# Environmental severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contamination in hospital rooms during the first and third coronavirus disease 2019 (COVID-19) waves

Killian Le Neindre, Jeanne Couturier, Aurélie Schnuriger, Sarah Jolivet, Cyril Gouot, Mickaël Majerholc, Pierre Supplisson, Céline Tan, Marine Perrier, Christelle Lazare, et al.

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Killian Le Neindre, Jeanne Couturier, Aurélie Schnuriger, Sarah Jolivet, Cyril Gouot, et al. Envi-ronmental severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contamination in hospital rooms during the first and third coronavirus disease 2019 (COVID-19) waves. Antimicrobial Steward-ship & Healthcare Epidemiology, 2022, 2 (1), pp.e82. 10.1017/ash.2022.228. hal-03948762

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Submitted on 20 Jan 2023

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Environmental severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contamination in hospital rooms during the first and third coronavirus disease 2019 (COVID-19) waves

Killian Le Neindre PhD<fnref n="1" /> ID, Jeanne Couturier PhD<fnref n="2" />, Aurélie Schnuriger PhD³,⁴ ID, Sarah Jolivet PhD<fnref n="2" />, Cyril Gouot BSc<fnref n="2" />, Mickaël Majerholc PharmD<fnref n="2" />, Pierre Supplisson PharmD<fnref n="2" />, Céline Tan PharmD<fnref n="2" />, Marine Perrier PhD³ ID, Christelle Lazare BSc<fnref n="1" />, Laurence Morand-Joubert PhD³,⁴ and Frédéric Barbut PhD<fnref n="1" />,<fnref n="2" />

Abstract

We investigated the frequency, distribution, and risk factors of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) environmental contamination around infected patients during the first and third wave of the coronavirus disease 2019 pandemic. The shedding of SARS-CoV-2 in rooms of infected patients was limited in our hospital setting.

(Received 4 March 2022; accepted 14 April 2022)

Since December 2019, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has disseminated worldwide, causing an unprecedented public health threat. SARS-CoV-2 is primarily transmitted directly from person to person by respiratory transmission.<fnref n="1" /> In the hospital setting, SARS-CoV-2 is rarely detected in air samples, and when it is detected the viral load is very low.<fnref n="2" /> Surfaces could also be involved in SARS-CoV-2 transmission. Surface contamination is highly variable across studies and wards,³ and SARS-CoV-2 has been shown to persist on different surfaces from several hours to days.⁴,⁵ The factors associated with environmental contamination are still unclear and require further investigation. In this study, the frequency of surfaces and air contamination by SARS-CoV-2 was evaluated in rooms of patients infected by SARS-CoV-2 and risk factors associated with the environmental contamination were investigated.

Methods

Study design

This prospective study was conducted in a university-affiliated,
650-bed, acute-care hospital in France during the end of the first

epidemic wave (April to August 2020) and during the third epidemic wave (February-May 2021). Symptomatic patients were screened for coronavirus disease 2019 (COVID-19) on admission except for 19 patients who came to the hospital with an external positive test. Rooms of patients infected by SARS-CoV-2 were sampled at least 4 hours after the cleaning procedure and within 15 days following the first positive biological diagnosis by reverse-transcription quantitative polymerase chain reaction (RT-qPCR). The cleaning procedure was manual using a detergent disinfectant product (containing quaternary ammonium compound). Environmental sampling included 4–10 surfaces (mainly door handle, respirator, syringe driver, bed rail, floor, bench, fluids ramp, bedside table, toilet seat, sink, and windowsill) and air.

For each patient, the following data were collected: age, sex,
symptoms at the time of sampling, type of ward (medicine or
intensive care unit), type of mechanical ventilation, delay between
positive diagnosis and environmental sampling, origin of SARS-
CoV-2 infection (community versus hospital acquired), type of
SARS-CoV-2 variant (when available) and cycle threshold (Ct)
values. We defined a hospital-acquired infection as a patient
admitted without symptoms suggestive of COVID-19 who had
SARS-CoV-2 RNA detected at least 3 days after admission.

**Author for correspondence:** Killian Le Neindre, PhD, Service de Microbiologie de l'Environnement, Hôpital Saint-Antoine, 184 Rue du Faubourg Saint-Antoine, 75012 Paris, France. E-mail: killian.le-neindre@parisdescartes.fr or killian.leneindre@aphp.fr

Cite this article: Le Neindre K, et al. (2022). Environmental severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contamination in hospital rooms during the first and third coronavirus disease 2019 (COVID-19) waves. *Antimicrobial Stewardship & Healthcare Epidemiology*, https://doi.org/10.1017/ash.2022.228

Environmental sampling and virology assays

Surfaces (10-cm × 10-cm) were sampled using premoistened swabs discharged in 500 µL viral medium transport VTM (Labomoderne, France). Air (1,000 L, 50 L/min flow) was sampled 1 m from the patient using the MD8 air sampler (Sartorius, Germany). Air was

<footnote n="1">¹Department of Environmental Microbiology, Saint-Antoine Hospital, Sorbonne Université, Assistance Publique – Hôpitaux de Paris (AP-HP), Paris, France,</footnote>

<footnote n="2">²Infection Control Unit, Saint-Antoine Hospital, Sorbonne Université, Assistance Publique – Hôpitaux de Paris (AP-HP), Paris, France, ³Department of Virology,</footnote>

<footnote>Saint-Antoine Tenon Trousseau Hospitals, Sorbonne Université, Assistance Publique – Hôpitaux de Paris (AP-HP), France and ⁴Department of Virology, Sorbonne</footnote>

<footnote>Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (iPLESP), Assistance Publique-Hôpitaux de Paris (AP-HP), Paris, France</footnote>
<table><caption>**Table 1.** Description of Data Collected From Rooms With Positive Environmental Samples</caption><thead><tr><th>Room</th><th>Date of Sampling</th><th>Ward</th><th>Ct Value (Clinical)</th><th>SARS-CoV-2 Variant</th><th>Mechanical Ventilation</th><th>Oxygen Flow, L/min</th><th>Positive Surface Sample/Total Surface</th><th>Ct per Sample (Environmental)</th></tr></thead><tbody><tr><td>1</td><td>04-20</td><td>ICU</td><td>Positive</td><td>NA</td><td>Intubation</td><td>NA</td><td>3/10</td><td>36.03/37.43/37.74</td></tr><tr><td>2</td><td>04-20</td><td>MD</td><td>26.04</td><td>NA</td><td>Nasal</td><td>2</td><td>3/10</td><td>39.03/35.52/37.84</td></tr><tr><td>3</td><td>06-20</td><td>MD</td><td>26.95</td><td>NA</td><td>Absence</td><td>No</td><td>4/10</td><td>38.72/36.97/37.1/38.6</td></tr><tr><td>7</td><td>07-20</td><td>MD</td><td>12.47</td><td>NA</td><td>Mask</td><td>13</td><td>2/10</td><td>36.77/38.16</td></tr><tr><td>15</td><td>02-21</td><td>MD</td><td>20.18</td><td>H</td><td>Nasal</td><td>2</td><td>2/4</td><td>38.33/38.08</td></tr><tr><td>16</td><td>02-21</td><td>MD</td><td>28.83</td><td>H</td><td>Absence</td><td>No</td><td>0/4</td><td>37.66<sup>a</sup></td></tr><tr><td>25</td><td>02-21</td><td>MD</td><td>19.97</td><td>α</td><td>Absence</td><td>No</td><td>1/4</td><td>30.35</td></tr><tr><td>29</td><td>03-21</td><td>MD</td><td>26.46</td><td>H</td><td>Absence</td><td>No</td><td>1/4</td><td>28.60</td></tr><tr><td>49</td><td>04-21</td><td>MD</td><td>23.62</td><td>α</td><td>Absence</td><td>No</td><td>1/5</td><td>40</td></tr><tr><td>52</td><td>04-21</td><td>MD</td><td>29.78</td><td>α</td><td>Absence</td><td>No</td><td>3/5</td><td>34.13/30.34/33.74</td></tr><tr><td>53</td><td>04-21</td><td>MD</td><td>33.02</td><td>β</td><td>Mask</td><td>7</td><td>1/5</td><td>39.18</td></tr><tr><td>54</td><td>04-21</td><td>MD</td><td>11.81</td><td>β</td><td>Nasal</td><td>5</td><td>1/5</td><td>35.66</td></tr><tr><td>55</td><td>04-21</td><td>MD</td><td>Positive</td><td>NA</td><td>Mask</td><td>12</td><td>1/5</td><td>35.74</td></tr><tr><td>61</td><td>05-21</td><td>ICU</td><td>20.78</td><td>β</td><td>Optiflow</td><td>60</td><td>1/5</td><td>38.07</td></tr></tbody></table>

Note: Ct, cycle threshold; MD, medicine; ICU, intensive care unit; M, male; F, female; Positive: positive test but the Ct value was not available; H, historical strain; NA, data not available.

collected onto gelatin membrane filters (Sartorius, Germany) that were dissolved in 10 mL sterile water at 37°C for 10 minutes. Environmental samples were analyzed by reverse-transcription quantitative polymerase chain reaction (RT-qPCR) using the Allplex SARS-CoV-2 Assay (Seegene, South Korea) or Bosphore Novel Coronavirus (2019-nCoV) Detection Kit v2 (Anatolia Genework, Turkey). The Allplex kit (SEE) amplified the *E* gene, *RdRP/S* gene, and *S* gene, whereas the Bosphore kit (BOS) amplified the *E* gene and *orf1ab* gene. Positive results were presented as the mean of PCR cycle threshold (Ct) values for each viral genes. Both techniques yield similar Ct values, as previously described.⁶

## Statistical analysis

A room was considered contaminated when at least 1 environmental sample was positive. Contaminated and noncontaminated rooms were compared using the Fisher exact test or the χ² for qualitative data and the Mann-Whitney test for quantitative data. Statistical tests were performed using GraphPad Prism version 7 software (GraphPad Software, San Diego, CA).

## Results

In total, 61 rooms were sampled, and 338 surfaces and 59 air samples were analyzed (Table 1). The median age of patients was 73 years (interquartile range [IQR], 61–84), and the sex ratio (M/F) was 2.2. The median delay between symptom onset and environmental sampling was 9 days (IQR, 6–13). Environmental sampling was conducted after a median delay of 4 days (IQR, 2–7) following admission and after a median delay of 3 days (IQR, 2–7) following positive RT-qPCR. Identification of variants was not performed during the first wave.

Of 61 rooms, 14 (22.9%) had at least 1 positive sample for SARS-CoV-2 (Table 1). Overall, 24 (7.1%) of 338 surfaces and 1 (1.7%) of 59 air samples were positive. Cycle threshold (Ct) values

ranged from 28.6 to 40. Of 25 positive samples, 20 (80%) had a Ct value >35. The 4 most contaminated surface samples were the floor (7 of 55, 12.7%), the windowsill (4 of 20, 20.0%), the toilet seat (4 of 44, 9.1%), and the door handle (3 of 62, 4.8%). Other positive surfaces included bedside table (n = 2), bed rail (n = 1), respirator (n = 1), bench (n = 1), and fluids ramp (n = 1). Between the first and the third epidemic waves, the frequency of contaminated surfaces slightly decreased (10.0% and 5.5%, respectively; Fisher exact test: *P* = .13).

Contaminated rooms were compared to noncontaminated rooms for the variables shown in Table 2. The frequency of contamination was significantly higher in rooms of older patients.

## Discussion

The frequency of surface contamination (7.1%) in our study was lower than that reported in other studies (533 of 3,077, 17.3%), although surface contamination has varied from 0% to 74.2% across published studies.³ Furthermore, most positive samples have high Ct values, suggesting a low genome level and a likely lack of infectivity in cell culture, as shown by others.³,⁷

Interestingly, 4 rooms (nos. 1, 2, 3, and 52) were more contaminated than others (at least 3 positive environmental samples per room), suggesting either the presence of a “super-spreader” patient or just a poor room cleaning (Table 1).

Only a single air sample was positive. This air contamination was lower than frequencies reported in other studies (82 of 471, 17.4%).² The variations (from 0% to 100%) of air contamination might be explained by differences of sampling across studies (volume of sampled air, sampling technique, distance from the patient, areas of sampling, etc).²,⁸

We investigated risk factors for the environmental spread of SARS-CoV-2 (Table 2). The only significant risk factor was patient age. This might be explained by the higher severity of patients who

<footnote>ªCt from positive air sample.</footnote>
<table><caption>**Table 2.** Variable Analysis According to Room Contamination</caption><thead><tr><th>Variable</th><th>Contaminated Rooms</th><th>Noncontaminated Rooms</th><th>Total</th><th>P Value</th></tr></thead><tbody><tr><td>Medical unit</td><td>12/14 (85.7)</td><td>32/47 (68.1)</td><td>44</td><td>.31</td></tr><tr><td>Intensive care unit</td><td>2/14 (14.3)</td><td>15/47 (31.9)</td><td>17</td><td></td></tr><tr><td>Age, median y (IQR)</td><td>84 (72.2-92.2)</td><td>71 (57-83)</td><td>61</td><td>.008</td></tr><tr><td>Sex, male</td><td>9/14 (64.3)</td><td>33/47 (70.2)</td><td>61</td><td>.75</td></tr><tr><td>Symptoms present</td><td>11/14 (78.5)</td><td>40/47 (85.1)</td><td>51</td><td>.68</td></tr><tr><td>Symptom delay, median d (IQR)</td><td>7 (5-10)</td><td>10 (6-14)</td><td>51</td><td>.11</td></tr><tr><td>Admission delay median d (IQR)</td><td>4.5 (3.7-12.5)</td><td>3 (2-6)</td><td>61</td><td>.09</td></tr><tr><td>Diagnosis delay median d (IQR)</td><td>3.5 (2-5.5)</td><td>3 (2-9)</td><td>61</td><td>.82</td></tr><tr><td>Ct of clinical sample, median (IQR)</td><td>24.83 (20.13-27.42)</td><td>23.79 (20.06-28.93)</td><td>54</td><td>.75</td></tr><tr><td>Historical strain</td><td>3/9 (33)</td><td>13/37 (35.1)</td><td>16</td><td>.34</td></tr><tr><td>α (alpha) variant</td><td>3/9 (33)</td><td>19/37 (51.4)</td><td>22</td><td></td></tr><tr><td>β (beta) variant</td><td>3/9 (33)</td><td>5/37 (13.5)</td><td>8</td><td></td></tr><tr><td>Mechanical ventilation</td><td>8/14 (57.1)</td><td>36/47 (76.1)</td><td>44</td><td>.18</td></tr><tr><td>Intubation</td><td>1/8 (12.5)</td><td>4/36 (11.1)</td><td>5</td><td>.87</td></tr><tr><td>Nasal</td><td>3/8 (37.5)</td><td>15/36 (41.7)</td><td>18</td><td></td></tr><tr><td>Mask</td><td>3/8 (37.5)</td><td>9/36 (25)</td><td>12</td><td></td></tr><tr><td>Optiflow</td><td>1/8 (12.5)</td><td>8/36 (22.2)</td><td>9</td><td></td></tr><tr><td>Flow ≥6 L oxygen</td><td>4/7 (57.1)</td><td>15/32 (46.9)</td><td>19</td><td>.69</td></tr><tr><td>Oxygen flow. L</td><td>7 (2-13)</td><td>4.5 (2.25-14.25)</td><td>39</td><td>.63</td></tr><tr><td>HA COVID-19</td><td>4/14 (28.6)</td><td>9/47 (19.1)</td><td>13</td><td>.47</td></tr></tbody></table>

Note. Ct, cycle threshold value; IQR, interquartile range; HA, hospital acquired. Quantitative values were calculated using the Mann-Whitney test as median (IQR) and qualitative values were calculated using the Fisher or $\chi^2$ test. Significant P value is shown in bold.

required heavier care load resulting in more contacts with the healthcare staff. We did not find any association between environmental contamination and type of ward (medical versus intensive care units) despite a higher frequency of aerosol-generating procedures in ICUs.

Since the first wave of COVID-19, many variants of SARS-CoV-2 have emerged. The third wave was characterized by the emergence of the α (alpha) variant (20I/501Y.V1; B.1.1.7).⁹ Nevertheless, the environmental contamination frequency between historical strain and variant was not significantly different. The high spread of the α variant does not seem to be explained by environmental contamination.

This study had several limitations. First, the correlation between the Ct values of clinical and environmental samples was difficult to interpret due to the different PCR assays used. Second, the viral culture for infectivity determination was not done, but previous studies failed to isolate the virus in cell culture in samples with Ct > 35.¹⁰ Only 2 of 16 studies have successfully isolated viable virus in cell culture from the environment.²,³ Third, we did not assess the quality of room cleaning, although we insured that sampling was performed at a distance from any cleaning procedure.

In conclusion, The frequency of surface and air contamination by SARS-CoV-2 was low and decreased by 50% when comparing the first epidemic wave to the third one, despite the emergence of more transmissible virus variants. The environmental spread of SARS-CoV-2 is likely limited in the hospital setting.

**Acknowledgments.** The authors thank Dr Muriel Ehmig for her helpful comments.

**Financial support.** This study is supported by the “Association Robert Debré pour la Recherche Médicale,” a nonprofit organization for the development of medical research.

**Conflicts of interest.** All authors report no conflicts of interest relevant to this article.

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