key: cord-1032740-17xz7dz8 authors: Ali, Banafsheh Haji; Shahin, Mohammad Sajjad; Sangani, Mohammad Mahdi Masoumi; Faghihinezhad, Mohsen; Baghdadi, Majid title: Wastewater Aerosols Produced during Flushing Toilets, WWTPs, and Irrigation with Reclaimed Municipal Wastewater as Indirect Exposure to SARS-CoV-2 date: 2021-08-13 journal: J Environ Chem Eng DOI: 10.1016/j.jece.2021.106201 sha: 5c5b29270fb265fdd11ff0c7a33624a068b2c7ce doc_id: 1032740 cord_uid: 17xz7dz8 The detection of SARS-CoV-2 RNA in raw and treated wastewater can open up a fresh perspective to waterborne and aerosolized wastewater as a new transmission route of SARS-CoV-2 RNA during the current pandemic. The aim of this paper is to discuss the potential transmission of SARS-CoV-2 RNA from wastewater aerosols formed during toilet flushing, plumbing failure, wastewater treatment plants, and municipal wastewater reuse for irrigation. Moreover, how these aerosols might increase the risk of exposure to this novel coronavirus (SARS-CoV-2 RNA). This article supplies a review of the literature on the presence of SARS-CoV-2 RNA in untreated wastewater, as well as the fate and stability of SARS-CoV-2 RNA in wastewater. We also reviewed the existing literatures on generation and transmission of aerosolized wastewater through flush a toilet, house's plumbing networks, WWTPs, wastewater reuse for irrigation of agricultural areas. Finally, the article briefly studies the potential risk of infection with exposure to the fecal bioaerosols of SARS-CoV-2 RNA for the people who might be exposed through flushing toilets or faulty building plumbing systems, operators/workers in wastewater treatment plants, and workers of fields irrigated with treated wastewater - based on current knowledge. Although this review highlights the indirect transmission of SARS-CoV-2 RNA through wastewater aerosols, no research has yet clearly demonstrated the role of aerosolized wastewater in disease transmission regarding the continuation of this pandemic. Therefore, there is a need for additional studies on wastewater aerosols in transmission of COVID-19. In light of the emergence of the new coronavirus in China caused worldwide pandemics, it seems necessary to have a greater understanding of the role of wastewater as a risk factor for public health. Emerging pathogens from humans can enter sewage systems. It could be because feces and urine from humans can contain a wide range of pathogens, which sewage systems can transport to wastewater treatment plants [1, 2] . The SARS-CoV-2 is enveloped RNA virus of the viral family Coronaviridae like SARS-CoV and MERS-CoV, which can cause respiratory illnesses known as COVID-19 [3] [4] [5] . J o u r n a l P r e -p r o o f reach wastewater treatment facilities, and therefore, they cannot be eliminated in wastewater systems [56] . The survivability of coronavirus is influenced by viral structure. The lifespan of enveloped viruses such as coronavirus is shorter than non-enveloped viruses. The reason would be the activity of detergents and proteolytic enzymes on the virus's outer layer of lipid envelope [48, 50] . Wastewater characteristics and the compounds available in wastewater can play a crucial role in the survival and inactivation of coronaviruses in sewage. Coronavirus can be quickly inactivated in wastewater due to the existence of chemical which has the anti-viral properties. Studying virus survival in wastewater has also shown that organic matter and suspended solids in wastewater may have a strong influence as helper contributions on the survival of viruses in sewage because they can afford protection to viruses which may result from adhering to these particles [54, 55] . More details of studies factor are tabulated in (Table S1) . Evidence from recent studies suggests that temperature has the most crucial effect on SARS-CoVs survival in wastewater [58, 59] . The result of these studies has also indicated that the persistence of SARS-CoVs in sewage reduces with an increase in temperature. Thus, SARS-CoVs can survive and remain infectious for long periods at low temperatures in wastewater (Table S2 ) [50, 60] . Likewise, preliminary research on SARS-CoV-2 illustrated that low survival times of SARS-CoV-2 RNA in sewage occur in temperatures more than 20°C [61] . The fecal pH has a significant effect on the survival of SARS-CoV-1 starting from three hours in newborn baby's stools, which is slightly acidic, to four days at pH up to 9 in diarrhea stool of grown-ups. Conversely, results from the initial research on SARS-CoV-2 have shown J o u r n a l P r e -p r o o f that SARS-CoV-2 in suspension does not considerably decrease following titration (60 minutes) at various pHs (3) (4) (5) (6) (7) (8) (9) (10) [54, 62] . Above all, it should be noted that amount of SARS-CoVs in wastewater may change seasonally and daily according to wide-ranging factors, including the prevalence of disease during outbreaks in communities and the rate of viral shedding (i.e., feces, urine, vomit), which discharges into the wastewater system and meteorological and environmental conditions. The temperature of wastewater changes seasonally because seasonal variation in weather and soil temperature impacts the heat transfer between wastewater and the surrounding ambient [14] . Based on data from different studies, SARS-CoV-1 and MERS-CoV can maintain viability under various ecological conditions in wastewater. In view of the evidence, the survival period of SARS-CoV-1 in sewage is 14 days at 4°C or 2 days at 20°C, which is considerably related to numerous environmental factors (such as temperature, solar, or UV inactivation) and their association with biofilms. Furthermore, its RNA could still be detected in the sewage after 8 days, even if the virus was inactive [63, 64] . Similarly, a recent analysis of the wastewater in Arizona, USA for SARS-CoV-2 RNA revealed that at 20 °C, the remains of virus in sewage is estimated to be at least 25%, even in a circumstance where the average time of travel of sewage through the sewer network is 10 hours and the stability of virus is reasonably low [65] . strong upwards airflows, inadequate traps, and non-functional water seals [66] [67] [68] [69] , and fecal aerosols seem to be formed during the wastewater treatment processes on a large scale. Further, aerosol transmissibility of viruses is likely to be feasible during irrigation by treated wastewater because a conventional WWTP without disinfection can usually provide a substantial reduction of viruses but not complete removal. This issue becomes even more important if high viral loads in the influent wastewater arrive at WWTPs during pandemics, which could cause the inadequate decrease of viruses before discharge [54] . In research, Ali et al. investigated the reduction of SARS-CoV-2 RNA concentration in different sectors of two WWTPs in Israel. Their findings highlight the potential and shortcomings of traditional wastewater treatment in lowering SARS-CoV-2 RNA concentrations, as well as early evidence for the necessity of tertiary treatment and disinfection by chlorine in preventing viral spread to the environmental surroundings [70] . It is noteworthy that fecal bioaerosols can consist of viable and dead pathogens, which can pose a severe risk to public health, especially exposed people (e.g., residents of buildings, workers of sewage treatment plants, or residents of neighboring areas, and farmers) [71] . Observations to date indicate that microbial bioaerosols exposure has been connected to a wide range of health effects involving infectious diseases, acute toxic effects, allergies, and cancer [72, 73] . Interestingly, these negative health effects of fecal bioaerosols on humans come from their size, which can easily be transmitted by inhalation, ingestion, and skin contact. Therefore, they are usually capable of causing infections among people [74] [75] [76] . through fecal bioaerosols or not. To this end, the authors have proposed studying the possibility of fecal bioaerosols as a route of transmission for SARS-CoV-2 in the prevention and proliferation of a recent pandemic [77] . Based on the prior literature, during the SARS-CoV-1 outbreak in Hong Kong (2003), wastewater-associated bioaerosols were reported as a key mechanism of SARS-CoV-1 transmission among individual homes and buildings [66, 78] . On balance, authors stated that the survival of the virus in sewage due to the lack of adequate disinfection could increase contagion risk [79] . For instance, Yu's study group, in 2003, illustrated that fecal aerosols, which had been transmitted via sewer systems, were involved in the spread of the virus of SARS-CoV to more than 300 residents in the Amoy Gardens outbreak [80] . It should be mentioned that SARS-CoV concentrations in sewage and their dissemination potential are directly affected by the number of households connected to the urban wastewater collecting system [81, 82] . Consequently, according to the similarities between SARS-CoV-1 and SARS-CoV-2, it is suspected that the SARS-CoV-2 RNA can be transmitted through sewage aerosolization in the current outbreak [54, 83] . A recent study by Gormley et al. (2020) explained that the wastewater system, in some circumstances, can enable airborne transmission of SARS-CoV-2 [84] . Similarly, McDermott's research group confirmed that fecal bioaerosols are among the significant routes transmitting SARS-CoV-2 in hospitals [85] . This result was in line with findings reported by , indicating the high concentration of airborne SARS-CoV-2 inside the patient mobile toilet room Fangcang Hospital due to aerosolization from feces and urine [86] . Later, further information was given by Xu et al. (2020) that the sewage system was suggested as one possible transmission route of SARS-CoV-2 in the Diamond Princess Cruise ship [87] . Furthermore, Zhang et al. considered three hospitals in Wuhan and detected SARS-J o u r n a l P r e -p r o o f CoV-2 RNA in the form of aerosol with a concentration of (285-1,130 copies/m 3 ). Their results indicated a substantial viral spillover in the hospital outside settings, which might have been produced by infected people exhalation or bioaerosols from wastewater that carries SARS-CoV-2 RNA [88] . Another point in which EC et al. reported research on SARS-CoV-2 transmission in flight, covering 130 individual flights and two studies on airplane wastewater. PCR-positive SARS-CoV-2 samples were found in two wastewater investigations, but with Cycle threshold values (C t ) varying from 36 to 40 [89] . Thus, at the same time, when transmission routes of novel coronavirus are attracted the world's attention, the chance of spreading SARS-CoV-2 via the fecal-oral route must not be ignored [45] . There is no doubt that the continuous pandemic of COVID-19 emphasizes the need for more excellent knowledge associated with transmission routes through wastewater exposure pathways; however, fecal bioaerosol is another route of SARS-CoV-2 transmission that has been less investigated [83] . The relationship between inhalable airborne particles generated from disturbed sewage and infection spread has been suggested for a century. Therefore, scientists have formerly been concerned that toilet plumes pose a risk for transmitting infectious diseases because a large number of bioaerosols can be generated by the high pressure and turbulence of toilet flushing [90] . Nevertheless, a few research studies have been dedicated to this issue [80, 93] . A recent study by Li et al. (2020) was conducted on the effects of toilet flushing on virus transmission promotion. The results indicated a high fluctuation in both types of toilets (single-inlet flushing and annular flushing) tends to create a 5 m/s upward velocity that can throw the aerosol away from the toilet bowl. Besides, they observed that 40-60% of aerosols rose from the toilet seat to a height of 106.5 cm above the floor [91] . Another study by D. Knowlton et al. (2018) investigated the quantity of particulate matter and bioaerosols in the hospital toilet in three distinct scenarios, including neither waste nor flushing, flushing without waste, and fecal matter with flushing a J o u r n a l P r e -p r o o f toilet. The bioaerosols concentration was measured 0.15 m, 0.5 m, and 1.0 m from the rims of the toilet, with 5, 10, and 15 min before and after flushing the toilet. The findings suggested that the amount of bioaerosol increased in the toilet space in the case of feces flushing. Additionally, the contrast in the concentrations of bioaerosol was not found at various points in time, indicating that bioaerosols existed in indoor air environments for a prolonged retention time (longer than 30 min after flushing). They summarized that feces and flushing in nosocomial toilets exacerbated the condition towards bioaerosol increase [94] . Besides, Gormley et al. employed two techniques to assess the emission of bioaerosols occurrence. The Aerodynamic Particle Sizer (APS) data indicated that the majority of particles (>99.5%) were smaller than 5 μm and hence classified as aerosols. During system defect conditions, particles created inside the municipal plumbing system in the form of a flushing toilet cause emissions into the building, including an equivalency by somebody speaking loudly for almost 6 and a half minutes. The result also demonstrated that no particles larger than 11 μm were found in the whole system. The volume of toilet flushing correlated to the population of particles. However, there was a lack of information on the impact of airflow rate on particle count. The number of particles for a 6 L toilet flush was between 3000-4000, while for a 1.2 L toilet flush, the total particle was 886-1045. As a result, the decrease in particle size is proportional to the decrease in toilet flush amount [69] . Similarly, Wu et al. introduced a new toilet consisting of a liquid-curtain technique and demonstrated its benefits using computational fluid dynamics. The results showed that the liquid curtain could inhibit upward viral particles in order that a mere 1% of viruses can go up above the toilet, and the water flow can properly transport viruses into the wastewater [95] . Hence, existing scientific research on sewage aerosolization from toilets has demonstrated that flushing a toilet may cause to release of up to 80,000 bioaerosols into the J o u r n a l P r e -p r o o f toilet's indoor air and the toilet environment, floating above the toilet area around one meter (3.2ft) . Therefore, these bioaerosols can easily be floating in air currents due to their low settling velocity, finally depositing on the toilet seat, toilet lid, and the toilet's surface surroundings [90, 92, 96] . Other authors like Mendes and Lynch observed similar findings. The results of their investigations in public toilets in schools, shops, factories, offices, railway sites, and hospitals showed that bioaerosols were found in the seat of the toilet, washbasin, faucet handle, and floor surrounding toilet. The remarkable point was that some toilet surfaces like the seat of the toilet, the handle of the faucet, washbasin, and interior door knobs were more contaminated than other surfaces [97] . Another issue is that wastewater drainage systems in buildings can serve as a potential reservoir for many bacterial and viral pathogens. Based on recent facts, it seems likely that viruscontaining fecal aerosols may be airborne in drainage systems and vents for several hours, regarding being sufficiently small in size. Generally, the hydraulic interactions of toilet wastewater inside vertical building drainage stacks can generate virus-laden aerosol particles after flushing a toilet. The buoyancy (chimney) effect, as well as falling wastewater, can act as a driving force for these virus-contaminated aerosols to move in the drainage stacks and vents transiently. The buoyancy effect is a common phenomenon in high-rise buildings when there is a difference between the air temperature and humidity in the drainage pipes and the indoor air, especially the air in the bathrooms [77, 98] . Additionally, the suction flow rate is related to the negative pressure on each flat. A negative pressure situation can result from both toilet or bathroom exhaust fans and a northerly wind that generates a wake flow when there is a bathroom window onto a balcony. In a better word, the negative pressure can cause the generated fecal aerosols in the drainage pipe to suck into bathrooms, and consequently, they can settle on some surfaces; after that, it is possible to spread to other residents or surfaces by touching them [77, 98] . For the first time, the hypothesis of a transmission pathway for SARS-CoV-1 through dried-up U-traps, which allowed the virusladen fecal aerosols to enter households from the plumbing system, was suggested WHO in the According to recent infectious disease outbreaks worldwide, more scientific attention has been paid to the fecal aerosols containing the virus as a transmission route of infectious diseases in buildings, especially in high-rise residential buildings with dense populations [98] . As mentioned, a wide variety of transmissible pathogens can be carried by human excreta. They enter into sanitation facilities, and building drainage systems can cause a range of potential transport pathways. Research suggests toilets seem to be among the most probable infection and disease transmission spots especially shared toilets in hospitals and workplaces [99] . Since the transmission of SARS-CoV-2 RNA through fecal aerosols was suspected, various studies have investigated whether fecal aerosol may play a role in the transmission of COVID-19 and lead to potential community risks of COVID-19 outbreak. An early report by Ong and al. (2020) indicated that the total positive results of surface samples in the patient's J o u r n a l P r e -p r o o f bathroom was 60%, which highlighted fecal shedding of viral RNA by patients as a possible way of transmission of the virus [100] . In another recent case report at Fangcang Hospital in China, the highest concentration of SARS-CoV-2 aerosols appeared in a patient mobile toilet room, which was a portable single toilet unit of an approximate one square meter area without ventilation. Therefore, the authors proposed that airborne SARS-CoV-2 could generate during the patient's breath or aerosolized the urine and feces of patients [86] . Results obtained in samples from bathrooms of four isolation rooms in The Second Hospital of Nanjing in China indicated that a tremendous amount of SARS-CoV-2 virus was detected in patients' toilets in the hospital. Thus, the authors suggested the toilet as the most contaminated area of the hospital compared to other parts. In addition, the tested positive surface samples on the ceiling-exhaust grille and the toilet-exhaust louvers recommended the possible existence of fine airborne aerosols in the patient bathrooms due to patients' exhalation or fecal-derived aerosols during flushing a toilet ( Fig. 3 ) [101] . Similarly, it has been assessed the SARS-CoV-2 stability in the quarantine hotel environment. It was indicated the contamination rates of hand sinks (12.82%), toilet seats and flushes (7.89%), and floor drains (5.41%) [103] . In another study, Cheng et al. believe that the airborne path is not the most common way for SARS-CoV-2 to spread. They examined the air and surface sample of an isolated room that the patient was there. All air samples for SARS-CoV-2 were negative, but SARS-CoV-2 RNA was found in contaminated environments such as patients' J o u r n a l P r e -p r o o f mobile phones (6 of 77, 7.8%) and toilet door handles (4 of 76, 5.3%) with a median concentration of 9.2 × 10 2 copies. mL -1 (range, 1.1 × 10 2 to 9.4 × 10 4 copies. mL -1 ) [104] . conversely, the results in further research on SARS-CoV-2 air contamination evaluation in medical settings showed that the positivity rating was 5 of 21 air samples (23.8%) with concentrations of 9.7 × 10 3 copies. m -3 (5.1 × 10 3 to 14.3 × 10 3 copies. m -3 ) in the air of toilets or bathrooms [105] . It should be mentioned that the SARS-CoV-2 contamination in the six bathrooms of the non-ICU isolation ward in West China Hospital before routine cleaning was not remarkable and high, while in several studies, the contaminated areas in the toilet have been seen in dedicated COVID-19 hospitals [106] . The authors mentioned flushing a toilet daily with 2000 mg/L chlorine solutions as a possible reason [107] . Evidence from other studies, as tabulated in Table S3 ( in Guangzhou, China in a period of social distancing (Fig. 4b) . It should be mentioned that block X was a 29-floor residential high-rise with 3-unit apartments (flats 01, 02, and 03) on each floor except the 29th floor ( Fig. 4a) . Two similar cases were also observed in Hong Kong. Those results seem to indirectly confirm the possibility of vertically aligned flats to facilitate the spread of SARS-CoV-2 within, or even between, apartments because of the common vertical drainage pipes [109, 110] . (2020) found that SARS-CoV-2 could be alive on different surfaces for 3 days; the results suggest that the SARS-CoV-2 seems to be more stable on plastic, stainless steel, which was detected viable after 2-3 days [111] . Another study also estimated SARS-CoV-2 half-lives on indoor surfaces, including cloth, styrofoam, cardboard, concrete, rubber, glass, stainless steel, galvanized, and steel under three seasonal conditions, which are exhibited in Table 3 . J o u r n a l P r e -p r o o f The Table 4 ). The results showed that the median risks of illness with COVID-19 per exposure for a single day were estimated 1.11 × 10 -10 and 3.52 × 10 -11 for two scenarios of flushing a toilet and faulty drainage, respectively. Moreover, the estimated exposure of the worst-case scenario, highly polluted aerosols after flushing toilets in the hospital bathroom, was reported 1.9 × 10 −6 per person per event [113] . J o u r n a l P r e -p r o o f As mentioned, one of the most important elements of particular matter (PM), which plays a crucial role in human health, is bioaerosol. It provokes cancer, severe toxic effects, diabetes, neurological diseases, hypertension, cardiovascular diseases, infectious diseases, and allergies. Most bacterial diseases like Tuberculosis, Anthrax, and Legionellosis are caused by long-term or even short-term exposure to bioaerosols [115, 116] . It can be argued that some bacteria, viruses, and fungi in the form of aerosols have prolonged stability in the air. It has been reported that bacteria with the size of (PM 10-2.5 ) remain in the air. The aerosols in which humans breathe into the respiratory tract deposit in different locations of the respiratory system with various precipitation rates (Fig. 5 ) [116] . for this reason, it gives significant indications for analyzing the possible health consequences of aerosols. J o u r n a l P r e -p r o o f Overall, bioaerosols structures are positively correlated with relative humidity due to pathogens' survival, influenced by relative humidity levels in the air [123] . Results obtained by Jones and Harrison (2004) indicated that an RH of 70-80% and a temperature of 12-15 °C led to the survival and proliferation of bioaerosols in the air [130] . In another study by Wang and et al. (2018), the RH range and the ideal temperature were 37.8% to 57.4% and 27.6 °C to 40.1 °C in the summer, respectively, to have maximum numbers of pathogens in the air. Also, Wang's results agreed with Han's research report, which suggested that RH of 40-60% and temperature of 20-37 °C led to the survival and proliferation of bioaerosols in the air [123, 125] . It seems that the survival of viable airborne pathogens is greatly negatively affected by solar illumination because pathogens can be inactivated by ultraviolet irradiation. As a result, this causes the death of pathogens in bioaerosols. It was also observed that high temperatures could control the number of airborne pathogens at WWTPs [118] . [119, 124] . In fact, the continuous flow of raw wastewater into the grit chamber can cause to generate a great number of bioaerosol droplets that the wind can scatter. Besides, screw conveyors, which can carry sludge or solid matter from the grit chamber to a storage container, are another one to generate and emit bioaerosols in the air in the WWTP's area [133] . In bar screens, which are required routine cleaning manually or mechanically, the water surface is regularly disturbed by mechanical devices such as the drive chains or by hand cleaned racks that result in the release of bioaerosols in the air surroundings of the WWTPs [76, 118] . Similarly, the authors stated that biological treatment, based on the aeration of wastewater, either by mechanical agitation or air bubbling, was determined as one of the main bioaerosol emission sources at a WWTP. In general, aeration, which provides oxidative processes in the aeration tanks, is achieved with diffused aeration located at the bioreactor bottom or mechanical surface aeration. Therefore, bioaerosol emission from aeration basins is associated with types of aeration and mixing of liquids by mechanical agitation [71] [72] [73] 124, 132] . In better words, bioaerosols release from aeration basins because of the intense mixing and turbulence; it means that a large number of pathogens can be entered into ambient air during aeration as a consequence of splashing and bubble bursting. The study of Sánchez-Monedero et al. (2008) showed that the bioaerosol levels generated were influenced by the type of aeration utilized in the biological process. Consequently, this may result in different exposure risk levels of site workers. In addition, the results of similar studies confirmed that mechanical aerationespecially horizontal rotors and surface turbine aerators -has generated higher levels of bioaerosols than bubble diffused air systems. However, a tremendous amount of respirable bioaerosols is generally generated by bubble aeration [72, 76, 118, 132] . Likewise, airborne pathogens emitted from different types of sludge treatment was observed in sludge storage site and various units of sludge treatment such as the sludge dewatering chamber, thickening basin, and the sludge centrifugation process [71, 125] . Obtained results of different WWTP stages also revealed that the highest amount of airborne pathogens was detected in the pretreatment process compared to other parts. This might result from the worse microbiological quality of wastewater in the first stages since the wastewater is still wholly untreated. The decrease in pathogen contamination levels might be due to the sewage become more treated [124, 126] . Thus, microbial bioaerosol emission levels will begin to decline from pretreatment to primary, secondary slowly, and the advanced treatment methods of wastewater [92] . Generally, virus removal in secondary treatment technologies is approximately 90% from raw sewage. However, conventional wastewater treatment processes cannot remove all viruses from sewage [64] . stage. The authors also reported similar results for the sludge processors due to becoming inactive a significant percentage of pathogens [126] . Infectious agents, such as respiratory viruses, could be transported in aerosols formed during processes of wastewater treatment, and thus, the aerosolization of the virus-infected droplets is likely to spread viral infections to workers in wastewater treatment plants [54, 134] . As has been illustrated by Khuder et al. (1998) and other studies, there is a critical correlation J o u r n a l P r e -p r o o f between exposure to wastewater bioaerosols and the occurrence of infectious diseases at WWTPs. In the better word, site workers may be exposed to fecal bioaerosols containing various infectious agents, including viruses, through inhalation or by the hand-to-mouth route. Hence, fecal aerosols could be considered a possible transmission route and pose a risk to workers at WWTPs and groups living around them. There is much evidence of this fecal-aerosol transmission pathway for various enteric viruses and bacteria. In other words, exposure to fecal bioaerosols, due to a potential risk of spreading the virus among the sewer workers, has been regarded. It becomes crucial when the infective pathogens can be easily transmitted from the exposed workers to their family members and other friends and relatives [118, 135] . Literature data demonstrated that wastewater operators -depending on kinds of facilities, carried out work, and climate conditions -may be exposed to the unpredictable amount of fecal bioaerosols [117, 124, [136] [137] [138] . In particular, the inflow chambers and treating processes that produce splashing, bubbling, and spraying have proved to be of greatest potential risk for site workers. Thus, during wastewater treatment processes, a significant exposure risk for sewage workers is [132] . In another study, Tang found that the outdoor wastewater treatment processes such as aeration parts and a sludge dewatering room generated various microbial aerosols that may cause high exposure risks microbial aerosols at WWTPs [122] . In view of processes, they differ mainly in being performed outdoors or indoor. In indoor sewage treatments where the main stages of the processes occur in a closed area, the concentrations of airborne pathogens in the ambient air are higher due to inadequate ventilation and a low rate of die-off from the absence of sunlight. Moreover, pathogens could survive better in indoor WWTPs. Therefore, the working areas of indoor wastewater treatment plants pose a higher risk of exposure to bioaerosols [118] . In fact, there is wide variation in the number of bioaerosols across different studies because of differences in methods of wastewater treatment and sludge, meteorological conditions as well as in sampling and statistical methods [137, 139, 140] ; consequently, no uniform international standard have been set up for estimating indoor and outdoor airborne pathogen concentrations [119] . Furthermore, in recent years, there is a great concern about the particle size of bioaerosols, which influences the exposure risk of respiratory diseases to the staff at a WWTP and surrounding residents [125] . Previous studies have reported that most bioaerosol particles in the air of a WWTP are in the respirable size range, diameter below 4.7 µm, which can raise the risk of penetrating deep into the lungs of exposed site workers. It has also been reported that the size distribution of bioaerosols significantly differs in various seasons. Of note, these small bioaerosols can be transported by the wind to distances between a few hundred meters to several kilometers. Therefore, they can pose a serious biohazard threat not only to sewage workers but also to neighborhood residents. Another issue of great concern is linked to the high ability of J o u r n a l P r e -p r o o f viruses to infect humans. This means that the minimum infective dose of airborne viruses is enough to cause an infection [71, 73, 75] . In the case of SARS-CoV-2, knowledge on SARS-CoV-2 aerosolization within WWTPs is limited, even though recent initial studies have confirmed the presence of SARS-CoV-2 RNA in wastewater influents of a WWTP in the Netherlands, France, Germany, Spain, Italy, Australia, the United States, and Turkey ( Table 5 ). [142] Gandhinagar, India Untreated wastewater RT-qPCR August: 897.5 September: 924.5 [143] Murcia, Spain Untreated wastewater RT-PCR 1.48 × 10 5 -3.9 × 10 5 [144] Virginia, USA Untreated wastewater RT-ddPCR 10 1 -10 4 [33] Australia Untreated wastewater RT-qPCR 19-120 [145] Yamanashi Prefecture, Japan Untreated wastewater RT-qPCR Not detected [36] Louisiana, USA Untreated wastewater RT-qPCR 3.5 × 10 3 -7.5 × 10 3 [18] Milan, Italy Untreated wastewater RT-PCR No data available [24] Ahmedabad, India Untreated wastewater RT-PCR 5.6 × 10 1 -3.5 × 10 2 [146, 147] Italy Untreated wastewater RT-PCR, RT-qPCR Below the identification threshold to 5.6 × 10 4 [26] Japan Untreated wastewater RT-qPCR 1.2 × 10 4 -4.4 × 10 4 [148] Belgrade, Serbia Untreated wastewater RT-qPCR 5.97 × 10 3 -1.32 × 10 4 [149] Valencia, Spain Untreated wastewater RT-qPCR 1.66 × 10 5 -9.77 × 10 5 [28] Montana, USA Untreated wastewater RT-qPCR > 10 4 [150] Canada Untreated wastewater RT-ddPCR, RT-qPCR PGS (RT-ddPCR): 1.24 × 10 6 -1.42 × 10 6 PGS (RT-qPCR): 1.42 × 10 6 -1.93 × 10 6 PCS (RT-qPCR): 1.10 × 10 6 -1.51 × 10 6 PCS (RT-ddPCR): 2.74 × 10 5 -3.93 × 10 5 [17] Rio de Janeiro, Brazil Untreated wastewater RT-qPCR Cycle threshold numbers are reported [151] Paris, France Untreated wastewater RT-qPCR 5 × 10 4 -3 × 10 6 [152] Israel Untreated wastewater RT-qPCR Cycle threshold numbers are reported [19] Istanbul, Turkey Untreated wastewater RT-PCR Primary and activated sludge: 1.17 × 10 4 -4.02 × 10 4 [153] USA Untreated wastewater RT-qPCR Primary sludge: 1.7 × 10 6 -4.6 × 10 8 [154] New York, USA Untreated wastewater RT-qPCR Below the identification threshold to 1.12 × 10 5 [155] Ourense, Spain Untreated wastewater RT-qPCR 7.5 × 10 3 -4 × 10 4 [156] Pakistan Untreated wastewater RT-qPCR Quantification cycle numbers are reported [157] Barcelona, Spain Untreated wastewater RT-qPCR < 10 5 [158] Santa Catalina, Brazil Untreated wastewater RT-qPCR 2.95 × 10 5 -5.01 × 10 6 [159] Santiago, Chile Untreated wastewater RT-qPCR Not detected-4.81 × 10 6 [160] Jeddah, Saudi Arabia Untreated wastewater RT-qPCR Septic tank: 1.74 × 10 2 -1.33 × 10 3 Hospital: 1.4 × 10 4 [161] San Francisco, USA Untreated wastewater RT-qPCR 1.07 × 10 3 -1.02 × 10 6 [162] USA Untreated wastewater RT-qPCR Nine states higher than 100 [163] Florida, USA Untreated wastewater RT-ddPCR 9.47 × 10 5 -2.19 × 10 6 [169] Lahore, Pakistan Untreated wastewater RT-qPCR Not detected-3.55 × 10 7 [170] Hyderabad, India Untreated wastewater RT-PCR Cycle threshold number are reported [171] Wuhan, China Untreated wastewater RT-qPCR Municipal: 7.4 × 10 3 Hospital: 3.8 × 10 3 -9.3 × 10 3 [172] Canada Untreated wastewater MPAD Cycle threshold number are reported [173] Rajasthan, India Untreated wastewater RT-qPCR Cycle threshold number are reported [174] Frankfurt, Germany Untreated wastewater RT-qPCR WWTP, Niederrad: 4 × 10 11 -1 × 10 15 copies/day WWTP, Sindlingen: 1 × 10 11 -2 × 10 14 copies/day [175] Hangzhou, China Untreated wastewater RT-qPCR Septic tank influent: 5.89 × 10 5 J o u r n a l P r e -p r o o f [176] Germany Untreated wastewater RT-qPCR Solid stage: 2.5 × 10 4 Aqueous stage: 3 × 10 3-2 × 10 4 In particular, the WHO has published brief literature data, which are in agreement with these reports. Despite these reports, there is a lack of enough information about the fate of SARS-CoV-2 during the different wastewater treatment stages, and consequently, few reports investigated aerosolization of SARS-CoV-2 during different stages of wastewater collection and treatment [3] . Thus, based on previous evidence, it can be concluded that the risks of aerosolization of SARS-CoV-2 can be extremely significant during pumping wastewater, discharge from sewerage networks, and in uncovered aerobic treatment units. This is why raising concerns about the exposure to SARS-CoV-2 emerged from the fecal-derived aerosols due to the considerable SARS-CoV-2 RNA load to WWTPs [54, 152] . Therefore, in few recent studies (Table 6) (0.036, 0.32, and 3.21 illness cases per 1000 exposed workers at WWTP, respectively, for low grade, moderate and aggressive scenarios) [178] . The later study of QMRA by Gholipour et al. (2021) found that the range of infection risks of SARS-CoV-2 for WWTP operators was from 1.1 × 10 -2 to 2.3 × 10 -2 pppy by analyzing the viral SARS-CoV-2 RNA presence in raw sewage and samples of air from surroundings of the WWTPs. The obtained results by the authors indicated that The estimated level of the annual risk of infection for treatment plant operators was greater than the suggested level by WHO (10 -3 pppy). Therefore, according to a few current investigations in the risk exposure assessment, further research is quite necessary [179] . 2% of population f 3% of infected individuals presumed to be responsible for SARS-CoV-2 stool shedding in wastewater g 0.3% of infected individuals presumed to be responsible for SARS-CoV-2 stool shedding in wastewater h 0.03% of infected individuals presumed to be responsible for SARS-CoV-2 stool shedding in wastewater i per person per year j illness case per exposed WWTP operators k South of Isfahan (Iran) L North of Isfahan (Iran) In the 21 st century, the role of water reclamation in order to protect water resources and the environment is not negligible. Generally, water reclamation is the reuse of wastewater after the suitable treatment process for the purposes such as the irrigation of agricultural land or replenishing surface water and groundwater. It is noteworthy that wastewater reuse is highly beneficial for the environment, as it can provide opportunities for communities to have the ability to access enough water for different purposes without requiring to utilize plenty of potable water [180] . Therefore, regarding the increasing need for wastewater reuse globally, it is critical to consider the role and potential of reclaimed wastewater in the spread of infections, especially in recent outbreaks [3] . J o u r n a l P r e -p r o o f As noted above, conventional wastewater treatment plants are not specifically designed to remove viruses in full-scale. On the whole, typical secondary wastewater treatment processes are usually able to achieve an average of 90 percent removal for viruses [54] . For instance, Prevost and colleagues observed that the viral load of treated wastewater effluents was decreased by 100 times compared to raw wastewater [151, 181] . Even though the level of removals seems widely changeable (ranging from inconsiderable to a 99 percent reduction), some supplementary wastewater treatment units (such as filtration and disinfection processes) are most commonly added to the current secondary treatment system to decrease the amount of virus in order to provide acceptable levels of effluent quality for discharging into the environment [180, 182] . The findings of previous studies on the presence of enteric viruses of humans in raw and treated wastewater indicated that the amount of virus in them was strongly related to the state of an epidemic in the population. A recent study conducted in Paris confirmed that the increase in COVID-19 cases is accurately impacted by the increase of genome units in untreated wastewater. This observation was very similar to another study in the Murcia Region (Spain). The results showed a close relationship between the prevalence of viruses and the load of viruses in the WWTP influents [29, 143] . Therefore, the issue is expected to significantly influence treated wastewater when wastewater viral loads during a pandemic outbreak are higher than expected. Consequently, the influent of high viral load cause inefficiency in the virus reduction of effluent before discharge. Thus, wastewater treatment facilities would be required quick responses to minimize the risk of spreading infection during outbreaks since remain viruses in wastewater effluents would perhaps affect recreation, irrigation, and drinking waters [183, 184] . Of note, this issue can become one of the main challenging issues that most developing countries face now because they could not be easily equipped with the advanced wastewater J o u r n a l P r e -p r o o f treatment technologies regarding inadequate infrastructure compared with developed countries. Accordingly, modern wastewater treatment plants significantly protect public health [151, 182] . For example, the results acquired in case of sampling and analysis from the WWTPs effluents in the United Arab Emirates demonstrated that none of wastewater effluent samples tested positive for SARS-CoV-2 RNA because of implementing a sequence of treatment technologies inclusive of preliminary, primary, secondary (activated sludge process /clarification), and tertiary (sand filtration, disinfection, chlorination). Indeed, these advanced technologies allow them to provide the safety of reusing treated wastewater for irrigating agricultural lands and urban green spaces [165] . Currently, most of the recently published data on SARS-CoV-2 only exist from detection in human stool and municipal wastewater. Nevertheless, the existence of SARS-CoV-2 RNA in the effluents of wastewater treatment plants and at various stages in the wastewater treatment plants has been analyzed by limited studies (Table S4) . J o u r n a l P r e -p r o o f Over the past decades, treated wastewater, owing to inadequate water resources, is most widely used to irrigate agriculture fields and the cities' green areas in numerous countries. Presently, in developing countries (such as Pakistan, India, Mexico, Iran, etc.), it is ubiquitous in reusing approximately 80% of untreated or partially treated domestic wastewater for irrigation. Accessing sufficient water quality for irrigation is essential to slow the spread of the disease and protect people's health. According to published papers ( the source of the emissions and the rising wind speed. Hence, the dispersion of these aerosols on regional scales can expose farm workers, and people located close to areas irrigated with reuse wastewater and high infection levels [187, 190] . Undoubtedly, the possibility of SARS-CoV-2 transmission via exposure pathways linked with agricultural reuse of wastewater needs a crucial consideration and requires further research during a current outbreak. The current pandemic has had repercussions for the whole social community. The media's coverage of the soaring number of mortality, as well as public limitations and quarantine, is likely to exacerbate anxiety, which might have significant consequences for social and mental health worldwide [191] . For instance, it has been reported that the COVID-19 J o u r n a l P r e -p r o o f pandemic caused dread in half of the overall population, although it generally had a stressful light effect [192] . Another report by Junling et al. showed a 48 % prevalence of sadness and anxiety among the Chinese populace, which was linked to social media usage [193] . Similarly, due to the outbreak, up to 38% of people were forced to work from home. A quarter of the people quit working, and their mental and physical health suffered as a result. Physically active people were more susceptible to mental health problems [194] . Also, Psychological discomfort was reported by up to 35% of those under lockdown. It has been demonstrated that young people, women, high-educated people, and the elderly were considerably more likely to acquire Post-traumatic stress disorder (PTSD) [195] . In health care settings, anxiety was mentioned by 23.2 percent of medical professionals, insomnia by 38.9%, and depression by 22.8 percent. Emotional symptoms were more common in female workers and nurses [196] . Furthermore, a recent study by Kang et al. showed that 22.4 percent of medical and nursing personnel in Wuhan had mild and 6.2 percent severe disruptions in the aftermath of the pandemic, particularly among young women. A total of 17.5 percent were required to attend counseling or psychotherapy [197] . For nearly more than one year, the world has got involved in the pandemic of the COVID-19 outbreak. Scientists worldwide study how to combat (identification, probable transmission routes, and disinfection) this disease. Hence, a mass of data is being produced every day. One of the main issues that have garnered worldwide attention at this crucial moment is the spread of SARS-CoV-2 through environmental media. via aerosol in WWTPs and toilets must be more investigated. Also, the survival of SARS-CoV-2 RNA in wastewater and the influential factors for the survival of SARS-CoV-2 RNA in wastewater were discussed. In the meantime, it can be helpful that the governments invest in monitoring wastewater to estimate the extent of the current pandemic or any other ones in the future. As presented, bioaerosols carry viruses, pathogens, and pathogens. The review demonstrates the bioaerosols generation in WWTPs and toilets. Based on results, the number of bioaerosols in toilets depends on toilet types, the energy of flushing, and distances from the toilet. Based on recent studies, there are some measurements to limit the transmission of viruses via bioaerosols. Providing a distinct wastewater collection network for feces and vomits of patients is suggested to prevent bioaerosol generation. Also, closing the toilet lid before flushing and using the extractor fan effectively reduces airborne pathogens in the bathroom. Moreover, consider a disinfection system before flushing can remove the bioaerosols effectively. Furthermore, the bioaerosols generated in WWTPs contain toxic compounds of pathogens that are harmful to personnel there. In the current pandemic, the infected people enter the mass of viral loads in wastewater that threaten transmitting the disease. 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