key: cord-1049617-f1mu7hwf authors: Mello, V. M.; Eller, C. M.; Salvio, A. L.; Nascimento, F. F.; Figueiredo, C. M.; Silva, E. S. R. F.; Sousa, P. S. F.; Costa, P. F.; Paiva, A. A. P.; Mares-Guias, M. A. M. M.; Lemos, E. R. S.; Horta, M. A. P. title: Effectiveness of Face Masks in Blocking the Transmission of SARS-CoV-2: a Preliminary Evaluation of Masks Used by SARS-CoV-2-Infected Individuals date: 2021-06-27 journal: nan DOI: 10.1101/2021.06.20.21259167 sha: 69be84ebfb05e82dbc84e1d1f54947afa88579ba doc_id: 1049617 cord_uid: f1mu7hwf In 2019, a novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is trans-mitted via airborne route, caused a new pandemic namely, 'coronavirus disease 2019' (COVID-19). Although it is still debated whether the use of masks can prevent the transmission of SARS-CoV-2, no study has evaluated the virus-blocking efficacy of masks used by patients. We aimed to evaluate this efficacy of masks used by SARS-CoV-2-infected individuals. Data, masks used, and nasopharyngeal swab samples were obtained from these patients. Forty-five paired samples of nasopharyngeal swabs and masks were obtained and processed; the majority of masks were woven. Viral RNAs were amplified using quantitative reverse-transcription polymerase chain reaction and detected only on the inner parts of masks. Median cycle threshold (CT) values of swabs and masks were 28.41 and 37.95, respectively. Statistically, there was a difference of ap-proximately 10 CT values between swabs and masks and no significant difference in CT values among different types of masks. There were statistically significant differences in CT values be-tween men and women and symptomatic and asymptomatic patients. Our findings suggest the blocking of the transmission of the virus by different types of masks and reinforce the use of masks by both infected and non-infected individuals. In 2019, a new respiratory coronavirus namely, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is transmitted via the airborne route, primarily through respiratory droplets and aerosols, caused the new global pandemic associated with a respiratory syndrome namely, the "coronavirus disease 2019" (COVID-19) that has resulted in millions of deaths [1, 2] Some studies suggest that the use of a mask can possibly prevent the transmission of several respiratory viruses, such as influenza and rhinovirus, in addition to the new coronavirus [3] [4] [5] . Although there has been much discussion regarding whether masks should be used to prevent viral transmission during the initial period of the COVID-19 pandemic, now there is a global understanding of the importance of using masks for preventing SARS-CoV-2 infection. It has been reported that a mask not only protects the person who is wearing it but also reduces the likelihood of transmission of the disease from the person wearing the mask to another person [6] . Current epidemiological data indicate that wearing a mask can reduce the emission of SARS-CoV-2 particles into the environment [7] . The surgical mask (nonwoven mask) had a greater filtration efficiency for viral aerosols; however, the filtration efficiency was inferior to that of an N95 mask [8] [9] [10] [11] . With the worsening of the pandemic in some countries, especially the developing ones, countries have suffered from the non-availability of surgical masks [8, 12, 13] . As a great alternative, homemade fabric masks have become very popular in several affected countries, mainly in Brazil [14] [15] [16] [17] [18] [19] [20] . Although fabric masks provide less protection and have low filtering efficiency when compared with surgical masks, they may have some effectiveness in preventing the transmission of SARS-CoV-2 [8] [9] [10] [11] . Nevertheless, these homemade masks are produced by small-scale fashion productions and do not have quality certifications from health authorities [14] [15] [16] [17] [18] [19] [20] . Despite the World Health Organization recommendations about the use of face masks, it is still controversial whether it reduces the risk of transmission of SARS-CoV-2 [21] . No study has evaluated the presence of retained viruses on the masks, which are made using different materials and used by SARS-CoV-2-infected individuals, and the effectiveness of these masks in preventing viral transmission. Considering the heterogeneity of cloth masks that are sold in Brazil, it is still unclear whether these homemade masks are effective in blocking the transmission of virus. Considering these . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted June 27, 2021. ; https://doi.org/10.1101/2021.06.20.21259167 doi: medRxiv preprint points, in the present study, we aimed to evaluate the virus-blocking efficacy of masks used by SARS-CoV-2-infected individuals. The results presented here suggest that the use of masks helps to block viral transmission by SARS-CoV-2-infected individuals and reinforce the importance of using masks as a preventive measure against the viral transmission. The Oswaldo Cruz Institute/IOC/FIOCRUZ Research Ethics Committee approved this study (number: CAAE 37142520.0.0000.5248). All procedures were performed in accordance with the ethical standards of the responsible committees on human experimentation (institutional and national) and the Helsinki Declaration of 1975, as revised in 2008. All patients who were included in the study agreed with their participation in the research by signing the informed consent. Nasopharyngeal swab samples and masks were collected from patients who were suspected to be infected by SARS-CoV-2 and attended the Municipal Theatre and Benjamin Constant Institute survey, conducted in the city of Rio de Janeiro, Brazil, according to medical decision and after obtaining permissions from the patients. Samples were collected as follows: a nasopharyngeal swab was inserted in the nostril until it hit an obstacle (the inferior concha or the back of the nasopharyngeal cavity), rotated, and removed. The test was conducted in two nostrils per patient. After sampling, the nasopharyngeal swab was inserted into a vial containing 3 mL of a viral transport medium (VTM; Xpert nasopharyngeal sample collection kit, Cepheid, Sunnyvale, CA, USA). After the collection of swab samples, the masks used for 2-3 h by the participants were placed inside a clean plastic bag and they were provided clean, new masks for use. Furthermore, data, including the biological sex and age of these patients were collected. . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) Nasopharyngeal swab specimen was collected and immediately resuspended in 3 mL of the VTM. For mask samples, immediately after the collection of masks, pieces were cut based on the following reference measures: the right side and left side areas with a width of 2 cm each, obtained after removing side seam using the entire height of the mask; the nose area (N) with a height of 5 cm and width of 5 cm; and the mouth area (M) with a height of 5 cm and width of 8 cm, and subsequently, these pieces were added to the VTM. In cases of samples with double or triple layers of the material, these areas were . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 27, 2021. ; https://doi.org/10.1101/2021.06.20.21259167 doi: medRxiv preprint Between resuspension and the processing of each sample (swabs and mask pieces), incubation at 4 °C for a minimum of 30 minutes and a maximum of 12 hours was be performed. Subsequently, the samples were processed through vortex homogenization and transferred from the medium to a previously identified 1.5-mL tube using a Pasteur pipette (2 mL). Then, swabs and masks were discarded, and the final sample in the medium was stored at -80 °C. Nucleic acids from all the samples were extracted and purified using the DNA/RNA 300 kit H96 in the Janus G3 and Janus Chemagic automatic extractor (Perkin-Elmer, Waltham, USA). The Janus 360 system is based on magnetic spheres for extracting viral nucleic acids from 300 uL of the sample. The operation of the equipment and the use of the commercial kit were in accordance with the manufacturer's instructions. For SARS-CoV-2 genome amplification, we used a molecular kit for the E region . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 27, 2021. ; https://doi.org/10.1101/2021.06.20.21259167 doi: medRxiv preprint The results of descriptive statistical analyses are presented using frequency tabulations and percentages. Medians are presented with interquartile range (IQR) values. The Mann-Whitney U test was used to compare the differences in Ct values between the independent groups of masks and swabs. Statistical significance was set at a p-value ≤ 0.05. All analyses were performed using R software version 4.1.0 (The R Foundation for Statistical Computing, Vienna, Austria). Forty-five swab samples with their paired respective masks were collected. The masks were classified as woven masks (30/45; 66.7%) and surgical non-woven masks respectively. The descriptive information can be seen in Table 1 . . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. Women and men presented significant differences in median Ct values of masks (U = . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. Table 2 . After 1 year of the COVID-19 pandemic, the Americas have become the epicenter of COVID-19 cases and deaths; especially in Brazil, there has been an increase in the average number of deaths [1] . This may be associated with late interventions against the pandemic and adherence to scientific negationism, for example, not wearing protective masks, among other factors [22, 23] . The results in this study, reinforce the evidence that in general, wearing masks can be beneficial to the community and that this beneficial effect is derived from the ability of masks to block the exhalation and inhalation of infectious viruses, regardless of the type of mask used, as shown in a review by Brooks and Butler (2021) [24] Data from different studies conducted in several countries have shown that the use of masks together with social distancing can reduce the transmission of SARS-CoV-2 and the number of cases of SARS-CoV-2 infection [11, [25] [26] [27] [28] . A study performed by Ma et al. (2020) , which used an automated system that mimicked human breathing, showed that the virus-blocking rates of surgical and homemade masks were approximately 97% and 95%, respectively [29] . Another study performed by Morais et al. (2021) , which used a . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 27, 2021. similar methodology for evaluating different mask types, demonstrated similar results, where surgical masks had a filtration rate of 89% and homemade masks had filtration rates ranging from 40% to 83%, depending on the type of the fabric [30] . In the present study, we observed a reduction by approximately 10 Ct values (≅ 3 logs [≅ x1000 RNA copies/mL]) for masks compared with the paired swabs collected from the same individual. These findings corroborate with data from the previous studies [29, 30] (2020) that evaluated the effectiveness of the materials of cloth masks, which were used to reduce the transmission of SARS-CoV-2, in the filtration of nanoscale aerosols and showed that no cloth mask performed similar to an N95 mask. However, woven, and nonwoven cloth masks may be used to reduce the transmission of SARS-CoV-2 and to filter viral particles [31] . In this context, in a country like Brazil, where it is impossible to totally adopt measures of social distancing, mainly in socially vulnerable populations in peripheral areas and slums, the use of masks seems essential. Moreover, the use of masks could be beneficial to those individuals who still need to use public transport, such as buses, trains, and/or subways, which are often crowded [21, [32] [33] [34] . The use of masks, especially . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 27, 2021. ; https://doi.org/10.1101/2021.06.20.21259167 doi: medRxiv preprint woven ones, is extremely relevant as an additional protective measure for reducing the increasing number of cases and deaths due to COVID-19 in Brazil [12] . Statistically significant results (p = 0.02 and p = 0.01, respectively) were obtained on comparing Ct values between swabs and masks from men and women. This may be directly associated with the sex-a hypothesis to be considered is a greater release of viral particles by males. Some studies have shown that males have a significantly high risk of severe disease, mainly due to differences in inflammatory responses to viral infections, and genetic and hormonal regulation [35] [36] [37] . However, more studies are needed to understand the underlying biological phenomena. Some studies suggest that the viral load found in asymptomatic patients is similar to that found in symptomatic patients [38] [39] [40] . However, we identified lower Ct values in symptomatic patients than those in asymptomatic patients, and this difference was statistically significant (p = 0.004), indicating an elevated viral load mainly in swab samples. This was a preliminary study and has some limitations. The sample size was relatively small, and it did not evaluate the filtering efficiency of the masks as performed in some other studies [28, 29] . Furthermore, this study only evaluated masks from SARS However, our results provided real-life evidence regarding blocking of viral transmission by masks used by individuals infected by SARS-CoV-2. Furthermore, the results also reinforce the suggestion to use a mask by everyone, regardless of whether the individual is infected or not. This is important since there are asymptomatic cases of infection and evidence of transmission of the virus even before the appearance of the first symptoms, as reported by some studies [38, 41] . . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 27, 2021. ; https://doi.org/10.1101/2021.06.20.21259167 doi: medRxiv preprint The study results shed light on the importance of using masks by individuals infected with SARS-CoV-2 and shows that different types of masks can help block viral transmission. Moreover, our findings also reinforce the importance of using masks as a preventive measure against the viral transmission, regardless of the type of mask used, in addition to social distancing and personal hygiene measures. All patients in the study were aware of and according to their par-ticipation in the research, after signing the informed consent. We are also grateful to the Rio de Janeiro Municipal Theatre team and Ben-jamin Constant Institute team for our collaboration. We especially thank all people who made themselves available to participate in this study through the swab and mask collections. . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted June 27, 2021. ; https://doi.org/10.1101/2021.06.20.21259167 doi: medRxiv preprint World Health Organization. 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The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.