A Sensitive and Rapid Wastewater Test for SARS-COV-2 and Its Use for the Early Detection of a Cluster of Cases in a Remote Community

Abstract

Throughout the coronavirus disease 2019 (COVID-19) pandemic, wastewater surveillance has been used to monitor trends in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) prevalence in the community. A major challenge in establishing wastewater surveillance programs, especially in remote areas, is the need for a well-equipped laboratory for sample analysis. Currently, no options exist for rapid, sensitive, mobile, and easy-to-use wastewater tests for SARS-CoV-2. The performance of the GeneXpert system, which offers cartridge-based, rapid molecular clinical testing for SARS-CoV-2 in a portable platform, was evaluated using wastewater as the input. The GeneXpert demonstrated a SARS-CoV-2 limit of detection in wastewater below 32 copies/mL with a sample processing time of less than an hour. Using wastewater samples collected from multiple sites across Canada during February and March 2021, a high overall agreement (97.8%) was observed between the GeneXpert assay and laboratory-developed tests regarding the presence or absence of SARS-CoV-2. Additionally, with the use of centrifugal filters, the detection threshold of the GeneXpert system was improved to <10 copies/mL in wastewater. Finally, to support on-site wastewater surveillance, GeneXpert testing was implemented in Yellowknife, a remote community in Northern Canada, where its use successfully alerted public health authorities to undetected transmission of COVID-19. The identification of SARS-CoV-2 in wastewater triggered clinical testing of recent travelers and identification of new COVID-19 cases/clusters. Taken together, these results suggest that GeneXpert is a viable option for surveillance of SARS-CoV-2 in wastewater in locations that do not have access to established testing laboratories. IMPORTANCE Wastewater-based surveillance is a powerful tool that provides an unbiased measure of COVID-19 prevalence in a community. This work describes a sensitive wastewater rapid test for SARS-CoV-2 based on a widely distributed technology, the GeneXpert. The advantages of an easy-to-use wastewater test for SARS-CoV-2 are clear: it supports surveillance in remote communities, improves access to testing, and provides faster results allowing for an immediate public health response. The application of wastewater rapid testing in a remote community facilitated the detection of a COVID-19 cluster and triggered public health action, clearly demonstrating the utility of this technology. Wastewater surveillance will become increasingly important in the postvaccination pandemic landscape as individuals with asymptomatic/mild infections continue transmitting SARS-CoV-2 but are unlikely to be tested.

Keywords: COVID-19; GeneXpert; SARS-CoV-2; remote; wastewater rapid testing; wastewater surveillance; wastewater-based epidemiology.

FIG 1

SARS-CoV-2 concentration in Canadian wastewater samples and GeneXpert rapid test results. SARS-CoV-2 concentration was determined using a laboratory-developed, solids-based extraction and RT-qPCR test targeting N1 and N2. Value shown is the average of N1 and N2 targets. For a subset of samples, SARS-CoV-2 concentration was measured in the liquid fraction with a laboratory-developed test. Locations listed multiple times refer to unique samples collected on different days. All 15 samples on the far right of the graph (negative for SARS-CoV-2) were collected from communities with no active SARS-CoV-2 cases. GeneXpert results are either positive (+), weakly positive as determined by endpoint fluorescence (w), or negative (−). EGB, Edmonton Goldbar; HAD, Halifax Dartmouth; HHA, Halifax Halifax; HMC, Halifax Millcove; MMN, Montreal North; MMS, Montreal South; TAB, Toronto Ashbridges Bay; THC, Toronto Highland Creek; THU, Toronto Humber; TNT, Toronto North Toronto; VAI, Vancouver Annacis Island; VII, Vancouver Iona Island; VLG, Vancouver Lions Gate; VLI, Vancouver Lulu Island; VNL, Vancouver Northwest Langley; YK5, Yellowknife lift station 5; YK6, Yellowknife lift station 6; HR, Hay River; FSM, Fort Smith; FSI, Fort Simpson; IN, Inuvik; INT, institutional sample; NW, Norman Wells sewer.

FIG 2

Use of Amicon centrifugal filters to increase sensitivity of the GeneXpert SARS-CoV-2/Flu/RSV assay. (A) The effect of centrifugal filtration on GeneXpert C_T values. Left white bars show C_T values without the use of centrifugal filters; right gray bars show values with the use of centrifugal filters. (B) Quantification of SARS-CoV-2 concentration in wastewater using centrifugal filters and GeneXpert standard curve (light gray) compared to results from laboratory-developed solids assay (dark gray). TAB, Toronto Ashbridges Bay; THC, Toronto Highland Creek; THU, Toronto Humber; VLG, Vancouver Lions Gate; VLI, Vancouver Lulu Island; VNL, Vancouver Northwest Langley; YK5, Yellowknife lift station 5 (multiple time points).

FIG 3

Wastewater surveillance of two Yellowknife lift stations with the GeneXpert and laboratory-developed solids assay. (A) Measured SARS-CoV-2 concentration in lift station 5 wastewater using the GeneXpert-Amicon rapid test (solid gray line) or laboratory-developed solids assay at the National Microbiology Laboratory (dotted black line). Collection frequency increased after first detection. (B) Measured SARS-CoV-2 concentration in wastewater collected at lift station 6. (C) New cases identified in Yellowknife between 26 March and 2 May. One travel-related case was identified on 6 April, and a second independent cluster was identified on 21 to 25 April.

FIG 4

Timeline of events during the GeneXpert wastewater surveillance pilot in Yellowknife, Canada. (A) Map showing locations of Yellowknife (green) and PHAC-NML laboratory in Winnipeg (blue). (B) Two main lift stations (no. 5 and no. 6) in Yellowknife and their corresponding catchments covering >85% of the Yellowknife population. (C) Timeline of events leading to the identification of a SARS-CoV-2 outbreak in Yellowknife. Timing of wastewater test results and public health actions in Yellowknife are shown in green, sample collection and transport events are shown in red, and at PHAC-NML testing is shown in blue.