovery from steel carriers of 5 patients (38.5 %). Conclusions: Transmission of infectious SARS-CoV-2 via fomites is possible upon extensive moistening, but unlikely to occur in real-life scenarios and from droplet-contaminated fomites. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted March 23, 2022. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The initial viral load was determined using Allplex™ 2019-nCoV Assay (Seegene Inc., Seoul, Republic 88 of Korea) targeting three SARS-CoV-2 specific genes (E gene, RdRP gene and N gene) with a sensitivity 89 of 100 copies per run. Exclusion criteria were acute myocardial infarction, current need of ventilation 90 support systems (e.g., high flow-or non-invasive ventilation), current treatment in an intensive care unit, After written informed consent, two combined nasopharyngeal and oropharyngeal swabs were collected 100 from each patient. Then, patients were asked to forcefully cough two-times on a pre-defined surface 101 area containing nine standardized steel-carriers, each with a one-centimeter diameter ("cough"), using 102 a specially designed tripod with a defined distance of 15 centimeters (Appendix Figure 1 ). In addition, patients were asked to moisten nine steel carriers with saliva for ten seconds within their mouth 104 ("moisten"). After defined time points at room temperature (1 min, 5 min, 15 min, 30 min, 45 min, 90 105 min, 120 min and 240 min), the steel-carriers were placed in containers containing 2 mL cold Dulbecco's 106 modified Eagle's medium (DMEM complete, supplemented with 10 % (v/v) fetal calf serum, 1 % non-107 essential amino acids, 100 IU/mL penicillin, 100 µg/mL streptomycin and 2 mM L-Glutamine) and is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint VeroE6 cells were seeded at 3×10 5 cells/well in a six well cell culture plate and incubated for at least 112 four hours at 37 °C and 5 % CO2. Hereafter, the medium was replaced with 1·8 mL of patient swabs, 113 "cough" samples or "moisten" samples and 2·5 µg/mL amphotericin B was added. Over a maximum 114 period of ten days, cells were monitored daily for the appearance of a cytopathic effect (CPE), indicating www.graphpad.com). Sample size calculation was performed using G*Power Version 3.1.9.6 for 136 windows [12] . is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint (Table 2 ). In addition, variants in spike domains were identified and annotated using Geneious is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint Two patient samples were excluded from the study due to bacterial/fungal contamination within the 191 cultures (Appendix Table 1 ). Viral RNA could be detected from all (n=13/13) combined nasopharyngeal 192 and oropharyngeal swabs (inoculum) and viral RNA could be successfully detected within the 193 inoculated cell cultures (Figure 2, Appendix Figure 3 ). After inoculation with the "swabs", viral loads 194 in the cells ranged from 2·23×10 1 to 2·03×10 9 RNA copies/50 ng and in the supernatant from not 195 detectable to 6·58×10 7 RNA copies/mL (Appendix Table 1 is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint However, the present cohort is characterized by high viral load. Hence, a high transmission rate can be 244 assumed [24], supporting the main conclusion of the study. Of note, several laboratory parameters on 245 admission significantly correlated with a severe outcome. However, the present study was not powered 246 for this analysis and therefore, these correlations need to be considered exploratory. Infectious virus could be recovered from the combined nasopharyngeal and oropharyngeal swabs and dose, which subsequently needs to be exposed towards susceptible tissues (e.g., mucosa, eyes). is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The present study provides evidence that fomites may not be as critical in the transmission of SARS- CoV-2 as initially suspected. However, the present study also provides evidence that infectious SARS- CoV-2 can be found on some fomites after contamination with extensive amounts of saliva. Therefore, is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint . CC-BY-NC-ND 4.0 International license It is made available under a perpetuity. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint . CC-BY-NC-ND 4.0 International license It is made available under a perpetuity. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint Airborne transmission of respiratory viruses Temperature-dependent surface stability of SARS-CoV-2 Aerosol and Surface Stability of SARS-CoV-327 2 as Compared with SARS-CoV-1 Stability of SARS-CoV-2 in different environmental conditions Centers of Disease Control and Prevention. Science Brief: SARS-CoV-2 and Surface (Fomite) 333 Transmission for Indoor Community Environments Exaggerated risk of transmission of COVID-19 by fomites. The Lancet Infectious 337 Therapeutics and COVID-19: living guideline. World Health Organization Power 3: a flexible statistical power analysis 348 program for the social, behavioral, and biomedical sciences Assignment of epidemiological lineages in an emerging 351 pandemic using the pangolin tool Epidemiologischer Steckbrief zu SARS-CoV-2 und COVID Risk Factors for Severe Disease and Efficacy 355 of Treatment in Patients Infected With COVID-19: A Systematic Review Meta-Regression Analysis Bericht zu Virusvarianten von SARS-CoV-2 in Deutschland Fomite-mediated transmission as a 362 sufficient pathway: a comparative analysis across three viral pathogens Significance of fomites in the spread of respiratory and enteric viral disease Transmissibility and transmission of respiratory viruses A realistic transfer method reveals low risk of SARS-CoV-369 2 transmission via contaminated euro coins and banknotes Risk Factors for Mortality in Patients with COVID-19 371 in New York City Deliberation, Dissent, and Distrust: Understanding distinct is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprintThe copyright holder for this this version posted March 23, 2022. ; https://doi.org/10.1101/2022.03.22.22272773 doi: medRxiv preprint