Water Research

Wastewater surveillance offers an objective, comprehensive, and cost-effective means of monitoring the prevalence and genomic heterogeneity of pathogens circulating in a community. Here, a novel two-step extraction procedure for the direct capture of SARS-CoV-2 RNA from raw wastewater is presented. Combined with reverse transcription-droplet digital polymerase chain reaction (RT-ddPCR) detection, the method provides a fast and sensitive method for measuring viral RNA concentrations in wastewater...

There is increasing interest to use wastewater-based surveillance of SARS-CoV-2 as an early warning of the outbreak within a community. Despite successful detection of SARS-CoV-2 in wastewaters sampled from multiple locations, there is still no clear idea on the minimal number of cases needed in a community to result in a positive detection of the virus in wastewaters. To address this knowledge gap, we sampled wastewaters from a septic tank and biological activated sludge tank located on-site of...

The use of wastewater for SARS-CoV-2 surveillance is a useful complementary tool to clinical surveillance. The aims of this study were to characterize SARS-CoV-2 from wastewater samples, and to identify variants of concern present in samples collected from wastewater treatment plants in South African urban metros from April 2021 to January 2022. A total of 325 samples were collected from 15 wastewater treatment plants. Nucleic acids were extracted from concentrated samples, and subjected to ampl...

Wastewater surveillance is useful for monitoring the prevalence of hepatitis A virus (HAV). We developed and optimized HAV detection and quantification methods for wastewater samples, and applied them to samples collected through a national wastewater surveillance program. Previously identified 5-untranslated region-targeting primers and probes were used to develop the assay. Serial dilutions of HAV-positive clinical samples were used to validate and determine limits of quantification (LOQ). Ret...

Wastewater surveillance using RT-qPCR has now been widely adopted to track circulating levels of SARS-CoV-2 virus in many sewer sheds. The CDC qPCR assays targeting two regions (N1 and N2) within the N gene are commonly used, but a discrepancy between the two biomarkers has been noticed by many groups using this method since late 2021. The reason is presumed to be due to mutations in regions targeted by the qPCR probe. In this study, we systematically investigated and unequivocally confirmed tha...

Municipal wastewater-based surveillance (WBS) programs for SARS-CoV-2 were valuable tools for epidemiological modelling and informing COVID-19 health policy during the pandemic. We conducted a "near to source" study to assess the capacity and performance of SARS-CoV-2 WBS programs in schools relative to municipal wastewater treatment plants (WWTP). Only 4/17 schools screened had plumbing systems that were amenable to WBS. From December 2020 - March 2021 composite wastewater collected 2X/week fro...

Wastewater-based epidemiology (WBE) is an effective surveillance approach for monitoring viruses of public health relevance at the community level, complementing clinical surveillance systems. Molecular methods such as PCR/qPCR are widely used for targeted detection, while next-generation sequencing (NGS) with targeted enrichment panels has emerged as a complementary strategy for broader viral detection and genomic characterization. This study comparatively evaluated conventional PCR/qPCR and a ...

In the absence of an effective vaccine to prevent COVID-19 it is important to be able to track community infections to inform public health interventions aimed at reducing the spread and therefore reduce pressures on health-care units, improve health outcomes and reduce economic uncertainty. Wastewater surveillance has rapidly emerged as a potential tool to effectively monitor community infections for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), through measuring trends of viral...

Wastewater-based surveillance (WBS) has been established as a powerful tool that can guide health policy at multiple levels of government. However, this technology has not been well assessed at more granular scales, including large work sites such as University campuses. Between August 2021-April 2022, we explored the occurrence of SARS-CoV-2 RNA in wastewater from multiple complimentary sewer catchments and residential buildings spanning the University of Calgarys campus and how this compared t...

During the SARS-CoV-2 pandemic, genome-based wastewater surveillance sequencing has been a powerful tool for public health to monitor circulating and emerging viral variants. As a medium, wastewater is very complex because of its mixed matrix nature, which makes the deconvolution of wastewater samples more difficult. Here we introduce a gold standard dataset constructed from synthetic viral control mixtures of known composition, spiked into a wastewater RNA matrix and sequenced on the Oxford Nan...

The sequencing of human virus genomes from wastewater samples is an efficient method for tracking viral transmission and evolution at the community level. However, this requires the recovery of viral nucleic acids of high quality. We developed a reusable tangential-flow filtration system to concentrate and purify viruses from wastewater for whole-genome sequencing. A pilot study was conducted with 94 wastewater samples from four local sewersheds, from which viral nucleic acids were extracted, an...

Wastewater-based epidemiology is experiencing exponential development. Despite undeniable advantages compared to patient-centered approaches (cost, anonymity, survey of large populations without bias, detection of asymptomatic infected peoples...), major technical limitations persist. Among them is the low sensitivity of the current methods used for quantifying and sequencing viral genomes from wastewater. In situations of low viral circulation, during initial stages of viral emergences, or in c...

After the COVID-19 pandemic, wastewater monitoring is increasingly used for infectious disease surveillance. Using the data from a controlled experimental hospital setting, this paper examines the association wastewater SARS-CoV-2 with COVID-19 hospital admission and mortality, and whether this association varies by patients characteristics. Weekly wastewater samples were collected from the University of Miami (UM) hospitals where COVID-19 patients were admitted from February 2020 to October 202...

Sequencing of SARS-CoV-2 in wastewater provides a key opportunity to monitor the prevalence of variants spatiotemporally, potentially facilitating their detection simultaneously with, or even prior to, observation through clinical testing. However, there are multiple sequencing methodologies available. This study aimed to evaluate the performance of alternative protocols for detecting SARS-CoV-2 variants. We tested the detection of two synthetic RNA SARS-CoV-2 genomes in a wide range of ratios a...

In this communication, we report on the genomic surveillance of SARS-CoV-2 using wastewater samples in Jefferson County, KY. In February 2021, we analyzed seven wastewater samples for SARS-CoV-2 genomic surveillance. Variants observed in smaller catchment areas, such as neighborhood manhole locations, were not necessarily consistent when compared to associated variant results in downstream treatment plants, suggesting catchment size or population could impact the ability to detect diversity.

Laboratory-based wastewater surveillance for SARS-CoV-2, the causative agent of the ongoing COVID-19 pandemic, can be conducted using RT-qPCR-based screening of municipal wastewater samples. Although it provides rapid viral detection and can inform SARS-CoV-2 abundance in wastewater, this approach lacks the resolution required for viral genotyping and does not support tracking of viral genome evolution. The recent emergence of several variants of concern, a result of mutations across the genome ...

The spatial and temporal changes of the COVID-19 pandemic have been monitored with wastewater-based surveillance, which many countries have applied to their national public health monitoring measures. The most commonly used methods for the detection of SARS-CoV-2 in wastewater are RT-qPCR and RT-ddPCR. Previous comparisons of the two methods have produced conflicting results; some found RT-ddPCR to be more sensitive, one found RT-qPCR to be more sensitive, and others found them to be equal in se...

Evaluating drug use within populations in the United States poses significant challenges due to various social, ethical, and legal constraints, often impeding the collection of accurate and timely data. Here, we aimed to overcome these barriers by conducting a comprehensive analysis of drug consumption trends and measuring their association with socioeconomic and demographic factors. From May 2022 to April 2023, we analyzed 208 wastewater samples from eight sampling locations across six wastewat...

Wastewater-based epidemiology (WBE) may be useful for informing public health response to viral diseases like COVID-19 caused by SARS-CoV-2. We quantified SARS-CoV-2 RNA in wastewater influent and primary settled solids in two wastewater treatment plants to inform the pre-analytical and analytical approaches, and to assess whether influent or solids harbored more viral targets. The primary settled solids samples resulted in higher SARS-CoV-2 detection frequencies than the corresponding influent ...

The spread of SARS-CoV-2 has been studied at unprecedented levels worldwide. In jurisdictions where molecular analysis was performed on large scales, the emergence and competition of numerous SARS-CoV-2 lineages has been observed in near real-time. Lineage identification, traditionally performed from clinical samples, can also be determined by sampling wastewater from sewersheds serving populations of interest. Of particular interest are variants of concern (VOCs), SARS-CoV-2 lineages that are a...