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. 2023 Aug 30:11:1186525.
doi: 10.3389/fpubh.2023.1186525. eCollection 2023.

Assessment of seasonality and normalization techniques for wastewater-based surveillance in Ontario, Canada

Hadi A Dhiyebi  1 , Joud Abu Farah  1 , Heather Ikert  1 , Nivetha Srikanthan  1 , Samina Hayat  1 , Leslie M Bragg  1 , Asim Qasim  2 , Mark Payne  2 , Linda Kaleis  2 , Caitlyn Paget  2 , Dominika Celmer-Repin  3 , Arianne Folkema  3 , Stephen Drew  3 , Robert Delatolla  4 , John P Giesy  5   6 , Mark R Servos  1
Affiliations

Assessment of seasonality and normalization techniques for wastewater-based surveillance in Ontario, Canada

Hadi A Dhiyebi et al. Front Public Health. .

Abstract

Introduction: Wastewater-based surveillance is at the forefront of monitoring for community prevalence of COVID-19, however, continued uncertainty exists regarding the use of fecal indicators for normalization of the SARS-CoV-2 virus in wastewater. Using three communities in Ontario, sampled from 2021-2023, the seasonality of a viral fecal indicator (pepper mild mottle virus, PMMoV) and the utility of normalization of data to improve correlations with clinical cases was examined.

Methods: Wastewater samples from Warden, the Humber Air Management Facility (AMF), and Kitchener were analyzed for SARS-CoV-2, PMMoV, and crAssphage. The seasonality of PMMoV and flow rates were examined and compared by Season-Trend-Loess decomposition analysis. The effects of normalization using PMMoV, crAssphage, and flow rates were analyzed by comparing the correlations to clinical cases by episode date (CBED) during 2021.

Results: Seasonal analysis demonstrated that PMMoV had similar trends at Humber AMF and Kitchener with peaks in January and April 2022 and low concentrations (troughs) in the summer months. Warden had similar trends but was more sporadic between the peaks and troughs for PMMoV concentrations. Flow demonstrated similar trends but was not correlated to PMMoV concentrations at Humber AMF and was very weak at Kitchener (r = 0.12). Despite the differences among the sewersheds, unnormalized SARS-CoV-2 (raw N1-N2) concentration in wastewater (n = 99-191) was strongly correlated to the CBED in the communities (r = 0.620-0.854) during 2021. Additionally, normalization with PMMoV did not improve the correlations at Warden and significantly reduced the correlations at Humber AMF and Kitchener. Flow normalization (n = 99-191) at Humber AMF and Kitchener and crAssphage normalization (n = 29-57) correlations at all three sites were not significantly different from raw N1-N2 correlations with CBED.

Discussion: Differences in seasonal trends in viral biomarkers caused by differences in sewershed characteristics (flow, input, etc.) may play a role in determining how effective normalization may be for improving correlations (or not). This study highlights the importance of assessing the influence of viral fecal indicators on normalized SARS-CoV-2 or other viruses of concern. Fecal indicators used to normalize the target of interest may help or hinder establishing trends with clinical outcomes of interest in wastewater-based surveillance and needs to be considered carefully across seasons and sites.

Keywords: PMMoV; SARS-CoV-2; crAssphage; fecal indicators; normalization; seasonality.

PubMed Disclaimer

Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Monthly log2 PMMoV concentrations (copies/mL) from January 2021-Februrary 2023 at the Warden (A), Humber AMF (B), and Kitchener (C) wastewater sampling sites.
Figure 2
Figure 2
PMMoV Seasonal Trend Loess (STL) decomposition plots for the Warden (A), Humber AMF (B), and Kitchener (C) wastewater sampling sites. The frequency was set as 1 week and the trend length was 3 months.
Figure 3
Figure 3
Violin distribution plots for the Warden, Humber AMF, and Kitchener wastewater sampling sites. Warden collections were from April 2021 to February 2023, and Humber AMF and Kitchener collections were from January 2021 to February 2023.
Figure 4
Figure 4
Monthly wastewater flow (ML/d) from January 2021 to February 2023 at the Humber AMF (A) and Kitchener (B) wastewater sampling sites.
Figure 5
Figure 5
Flow Seasonal Trend Loess (STL) decomposition plots for the Humber AMF (A) and Kitchener (B) wastewater sampling sites. The frequency was set as 1 week and the trend length was 3 months.
Figure 6
Figure 6
Clinical cases by episode date (purple circle) and wastewater SARS-CoV-2 in raw (green triangle) and PMMoV normalized (yellow square) N1–N2 concentrations from the Warden (A), Humber AMF (B) and Kitchener (C) Wastewater sampling sites. Warden sample dates were from April to December 2021. Humber AMF and Kitchener sample dates were from January to December 2021.
Figure 7
Figure 7
Scatterplots of new cases by episode date (CBED) and the raw (N1–N2), crAssphage normalized (N1–N2/crAssphage), and pepper mild mottle virus normalized (N1–N2/PMMoV) wastewater concentrations at Warden (n = 32, A), Humber AMF (n = 29–31, B) and Kitchener (n = 57, C).
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