key: cord-0792753-lwjutzdv
authors: Ahmed, Warish; Bivins, Aaron; Simpson, Stuart L.; Bertsch, Paul M.; Ehret, John; Hosegood, Ian; Metcalfe, Suzanne; Smith, Wendy J.M.; Thomas, Kevin V.; Tynan, Josh; Mueller, Jochen F.
title: Wastewater surveillance demonstrates high predictive value for COVID-19 infection on board repatriation flights to Australia
date: 2021-10-14
journal: Environ Int
DOI: 10.1016/j.envint.2021.106938
sha: b99f1b4dedecaafe01989c26fdb0fd61cb100ddf
doc_id: 792753
cord_uid: lwjutzdv

Controlling importation and transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from overseas travelers is essential for countries, such as Australia, New Zealand, and other island nations, that have adopted a suppression strategy to manage very low community transmission. Wastewater surveillance of SARS-CoV-2 RNA has emerged as a promising tool employed in public health response in many countries globally. This study aimed to establish whether the surveillance of aircraft wastewater can be used to provide an additional layer of information to augment individual clinical surveillance testing. Wastewater from 37 long-haul flights chartered to repatriate Australians was tested for the presence of SARS-CoV-2 RNA. Children 5 years or older on these flights tested negative for coronavirus disease 19 (COVID-19) (deep nasal and oropharyngeal reverse-transcription (RT)-PCR swab) 48 h before departure. All passengers underwent mandatory quarantine for 14-day post arrival in Australia. Wastewater from 24 (64.9%) of the 37 flights tested positive for SARS-CoV-2 RNA. Clinical surveillance identified 112 cases of COVID-19 during the 14-day mandatory quarantine. Surveillance for SARS-CoV-2 RNA in repatriation flight wastewater using pooled results from all three RT-qPCR assays demonstrated a positive predictive value (PPV) of 87.5% and a negative predictive value (NPV) of 76.9% for COVID-19 cases during the post-arrival 14-day quarantine period (83.7% accuracy). The study successfully demonstrates that the surveillance of wastewater from aircraft for SARS-CoV-2 can provide an additional and effective tool for informing the management of returning overseas travelers and for monitoring the importation of SARS CoV-2 and other clinically significant pathogens.

Wastewater surveillance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genetic fragments (i.e., RNA) has proven to be an effective tool for assessing infections at the community level ( Australia historically has a low risk of importation of infectious diseases due to its geographic remoteness and, as an island nation, the ability to apply strict controls at entry points, including the use of mandatory quarantine, which has been an important part of minimizing COVID-19 impacts in Australia (Fotheringham et al., 2021) .

However, the COVID-19 pandemic has revealed Australia's vulnerability despite its geographic isolation and caused significant economic and social damages despite low national case numbers to date. The stringent border measures, including border closures and long periods of quarantine on arrival, contribute to these direct impacts, thereby causing significant disruption to the travel, tourism, and education sectors. Australia's policies for suppressing all community transmissions of COVID-19 show that the identification of even a single new case of community transmission is epidemiologically significant. In the absence of broad immunity, one of the best lines of defense against COVID-19 is preparedness, early detection, and aggressive response to isolating the identified cases.

Currently, international travel to Australia is limited. Many Australians are being repatriated via facilitated commercial flights operated by the airline Qantas on behalf of the Government. Upon arrival in Australia, passengers on these international flights must undertake a mandatory 14-day quarantine in a facility designated by the Government. Under the current program, to be accepted onto these flights, potential travelers are required to provide verified negative test results for COVID-19 (deep nasal and oropharyngeal RT-PCR swabs) 48 h before departure. Children under five are exempt. Strict application of personal protective equipment (PPE) and other measures are required during the flight. Each passenger is provided with a sanitizer, face masks, and a disposal bag for used masks. Face masks must be worn during the flight, and every passenger must change masks every two hours throughout the flight.

In this study, we describe the data collected from flight passengers upon arrival at dedicated facilities in Darwin, Northern Territory, Australia, where they were quarantined. We report the application of wastewater surveillance for SARS-CoV-2 RNA in wastewater from non-identifiable repatriation flights and its correlation with non-identifiable clinical data for subsequent detection of SARS-CoV-2 among passengers during the mandatory quarantine period (usually 14 days). Typically, COVID-19 clinical testing for travelers during quarantine includes an early deep nasal and oropharyngeal RT-PCR swab test (days 0-3 of quarantine), and a late deep nasal and oropharyngeal swab test (day 11 or 12). For some repatriating passengers, additional tests were conducted between days 3 and 11 for public health purposes. Once the international travel restrictions ease, and the testing of individuals may no longer be required, wastewater analysis may be used for surveillance for the presence of SARS-CoV-2 infections, information on novel variants, and COVID-19 case importation.

Wastewater sampling from the lavatory A total of 37 aggregated wastewater samples (500 mL to 1 L in volume) were collected from the wastewater exiting lavatories of 37 repatriation flights landing at Darwin International Airport (DRW), Northern Territory, Australia, between 17/12/2020 and 27/03/2021.

Wastewater samples were collected from flights exceeding 7 h duration. This duration was selected with the expectation that the repatriation passengers would consume food on one or more occasions while on these long-haul flight-legs (arriving at Darwin, Australia), and were more likely to use the lavatory during the flight. Wastewater was sampled using a specifically designed sample trap capable of retaining an aliquot of bulk wastewater sample exiting the aircraft lavatory before entering a waste service truck system ( Supplementary Fig. SF1 ). This 

To assess whether SARS-CoV-2 remained in aircraft wastewater tanks (after positive wastewater detection), we collected 28 lavatory wash samples (i.e., post-lavatory wash) from pre-departure flights to confirm the presence/absence of SARS-CoV-2 RNA. After emptying the wastewater tanks, the waste drain valve was left open, and approximately 200 L of water was pumped into each tank and drained into the lavatory waste truck. The waste drain valve was then closed, and an additional 40 L of water was pumped into each tank, followed by opening the waste drain valve to flush out 40 L of water into the lavatory waste truck. An aliquot (i.e., 500 mL to 1 L) of wash water sample was collected from the sampling trap, as wash water exited the aircraft into a lavatory waste service truck system. After wash water sampling, a sanitizing line was connected to the aircraft to dose each tank with aircraft toilet deodorant and viricidal/bactericidal (Novirusac Gel Bulk, Aero Defence Pty. Ltd, Southport, Queensland, Australia) before departure. PPE (face masks, gloves, goggles, and safety boots) was used during sampling. Samples were transported on ice to the laboratory, stored at 4ºC, and processed within 24-72 h after collection.

Before the sample concentration, known numbers (10 5 gene copies (GC)/50 mL) of murine hepatitis virus (MHV) were seeded into each wastewater sample as a process control (Ahmed et al., 2020c) . The same numbers of MHV were added to distilled water and subjected to RNA extraction. The reference quantification cycle (Cq) values obtained for the MHV-seeded distilled water were compared with the Cq values of the MHV-seeded wastewater to obtain information on potential RT-qPCR inhibition. If the Cq value of the RNA sample was < 2−Cq value difference compared to the reference Cq value for distilled water, the sample was considered to have PCR inhibitors (Ahmed et al., 2020b) . The presence of PCR inhibition in RNA/DNA samples extracted from wastewater was assessed using an MHV RT-PCR assay detailed elsewhere (Besselsen et al., 2002) . All samples were analyzed alongside three nontemplate controls.

Viruses were concentrated from each aggregated wastewater sample (50 mL) using an automated, rapid concentrator instrument (Concentrating Pipette Select™, CP Select™, InnovaPrep, Drexel, MO) designed for concentrating bacteria, protozoa, and viruses from water matrices simultaneously. The rationale for using 50 mL wastewater is that aircraft wastewater is more concentrated than primary influent samples of community wastewater treatment plants since in aircraft much less water is used for flushing and per person water use is very low. Hence, 50 mL wastewater sample was used for virus concentration. This instrument was used because it provides a faster concentration step than most other methods with a minor modification. In the final step, a 100 μL volume of buffer AVE was used to elute the RNA instead of 60 µL. All RNA samples were stored at −80°C for 3-5 days and subjected to RT-qPCR analysis. For US CDC N1, N2, and China CDC N assays, the assay limit of detection (ALOD) is defined as the minimum GC number with a 95% probability of detection and determined as previously described (Verbyla et al., 2016) . The sample limit of detection (SLOD) was calculated by dividing the ALOD by the RNA template volume added to the PCR well and then multiplying this number by the total volume of RNA extracted from each sample to yield the total RNA gene copies that could be detected with 95% probability. This number was then normalized to the total sample volume processed to yield the SLOD of SARS-CoV RNA GC/50 mL.

To minimize RT-qPCR contamination, RNA extraction and RT-qPCR setup were performed in separate laboratories. A method negative control was included for each batch of wastewater samples. A reagent negative control was also included during RNA extraction to account for any contamination during extraction. For each RT-qPCR run, triplicate negative controls were included. All method, reagent and RT-qPCR negative controls were negative for the targets analyzed.

Positive and negative predictive values (PPV and NPV) were estimated for wastewater surveillance of repatriation flights and subsequent COVID-19 cases during a mandatory 14day quarantine period (Parikh et al., 2008) . In this case, PPV is the probability of an incident COVID-19 case among the passengers of an aircraft following a positive wastewater result, and NPV is the probability of no incident COVID-19 cases following a negative wastewater result. We also calculated accuracy: the overall probability that wastewater samples correctly classified the presence or absence of COVID-19 clinical cases in quarantine. In addition to predictive values and accuracies, the probability of an RT-qPCR-positive wastewater sample for a given number of cumulative COVID-19 cases during the quarantine period was described by fitting a cumulative Gaussian distribution to paired wastewater RT-qPCR replicates and subsequent COVID-19 cases from each flight. This is synonymous with the methods previously described to determine 95% ALOD (Bivins et al., 2021) . All graphs and statistical analyses were prepared and performed using GraphPad Prism Version 9.0.0 (GraphPad Software, LaJolla, CA, USA).

Low-risk approval as defined by the National Statement on Ethical Conduct in Human Research was obtained from the CSIRO Ethics Committee (reference number 2020_031_LR). Flight numbers and passenger information were non-identifiable.

All RNA samples were free from PCR inhibition as determined by Cq values from the MHV RT-qPCR assay and were therefore used for downstream RT-qPCR analysis for SARS-CoV- (Table 1) . Table 2 ). The mean concentrations by US CDC N2 and China CDC were also similar-3.56  0.97 (CDC N2) to 3.22  0.81 (China CDC N) log 10 GC/50 mL wastewater.

Among the 28 post-lavatory wash samples collected, only two samples (7.14%) were RT-qPCR positive for SARS-CoV-2 RNA by the US CDC N1 assay, but they were below the quantifiable range. US CDC N2 and China CDC assays yielded non-detections in all postlavatory wash samples (Supplementary Table ST2 ). A cumulative Gaussian distribution was fit to the proportion of positive RT-qPCR replicates given an observed number of COVID-19 cases to estimate the analytical sensitivity of wastewater surveillance using RT-qPCR assay data obtained in this study (Fig. 2) . This 

Until global COVID-19 cases reach a manageable level, restrictions on international travel to certain countries may continue for an extended period of time. Such restrictions, along with reduction in demand for passenger travel, will continue to significantly impact tourism and aviation industries that rely heavily on people moving across national and international borders. International airports are important COVID-19 control points, specifically for nations such as Australia and New Zealand, but also for many other small island nations that are following an elimination strategy. Controlling transmission from overseas is essential for island nations that are committed to preventing community transmission. The majority of COVID-19 infections recorded in Australia have been acquired overseas and detected in quarantine (Price et al., 2020) . It is therefore critical to screen incoming passengers for COVID-19 at points of entry.

In our previous proof-of-concept study, we demonstrated that SARS-CoV-2 RNA could be detected in aircraft wastewater and could provide information on the presence of COVID-19 on-board passengers. However, there were some important limitations in the previous work (Ahmed et al., 2020b) , including that wastewater was sampled from a lavatory waste truck that collected wastewater from multiple aircraft; therefore, the likelihood of crosscontamination could not be ruled out. Most importantly, in the absence of clinical surveillance data (i.e., deep nasal and oropharyngeal) for on-board passengers, it was not possible to establish a link between the wastewater surveillance data and COVID-19 infected passengers. Such information is critical for determining the usefulness of wastewater surveillance as a complementary tool to clinical surveillance. In this study, we addressed these knowledge gaps.

To avoid possible carryover contamination from other flights due to mixing in the lavatory waste truck, we designed a new sample trap allowing direct wastewater sampling from the aircraft before it enters the lavatory waste truck, thereby reducing the likelihood of crosscontamination from other aircrafts. If the lavatory of a positive wastewater aircraft is not cleaned after landing, it is possible that traces of SARS-CoV-2 RNA may remain in the wastewater chambers and will yield false-positive results for the next journey. To obtain information on the likelihood of false-positive results occurring between destinations, many aircraft waste tanks were cleaned twice before departure, and the wash samples were also analyzed for the presence of SARS-CoV-2 RNA. Among the 28 wash samples tested, two aircraft were positive for the US CDC N1 gene, but these samples were not quantifiable and not all replicates were positive. These samples were also RT-qPCR negative for US CDC N2 and China CDC, suggesting that a low level of cross-contamination may have occurred. The application of multiple RT-qPCR assays, such as those used in this study, and increased number of replicates (n = 6 replicates if possible) will be needed to avoid carryover contamination from the previous flight and to interpret true positive results. Consequently, we recommend that each aircraft tank be cleaned, and wash samples be archived and analyzed later if the returning flight yields RT-qPCR positive signals. If the returning flight is RT-qPCR negative, then analyzing the wash samples will not be required. If wash samples (departing flight) and wastewater samples (returning flight) both yield RT-qPCR positive signals, then the magnitude of Cq values (i.e., 3 Cq difference) or GC numbers (i.e., one log difference) need to be examined carefully to determine a sample positive or negative.

We used three different assays in parallel to increase the analytical sensitivity and reduce the likelihood of negative results near the ALOD. This is because, when the concentration of SARS-CoV-2 RNA is low in wastewater (due to dilution and excretion dynamics by infected individuals), SARS-CoV-2 RNA will be difficult to detect consistently with one RT-qPCR assay, which amplifies a sub-sample of RNA (Ahmed et al., 2021) . Unlike domestic toilets, which use several liters of water per flush, aircraft lavatories use a substantially reduced volume of water per flush, resulting in less dilution, which could be an added advantage for detecting small quantities of SARS-CoV-2 RNA.

In this study, even with the reduced dilution associated with aircraft sanitation systems, three of 37 wastewater samples were RT-qPCR negative for all three assays, but clinical surveillance identified COVID-19-positive cases in the mandatory 14-day quarantine. This is not completely unexpected since not every infected individual will shed SARS-CoV-2 RNA in their feces or through other body fluids (Cevik et al., 2020) . Additionally, there is a nonnegligible likelihood that passengers on an international flight may not avail themselves of the lavatory. The lavatory behavior of passengers on international and long-haul flights is understudied. Three of 37 wastewater samples were also RT-qPCR positive, but clinical surveillance did not detect any COVID-19 positive cases in passengers from these flights in pre-screening (before boarding) and in the mandatory 14-day quarantine. No SARS-CoV-2 RNA was detected in post lavatory wash samples collected from these flights. The reason for such discrepancies is unclear. It has been reported that SARS-CoV-2 viral load in the upper respiratory tract appears to peak in the first week of illness (Cevik et al., 2021) , and people can shed the virus in feces for five to seven weeks (Gupta et al., 2020) . Therefore, it is possible that asymptomatic or recovered individuals shed the virus in the lavatory at a detectable level, but the viral load in the deep nasal and oropharyngeal samples was too low to be detected (Chen et al., 2020; Gupta et al., 2020) . It is also possible that children four years old and younger may have shed the virus who are exempt from the pre-departure clinical testing.

Overall, the three RT-qPCR assays collectively achieved an accuracy of 83.7% for detecting COVID-19 cases in quarantine using aircraft wastewater, which was 8 to 21.5% better than any single assay performed alone. The 87.5% PPV and 76.9% NPV (for all the three assays) were 6% greater and 12% lower, respectively, than those reported during wastewater surveillance of college dormitories for COVID-19 using the US CDC N1 and CDC N2 assays (Betancourt et al., 2021) . This demonstrates the superior performance of multiple assays in parallel rather than a single assay to maximize the predictive value of wastewater surveillance. Assay sensitivity issues appear to occur when SARS-CoV-2 RNA concentrations are at or below 10 GC/µL of RNA eluate (Vogels et al., 2020) . We recommend the application of multiple assays and RT-qPCR replicates to improve detection sensitivity.

A Gaussian model was used to determine the analytical sensitivity of the RT-qPCR data obtained in this study. Regarding the Gaussian model results, a greater than 95% probability of a positive RT-qPCR reaction for US CDC N2 and China CDC N assays would require 19.5 and 22.2 COVID-19 cases, respectively. These probabilities are premised on a single RT-qPCR reaction. However, in this study, all assays were run in triplicate. The probability of detection estimated for a single reaction can be used to assess the probability of detection by triplicate reactions by cubing the probability of non-detection in a single reaction. For example, for the US CDC N1 assay, there is a 50% (0.5) probability of a non-detection in a single reaction at 2.8 COVID-19 cases; thus, the probability of non-detection associated with triplicate reactions would be 0.5 3 (0.125 or 12.5%), and the probability of detection would be 0.875 (87.5%). Using this mathematical basis, the probability of detecting a single COVID-19 case via aircraft wastewater for US CDC N1, N2, and China CDC, and all assays performed in triplicate are 70.4%, 58.6%, 67.6%, and 73.2%, respectively. If the number of replicates for each assay were doubled to six, the probabilities of detection would be 91.2%, 82.8%, 89.4%, and 92.8%, respectively.

These probabilities are based on the current detection sensitivity of the RT-qPCR method.

The analytical sensitivity of the SARS-CoV-2 assays when applied to wastewater could be further improved by increasing the sample volume from 50 mL to 200 mL wastewater, increasing the number of RT-qPCR replicates, and/or increasing the volume of RNA template added to each RT-qPCR reaction or using digital PCR. Droplet digital PCR offered substantially greater analytical sensitivity for the detection of SARS-CoV-2 in wastewater, yielding fewer false-negative results compared to RT-qPCR (Ciesielski et al., 2021) . However, the effects of increasing the sample volume or template volume on RT-PCR inhibition should be carefully monitored.

Importantly, all repatriation flight passengers (age > 5 years) tested negative for COVID-19 via both deep nasal and oropharyngeal testing during the 48 h prior to the flight. Yet, there were 108 COVID-19 cases during the mandatory 14-day quarantine, many of which were predicted by wastewater surveillance. There is a strong argument that wastewater surveillance should be used as part of an efficient clinical surveillance and quarantine system.

Only clinical testing and quarantining are unlikely to control the overseas introduction of COVID-19. While not every passenger will defecate on a long-haul flight, and not every infected passenger will shed RNA in their feces, most passengers will urinate in the lavatory on these flights, and nasal secretions of SARS-CoV-2 RNA via sputum and cough may also enter wastewater via the lavatory sink. SARS-CoV-2 RNA has been detected in all these bodily fluid samples (Peng et al., 2020) . It has been reported that there is the possibility of a lag in viral detection in deep nasal and oropharyngeal samples; therefore, the inclusion of wastewater surveillance will enhance detection sensitivity (Lo et al., 2020) . Analysis of aircraft wastewater samples will provide valuable information regarding the emergence of imported cases, especially for countries with a high prevalence of COVID-19 and minimal quarantine requirements.

SARS-CoV-2 RNA surveillance in aircraft wastewater has demonstrated the potential to detect onboard infections and prioritize clinical testing of all passengers to maximize the efficient use of resources. Although not performed in this study, there is also the possibility of screening wastewater for variants of concern via sequencing or RT-qPCR assays However, the approach presented in this study is valuable alongside clinical testing to provide multiple lines of evidence of the COVID-19 infection status of passengers during international travel. New approaches, such as wastewater surveillance applied to transportation-based sanitation systems, provide an additional layer of data that can be integrated with clinical testing, travel, border restrictions, and quarantine to robustly manage SARS-CoV-2 transmission during the COVID-19 pandemic. Surveillance of wastewater from aircraft offers an effective, convenient, and cost-effective means of monitoring infectious agents that could be globally scaled to manage the importation of disease during pandemics. However, the application of wastewater monitoring at airports requires further method optimization, seeking to achieve rapid on-site analysis, and we are assessing the feasibility of these requirements. Table 1 RT-qPCR performance characteristics and assay limit of detection (ALOD) and sample limit of detection (SLOD)

Performance characteristic (range) Assay

Efficiency (E) (%) Linearity (R 2 ) Slope Y-intercept ALOD for SARS-CoV-2 RNA (GC/µL reaction) Positive predictive value (PPV) and negative predictive value (NPV) of wastewater surveillance of SARS-CoV-2 RNA by RT-qPCR for incident cases of COVID-19 during the 14-day mandatory quarantine period following repatriation flights for CDC N1 assay, CDC N2 assay, China CDC N assay, and all assays.

The probability of repatriation flight wastewater sample positive being positive by a single RT-qPCR for CDC N1 (A), CDC N2 (B), China CDC N (C), or All Assays pooled (D) versus the total number of incidents of COVID-19 cases during the mandatory 14-day quarantine. The data for each assay were fit with a cumulative Gaussian distribution, which was then used to estimate the number of infections required for the 95% and 50% probabilities that a single RT-qPCR reaction is positive

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Analytical sensitivity and efficiency comparisons of SARS-CoV-2 RT-qPCR primer-probe sets

Rapid Increase of SARS-CoV-2 Variant B.1.1.7 Detected in Sewage Samples from England between

We thank Qantas ground staff for collecting wastewater samples. Our thanks to Prof. Martyn Kirk from Australian National University for providing feedback on the manuscript draft.

The authors have declared no conflicts of interest.

The authors did not receive any funding for this project. 

Detection of SARS-CoV-2 RNA in wastewater samples collected from repatriation flights at Darwin International Airport (DRW) Mean  SD log 10 CDG: Charles-de-Gaulle Airport; DEL: Indira Gandhi International Airport; FRA: Frankfurt Airport; LHR: London Heathrow Airport; MAA: Chennai International Airport; YVR: Vancouver International Airport; DNQ: Did not quantify; ALOD: Assay limit of detection; *all samples were collected from DRW. Green color denotes aircraft wastewater positive and the positive incidence of COVID-19 during the 14-day mandatory quarantine period; Light blue color denotes aircraft wastewater negative and the negative incidence of COVID-19; light orange color denotes aircraft wastewater negative and the positive incidence of COVID-19; grey color denotes wastewater positive and the negative incidence of COVID-19.

Low-risk approval as defined by the National Statement on Ethical Conduct in Human Research was obtained from the CSIRO Ethics Committee (reference number 2020_031_LR). Flight numbers and passenger information were non-identifiable.

☒ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.☐The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: