key: cord-0771163-czgdedtb
authors: Elbadawy, Hossein M.; Khattab, Amin; Alalawi, Ali; Dakilallah Aljohani, Fahad; Sundogji, Hamza; Mahmoud, Ameira S.; Abouzied, Meky; Eltahir, Heba M.; Alahmadey, Ziab; Bahashwan, Saleh; Suliman, Bandar A.
title: The detection of SARS‐CoV‐2 in outpatient clinics and public facilities during the COVID‐19 pandemic
date: 2021-02-10
journal: J Med Virol
DOI: 10.1002/jmv.26819
sha: be4228e3e6fc602e5a9b9ae076ef0d7e019c34c3
doc_id: 771163
cord_uid: czgdedtb

The transmission of the severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) can occur through an airborne route, in addition to contaminated surfaces and objects. In hospitals, it has been confirmed by several studies that SARS‐CoV‐2 can contaminate surfaces and medical equipment especially in hospitals dedicated to coronavirus disease 2019 (COVID‐19) patients. The aim of this study was to detect the contamination of hands, objects, and surfaces in isolation rooms and also in outpatients' clinics in hospitals and polyclinics. Environmental contamination of public high‐touch surfaces in public facilities was also investigated during an active COVID‐19 pandemic. Random swabs were also taken from public shops, pharmacies, bakeries, groceries, banknotes, and automated teller machines (ATMs). Samples were analyzed for SARS‐CoV‐2 positivity using real‐time polymerase chain reaction. In the COVID‐19 regional reference hospital, only 3 out of 20 samples were positive for SARS‐CoV‐2 RNA. Hand swabs from SARS‐CoV‐2‐positive patients in isolation rooms were occasionally positive for viral RNA. In outpatients' clinics, door handles were the most contaminated surfaces. Dental chairs, sinks, keyboards, ophthalmoscopes, and laboratory equipment were also contaminated. Although no positive swabs were found in shops and public facilities, random ATM swabs returned a positive result for SARS‐CoV‐2. Although there is no longer a focus on COVID‐19 wards and isolation hospitals, more attention is required to decontaminate frequently touched surfaces in health‐care facilities used by patients not diagnosed with COVID‐19. Additionally, high‐touch public surfaces such as ATMs require further disinfection procedures to limit the transmission of the infection.

during the first week of illness. 9 Another investigation showed that the air was clean in three tested rooms, where SARS-CoV-2 patients were staying, however, air vents and surfaces were mostly positive for the detection of the virus, showing that airflow systems can be shedding viral particles onto surfaces. 4 Importantly, data on the spread of viruses on surfaces are not redundant. One study, involved only three patients, while another study in Korea showed results from hospital rooms occupied with 13 patients only. 10 Additionally, very limited information is available regarding the transmission of the virus on surfaces in outpatients' clinics or in public high-touch surfaces. The virus was detected on patients' hands in one recent study 11 ; however, the debate is ongoing regarding the major route of transmission, whether it can be transmitted more through touching contaminated surfaces or by airborne droplets. Thus, more studies are required to confirm or oppose these findings and to highlight the precautions which should be taken to better control the spread of the infection. Information regarding the spread of the virus on surfaces and objects is essential for controlling the outbreak and is, therefore, the main target of this study. 

Collected swabs were processed for viral RNA extraction using the Viral Nucleic Acid Extraction Kit II (Geneaid Biotechnology).

Briefly, 400 µl of VB buffer and 200 µl of phosphate-buffered saline swab-elutes were added to each tube and then incubated for 10 min at 35°C. The mixture was centrifuged at 16,000g for 2 min and the supernatant was transferred to a new 1.5 ml microcentrifuge tube. A volume of 400 µl of washing buffer (AD buffer) was added to each tube and mixed gently for 10 s. Afterwards, 200 µl of absolute ethanol was then added to the mixture before transferring the entire contents to a extraction column.

The columns were centrifuged at 16,000g for 2 min to discard the flow-through and the column was then transferred to a new 2 ml collection tube. The column was then washed twice using 400 and 600 µl of W1 buffer. Finally, 50 µl of preheated elution buffer was added to the extraction columns and left standing for 3 min at room temperature to allow the elution buffer to be completely absorbed. RNA was then eluted from the extraction columns by centrifuging the columns at 16,000g for 30 s. RNA concentration and purity for extracted samples were assessed by analyzing 1 µl of the elute using a NanoDrop 1000 UV-VIS Spectrophotometer (Thermo Fisher Scientific).

For real-time PCR analysis, the center for disease control and prevention quantitative PCR (qPCR) Probe Assay was used. 12 The qRT-PCR reaction was prepared by adding 1.5 µl of primer/probe mix (100pM pH8) to 8.5 µl of Nuclease-free water (VWR International), and 5 µl of the Oasig Lyophilised OneStep RT-qPCR Master Mix (PrimerDesign Ltd.). Thermal cycling was performed on the ABI-7500

Fast 96-well Real-Time PCR System (Applied Biosystems) with an initial reverse-transcription step of 10 min at 55°C step, followed by an enzyme activation step at 95°C for 2 min, then followed by 45 cycles of denaturation at 95°C for 10 s, then annealing and data acquisition at 60°C for 10 s. CT and cut-off analysis to determine the positive and negative samples were performed using the internal software of the ABI-7500 system (software version 2.0.4).

For taking hand swabs, patients who are SARS-CoV-2 positive within 2 days and who agreed to participate in the study were recruited after taking signed consent. For ER patients presenting in the emergency room (ER) as suspected COVID-19 cases, swabs of hands were taken, and the patients' records were followed up later to select those with PCR positive results and include their hand swab results in this study.

A questionnaire was distributed to eight patients who agreed to participate and signed informed consent. The questionnaire was composed of questions about age, sex, days of home isolation before admission to hospital, hand wash hygiene, and adhering to home isolation rules.

Swabs were taken from COVID-19 isolation hospital and from outpatients' hospitals and polyclinics. High-touch surfaces were swabbed, and samples were sent for real-time PCR analysis following the protocol above. The investigation also included cell phones, automated teller machine (ATM) machines keypads, banknotes, and public shops, shopping carts, pharmacies, and bakeries.

Hand swabs were taken from patients admitted to the intensive care unit (ICU) and patients in home isolation or in isolation rooms in hospitals. All 16 patients were PCR positive for SARS-CoV-2. Of these patients, 12.5% showed positive hand contamination with SARS-CoV-2 (Table 1) , found only in patients in isolation with moderate to severe symptoms, however, under ICU settings, all hand swabs were negative. The hands of two patients at home isolation and another three patients presenting at visual triage in the hospital ER were all negative. Eight of these patients agreed to fill a brief questionnaire; six males and two females. The average age of the participants was 62.75%, the youngest being 50 years old while the oldest was 80 years old. It was found that the two patients who did not observe regular hand hygiene were positive when hand swabs were taken from them. Although those patients were SARS-CoV-2 positive for 8.6 days before being hospitalized, during the stay at home only 25% (two patients) were strictly adhering to home isolation rules. The other six patients reported visiting mosques, shopping malls, and going to work while having symptoms.

To detect the presence of the SARS-CoV-2 virus in the COVID-19 reference hospital, 20 swabs were taken from different locations in the hospital and sent for real-time PCR analysis. Starting with wards dedicated for COVID-19 inpatient, several sites were swabbed (Table 2 ). Three out of 20 swabs were positive, including swabs taken from the pharmacy office (office, door, and chair), from the laboratory station where samples are being processed, and in the nursing station at the ICU unit. On the other hand, all swabs taken from isolation wards and other hospital facilities were negative ( Table 2 ).

The contamination of surfaces and objects was investigated inside isolation rooms, at hospital or at home isolation. A total of 32 swabs were taken from banknotes, mobile phones, air condition filters, door handles, and sink faucets in isolation rooms in the hospital and at home, as detailed in Table 3 . To investigate the contamination of surfaces and objects in self-isolated and inpatients rooms, swabs were taken from door handles and sink faucets of rooms occupied by SARS-CoV-2-positive patients at home or in hospital. The air condition filter in two home isolation patients who were positive within 2 days from taking the swabs was negative.

Mobile phones and banknotes from patients at home or hospital isolation were also negative for SARS-CoV-2. Five random banknotes were taken from pharmacies, grocery shops, and fruit and vegetable shops were all negative for SARS-CoV-2 (Table 3 ).

To study whether public high-touch surfaces can represent a possible route of transmission for SARS-CoV-2 infection, swabs (Table 4 ).

Previous studies investigated the contamination of COVID-19 hospital wards; however, the focus of this study was to inspect the 

The contamination of surfaces and objects with SARS-CoV-2 has been proposed as one of the main routes of transmission of the infection. 4, 9, 13 It was recently reported that the detection of the virus always declines after the first week of the disease time course. 16 This can be the cause of not detecting viral contamination in patients in self-isolation or in ICU. Additionally, viral detection in rooms occupied by patients with severe pneumonia was reported to be small. 10 Viral load was reportedly lower in patients with progressive pneumonia. 17 For this, SARS-CoV-2, being undetectable in ICU, can be rationalized as due to lower viral load, regular disinfection, and limited movement in the room as patients are mostly on external oxygen support or mechanical ventilation.

Regarding dental clinics, it was shown that dental chairs and sinks in the dentistry clinic can be positive for SARS-CoV-2.

Although viral contamination was not previously detected in dental clinics, it was shown that saliva is a source of SARS-CoV-2

infection. 18 Dentistry was proposed as one of the major medical fields affected by the pandemic and strict measures should be taken to prevent infection. 19 

The SARS-CoV-2 RNA was detected mainly in nonclean areas, while ICU units were mostly clean. Under health-care settings, door handles were the most contaminated objects, while medical equipment such as dental chairs and ophthalmoscopes were also positive for SARS-CoV-2 RNA. Hand swabs taken from patients in ICU were negative, while the viral RNA was detectable on the hands of patients in isolation rooms. Random samples from mobile phones and banknotes were negative; however, the ATM returned some positive results, showing its possible role in the spread of infection. The results showed the necessity to take more protective measures to prevent the spread of the infection through door handles and medical instruments. Additionally, it is confirmed that the ATM can be a source of infection.

The author declares that there are no conflict of interests.

Study concept and design: Hossein M. Elbadawy and Amin K. Khattab. 

The peer review history for this article is available at https://publons. com/publon/10.1002/jmv.26819

The data that support the findings of this study are available from the corresponding author upon reasonable request.

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